19th Congress of theEuropean Society
for Photobiology
30 August – 3 September 2021
World Wide Web and Salzburg, Austria

BOOK OF ABSTRACTS

ARIS
ODRON
NIVERSITYALZBURG

19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

Programme-at-a-Glance
MONDAY, AUGUST 30

THURSDAY, SEPTEMBER 2

18:00 - 18:10

Opening ceremony

08:00 - 08:30

Plenary lecture 8

18:10 - 18:20

Welcome concert part I

08:30 - 09:00

Plenary lecture 9

18:20 - 18:35

Greetings from the Governor

09:00 - 09:10

Short break

18:35 - 18:50

Greetings from the Rector

09:10 - 10:40

Symposia 5.1 - 5.4 part 1

18:50 - 19:10

Welcome concert part II

10:40 - 11:00

Coffee break

19:10 - 19:40

Plenary lecture 1

11:00 - 12:30

Symposia 5.1 - 5.4 part 2

19:40 - 20:10

Welcome concert part III

12:30 - 12:40

Short break

12:40 - 13:40

Sponsored symp.
(La Roche-Posay)

13:40 - 13:50

Short break

TUESDAY, AUGUST 31
08:00 - 08:30

13:50 - 15:20

Symposia 6.1 - 6.4 part 1

Plenary lecture 2

08:30 - 09:00

15:20 - 15:40

Coffee break

Plenary lecture 3

09:00 - 09:10

Short break

15:40 - 17:10

Symposia 6.1 - 6.4 part 2

09:10 - 10:40

17:10 - 17:20

Short breakSymposia 1.1 - 1.4 part 1

10:40 - 11:00

Coffee break

17:20 - 18:00

ESP General Assembly

11:00 - 12:30

Symposia 1.1 - 1.4 part 2

12:30 - 13:30

Lunch break

13:30 - 14:00

Plenary lecture 4

14:00 - 14:30

Plenary lecture 5

08:00 - 08:30

Plenary lecture 10

14:30 - 14:40

Short break

08:30 - 09:00

Plenary lecture 11

14:40 - 16:10

Symposia 2.1 - 2.4 part 1

09:00 - 09:10

Short break

16:10 - 16:30

Coffee break

09:10 - 10:40

Symposia 5.1 - 5.4 part 1

16:30 - 18:00

Symposia 2.1 - 2.4 part 2

10:40 - 11:00

Coffee break

11:00 - 12:30

Symposia 5.1 - 5.4 part 2

12:30 - 12:40

Short break

12:40 - 13:40

Sponsored symp. (Therakos)

WEDNESDAY, SEPTEMBER 1

FRIDAY, SEPTEMBER 3

08:00 - 08:30

13:40 - 13:50

Short break

Plenary lecture 6.1 & 6.2

08:30 - 09:00

13:50 - 15:20

Symposia 6.1 - 6.4 part 1

Plenary lecture 7

09:00 - 09:10

Short break

15:20 - 15:40

Coffee break

09:10 - 10:40

15:40 - 17:10

Symposia 6.1 - 6.4 part 2

Symposia 3.1 - 3.4 part 1

10:40 - 11:00

Coffee break

17:10 - 17:20

Closing ceremony

11:00 - 12:30

Symposia 3.1 - 3.4 part 2

12:30 - 13:30

Lunch break

13:30 - 14:40

Poster presentation

14:40 - 16:10

Symposia 4.1 - 4.4 part 1

16:10 - 16:30

Coffee break

16:30 - 18:00

Symposia 4.1 - 4.4 part 2

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

BOOK OF ABSTRACTS
19th Congress of theEuropean Society
for Photobiology
30 August – 3 September 2021
World Wide Web and Salzburg, Austria
ISBN 978-3-200-07802-4

Contents
f Tuesday, August 31 _______________________________________________________________ 4–117
f Wednesday, September 31 ______________________________________________________118–203
f Thursday, September 2 ________________________________________________________204–302
f Friday, September 3 ___________________________________________________________ 313–386
f Author Index_____________________________________________________________________ 387
Cover: Panorama of Salzburg during summer season from Kapuzinerberg © Salzburg Tourismus

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

PL-3
Theoretical and Computational Photobiology: The Chemiexcitation Phenomenon in
Biology and Medicine
Daniel Roca-Sanjuán1
1) University of Valencia, Institute of Molecular Science, València, Spain
Light and biomolecules exchange energy within the framework of electronic-states interactions giving rise to aplethora of phenomena of relevance for Life and Medicine. Visible photons induce highly-energetic isomerizationsof the double bonds of retinal in the process of vision. UV light produces [2+2] photocycloadditions of pyrimidines
forming cyclobutane pyrimidine dimers (CPD) and by this means introducing lesions in the DNA. Meanwhile,thermal reactions with especial chemical functionalities (1,2-dioxetanes, dioxetanones, 1,2-dioxins) originate highly
energetic species that decay by irradiating light in a phenomenon called chemiluminescence (or bioluminescencewhen it occurs in a living organism). Interestingly, this type of thermal transformations involving peroxide bonds canalso activate the retinal isomerization1 and CPD production in the dark (without visible or UV light)2.
Theoretical and computational photobiology allows monitoring the mentioned phenomena of the light-biomatterinteraction with a molecular and short timescale resolution, thus capturing the chemical mechanisms at the bottomof the biological and medical observations. The quantum nature of the electronic excitations (induced by either lightor chemical reactions) requires quantum chemistry methodologies, and to model the geometrical and electrostatic
constraints of the macromolecular biological systems, hybrid approaches combining quantum chemistry for thephoto/chemi-excitation region and classical mechanics for the environment are highly efficient and convenient. In
this contribution, we will show the findings obtained during the last years within such theoretical and computational
photobiology framework, focusing on the phenomena of electronic excitation in the dark, which has been less
explored in the photobiology community as compared to the processes induced by UV and visible light.

References
1) S. Premi, et al., Chemiexcitation of Melanin Derivatives Induces DNA Photoproducts Long after UV Exposure.
Science 347, 842-847, 2015.
2) M. Marazzi, et al., Induced Night Vision by Singlet-Oxygen-Mediated Activation of Rhodopsin. J. Phys. Chem.
Lett. 10, 7133-7140, 2019.
Acknowledgements. This work was supported by the Spanish ‘‘Ministerio de Ciencia e Innovación (MICINN)’’(Project Ref. CTQ2017-87054-C2-2-P and grant RYC-2015-19234), a 2019 Leonardo Grant for Researchers andCultural Creators, BBVA Foundation. The Foundation takes no responsibility for the opinions, statements, andcontents of this project, which are entirely the responsibility of its authors.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.1.1

Engineering Cells, Membranes and Light Harvesting/Photosystem II SuperComplexes in Bio-Photoelectrochemical Cells
Noam Adir
Schulich Faculty of Chemistry, Technion, Haifa Israel; noam@ch.technion.ac.il
Photosynthetic organisms, membranes and complexes are attractive starting materials for solar energy conversion(SEC). Our overall goal is to develop methods to perform SEC using these materials in simple, inexpensive and afashion that will be non-polluting and will not compete with the growth of food materials. I will describe here how
the remarkable photocatalytic activity of the photosynthetic apparatus can provide overall water splitting withoxygen and hydrogen production in Bio-Photo-Electro-Chemical (BPEC) cells via the simplest and cleanest ofprocesses wither in the absence or presence of added electron transport molecules. With plant thylakoids, electrons
are shuttled by FeCN to a transparent FTO electrode, yielding a photocurrent density of 0.5 mA·cm-2. Hydrogen
evolution occurs at the cathode at a bias as low as 0.8 V. A tandem cell comprising the BPEC cell with thethylakoid membranes and a Si photovoltaic module achieves overall water splitting with solar to hydrogen
conversion efficiency of 0.3%1. With cyanobacterial cells, electrons from the respiratory system are energized by
the photosynthetic system and transferred directly to a graphite electrode, utilizing endogenous electron carriers,
especially NADPH2,3. The current produced can be used for hydrogen production at low additional bias for
significantly longer durations than the plant thylakoids.
Another route is to use isolated photosynthetic complexes and chemically connect them to the electrochemical cell.
One of the issues with this strategy is the small number of light absorbing chromophores connected to the complex,
lowering the photochemical efficiency. Connection of isolated light harvesting (LH) complexes can help toovercome this deficiency. Photosystem II (PSII) is the only enzyme that catalyzes light-induced water oxidation
being the basis for its application as a biophotoanode in various bio-photovoltaics and photo-bioelectrochemicalcells. However, the absorption spectrum of PSII limits the quantum efficiency in the range of visible light, due to a
gap in the green absorption region of chlorophylls (500 - 600 nm). To overcome this limitation, we have used two
strategies: stabilizing the interaction between PSII and Phycobilisomes (PBSs) –LH complex or direct connection
to modified gold-nanoparticles (Au-NPs) that serve both as LH and electron conduit. Integration of the PBS-PSII
super-complexes within an Os-complex-modified hydrogel on macro-porous indium tin oxide electrodes (MP-ITO)
resulted in notably improved, wavelength dependent, incident photon-to-electron conversion efficiencies4. With the
Au-NPs, record currents were obtained, with strong improvement in the SPR range of wavelengths that are notabsorbed by chlorophylls5.
1)
2)
3)
4)
5)

Pinhassi et al. Nature Communications 2016.
Saper et al. Nature Communications 2018.
Shlosberg et al. iScience 2020.
Hartmann et al. J. Mat. Chem. A. 2020.
Shoyet et al. Submitted 2021.

Acknowledgements. Funding was provided by a “Nevet” grant from the Grand Technion Energy Program (GTEP)
and a Technion VPR Berman Grant for Energy Research, by a grant from the Deutsche Forschungsgemeinschaft(DIP project LU315/17-1) and the Israeli Ministry of National Infrastructures, Energy and Water Resources (grant
number 218-11-044). Some of the results reported in this work were obtained using central facilities at theTechnion’s Hydrogen Technologies Research Laboratory (HTRL) supported by the Nancy & Stephen Grand
Technion Energy Program (GTEP), the ADELIS Foundation and the Solar Fuels I-CORE.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.1.2
Photosynthetic entities in biohybrid systems for environmental monitoring
Matteo Grattieri1,21) Università degli Studi di Bari “Aldo Moro”, Dipartimento di Chimica, Bari 70125, Italy; 2) Consiglio
Nazionale delle Ricerche, IPCF-CNR Istituto per i Processi Chimico Fisici, Bari 70125, Italy
Interfacing photosynthetic entities with electrode surfaces enables converting solar energy into electrical energy,
obtaining biohybrid electrochemical systems. Various biological entities have been explored, spanning fromisolated photosynthetic apparatus to intact organisms. Utilizing isolated photosynthetic apparatus, such as
photosystem II and I, thylakoid membranes, and the reaction center from purple bacteria allowed a simplified
photo-excited electron transfer process, resulting in higher photocurrents1,2. Conversely, utilizing more complex
apparatus as intact chloroplasts, or intact organisms such as cyanobacteria, purple bacteria, and algae, pose
additional challenges to transfer electrons from the biocatalysts to the electrode surface due to the additional
membranes that physically separate the photosynthetic apparatus from the electrode surface. However, suchorganisms provide all the necessary enzymatic machinery and photoprotection mechanisms that could make longterm applications feasible3. Accordingly, research efforts have been focused on developing artificial approaches tofacilitate the transfer of photoexcited electrons from these organisms to the electrode surface (and vice versa)4-6.
Once the biohybrid systems is obtained, its applications include micro/low power generation, photobioelectrosynthesis of valuable chemicals and fuels, and biosensing of toxic compounds and heavy metals. The
latter has attracted particular interest in recent years, as it would enable performing sunlight-powered monitoring ofdifferent environments (i.e., fresh and salt waters, remote areas), thus providing a sustainable approach for the earlymonitoring of environmental and human hazards.
Here, photo-bioelectrochemical sensors will be presented, showing approaches for establishing the photoexcitedelectron transfer process. Furthermore, the possibility to obtain self-powered biosensors will be discussed, enabling
the on-line monitoring of widely utilized compounds that are hazardous for the environment and toxic for humans7.
References
1) K.P. Sokol, D. Mersch, V. Hartmann, J.Z. Zhang, M.M. Nowaczyk, M. Rögner, A. Ruff, W. Schuhmann, N.
Plumeré, E. Reisner, Rational wiring of photosystem II to hierarchical indium tin oxide electrodes using redox
polymers. Energy and Environmental Science, 2016, 9, 3698-3709.
2) G. Buscemi, D. Vona, R. Ragni, R. Comparelli, M. Trotta, F. Milano, G.M. Farinola,
Polydopamine/Ethylenediamine Nanoparticles Embedding a Photosynthetic Bacterial Reaction Center for Efficient
Photocurrent Generation. Advanced Sustainable Systems, 2021, 2000303.
3) M. Grattieri, K. Beaver, E.M. Gaffney, F. Dong, S.D. Minteer, Advancing the fundamental understanding andpractical applications of photo-bioelectrocatalysis. Chemical Communications, 2020, 56, 8553-8568.
4) K. Hasan, K.V.R. Reddy, V. Eßmann, Ka. Górecki, P.Ó. Conghaile, W. Schuhmann, D. Leech, C. Hägerhäll, L.
Gorton, Electrochemical Communication Between Electrodes and Rhodobacter capsulatus Grown in Different
Metabolic Modes. Electroanalysis, 2015, 27, 118-127.
5) M. Grattieri, Z. Rhodes, D.P. Hickey, K. Beaver, S.D. Minteer, Understanding Biophotocurrent Generation in
Photosynthetic Purple Bacteria. ACS Catalysis, 2019, 9, 867-873.
6) M. Grattieri, S. Patterson, J. Copeland, K. Klunder, S.D. Minteer, Purple Bacteria and 3D Redox Hydrogels for
Bioinspired Photo-bioelectrocatalysis, ChemSusChem, 2020, 13, 230-237.
7) M. Grattieri, H. Chen, S. D. Minteer, Chloroplast biosolar cell and self-powered herbicide monitoring. Chemical
Communications, 2020, 56, 13161-13164.
Acknowledgements. This work is funded from Fondazione CON IL SUD, Grant “Brains to South 2018”, project
number 2018-PDR-00914.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.1.3
Engineering photosynthetic microorganisms for direct solar chemical and fuelproduction from carbon dioxide, example butanol
Peter Lindblad1
1) Microbial Chemistry, Department of Chemistry-Ångström, Uppsala University, SE-751 20 Uppsala, Sweden
Cyanobacteria, photosynthetic microorganisms with the same type of photosynthesis as plants and algae, can beengineered to produce solar chemicals and solar fuels in direct processes from carbon dioxide1. This presentation will
outline our strategies to engineer cyanobacteria to produce the alcohol butanol. Butanol is a four-carbon alcohol(C4H9OH) occurring in four structural isoforms: 1-butanol, 2-butanol, isobutanol and tert-butanol. It is an importantbulk chemical, as solvent or intermediate in chemical synthesis, and excellent blend-in fuel. The absolute majorityof butanol is presently a produced from fossil resources. Additionally, there are biological routes for fermentative
butanol production, mainly to produce 1-butanol and isobutanol. Fermentative 1-butanol is produced from starch,sugar, or cellulose such as wheat, beet, corn and wood. Products of these fermentation processes also include acetone
and ethanol. Cyanobacteria do not produce butanol naturally, they lack the butanol biosynthetic pathways andcorresponding relevant genes.
Introduction of a single gene encoding KivD resulted in isobutanol producing strains of Synechocystis PCC 68032.
Knowledge based modelling of the identified bottleneck KivD resulted in strains with significantly increased
isobutanol production3. Using our best isobutanol strain in long-term experiments a cumulative titer of 911 mg per L
was observed with a maximal rate of 43.6 mg per L and day4. A similar approach to systematically engineer
cyanobacteria to produce 1-butanol resulted in cells with a cumulative titer of 4.8 g per L and a maximal rate of 302
mg per L and day5, recently doubled to 600 mg 1-butanol per L and day and a carbon partitioning of 60%6.
Photosynthetic butanol is the most sustainable and CO2 neutral butanol synthetized from CO2 and solar energy.
References
1) Miao, R., Wegelius, A., Durall, C., Liang, F., Khanna, N., Lindblad, P., Engineering cyanobacteria for biofuel
production. Modern Topics in Phototroph Prokaryotes 11: 351-393, 2017.
2) Miao, R., Liu, X., Englund, E., Lindberg, P., Lindblad, P., Isobutanol production using Synechocystis PCC 6803
endogenous alcohol dehydrogenases. Metabolic Engineering Communications 5: 45-53, 2017.
3) Miao, R., Xie, H., Ho, F.M., Lindblad, P., Protein engineering of a-ketoisovalerate decarboxylase for improved
isobutanol production in Synechocystis PCC 6803. Metabolic Engineering 47: 42-48, 2018.
4) Miao, R., Xie, H., Lindblad, P,. Enhancement of photosynthetic isobutanol production in Synechocystis PCC
6803.Biotechnolgy for Biofuels 11: 267, 2018.
5) Liu, X., Miao, R., Lindberg, P., Lindblad, P., Modular Engineering for Photosynthetic 1-Butanol Production in
Cyanobacteria. Energy & Environmental Science 12: 2765-2777, 2019.
6) Wichmann, J., Lauersen, K.J., Biondi, N., Christensen, M., Guerra, T., Hellgardt, K., Kuronen, M., Lindberg, P.,
Rösch, C., Yunus, I.S., Jones, P., Lindblad, P., Kruse, O., Engineering Biocatalytic Solar Fuel Production: The
PHOTOFUEL Consortium. Trends in Biotechnology 39: 323-327, 2021.
Acknowledgements. This work was supported by the European Union Horizon 2020 Framework Program under the
grant agreement number 640720 (Photofuel), the Kamprad Family Foundation for Entrepreneurship, Research &Charity (Photosynthetic butanol), the NordForsk NCoE program “NordAqua” (project number 82845), and theSwedish Energy Agency (CyanoFuels, project number P46607-1).

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-1.1.4

Predicting the structural, electronic and spectroscopic properties of chromophore –protein assemblies towards efficient charge-separation
Mariano Curti1, Carles Curutchet2, Elisabet Romero1
1) Institute of Chemical Research of Catalonia (ICIQ), Barcelona Institute of Science and Technology (BIST),
Avinguda dels Països Catalans 16, 43007 Tarragona, Spain; 2) Department of Pharmacy and Pharmaceutical
Technology and Physical-Chemistry and Institute of Theoretical and Computational Chemistry (IQTCUB), Schoolof Pharmacy and Food Sciences, University of Barcelona, Joan XXIII, 27–31, 08028 Barcelona, Spain
In photosynthetic reaction centres, charge separation proceeds with near unitary quantum yield.[1] Plants, algae andphotosynthetic bacteria achieve this impressive efficiency by exploiting the “design principles of photosynthetic
charge separation”, that can be summarized as follows:[1](i) The excited states possess a large collective (excitonic) character, increasing light absorption and reducing thenumber of energy transfer steps. Moreover, there is a coherent mixing between excitonic and charge-transfer statesthat promotes ultrafast electron transfer.
(ii) Intramolecular vibrations, with energies close to the gap between excitonic and charge-transfer states (“resonant
vibrations”), ensure a strong mixing of these states.
(iii) Multiple charge separation routes offer flexibility and avoid energy losses caused by disorder.
(iv) The protein scaffold acts as a smart matrix, selecting the appropriate charge separation route and providing
decoherence channels for the irreversible formation of charge-separated states.
Efficient as it is, the photosynthetic machinery is also extremely complex, and thus not amenable to modifications
aiming to tune the process, for instance to perform other photochemical reactions. As an alternative, the de novo
design of proteins with tailored functions has seen substantial progress in recent years. In particular, a family of
structures termed “maquette proteins” provide much simpler scaffolds, and once loaded with cofactors are still able
to perform light absorption and energy transfer roles (although with low charge separation efficiency, so far).[2, 3]
Of particular interest are maquette proteins that consist of water-soluble four alpha helix bundles. By means ofappropriately placed histidine residues, they can readily bind metalloporphyrins in their interior, providing excellent
model systems in which to implement the design principles of charge separation.

Fig. 1 – Left: Chemical structure of Zn methyl-132-OH-pheophorbide a (mZnPPAOH). Right: Snapshot of the α-HP protein
structure with four bound chromophores, obtained from molecular dynamics simulations.

In this contribution we present a computational study on a maquette protein class termed “HP”,[3] and theircomplexes with the chromophore Zn methyl-132-OH-pheophorbide a (mZnPPAOH; Fig. 1, left). The base protein
design (“α-HP”) has four binding sites distributed by pairs: one in the upper half of the protein, and the other in thebottom half (Fig. 1, right). We analyze this structure both in its apo and in different holo states, considering as wellBook of Abstracts ������������������������������������������������������������������������������������������������� 8

19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

different binding modes (e.g. through histidine’s δN or εN), folding conformations, and mutated structures withadditional charged amino acids. By performing simulations that combine long (>1 µs) molecular dynamics with
QM/MM calculations, we evaluate the binding energy, site energies, electronic couplings, and spectral density,
together with the absorption, circular- and linear-dichroism spectra of maquette-bound mZnPPAOH. Furthermore,
we performed a preliminary analysis of charge transfer states. Altogether, this information can be used to model thetime-resolved dynamics upon light absorption.
Some of our main findings are:1- The binding energy depends on the relative position of the chromophores. Taking as reference the binding energyof a single chromophore in the top half of the protein, binding a second one in the bottom proceeds with a similarenergy. However, the binding of a second chromophore in the same (top) half of the protein results in a significant
energy penalty due to steric hindrance.
2- The electronic coupling between chromophores in opposite halves of the protein is relatively small (~ 4 cm-1); in
comparison, for chromophores in the same half it is ~ 40 cm-1. Consequently, the latter show strong excitonic signals
in circular dichroism spectra, while the former display only weak signals.
3- The site energies are virtually unaffected by the successive binding of chromophores, consistent with the symmetryof the protein sequence. However, by mutating the protein and adding positively charged amino acids in regions
nearby chromophores, we observe a slight red shift in their absorption spectra.
4- Charge-transfer states are significantly (~1 eV) higher in energy with respect to the lowest lying (excitonic) excitedstate. It is possible to red shift the charge-transfer states by including positively charged amino acids nearby.
The results obtained so far are generally in good agreement with ongoing experimental work. Furthermore, they
represent a solid step towards the implementation of the Design principles of charge separation in an artificial system.
References
1) E. Romero, V. I. Novoderezhkin and R. van Grondelle, Quantum design of photosynthesis for bio-inspired solarenergy conversion. Nature, 543, 355–365, 2017.
2) T.A Farid, G. Kodali, L.A. Solomon, B.R. Lichtenstein, M.M. Sheehan, B.A. Fry, C. Bialas, N.M. Ennist, J.A.
Siedlecki, Z. Zhao, M.A. Stetz, K.G. Valentine, J.L.R. Anderson, A.J. Wand, B.M. Discher, C.C. Moser and P.L.
Dutton, Elementary Tetrahelical Protein Design for Diverse Oxidoreductase Functions. Nature Chemical Biology, 9,
826–833, 2013.
3) I. Cohen-Ofri, M. van Gastel, J. Grzyb, A. Brandis, I. Pinkas, W. Lubitz, and D. Noy, Zinc-Bacteriochlorophyllide
Dimers in de Novo Designed Four-Helix Bundle Proteins. A Model System for Natural Light Energy Harvesting andDissipation. Journal of the American Chemical Society, 133, 9526–9535, 2011.
Acknowledgements. MC and ER acknowledge the funding from the European Union’s Horizon 2020 research andinnovation programme under the Marie Skłodowska-Curie grant agreement No 801474 and from the State Research
Agency/Spanish Ministry of Science and Innovation (AEI/MICINN) through the Severo Ochoa Excellence
Accreditation CEX2019-000925-S. ER is supported by a Starting Grant from the European Research Council (No.
805524, BioInspired_SolarH2). CC acknowledges support from the AEI/MICINN (Grants CTQ2017-89924-P and
MDM-2017-0767). The authors thankfully acknowledge the computer resources at Pirineus and CTE-POWER andthe technical support provided by the Barcelona Supercomputing Center and the CSUC (QSB-2020-3-0002 and QSB2021-1-0019).

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.1.5
The Metabolism of H2 in green algae – how can it be used to redesign photosynthesis?
Prof. Iftach Yacoby, PhD
Tel Aviv University, School of Plant Sciences and Food security Faculty of Life Sciences, Tel Aviv, Israel
The natural ability of photosynthesis to function as a factory of chemicals and fuels is the basis for life on earth;sunlight energy has been used for 1.5 billion years to power electron transfer towards the assimilation of inorganiccarbon dioxide into organic matter. Microalgae (a group of photosynthetic unicellular organisms) can also use thisenergy to power other potentially useful but inefficient processes, such as H2 production via the enzyme Hydrogenase.
Improving this process could open a new era in the agricultural history, this time dedicated for clean energy
production. Yet, attempting to upscale it proved challenging. Collective evidence suggest that the inefficiency ofalternative processes other than carbon assimilation stems from a non-optimal location, i.e., lack of docking
capabilities to photosynthetic reaction centers, which limits access to the energy supply. To date, the scientificefforts mainly focused on improving natural photosynthesis, rather than redesigning it to secure efficientelectron transfer to enzymes of interest. Photosynthetic improvement has been targeted by studying libraries ofmutants and screening for improved features; for example, faster growth or faster H2 production. The knowledge
obtained from studying mutants along with structural insights into the photosynthetic reaction centers, constitute theplatform for redesigning photosynthesis. Unlike photosynthesis improvement, photosynthesis redesigning attempts
are usually based on bottom-up strategies; for example, targeting enzymes towards hotspots, where they could benefit
from plenty of energy, low oxygen or other supporting features. To divert the photosynthetic electrons towards
reactions other than carbon fixation, the enzymatic machinery of choice needs to be efficiently hooked up to thephotosynthetic machinery. Therefore, the key for optimally driving other processes is to find strategic hotspots fromwhich the electrons can be “hijacked”; for example, by docking Hydrogenase directly to photosynthetic reaction
centers. This approach was successfully applied by tethering Hydrogenase to PSI via fusions to Ferredoxin (Fd) 1,2,
Superoxide dismutase (SOD)3 or directly to Photosystem I (PSI)4; these resulted in significant improvement to H2
production, in vivo. In this lecture, I will describe the basic findings and how they were used for engineering.
Fig. 1Illustration of Hydrogenase
Fused to Photosystem I
From Kanygin et al 2020

References
1.
Yacoby, I., et al., Photosynthetic electron partitioning between [FeFe]-hydrogenase andferredoxin:NADP+-oxidoreductase (FNR) enzymes in vitro. Proc Natl Acad Sci U S A, 2011. 108(23): p.
9396-401.
2.
Eilenberg, H., et al., The dual effect of a ferredoxin-hydrogenase fusion protein in vivo: successful divergenceof the photosynthetic electron flux towards hydrogen production and elevated oxygen tolerance. Biotechnol
Biofuels, 2016. 9(1): p. 182.
3.
Ben Zvi, O. and I. Yacoby, The in-vitro enhancement of FeFe hydrogenase activity by superoxide dismutase.
International Journal of Hydrogen Energy, 2016. 41(39): p. 17274-17282.
4.
Kanygin, A., et al., Rewiring photosynthesis: a photosystem I-hydrogenase chimera that makes H-2 in vivo.
Energy & Environmental Science, 2020. 13(9): p. 2903-2914.
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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.1.6
Bio-Inspired Chromophore-Protein Assemblies for the Generation of Solar Fuels
Valentin Maffeis1, Saeed Shareef1, Mariano Curti1, Sharmistha Chatterjee1, Dominik Bäuerle1, Elisabet Romero1
1) Institute of Chemical Research of Catalonia (ICIQ), Barcelona Institute of Science and Technology (BIST),
Avinguda dels Països Catalans 16, 43007 Tarragona, Spain
Photosynthesis is the fundamental biological process by which solar energy is converted into fuel in four basic steps:
light harvesting, charge separation, water splitting and fuel generation. At the heart of Photosynthesis, the reaction
center pigment-protein complexes perform charge separation with near unity quantum efficiency despite their highly
disordered energy landscape; they realise the first solar-energy conversion step in Photosynthesis by transforming
sunlight to electrochemical energy.
To achieve this amazing feat, the reaction centers exploit The Quantum Design Principles of Photosynthesis1,
complementary and interrelated solutions to ensure ultrafast and irreversible transfer of energy and electrons withina fluctuating environment. These Principles provide a guide for the rational design and construction of systems able
to transfer energy and electrons with high efficiency and in the right direction.
During my talk I will briefly present these Principles and focus on how we are putting all this knowledge into action
to Design and Construct Bio-Inspired Chromophore-Protein Assemblies2 composed by abundant and biodegradable
materials in order to maximize their capacity to absorb, transfer and convert solar to electrochemical energy.
Ultimately, these assemblies will be coupled to molecular catalysts to achieve sustainable and cost-effective solarenergy conversion to fuel.

-+Light Harvesting

Units

Charge Separation

Bio-Inspired Chromophore-Protein Assemblies
Fig. 1 – Schematic representation of the Bio-Inspired Chromophore-Protein Assemblies under investigation

References
1) E. Romero, V. I. Novoderezhkin and R. van Grondelle, Quantum design of photosynthesis for bio-inspired solarenergy conversion. Nature, 543, 355–365, 2017.
2) T.A Farid, G. Kodali, L.A. Solomon, B.R. Lichtenstein, M.M. Sheehan, B.A. Fry, C. Bialas, N.M. Ennist, J.A.
Siedlecki, Z. Zhao, M.A. Stetz, K.G. Valentine, J.L.R. Anderson, A.J. Wand, B.M. Discher, C.C. Moser and P.L.
Dutton, Elementary Tetrahelical Protein Design for Diverse Oxidoreductase Functions. Nature Chemical Biology, 9,
826–833, 2013.
Acknowledgements. VM thanks the European Union’s Horizon2020 Research and Innovation Program under theMarie Skłodowska-Curie grant agreement No. 754510. SS, SC and ER are supported by ER´s Starting Grant fromthe European Research Council (No. 805524, BioInspired_SolarH2). MC acknowledge the support of the European
Commission under the Marie Sklodowska Curie grant agreement No. 801474. MC, DB and ER thank the Ministeriode Ciencia e Innovación-Agencia Estatal de Investigación for financial support through Severo Ochoa Excellence
Accreditation 2020-2023 (CEX2019-000925-S) and FPI predoctoral fellowship to DB (PRE2020-095271).

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.1.7

Do photosynthetic bacteria dream of heavy metals?
Massimo Trotta1, Livia Giotta2, Luigi R. Ceci3, Maria Varsalona1,4, Rossella Labarile1,4, Francesco Milano5,
Maria Teresa Volpicella4, Matteo Grattieri1,4
Consiglio Nazionale delle Ricerche – Istituto per i Processi Chimico Fisici; 2 Università del Salento Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali; 3 Consiglio Nazionale delle Ricerche – Istituto di
Biomembrane, Bioenergetica e Biotecnologie Molecolari; 4 Università di Bari – Dipartimento di Chimica; 5Consiglio Nazionale delle Ricerche – Istituto di Scienze delle Produzioni Alimentari.
1

Purple non sulphur (PNS) photosynthetic bacteria have a high affinity toward metal ions, being prone tobioaccumulate them and – in some case – lower their toxicity by catalysing the change of their redox states.
The most recent advancement on the topic of bioremediation of sites polluted withheavy metals by the use of PNS1-9. Promising applications in soil washing will also be presented.

Fig. 1 – A pop-art view of PNS bacteria

References
1. Calvano, C. D.; et al, The lipidome of the photosynthetic bacterium Rhodobacter sphaeroides R26 is affectedby cobalt and chromate ions stress. Biometals 2015, 28 (1), 229-229.
2. Volpicella, M.; et al, Rhodobacter sphaeroides adaptation to high concentrations of cobalt ions requires
energetic metabolism changes. FEMS Microbiol. Ecol. 2014, 88 (2), 345-357.
3. Belviso, B. D.; et al, Cobalt binding in the photosynthetic bacterium R. sphaeroides by X-ray absorption
spectroscopy. Biometals 2013, 26 (5), 693-703.
4. Italiano, F.; et al., Changes in morphology, cell wall composition and soluble proteome in Rhodobacter
sphaeroides cells exposed to chromate. Biometals 2012, 25(5), 939-949.
5. Emese, A.; et al, The reaction center is the sensitive target of the mercury(II) ion in intact cells ofphotosynthetic bacteria. Photosynthesis Research 2012, 112 (2), 129-140.
6. Italiano, F.; et al, The photosynthetic membrane proteome of Rhodobacter sphaeroides R-26.1 exposed tocobalt. Res. Microbiol. 2011, 162 (5), 520-527.
7. Giotta, L.; et al, Reversible binding of metal ions onto bacterial layers revealed by protonation-induced ATRFTIR difference spectroscopy. Langmuir 2011, 27 (7), 3762-3773.
8. Asztalos, E.; et al Early detection of mercury contamination by fluorescence induction of photosynthetic
bacteria. Photochem Photobiol Sci 2010, 9 (9), 1218-23
9. Pisani, F.; et al., Soluble proteome investigation of cobalt effect on the carotenoidless mutant of Rhodobacter
sphaeroides. J. Appl.Microbiol. 2009, 106 (1), 338-349.
Acknowledgements. This work was partly funded by PON Energy for TARANTO project n. ARS01_00637.
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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-1.1.8
Coating photosynthetic bacteria with the versatile polydopamine
Rossella Labarile1,2, Danilo Vona1, Gabriella Buscemi1,2, Maria Varsalona1,2, Matteo Grattieri1,2, Gianluca Maria
Farinola1, Massimo Trotta2
1) Università degli Studi di Bari, Dipartimento di Chimica, Bari, Italy 2) CNR-IPCF Institute for PhysicalChemical Processes, Bari, Italy
The wild type and the carotenoidless mutant strains of Rhodobacter (R.) sphaeroides have been largely used asmodels for studying both the molecular mechanisms of photosynthesis and the metabolism of anoxygenic
photosynthesis. Beside of an efficient light harvesting system, the photochemical enzyme called reaction center(RC) plays a pivotal role, generating stable a charge-separated state across the membrane within a few hundred
microseconds from photon absorption with a conversion yield close to unit. These electron flows could beintercepted and funneled outside the cells and transferred to suitable material prone to collect such electrons andimpinge them in an electric/electronic circuit of a biohybrid device.
Polydopamine (PDA), a bioinspired synthetic polymer produced by the self-polymerization of dopamine, wasselected for R. sphaeroides coating because of its versatile adhesivity and low toxicity1. PDA possesses numerous
quinone groups that are redox-active functional ligands and further suitable for accelerating the transfer of electrons
during extracellular respiration.
The coating is assembled by introducing dopamine monomers into the biological feeding medium of bacteria2. The
PDA artificial conductive coating assembles around the cells, acting as soft functional matrix allowing thephotosynthetic organisms to thrive using light and ensuring the satisfactory electronic communication3 with theconductive surfaces, namely glassy carbon electrodes.
Electrochemical characterization was performed to investigate the electronic behavior of these biohybrids,
unveiling that the polymer layer on the bacterial cells does not hinder the diffusion of the mediator and its
capability to react at the electrode surface. The effects of dopamine concentration on the light-induced
photoresponse of the biohybrid system will be discussed and compared to bare bacteria.
References
1) Yang, S.H., et al., Mussel-inspired encapsulation and functionalization of individual yeast cells. J Am Chem
Soc, 2011. 133(9): p. 2795-7.
2) Vona, D., et al., Synthesis of (poly)gallic acid in a bacterial growth medium. MRS Advances, 2020. 5 (18-19): p.
957-963.
3) Buscemi, G., et al., Polydopamine/Ethylenediamine Nanoparticles Embedding a Photosynthetic Bacterial
Reaction Center for Efficient Photocurrent Generation. Advanced Sustainable Systems, 2021. 2000303
Acknowledgements. This work was supported by the European Union’s Horizon 2020 Research and Innovation
Programme under Grant Agreement No 800926 (HyPhOE, Hybrid Electronics Based on Photosynthetic
Organisms).

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.1.9

Biological approach in the synthesis of dopamine-based polymers: purple
photosynthetic bacteria as catalysers
Maria Varsalona1,2, Rossella Labarile1,2, Danilo Vona1, Gabriella Buscemi1,2, Matteo Grattieri1,2, Gianluca Maria
Farinola1, Massimo Trotta2
1

Università degli Studi di Bari, Dipartimento di Chimica, via Orabona 4, 70125, Bari, Italy; 2 CNR-IPCF Institutefor Physical-Chemical Processes, via Orabona 4, 70125, Bari, Italy

Rhodobacter (R.) sphaeroides is a non-sulfur bacterium able to growth either under aerobic andphoeterotropically conditions. The carotenoid-less mutant strain R26 grows phoeterotropically under strict
anoxygenic conditions.
Dopamine (DA) is a neurotransmitter that self-polymerizes into polydopamine (PDA), a synthetic analogueof melanin1 of great interest in organic and bioinorganic electronics.
The self-polymerization of dopamine is investigated in presence and in absence of growing R. sphaeroides
R26 under aerobic and phoeterotropically anaerobic environment.
The effects of the photosynthetic bacterium on the polymerization of dopamine solutions is investigatedunder a variety of conditions2, 3 and exposed to light under anaerobic conditions as previously reported.4, 5
The system was characterised via absorption spectroscopy showing that even in absence of the oxidising
species normally used to initiate polymerization, R. sphaeroides acts as a catalyst for the PDA formation.
References
1. d’Ischia M., et al., Polydopamine and eumelanin: From structure-property relationships to a unifield tailoring
strategy. Accounts of Chemical Research, 2014. 47(12): p. 3541-50.
2. Yang, S.H., et al., Mussel-inspired encapsulation and functionalization of individual yeast cells. J Am Chem
Soc, 2011. 133(9): p. 2795-7.
3. Giotta, L., et al., Heavy metal ion influence on the photosynthetic growth of Rhodobacter sphaeroides.
Chemosphere, 2006. 62(9): p. 1490-9.
4. Vona, D., et al., Synthesis of (poly)gallic acid in a bacterial growth medium. MRS Advances, 2020. 5(18-19): p.
957-963.
5. Buscemi, G., et al., Polydopamine/Ethylenediamine Nanoparticles Embedding a Photosynthetic Bacterial
Reaction Center for Efficient Photocurrent Generation. Advanced Sustainable Systems, 2021.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.1.10

Adhesive Polydopamine for Biohybrid Photoanodes with Photosynthetic Purple
Bacteria
Gabriella Buscemi1,2, Rossella Labarile1,2, Danilo Vona1, Pinalysa Cosma1,2, Angela Agostiano1,2, Massimo Trotta2,
Gianluca M. Farinola1, Matteo Grattieri1,21)Università degli Studi di Bari “Aldo Moro”, Dipartimento di Chimica, Bari, 70125, Italy;2) Consiglio Nazionale delle Ricerche, IPCF-CNR Istituto per i Processi Chimico Fisici, Bari, 70125, Italy.
Anoxygenic phototrophic purple bacteria are well-known microorganisms having a versatile metabolism, as they use
sunlight to oxidize various organic compounds. With the intent to mimic their natural processes, many attempts weremade to exploit the ability of energy storage and energy conversion into electrical (and/or chemical) energy inbiohybrid electrochemical systems. Such approaches allow developing an eco-friendly and scalable technology basedon self-repairing biocatalysts where sunlight is utilized as primary green energy source1.
In the application of living organism in bioelectronic devices, more than the “interface” between biotic and electronic
domains, the challenging aspect is related to the electrochemical communication between these two components.
Various approaches have been reported to overcome this problem, such as layer by layers techniques2 to entrap wholecell onto electrodes surfaces or the use of conductive material to burst electronic communication3.
In this context, we combined a biocompatible soft polymer, polydopamine (PDA), with the purple bacteria
Rhodobacter capsulatus as biocatalyst. Polydopamine, containing both catechol and amine groups, is a polymer
presenting similar characteristic to the adhesive plaque of mussel byssus, which has been recently utilized toencapsulate isolated photosynthetic apparatus4. PDA can be obtained by oxygenic self-assembly polymerization of
dopamine in water or by electro-polymerization. Moreover, the tunable conductive properties and the key feature ofbeing adhesive onto a variety of surfaces5 make this material suitable for bioelectrochemical devices. Here, we reporta biohybrid photoanode obtained by a one-step immobilization of the living whole cells into PDA matrix, while
entrapping a quinone-based redox mediator. The presented approach enabled a more stable adhesion of the bioactive
components onto the electrode surface over a prolonged period of time, while improving and stabilizing theelectrochemical response. Electrochemical and spectroscopic evidence for the obtained biohybrid photoanodes will
be discussed together with future research possibilities.
References
1) McCormick A.J., et al. Biophotovoltaics: oxygenic photosynthetic organisms in the world of bioelectrochemicalsystems. Energy Environ. Sci., (8), 1092-1109, 2015.
2) Grattieri M., et al. Non-Competitive Reversible Inhibition of Laccase by H2O2 in Osmium Mediated Layer-ByLayer Multilayer O2 Biocathodes. J. Electrochem. Soc. 162 (9) G82-G86.
3) Reggente, M, et al. Design of Optimized PEDOT‐Based Electrodes for Enhancing Performance of Living
Photovoltaics Based on Phototropic Bacteria. Adv. Mater. Technol. (5), 1900931, 2020.
4) Buscemi, G., et al. Polydopamine/Ethylenediamine Nanoparticles Embedding a Photosynthetic Bacterial Reaction
Center for Efficient Photocurrent Generation. Adv. Sustainable Syst., 2000303, 2021.
5) M. d'Ischia et al. Melanin Biopolymers: Tailoring Chemical Complexity for Materials Design, Angew. Chem. Int.
Ed.(59), 11196, 2020.
Acknowledgements. The authors would like to acknowledge the funding from Fondazione CON IL SUD, Grant“Brains to South 2018”, project number 2018-PDR-00914 and the European Union’s Horizon 2020 Research andInnovation Programme under grant agreement No 800926 (HyPhOE, Hybrid Electronics Based on Photosynthetic
Organisms).

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.1.11

Synthesis of meso-functionalized porphyrins as hole transport materials for perovskite
solar cells
Melani J.A. Reis1, Nuno M.M. Moura1, Ana Teresa Nogueira2, Ana Eulálio2, Ana M.V.M. Pereira2, Adélio Mendes2, M. Graça
P.M.S. Neves1

1)Department of Chemistry & LAQV-REQUIMTE, University of Aveiro, 3810-193 Aveiro, Portugal;
2)LEPABE - Laboratory for Process Engineering, Environment, Biotechnology and Energy, Faculty of Engineering, Universityof Porto, Rua Dr. Roberto Frias, 4200-465 Porto, Portugal.

The potential issues surrounding the use of fossil fuels have been leading to a crescent search for alternative energy
sources and it is nowadays a worldwide concern.1
Perovskite solar cells (PSCs, figure 1) are promising photovoltaic technologies and an alternative to traditional
silicon-based solar cells. The use of solid-state hole-transporting materials (HTMs) was easily foreseen, and SpiroOMeTAD was selected as the benchmark .2 However, the synthesis of Spiro-OMeTAD is associated with high costs,
since it involves a laborious and complex multistep synthetic route and a hard purification process.2
Porphyrins, due to their particular physicochemical features, namely high electronic delocalization, thermal stability,
susceptibility to electronic transfer and the ability to absorb light in a wide range of the visible spectrum, areparticularly attractive compounds to be used as HTMs for PSCs.3,4
In this communication, the synthetic pathways to obtain new porphyrin-based derivatives containing nitrogen donormoieties will be presented, as well as their efficiency in PSC devices.

Fig. 1 - Structure of a PSC device with the layout of the cargo transport process.

References
1) M. Ye, et al, Mater. Today, 2015, 18, 155.
2) M. Urbani et al. Chem. Soc. Rev, 2019, 48, 2738.
3) X. Lv, et al, Dyes Pigments, 2019, 160, 957-961.
4) Z. Shariatinia, Renew. Sust. Energ. Rev., 2020, 119, 109608.
Acknowledgements. The authors thank the University of Aveiro and the University of Porto and FCT/MCTES (PIDDAC) for

their financial support to the LAQV-REQUIMTE (UIDB/50006/2020) and LEPABE (UIDB/00511/2020) Research Units andPortuguese NMR Network. This work was financially supported by Project FCT POCI-01-0145-FEDER-030357 - “Taylor-madeporphyrinoids for emerging high-efficiency solar energy devices”, funded by FEDER funds through COMPETE2020 – Programa
Operacional Competitividade e Internacionalização (POCI) and by national funds (PIDDAC) through FCT/MCTES. M.J.A.
Reis, A.T. Nogueira, A. Eulálio, N.M.M. Moura (REF.-048-88-ARH/2018) and A.M.V.M. Pereira thanked the FCT and project
POCI-01-0145-FEDER-030357 for the research grants and concession agreements.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.1.12

In vivo Functionalization of Diatoms with a Transparent Polymer Matrix for
Biohybrid Photonic Systems
Cesar Vicente-Garcia1, Danilo Vona1, Roberta Ragni1, Stefania R. Cicco2, Gianluca Maria Farinola1
1) Università degli Studi di Bari, Department of Chemistry, Bari, Italy; 2) CNR-ICCOM, Department of Chemistry,
Bari, Italy
Diatom microalgae are known for generating highly nanostructured shells (frustules) by in vivo mineralization, madeof mesoporous silica with extraordinarily intricate photonic structures. These shells not only let diatoms maximize
light absorption for photosynthesis1, but also filter out potentially harmful near UV light wavelengths2. Moreover,upon incorporation into diatom cells, suitable organic fluorophores have been proven to harvest light in spectral
ranges where diatoms do not efficiently absorb, thus enhancing their photosynthesis3. Frustule properties haveattracted attention for their possible application in photonics and optoelectronics4. In addition, both covalent and noncovalent approaches to anchor functional moieties (drugs, dyes, antibodies, metals)5 are explored to afford biohybridnanostructures for biosensing6 and solar energy conversion7.
Here we report a versatile method of chemical modification of diatoms to embed these microalgae in a transparent
organic polyacrylate matrix, without compromising their viability. This method represents a straightforward strategy
to immobilize cells, paving the way to the formation of stable biofilms that can be further functionalized withfluorophores or other molecules for specific applications.
Characterization by electronic microscopy, FTIR-ATR and Raman spectroscopies, as well as functionalization witha suitable fluorescein-based dye, confirm the successful embedment of diatoms into the polymer.
Moreover, the embedded diatom cells can be subjected to an acidic treatment that removes their organic protoplasm,
this allowing to obtain frustules directly embedded in the organic matrix that is conversely preserved. This
immobilization strategy is promising to easily get transparent biosilica based films for photonics applications.
References
1) J. W. Goessling, M. Kühl, Structure-based optics of centric diatom frustules: modulation of the in vivo light field
for efficient diatom photosynthesis. New Phytologist, 219, 122-134, 2018.
2) M. M. Ghobara, R. Gordon, “On Light and Diatoms: A Photonic and Photobiology Review”, Diatoms:
Fundamentals and Applications, Scrivener Publishing LLC, 2019.
3) G. Leone, G. M. Farinola, Incorporating a molecular antenna in diatom microalgae cells enhances photosynthesis,
Scientific Reports, 11, 5209, 2021.
4) R. Ragni, G. M. Farinola, Multiple Routes to Smart Nanostructures Materials from Diatom Microalgae: A
Chemical Perspective, Adv. Mat. 30, 1704289, 20185) S. R. Cicco, G. M. Farinola, In vivo functionalization of diatom biosilica with sodium alendronate as osteoactive
material, Mater. Sci. Eng. C., 104, 109897, 2019.
6) J. Cai, D. Zhang, Au nanoparticle-grafted hierarchical pillars array replicated from diatom as reliable SERS
substrates, Appl. Surf. Sci., 541, 148374, 2021.
7) T. M. W. J. Bandara, B.-E. Mellander, Diatom frustules enhancing the efficiency of gel polymer electrolyte-baseddye-sensitized solar cells with multilayer photoelectrodes, Nanoscale Adv., 2, 199-209, 2020.
Acknowledgements. This work was supported by BEEP-ITN grant from MSCA-H2020.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.2.1

Interaction of different UV wavelengths present in natural sunlight: Impact on DNA
damage and apoptosis
Thomas Haarmann-Stemmann
IUF – Leibniz-Research Institute for Environmental Medicine, 40225 Düsseldorf, Germany
The skin aging exposome consists of all external and internal factors and their interactions, affecting a humanindividual from conception to death as well as the response of the human body to these factors, that lead tobiological and clinical signs of skin aging 1. In this presentation, I will focus on the interaction of two major
exposomal factors for premature skin aging, namely UVA (315 – 400 nm) and UVB (280 – 315 nm) radiation.
Historically, the vast majority of irradiation studies was carried out by exposing the test model (in vitro, in vivo, exvivo) to either UVA or UVB radiation alone or by applying a sequential irradiation protocol, i.e. the test model isfirst exposed to UVA radiation and in a second step irradiated with UVB, or vice versa 2. Given that our skin issimultaneously exposed to the whole electromagnetic spectrum of the sun, we hypothesize that the biologicalconsequences of a simultaneous exposure to UVA and UVB radiation is not adequately mimicked by a simple
addition of the outcome of the respective single or sequential exposure studies. In fact, we found that asimultaneous but not a sequential (UVA>UVB or UVB>UVA) exposure to UVA decreases UVB-induced
keratinocyte apoptosis, as measured on the level of caspase activity, Annexin V/propidium iodide-positive cells,
and subG1 fraction. DNA double-strand breaks (DSBs) are a major driver of UVB radiation-induced apoptosis and,in fact, by investigating the occurrence of DSBs (Comet Assay) and phosphorylated histone 2AX (γH2AX), we
found that a simultaneous but not a sequential exposure to UVA radiation reduced the amount of UVB radiationinduced DSBs. Further mechanistic analyses revealed that in contrast to sequential irradiation protocols, thesimultaneous exposure to UVA and UVB resulted in a differential modulation of heme oxygenase-1, which iscapable of modulating the activity of Ataxia telangiectasia and Rad3 related (ATR) and downstream DSB repair
systems 3. The data presented in this paper indicate that, when simultaneously applied, UVA radiation attenuates
the genotoxic effects of UVB radiation and thus challenge established concepts of skin photoprotection.
References
1) Krutmann J, Bouloc A, Sore G, Bernard BA, Passeron T (2017) The skin aging exposome. J Dermatol Sci 85,
152-61.
2) Krutmann J, Sondenheimer K, Grether-Beck S, Haarmann-Stemmann T (2018) Combined, simultaneous
exposure to radiation within and beyond the UV spectrum: A novel approach to better understand skin damage by
natural sunlight. In: Environment and Skin. H. Merk & J. Krutmann (eds). Springer International Publishing
Switzerland, ISBN: 978-3-319-43100-0.
3) Otterbein LE, Hedblom A, Harris C, Csizmadia E, Gallo D, Wegiel B (2011) Heme oxygenase-1 and carbon
monoxide modulate DNA repair through ataxia-telangiectasia mutated (ATM) protein. PNAS 108, 14491-6.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.2.2
Photo-AGEs and related reactive carbonyl species are molecular mediators of skinphotodamage, photoaging, and carcinogenesis
Georg T. Wondrak1
1) University of Arizona, Pharmacology and Toxicology, College of Pharmacy and UA Cancer Center, Tucson,
Arizona, USA
Recent research indicates that solar UV, in addition to genotoxic effects, causes proteotoxic effects through formationof photodamage-associated protein-epitopes that accumulate in skin as a consequence of chemical reactions including
photooxidation, lipid peroxidation, and glycation. We observed that acute exposure to sub-apoptogenic doses of solar
simulated UV is sufficient to cause an almost twenty-fold increase in photodamage-associated advanced glycation
endproduct protein-epitopes (‘photo-AGEs’) in healthy human skin. These photo-AGEs originate from carbonyl
stress (mediated by reactive carbonyl species) downstream of glycation and lipid peroxidation reactions forming
major lysine- and arginine-derived epitopes including CML (Nɛ-carboxymethyl-L-lysine), DHP (1,4dihydropyridine-3,5-dicarbaldehyde-L-lysine),andarg-pyrimidine{(2S)-2-Amino-5-[(5-hydroxy-4,6dimethylpyrimidin-2-yl)amino]pentanoic acid}, respectively. Likewise, photo-AGEs accumulate in chronically sunexposed skin in the context of photoaging. Remarkably, photo-AGEs are detectable in cutaneous precancerous
lesions including actinic keratosis, and subsequent tissue microarray (TMA) analysis revealed the pronounced
occurrence of photo-AGEs in human nonmelanoma and melanoma skin cancer patient tissue. Interestingly, we have
now identified the topical use of sunless tanning formulations containing glycation agents including
dihydroxyacetone as another significant exposure underlying epidermal photo-AGE formation. A functional role ofphoto-AGEs in skin inflammatory pathology and carcinogenesis is supported by the fact that these epitopes act as
potent endogenous ligands of specialized damage recognition receptors including RAGE and TLR4, potentially
impacting inflammatory dysregulation and both tumorigenic invasion and metastasis. Using genetic target
modulation employing CRISPR/Cas9 we have also substantiated the role of glyoxalase 1 (encoded by GLO1, aglutathione-dependent enzyme involved in detoxification of the reactive carbonyl species methylglyoxal) in
modulating cutaneous carbonyl stress, photo-AGE formation, and tumorigenesis.
Acknowledgements. Supported in part by grants from the National Institutes of Health (1R01CA229418,
1R03CA230949, 1R21ES029579, 1P01CA229112, ES007091, ES006694, and UA Cancer Center Support Grant
CA023074). The content is solely the responsibility of the author and does not necessarily represent the official viewsof the National Cancer Institute or the National Institutes of Health.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.2.3
UVB-induced senescence and its role in melanomagenesis
Eleni Fitsiou, Marco Demaria
European Research Institute for the biology of Ageing (ERIBA), University Medical Center Groningen (UMCG),
Groningen, Netherlands
A common feature of older organisms is the accumulation of senescent cells – cells that have irreversibly lost thecapacity to undergo replication. Senescent cells are characterized by an irreversible cell cycle arrest and by theSenescence-Associated Secretory Phenotype (SASP), which includes many tissue remodeling and pro-inflammatory
factors. Senescent cells are intermittently present during embryogenesis and in young organisms. On the contrary,senescent cells accumulate and persist in aging tissues. Importantly, accumulation of senescence is accelerated incontext of excessive genotoxic stress, such as in the skin exposed to UV radiation. Significantly, these persistentsenescent cells can drive low-grade chronic inflammation, and their genetic or pharmacological elimination is
sufficient to delay tissue dysfunciton. Here, I will discuss recent insights on how UVB can promote skin senescence,and general phenotypes of UVB-induced senescent cells. I will also describe the mechanisms by which accumulating
dermal senescent cells might promote melanoma initiation and progression, and the potential to reduce UVBassociated melanoma by targeting senescent cells.

Acknowledgements. This work was supported by a grant from the Dutch Cancer Foundation (KWF).

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-1.2.4

Cellular response of human keratinocytes to a photo-pollution stress
Alexe Grenier1,2, Patrick J Rochette1,3,*, Roxane Pouliot1,2,*
1) Centre de Recherche en Organogénèse Expérimentale de l’Université Laval/LOEX, Centre de recherche du CHU
de Québec-Université Laval, Québec, QC, G1J 1Z4, Canada; 2) Université Laval, Faculté de pharmacie, Québec,QC, G1V 0A6, Canada; 3) Université Laval, Département d’ophtalmologie et ORL-chirurgie cervico-faciale,
Faculté de médecine, Québec, QC, G1V 0A6, Canada; *These authors contributed equally to this work.
Skin aging is a natural process affected by several environmental factors, such as sun exposure and cigarette smoke1-3.
Studied separately, these two extrinsic factors are well-known to accelerate skin aging. However, their combined
effects have been poorly characterized. Those factors could although interact with one another in the skin sincecigarette smoke components can accumulate in the skin by contact or by systemic effect after inhalation, and solarrays can penetrate the epidermis and dermis. The aim of this study was to evaluate the cellular response of
keratinocytes to a photo-pollution stress generated by cigarette smoke and sunlight. A cigarette smoke extract (CSE)
was obtained by capturing the soluble fraction of cigarette smoke using an impinger and cigarettes (3R4F). Sun
exposure was performed using a solar simulator equipped with a CGA-345 filter (Newport) to block wavelengths
under 340 nm. Irradiation doses were 14.5 and 29 kJ/m2 of UVA, representing 15 and 30 minutes of zenith sun
exposure. Firstly, the CSE absorption spectrum and total antioxidant capacity (TAC) were assayed. The cellularresponse of keratinocytes was studied using ROS specific fluorescent markers, and MTS to evaluate cellular
metabolic activity. The implication of metabolic enzymes was assessed with RT-qPCR. The characterization of theCSE confirmed that it absorbs UV and visible light wavelengths, and the TAC test revealed an increasing antioxidant
capacity with increasing concentrations of CSE. Photo-oxidation results indicate a decrease in type I ROS and a slightincrease in type II ROS. CSE and solar irradiation alone showed no cytotoxicity up to the maximal doses tested.
However, CSE was highly phototoxic when irradiated. Indeed, when exposed to 5% CSE at 14.5 kJ/m2 and 29 kJ/m2
UVA, keratinocytes showed respectively a 39 ± 1% and 2 ± 17% cellular viability (p value < 0.001). Our results
showed that the photo-polluting stress causes a photo-oxidation and a decrease in cell viability on humankeratinocytes.
References
1) Krutmann, J., et al., The skin aging exposome. Journal of Dermatological Science, 2017.
2) Xu, Y.; Fisher, G. J., Ultraviolet (UV) light irradiation induced signal transduction in skin photoaging. Journal of
Dermatological Science Supplement, 2005.
3) Morita, A., Tobacco smoke causes premature skin aging. Journal of Dermatological Science, 2007.
Acknowledgements. This work was supported by grant from the Canadian Institutes of Health Research (CIHR)(MOP-311262). The student also received studentships from the “Fonds de Recherche Québec-Santé” (FRQS).

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.2.5

Skin Photoaging: a stress-induced cognitive misleading
Mauro Picardo, Enrica Flori, Federica Papaccio, Daniela Kovacs, Stefania Briganti, Giorgia Cardinali, Barbara
BelleiSan Gallicano Dermatological Institute, IRCCS, Cutaneous Physiopathology, Rome, Italy
Skin photoaging blends with the process of intrinsic aging. The severity of photodamage depends on bothconstitutional factors, such as skin phototype, intensity, and duration of sunlight/UV exposure, and the impact of
extrinsic factors, collectively called exposome, including, pollution, smoke or heat, nutrition, and stress, resulting
in an increased senescent cells number. Aging affects nearly every aspect of cutaneous biology, including
pigmentation. Clinically, the phenotype of older pigmented skin has a mottled, uneven color, primarily due to “agespots”, with or without hypopigmentation. Uneven pigmentation might be attributed to the hyperactivation ofmelanocytes, altered distribution of pigment, and turnover. Various skin cells play a role in inducing abnormal
pigmentation, with an aberrant paracrine regulation that impacts melanocyte function, and results in the formationof hyperpigmented lesions, such as solar lentigo and melasma. Not only keratinocytes and fibroblasts but also
endothelial cells and sebocytes, as well as immune inflammatory cells, contribute to the pigmentary changes
associated with aging. The generation of inflammatory mediators and alteration in growth factor production fromthe different cellular components is a biological marker of photo-damaged skin which produces a cognitive
misunderstanding1. Similar biological characteristics can be detected in non-melanoma skin cancer-associatedfibroblasts. Interestingly, age-independent, diffuse expression of senescence-associated markers in the dermal andepidermal compartment is also associated with vitiligo, suggesting that premature senescence plays an importantrole in the pathology 2,3.

References
1. Kang HY, Lee JW, Papaccio F, Bellei B, Picardo M. Alterations of the pigmentation system in the aging
process Pigment Cell Melanoma Res. 2021 May 28. doi: 10.1111/pcmr.12994.
2.

Seneschal J, Boniface K, D'Arino A, Picardo M. An update on Vitiligo pathogenesis Pigment Cell
Melanoma Res. 2021 Mar;34(2):236-243

3. Bellei B, Migliano E, Picardo M. A Framework of Major Tumor-Promoting Signal Transduction Pathways
Implicated in Melanoma-Fibroblast Dialogue. Cancers (Basel). 2020 Nov 17;12(11):3400

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IL-1.2.6

Relationship between intrinsic and extrinsic skin aging
S. Schneider1, M. Majora1, S. Faßbender1, M. Pollet1, A. Marini1, J. Hüsemann1, M. Knechten1, H. Schwender2, J.
Krutmann1
1) IUF - Leibniz Research Institute for Environmental Medicine, Duesseldorf, Germany; 2) Heinrich Heine
University Duesseldorf, Mathematical Institute, Duesseldorf, Germany
Skin aging is principally driven by the combination of intrinsic (e.g. genetic constitution) and extrinsic factors (e.g.
solar radiation). It is generally assumed that extrinsic skin aging superimposes intrinsic skin aging, neverthelessscientific evidence for this assumption is scarce. To assess this relationship more closely, we have focused on dermal
fibroblasts isolated from intrinsically (NHDFINT) vs extrinsically aged skin (NHDFEXT) of young (Y; 20-25 yr.),
middle-aged (M; 36-49 yr.) and old (O; 60-64 yr.) female volunteers. Proteome analysis via quantitative, label-freemass spectrometry revealed fundamentally different age-dependent trajectories of protein levels in NHDFINT vs
NHDFEXT. Accordingly, NHDFINT exhibited gradual, almost linear changes (Y<M<O and Y>M>O). In contrast,
NHDFEXT showed a non-linear kinetic with minor alterations in Y vs M probands (Y=M), but a marked difference
between M vs O donors (M<<O and M>>O) which outreached the changes found in NHDFINT. For example,
tricarboxylic acid (TCA) cycle-related proteins were increased in NHDFEXT vs NHDFINT of M donors. At the sametime, ADP/ATP ratios were elevated, NAD+ levels were reduced and histone deacetylation through NAD+-dependent
sirtuin-1 (SIRT1) was compromised. These observations indicate that NHDFEXT, in contrast to NHDFINT, show acompensatory behaviour to cope with an energy deficit. However, when NHDFEXT of M vs O donors were compared,
NHDFEXT of O donors seemed to have passed a ‘tipping point’, at which compensatory mechanisms failed.
Accordingly, we observed a dramatic decline of TCA cycle-associated proteins accompanied by a severe reductionof protein diversity and protein biosynthesis, i.e. two biological processes known to consume vast amounts of ATP.
Of note, treatment of NHDFEXT of M donors with the NAD+ precursor β-nicotinamide mononucleotide (NMN) could
rescue their mitochondrial phenotype and SIRT1-dependent histone deacetylation. These studies demonstrate thatthe relationship between intrinsic and extrinsic skin aging might be more complex than a simple superimposition,
because age-dependent proteomic/metabolic changes in NHDFEXT follow a substantially different kinetic comparedto NHDFINT. They identify restricted availability of NAD+ as a factor driving metabolic/epigenetic alterations
contributing to extrinsic skin aging.

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IL-1.2.7

Epilipidomic signatures of senescence and UV stress
Florian Gruber1,2,3, Marie-Sophie Narzt1,3, Vera Pils3,4, Christopher Kremslehner1,2,3,
Martina Marchetti-Deschmann2,5, Samuele Zoratto2,5, Markus Schosserer2,4, Ingo Lämmermann3,4,Johannes Grillari3,4,6
1) Medical University of Vienna, Department of Dermatology, Vienna, Austria; 2) Christian Doppler
Laboratory for Skin Multimodal Analytical Imaging of Aging and Senescence – SKINMAGINE-;
3) Christian Doppler Laboratory for Biotechnology of Skin Aging, Vienna, Austria; 4) University of Natural
Resources and Life Sciences Vienna, Department of Biotechnology, Vienna, Austria; 5) Institute ofChemical Technologies and Analytics, TU Wien, Vienna, Austria; 6) Ludwig Boltzmann Institute forExperimental and Clinical Traumatology, AUVA Research Center, Vienna, Austria
During aging, skin accumulates senescent cells. The transient presence of senescent cells, followed by theirclearance by the immune system, is important in tissue repair and homeostasis. The persistence of senescent cells
that evade clearance contributes to the age-related deterioration of the skin. The senescence-associated secretory
phenotype of these cells contains immunomodulatory molecules that facilitate clearance but also promote chronic
damage. Ultraviolet light is the dominant environmental oxidative skin stressor and a major skin aging factor. We
have been studying which oxidized phospholipid (OxPL) mediators (in their totality, the epi phospho-lipidome)
would be generated in human keratinocytes (KC) and fibroblasts (FB) upon exposure to ultraviolet radiation andinvestigated the contribution of OxPL to stress responses. We identified known and novel lipid species including
known bioactive and also potentially reactive carbonyl containing species. We found indication for selective
metabolism and degradation of selected reactive lipids. Exposure to both UVA and to in vitro UVA - oxidized
phospholipids activated, on transcriptome and proteome level, NRF2/antioxidant response signaling, lipid
metabolizing enzyme expression and unfolded protein response (UPR) signaling.
Furthermore, we have investigated the epilipidome of senescent dermal fibroblasts, because these molecules areamong the bioactive lipids that were recently identified as senescence-associated secretory phenotype factors.
Using replicative- and stressinduced senescence protocols, we identified lysophosphatidylcholines as universallyelevated in senescent fibroblasts, whereas other oxidized lipids displayed a pattern that was characteristic for theused senescence protocol. When we tested the lysophosphatidylcholines for senescence-associated secretory
phenotype activity, we found that they elicit chemokine release in nonsenescent fibroblasts but also interfere with
toll-like receptor 2 and 6/CD36 signaling and phagocytic capacity in macrophages.
Our data suggest that oxidized lipids can act as signaling mediators of extrinsic and intrinsic (senescence
associated) stress / SASP, and that the composition of the epilipidome influences the signaling andimmunomodulatory events associated with these cellular states in a fashion that is more specific than anticipated.
Furthermore, we have begun to localize these signaling mediators in human skin using mass spectrometry imaging,which allows us to understand spatial distributions of these novel signaling compounds the UV response and aging.
References
1) Narzt, et al. “A novel role for NUPR1 in the keratinocyte stress response to UV oxidized phospholipids”, Redox
Biology, 20192) Narzt, et al. “Epilipidomics of Senescent Dermal Fibroblasts Identify Lysophosphatidylcholines as Pleiotropic
Senescence-Associated Secretory Phenotype (SASP) Factors”, J. Invest. Dermatol. 2021
Acknowledgements. The work of the CDLs is funded by the Federal Ministry for Digital and Economic Affairs(BMWFW) of Austria and the National Foundation for Research, Technology, and Development of Austria and by
CHANEL Parfums et Beauté and the Herzfelder'sche Familienstiftung, Austria

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OC-1.2.8

Infrared and visible light in combination with UV increase skin cell stress markers
Catherine A Bonn1, L. Hudson1, E. Rashdan1, Bhaven Chavan2, Mark Birch-Machin1
1) Newcastle University, Institute of Translational and Clinical Research, Newcastle-upon-Tyne, UK;
2) Croda Europe Ltd, Snaith, East Yorkshire, UK
Introduction
While it is known that Ultraviolet (UV) light can have detrimental impacts to skin ageing, less is known about the
impacts that longer wavelengths have on skin biology. The impacts of infrared (IR) and visible lights on skin are
clearly different to that of UV, but still seem to affect the response of skin to incident light. While some have foundthat IR alone can cause damage to skin cells[1], in the experiments detailed here we have found that in combination
with UV and visible light it can increase damage markers in human skin cells[2].
Methods
Human skin fibroblast cell lines or primary fibroblasts and keratinocytes were cultured and irradiated with solarsimulated light or an IR lamp. In the “UV+Vis+IR” condition, cells were dosed with 2.16 Standard Erythemal
Doses of UV, equivalent to 2 hours of Mediterranean, noon, midsummer sun, with visible and infrared light alsopresent from the Newport Solar Simulator. In the UV-only treated cells, the cells were exposed to the same solar
simulator but with a filter allowing only the UV light through. The IR-only treated cells were irradiated with a doseof equivalent to a 2 hour dose of light from a Hydrosun IR lamp. Assays included ROS detection with ROS-Glo,mitochondrial DNA damage with long-range PCR and nuclear DNA damage with comet assays.
Results
UV light increases damage biomarkers such as ROS generation by at least 44% in both primary fibroblasts andkeratinocytes, (measured by ROS-Glo, P<0.05). In comparison, infrared light alone had no significant effect oneither measure. However, when treated with UV, visible and IR light, the ROS increased by 168%, though only infibroblasts (P<0.05). In keratinocytes, the additional visible and infrared lights had no effect, and the ROS
generation was not significantly different to that in UV-only irradiated cells.
This pattern of increased damage biomarkers in fibroblasts but not keratinocytes was found to be the same when
mitochondrial DNA damage was assayed. In fibroblasts, the damage in mitochondrial DNA when visible and IR
light were present was 32-fold higher than in unirradiated cells, whereas it was only 8-fold higher in cells irradiatedwith only UV. In keratinocytes the damage was similar in both conditions (3.8 and 4.7-fold respectively). The
pattern was the same in nuclear DNA damage, and was reflected in fibroblast and keratinocyte cell lines.
Furthermore, the sum of damage from the constituent parts of the solar light did not add up to the effect of UV,
visible and IR lights together, indicating a possible synergy of effects on skin.
Conclusion
Infrared and visible light seem to contribute to skin damage from light containing be able to affect the biologicaldamage caused by ultraviolet light, potentially synergistically increasing skin damage.
References
1.
Schroeder, P., et al., Cellular response to infrared radiation involves retrograde mitochondrial signaling.
Free Radical Biology and Medicine, 2007. 43(1): p. 128-135.
2.
Hudson, L., et al., Individual and combined effects of the infrared, visible, and ultraviolet light components
of solar radiation on damage biomarkers in human skin cells. The FASEB Journal, 2020. 34(3): p. 38743883.

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P-1.2.9

Photoprotective efficacy of PLGA-Curcumin versus PLGA-Piperine under ambient
UV-B and Sunlight Exposure
Sunil Kumar Patel1,2, Lipika Ray3, Ratan Singh Ray4, Ashish Dwivedi1,2*1) Food, Drug & Chemical Toxicology Group, CSIR-IITR, Lucknow; 2) Academy of Scientific and Innovative
Research (AcSIR), CSIR-IITR Campus, Lucknow; 3) Pharmaceutics & Pharmacokinetics Division, CSIR-CDRI,
Lucknow; 4) System Toxicology and Health Risk Assessment Group, CSIR-IITR, Lucknow.
Due to pollution and depletion of stratospheric ozone the intensity of UV-R has been augmented in tropical andsub-tropical countries. Chronic exposure of UV-R promotes skin damage, skin aging and skin cancer.
Phytochemicals are promising agents to counteract the negative impact of UV-R by quenching the ROS or
upregulating the molecular expression of cellular anti-oxidant enzymes. But photoinstability is major concern
which limits its long time protection efficacy under UV-R and Sunlight. Here, we have taken two very important
phytochemicals i.e., curcumin and piperine and their nanoformulations for photostability and photoprotection under
UV-B and Sunlight. Our photodegradation results suggest that both the phytochemicals are degraded under UV-B
and Sunlight. However, nanoformulations are stable under both UV-B and Sunlight exposures. In-vitro result alsosuggests that nanoformulations have more protective efficacy than normal forms. Moreover, comparison to
curcumin-PLGA, piperine-PLGA provides strong protection against UV-B and Sunlight mediated damage. These
results suggest that PLGA-piperine has strong photoprotection efficacy and may be used in preparations ofsunscreens and other skin protecting creams.
Keywords: Phototoxicity; Photodegradation; Nanoformulation; Curcumin; Apoptosis/Necrosis; Photoprotection.
Acknowledgements. This work was supported by CSIR-IITR, Lucknow and UGC New Delhi.

*Corresponding Author: Dr Ashish Dwivedi
Food, Drug and Chemical Toxicology Group,
CSIR-Indian Institute of Toxicology Research (CSIR-IITR), Vishvigyan Bhawan 31,
MG Marg, Lucknow, Uttar Pradesh-226001, India
Telephone: +917897944888,
E-mail: ashish.dwivedi1@iitr.res.inBook of Abstracts ������������������������������������������������������������������������������������������������ 26

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.3.1

Photoantimicrobials: What we know and do not know!
Tim Maisch1
1) Department of Dermatology, University Hospital Regensburg, Germany
The worldwide and continuous increase in antibiotic resistance requires alternative strategies to combat bacteria. A
promising complement to the conventional use of antibiotics is the principle of antimicrobial photodynamic
inactivation. In this procedure, a per se non-toxic dye, so-called photoantimicrobial1, are excited with visible lightin the presence of oxygen. Thereby it produces reactive oxygen species that irreversibly cause an oxidative damage
to biological molecules, which enables the non-specific killing of bacteria, including multi-resistant pathogens. In
this talk the current knowledge about antimicrobial photodynamic inactivation will be discussed.
References
1) Mark Wainwright, Tim Maisch, Santi Nonell, Kristjan Plaetzer, Adelaide Almeida, George P Tegos,
Michael R Hamblin, Photoantimicrobials—are we afraid of the light? Lancet Infect Dis. 2017 Feb; 17(2):e49–e55.

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19th Congress of the European Society
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.3.2
Adding Photodynamic Inactivation to the farmer's armamentarium against
phytopathogens
Michael Glueck1, Christoph Hamminger1, Annette Wimmer1, Michael Fefer2, Jun Liu2, Kristjan Plaetzer1
1) University of Salzburg, Department of Biosciences, Laboratory of Photodynamic Inactivation of
Microorganisms, Salzburg, Austria; 2) Suncor AgroScience, Mississauga Ontario, Canada
Access to sufficient and healthy food is a fundamental human right. Crop production has essential importance forfood safety, but yields are threatened by plant pathogens. This study investigates the efficacy of Photodynamic
Inactivation (PDI) based on two chlorin e6 derivatives, Sodium-Magnesium-Chlorophyllin (Chl, E140) and thecationic B17-0024 against bacterial and fungal plant pathogens, both, in vitro and in situ.
PDI against the Gram(+) phytopathogens Rhodococcus fascians and Clavibacter michiganensis as well as Gram(-)
Erwinia amylovora - susceptible and resistant to streptomycin - and Xanthomonas axonopodis was performed inliquid culture and using illumination with 395 nm LED light (26.6 J/cm2) or natural sunlight. CFU counting wasdone 24/48 h after phototreatment. The PDI efficiency was also tested on Solanum lycopersicum (tomato) plants,
which were contaminated with C. michiganensis. For the antifungal PDI against Alternaria solani and
Botrytis cinerea the growth of mycelial patches after photoactivation (395 nm, 106.6 J/cm2) of either Chl or
B17-0024 was determined seven days post treatment. Gram(-) and fungal phytopathogens were incubated with Chl
in combination with a cell wall permeabilizing agent. To investigate the effect of the PDI-treatment to host plants
the growth of Fragaria vesca (strawberry, BBCH stage 14) inoculated with the photosensitizers with or without
additives was monitored for 14 days.
Both compounds photokill (inactivation >6 log10 steps) R. fascians and C. michiganensis at 100 µM concentration.
Photoinactivation (>7 log10) of Gram(-) bacteria with 100 µM Chl requires addition of Na2EDTA (X. axonopodis)
or polyaspartic acid (E. amylovora).1 The cationic B17-0024 is phototoxic against both Gram(-) species at
10/100 µM without additives. E. amylovora resistant to streptomycin was photokilled with an efficiency
comparable to the wild type, and addition of 100 µg/ml streptomycin further enhanced the cytotoxicity. Sunlightactivation induces an antibacterial effect towards C. michiganensis after 30 minutes in situ on S. lycopersicum
leaves (100 µM B17-0024 integrated radiant exposure 64.1 J/cm²) and E. amylovora in liquid suspension after twohours (100 µM B17-0024, average irradiance 28 mW/cm²). Photoinactivation of C. michiganensis inoculated on
S. lycopersicum plants is effective (> 5 log steps reduction) using 100 µM B17-0024 (395 nm, 26.6 J/cm²). Bothchlorin e6 derivatives also act as very effective photofungicides, if Chl is applied with a cell wall permeabilizing
compound and B17-0024 without any additives. The PDI-treatment had no effects on the development and growthof F. vesca.
PDI based on the two chlorin e6 derivatives tested in this study is effective in killing bacterial and fungal
phytopathogens, both, in vitro and in situ and irrespective of resistance to conventional treatment. As no negative
effects to the host plants have been observed so far, the photodynamic approach will add to the grower's toolbox tominimize crop losses.
References
1) M. Glueck, C. Hamminger, M. Fefer, J. Liu and K. Plaetzer, Save the crop: Photodynamic Inactivation of plant
pathogens I: bacteria. Photochemical & Photobiological Sciences 18(7): 1700-1708, 2019.
Acknowledgements. The authors are grateful to Suncor AgroScience, Mississauga Ontario, Canada for support.

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IL-1.3.3

Anti-bacterial and anti-virulent photoinactivation of Staphylococcus aureus isolated
from atopic dermatitis patients: an in vitro and in vivo approach.
Joanna Nakonieczna1
1) Laboratory of Molecular Diagnostics, Intercollegiate Faculty of Biotechnology of University of Gdańsk andMedical University of Gdańsk, Gdańsk, Poland
Atopic dermatitis (AD) is the most common chronic inflammatory skin disease. It affects 5-20% of children
worldwide and 1-3% of adults. The literature describes the phenomenon of dysbiosis with respect to the skin ofpatients with AD. This state is defined as a change in the composition of the normal flora on the patient’s skin. In
particular, an increase in skin colonization of S. aureus species was observed in AD patients. The prevalence of S.
aureus on atopic skin is 80-100%, whereas in healthy populations it is estimated to be 5-20%. Currently, AD
treatment is primarily based on anti-inflammatory treatment with the use of topical corticosteroids (TCSs) andcalcineurin inhibitors (TCI) depending on the disease severity1. Additionally, antimicrobial therapy in eachexacerbation of AD can be applied, mainly with the use of mupirocin, fusidic acid, retapamulin. However, prolonged
use of antibiotic is not recommended due to significant rise in resistance. Published data suggest that the increased
severity of AD is attributed to S. aureus secreted virulence factors with superantigen properties. The best studied
superantigens are staphylococcal enterotoxins (SEA, SEB, SEC, SED) and toxic shock syndrome toxin-1 (TSST-1).
Several studies indicated a direct correlation between the occurrence of superantigens-producing S. aureus and severe
AD. In the in vitro conditions we verified the hypothesis that numerous strains of AD-derived S. aureus and thestaphylococcal virulence factors can be effectively photoinactivated, including those of MRSA strains. We checked
the effect of aPDI on the studied virulence factors at RNA and protein levels, which revealed interesting differencesin response to aPDI among various SEs. We found such conditions of photoinactivation wherein the host skin cells
remained intact, while effectively destroying bacterial cells and virulence factors produced. Currently, we aredeveloping in vivo model of atopic-like skin colonization by MRSA bioluminescent strain to assess the efficacy of
decolonization of lesional and non-lesional areas of skin. The results obtained so far allow to propose aPDI as a goodcandidate complementary method of skin decolonization and inactivation of virulence factors exacerbating flares ofAD.
References
1) Ogonowska P, Gilaberte Y, Baranska-Rybak W, Nakonieczna J, Colonization With Staphylococcus aureus in
Atopic Dermatitis Patients: Attempts to Reveal the Unknown, Frontiers in Microbiology, 2021.
Acknowledgements. This work was supported by grant no. 2017/27/B/NZ7/02323 from the National ScienceCentre, Poland.

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OC-1.3.4

aPDT Activity of Cationic Copolymers with Dual Function: Synergistic Effect ofPoly(oxanorbornene) and Phthalocyanine
Erem Ahmetali1, Pinar Sen2, N. Ceren Süer1, Burak Aksu3, Tebello Nyokong2, Tarik Eren1, M. Kasım Şener1
1) Yıldız Technical University, Department of Chemistry, Istanbul 34210, Turkey ; 2) Rhodes University,
Department of Chemistry, Grahamstown 6140, South Africa ; 3) Marmara University, Department of Medical
Microbiology, Istanbul 34854, Turkey
In today's world, microorganisms’ infections are serious problems for public health. The researchers have focused
how to overcome the problem of microbial drug resistance in recent years1. Antimicrobial photodynamic therapy(aPDT) is an alternative method for photo-inactivation of microorganisms through cytotoxic species produced inconjunction with photosensitizer (PS) and light. Phthalocyanine compounds are especially known as potential PS
that can produce high singlet oxygen in photodynamic process2.
On the other hand, amphiphilic poly(oxanorbornene)s are promising synthetic polymers that can show highantimicrobial functions against bacteria. Although different antimicrobial action mechanisms have been proposed
for amphiphilic poly(oxanorbornene)s, it is thought that the electrostatic interaction and the hydrophobic interaction
provide a lethal effect3.

Fig. 1 – Representation of Enhanced Light-Driven Antimicrobial Activity

In this study, poly(oxanorbornene)s bearing zinc(II) phthalocyanine were synthesized to combine the antimicrobial
mechanisms of cationic poly(oxanorbornene) with aPDT activity of zinc(II) phthalocyanine. The results suggestthat synergistic effect of Zn(II) phthalocyanine-triphenyl phosphonium based copolymer caused complete damageof bacterial strain with multifunctional mechanism, i.e., electrostatic interaction, hydrophobic interaction and theformation of reactive oxygen species4.
References
1) M. L. Agazzi, M. B. Ballatore, A. M. Durantini, E. N. Durantini, A. C. Tome, BODIPYs in antitumoral andantimicrobial photodynamic therapy: An integrating review. J. Photochem. Photobiol. C., 40, 21, 2019.
2) P. Sen, A. Sindelo, D. M. Mafukidze, T. Nyokong, Synthesis and photophysicochemical properties of novel
axially di-substituted silicon (IV) phthalocyanines and their photodynamic antimicrobial chemotherapy (PACT)
activity against Staphylococcus aureus. Synth. Met, 258, 116203, 2019.
3 ) N. C. Süer, C. Demir, N. A. Ünübol, Ö. Yalçın, T. Kocagöz, T. Eren, Antimicrobial activities of phosphonium
containing polynorbornenes. RSC Adv., 6, 86151, 2016.
4) E. Ahmetali, P. Sen, N. C. Süer, B. Aksu, T. Nyokong, T. Eren, Enhanced light-driven antimicrobial activity of
cationic poly(oxanorbornene)s by phthalocyanine incorporation into polymer as pendants. Macromol. Chem. Phys.,
221, 2000386, 2020.
Acknowledgements. The authors would like to acknowledge the financial support from The Scientific andTechnological Research Council of Turkey (TÜBİTAK) (Project Number: 219Z271).

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.3.5

LIGHT4LUNGS: Inhalable Aerosol Light Source for Controlling Drug-ResistantBacterial Lung Infections
Santi Nonell, on behalf of the Light4Lungs GroupS. Nonell, Institut Químic de Sarrià, Universitat Ramon Llull, Barcelona, Spain; G. Romano, University of
Florence, Florence, Italy; N. Henry, Sorbonne Université, Paris, France; E. Palomares, Fundacio Privada Institut
Catala d’Investigacio Quimica, Tarragona, Spain; E. Reddi, University of Padova, Padova, Italy; G. Taylor,
Cardiff Scintigraphics Ltd, Cardiff, UK; A. Kadioglu, The University of Liverpool, Liverpool, UK
Light4Lungs addresses the problem of antimicrobial resistance in the treatment of chronic lung infections, theleading cause of morbidity and mortality in patients with diseases such as cystic fibrosis and hospital-acquired lunginfections. The project aims to develop an alternative therapeutic scheme for the treatment of these infections: a
novel inhalable photodynamic therapy whereby antibiotics are replaced by inhalable light sources that will excite
bacterial endogenous photosensitizers, eliminating the pathogenic bacteria by the photodynamic effect irrespectiveof their multidrug resistance profile.
The project encompasses the development of breathable particles with persistent luminescence, the aerosol
technology for activation and delivery to the lungs, and the definition of the treatment parameters through toxicity
and efficacy tests in clinically relevant models of respiratory infections. Its results have the potential to go beyond
treatment of recalcitrant respiratory tract bacterial infections to other lung diseases and to other organs, enriching
fields of healthcare, nanomedicine, materials science, and nanotechnology.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-1.3.6

Photodynamic inactivation of pathogenic bacteria to decontaminate surfaces in foodindustry
Larissa Kalb1, Bernd Kramer2, Peter Muranyi2, Wulf Schneider-Brachert3, Wolfgang Bäumler1
Department of Dermatology, University of Regensburg, Germany, 2Fraunhofer Institute for Process Engineeringand Packaging IVV, Freising, Germany, 3Department of Infection Control and Infectious Diseases, University ofRegensburg, Germany

1

Introduction Despite established standards of cleaning and disinfection, decontamination of environmental surfaces
remains a big challenge in food processing. Contaminated surfaces in meat production are a source of bacteriatransmission from surface to surface of different equipment (e.g. conveyor belts, housings). The photodynamic
procedure is a promising, new technology that uses a special dye molecule (photosensitizer). Upon visible light
exposure, a photosensitizer generates reactive oxygen species, in particular singlet oxygen, which inactivates bacteria
regardless of its type and its resistance against diverse disinfection strategies. The photodynamic procedure can beapplied in suspension1 and as antimicrobial coating of surfaces.2 In the current study, the inactivation of food relevantbacteria Salmonella enterica and Listeria monocytogenes was investigated using different photosensitizers insuspension, antimicrobial coatings, and in micro-emulsions.
Material and Methods Overnight cultures of L. monocytogenes DMSZ 15675 or S. enterica DMSZ 11320 were resuspended to achieve up to 108 bacteria per mL. Curcumin, flavin, or perinapthenone photosensitizers were added tosuspension or micro-emulsions to yield concentrations of up to 500 µM. In case of the coating, photosensitizers wereadded to a lacquer material that was applied to PU-plastic (conveyor belt) or stainless steel (housings) samples.
Photosensitizers and coatings were purchased from TriOptoTec GmbH, Germany. In case of surface experiments,
50 µL of bacteria suspension was applied as droplets on the surface, allowing bacteria to dry for 60 minutes. Different
concentrations of protein and blood components were added to simulate the organic burden in meat processing.
Photosensitizers were activated by visible blue light of LEDs (DELO GmbH, Germany) at different light intensities
and irradiation times. After irradiation, the number of viable cells was determined by plating bacteria on agar andcounting the colony forming units. The results were calculated as log10 reduction using internal references.
Experiments were repeatedly performed yielding mean values ± standard deviation.
Results Regardless of the photosensitizer used, L. monocytogenes and S. enterica could be inactivated in suspension
experiments with increasing concentration of the respective photosensitizer. The maximal antimicrobial efficacy
showed the perinapthenone photosensitizers with a reduction of up to 6.8 log10 ± 0.1. After inoculation of bacteria on
coated stainless steel or PU-plastic, bacteria were inactivated on the surface upon light exposure yielding a reductionof maximal 4.8 ± 0.8 log10. After inoculation of bacteria on PU-plastic samples, the micro-emulsion containing
photosensitizer (perinapthenone or curcumin) was sprayed on the contaminated surfaces. Irradiation of sample
surfaces for five seconds could reduce the number of bacteria by 3.9 ± 0.8 log10.
Discussion The results of these experiments provide evidence that the photodynamic technology enables a very
efficient inactivation of typical food pathogens. On one hand, the antimicrobial coating can permanently andautonomously inactivate bacteria on surfaces of food processing equipment like housings and conveyor belts. On theother hand, the antimicrobial micro-emulsion can be sprayed on surfaces of food processing equipment to inactivate
food pathogens. The micro-emulsion contains only water, food or cosmetic approved emulsifiers, and harmlessphotosensitizer molecules, which are derivatives of bio-molecules or spices. The biocidal agent, which oxidatively
inactivates the bacteria, is mainly the short-lived singlet oxygen that is generated in situ upon exposure ofphotosensitizers to visible light.
References
1
Schreiner M, Bäumler W, et al, Photodynamic inactivation of bacteria to decolonize meticillin-resistant Staphylococcus aureus
from human skin, Br J Dermatol. 179(6):1358-1367, 20182
Eichner A, Holzmann T, et al Novel photodynamic coating reduces the bioburden on near-patient surfaces thereby reducing
the risk for onward pathogen transmission: a field study in two hospitals, J Hosp Infect 104(1):85-91, 2020
Acknowledgement. This work was supported by grant 281C101C18 from the Federal Ministry for Agriculture and Food,
GermanyBook of Abstracts ������������������������������������������������������������������������������������������������ 32

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OC-1.3.7

How long Escherichia coli can resist a resistance to antimicrobial blue light (aBL)treatment?
Aleksandra Rapacka-Zdończyk1, Agata Woźniak2, Beata Kruszewska1,2, Mariusz Grinholc2 and Krzysztof
Waleron1
1) Department of Pharmaceutical Microbiology, The Faculty of Pharmacy Medical University of Gdańsk,Poland;2) Laboratory of Molecular Diagnostics at Department of Biotechnology, Intercollegiate Faculty of
Biotechnology University of Gdańsk & Medical University of Gdańsk, Poland
Antimicrobial blue light (aBL) is considered a low risk treatment for the development of bacterial resistance or/and
tolerance due to their multi-targeted modes of action. In our previous published studies1 we assessed the tolerance
development to aBL by several Gram positive species, including Staphylococcus aureus, Enterococcus faecium andStreptococcus agalactiae.
The aim of our studies was to demonstrate whether tolerance development occurs in Gram negative bacteria. The
application of the exact protocol2 for the evaluation of resistance development as well as the investigation of umuCand recA mutants of Escherichia coli K-12 demonstrated that representative of Gram negative bacteria also may
develop tolerance to the aBL.
Reference strain of Escherichia coli K-12 was used in the experiments. In the studies cells were irradiated with blue
(415 nm) light. 15 repeated cycles of sub-lethal photoinactivation was followed by bacteria re-growth overnight. A
potential reduction in susceptibility was examined after 5th, 10th, and 15th consecutive cycles at higher light doses.
Reduction in susceptibility has been observed since the 5th cycle.
Lack of tolerance development in recA- lacking Escherichia coli K-12 mutants3 indicates a possible mechanism oftolerance development. Our studies indicate that aBL tolerance development is recA-dependent process confirmingthat photoinduced DNA damage occur under aBL treatment. However, aBL tolerance development demonstrated tobe umuC-independent process (contrary to S. aureus). The acquired tolerance was stable in the case of the wild typestrain and the umuC mutant.
References
1) Rapacka-Zdonczyk, A., et al. "Development of Staphylococcus aureus tolerance to antimicrobial photodynamic
inactivation and antimicrobial blue light upon sub-lethal treatment." Scientific reports 9.1 (2019): 1-18.
2) Rapacka-Zdonczyk, A., Wozniak, A., Nakonieczna, J., & Grinholc, M. (2021). Development of Antimicrobial
Phototreatment Tolerance: Why the Methodology Matters. International Journal of Molecular Sciences, 22(4), 2224.
3) Baba, T., Ara, T., Hasegawa, M., Takai, Y., Okumura, Y., Baba, M., ... & Mori, H. (2006). Construction ofEscherichia coli K‐12 in‐frame, single‐gene knockout mutants: the Keio collection. Molecular systems biology, 2(1),2006-0008.
Acknowledgements. This work was supported by grant No. 2020/36/C/NZ7/00061 from Polish National Science
Center; E. coli single-gene mutants were used courtesy of National BioResource Project (NIG, Japan).

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-1.3.8
Cationic Phthalocyanine Palladium complexes for Photodynamic Inactivation of
Pathogenic Bacteria on Farm Fishes
Vanya Mantareva1, Vesselin Kussovski2, İrem Kulu3, Ivan Angelov2, Mahmut Durmuş3, Petya Orozova4
1

Institute of Organic Chemistry with Centre of Phytochemistry, Bulgarian Academy of Sciences, Bld. 9,1113 Sofia, Bulgaria; 2The Stephan Angeloff Institute of Microbiology, Bulgarian Academy of Sciences,
Bld. 26, 1113 Sofia, Bulgaria; 3Gebze Technical University, Department of Chemistry, P.O. Box 141,
Gebze 41400, Kocaeli, Turkey; 4National Reference Laboratory for Fish, National Diagnostic Scienceand Research Veterinary Medical Institute, 1000 Sofia, Bulgaria
The increasing drug resistance of pathogenic bacteria leads to further development of new strategies to fightmicrobial resistance. The photodynamic inactivation (PDI) appears a prospective approach which takes
action by the light activation of a proper photosensitizer in the presence of oxygen atmosphere. The
effective generation of singlet oxygen predicted the efficacy of the proposed new palladium complexes forphotoinactivation of the pathogenic species. The study presents the PDI efficacy of two new cationic watersoluble palladium phthalocyanines (PdPcs) which are differing in the positions of quaternized 2hydroxypyridine groups to the Pc ring, namely on peripheral (pPdPc, 1) and nonperipheral (nPdPc, 2) incomparison to a highly effective methylpyridyloxy-substituted Zn(II)- phthalocyanine (ZnPcMe) which
was synthesized by the well-developed procedure for quaternized metallophthalocyanines, Scheme 1).

Scheme 1. Synthesis of quaternized MPcs.

Fig. 1. PDI of Aeromonas hydrophila (R).

The typical for farm fishes bacterial strains Flavobacterium psychrophilum is found in cold fresh
waters with an optimal growth below 160 ºC and the Gram(-) Aeromonas hydrophila which grow at 4 ֯C,
were evaluated with high sufficiency for inactivation with non-peripherally substituted Pd(II)Pc, 2 and with
ZnPcMe (Fig. 1). The observations obtained for the complexes of palladium (PdPcs) suggested that metal
ion can influence the photodynamic inactivation to keep under control the resistant bacterial species in the
farm’s fishes.
Acknowledgements. This work was supported by grant of the Bulgarian National Science Fund, KP-06-H29/11.
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OC-1.3.9

Optimisation of Streptococcus
for photoinactivation studies

agalactiae

biofilm

culture

as

a

model

Michał K. Pierański1, Mariusz S. Grinholc1
1) Laboratory of Molecular Diagnostics, Intercollegiate Faculty of Biotechnology University of Gdansk & Medical
University of Gdansk, Gdansk, POLAND
Streptococcus agalactiae is a relevant cause of neonatal mortality. About 10-30% of women are carrying
S. agalactiae in their vaginal tract, from where it can be transferred to infant and cause among others: meningitis,
pneumonia, arthritis or sepsis1. Serotypes the most commonly identified from vaginal tract infections are Ia, III
and V2. It is commonly believed that the cause of therapy ineffectiveness and recurrence of infection is the growthof bacteria in the form of a biofilm. This situation also takes place in the vaginal environment3. So far, several teams
have attempted to find a suitable medium for the cultivation of a Streptococcus biofilm.
Among others there was used simulated vaginal fluid. However, in the case of S. agalactiae, biofilm production
in simulated vaginal fluid is lower than in Tryptic Soy Broth4. We have already showed that S. agalactiae can be
successfully eradicated with rose bengal mediated aPDI in planktonic culture5, but there were no studies
about photoinactivation of biofilm culture yet.
The aim of this study was to optimize S. agalactiae biofilm culture conditions, so that it could be usedin photoinactivation studies.
We compared biofilm production by ATCC strain and three clinical strains representing most common serotypes
in four different broths with crystal violet staining method. Then we evaluated stationary biofilm culture in microtiterplates and biofilm grown on four types of coupons with use of CDC Biofilm Reactor® in continuous flow conditions.
We compared growth in CFU/ml and with confocal laser scanning microscopy. Then we applied rose bengal
mediated photoinactivation on both biofilm models.
We managed to find conditions allowing for stableand repetitive S. agalactiae biofilm growth, what was
confirmed with confocal laser scanning microscopy imaging. We believe that biofilm grown in continuous flow
conditions is more similar to the one grown in natural environment and can be successfully utilizedin photoinactivation studies.
References
1) Shet A, Ferrieri P. Neonatal & Maternal Group B Streptococcal Infections: A Comprehensive Review.
Vol 120.; 2004.
2) Ulett KB, Benjamin WH, Zhuo F, et al. Diversity of group B streptococcus serotypes causing urinary tract
infection in adults. J Clin Microbiol. 2009;47(7):2055-2060. doi:10.1128/JCM.00154-09
3) Muzny CA, Schwebke JR. Biofilms: An Underappreciated Mechanism of Treatment Failure and Recurrence
in Vaginal Infections: Table 1. Clin Infect Dis. 2015;61(4):601-606. doi:10.1093/cid/civ353
4) Borges S, Silva J, Teixeira P. Survival and biofilm formation by Group B streptococci in simulated vaginal
fluid at different pHs. Antonie van Leeuwenhoek, Int J Gen Mol Microbiol. 2012;101(3):677-682.
doi:10.1007/s10482-011-9666-y
5) Pieranski, M., Sitkiewicz, I., & Grinholc, M. (2020). Increased photoinactivation stress toleranceof Streptococcus agalactiae upon consecutive sublethal phototreatments. Free Radical Biology and Medicine,160, 657-669.
Acknowledgements. This work was supported by National Science Centre grant no. 2016/23/B/NZ7/03236

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SP-1.3.10, P-1.3.10

Maltohexaose-porphyrin conjugate for bacteria-targeted Photodynamic Therapy
Abdechakour ELKIHEL, Tan Sothéa OUK and Vincent SOL
Université de Limoges, Laboratoire PEIRENE, EA 7500, Faculté des Sciences et Techniques, 123 Avenue AlbertThomas, 87060 Limoges, France.
Although photodynamic therapy was discovered over a hundred years ago for its ability to kill microorganisms, it
has been developed mainly as an anti-cancer therapeutic treatment. Recently, with the increasing number of infections
caused by multidrug-resistant (MDR) bacteria, Photodynamic Antimicrobial ChemoTherapy (PACT) is considered
as a promising alternative antimicrobial approach whereby microbial cells will not be able to develop resistance 1.
Interesting results were achieved against Gram-positive bacteria, but it also appeared that Gram-negative strains wereless sensitive to PACT. The most used photosensitizers in PACT are porphyrins and their derivatives 2,3. However,these compounds often suffer from a lack of selectivity against bacterial cells. In this work, a novel porphyrinderivative has been synthetized which offers an improved activity and targeting than a usual porphyrin. Neutral mesotritolylporphyrin have been linked to a maltohexaose using a spacer and a cycloaddition (CuAAC) ‘click’ coupling.
Used recently as a bacterial targeting agent for medical imaging with promising results4,5, the maltohexaose enable a
rapid internalization of diverse conjugates through the maltodextrin transport pathway. This highly selective bindingof the photosensitizer to bacteria may increase its efficiency against bacterial strains. This new conjugate has been
investigated for their antibacterial properties against four bacterial strains: two Gram (+) strains (S. aureus CIP76.25; S. epidermidis CIP109.562) and two Gram (−) strains (E. coli CIP54.8T ; E. coli CIP 53.126). The tests have shownthat the coupling of the maltohexaose to neutral porphyrin significantly increases the photocytotoxicity against Gram(+) strains, while no cytotoxicity has been observed in dark condition. Moreover, the conjugate was able to inhibit
the formation of biofilms of S.aureus and S.epidermidis after light irradiation. In addition of this efficiency, flow
cytometry analyses have demonstrated that maltohexaose have increased the interaction of the PS on bacteria. In thenear future, our work may lead to extensive studies for the design of specific targeting photosensitizer, which may
reduce the photocytoxicity on human cells. Thus, it may be applied to the formulation of dermatological ointments
and creams.
References
1) Dai, T.; Huang, Y.-Y.; Hamblin, M. R. Photodynamic Therapy for Localized Infections—State of the Art.
Photodiagnosis Photodyn. Ther. 2009, 6 (3–4), 170–188.
2) Le Guern, F.; Sol, V.; Ouk, C.; Arnoux, P.; Frochot, C.; Ouk, T.-S. Enhanced Photobactericidal and Targeting
Properties of a Cationic Porphyrin Following the Attachment of Polymyxin B. Bioconjug. Chem. 2017, 28 (9),2493–2506.
3) Le Guern, F.; Ouk, T.-S.; Ouk, C.; Vanderesse, R.; Champavier, Y.; Pinault, E.; Sol, V. Lysine Analogue ofPolymyxin B as a Significant Opportunity for Photodynamic Antimicrobial Chemotherapy. ACS Med. Chem.
Lett. 2018, 9 (1).
4) Ning, X.; Lee, S.; Wang, Z.; Kim, D.; Stubblefield, B.; Gilbert, E.; Murthy, N. Maltodextrin-Based Imaging
Probes Detect Bacteria in Vivo with High Sensitivity and Specificity. Nat. Mater. 2011, 10 (8), 602–607.
5) Ning, X.; Seo, W.; Lee, S.; Takemiya, K.; Rafi, M.; Feng, X.; Weiss, D.; Wang, X.; Williams, L.; Camp, V.
M.; Eugene, M.; Taylor, W. R.; Goodman, M.; Murthy, N. Fluorine-18 Labeled Maltohexaose Images Bacterial
Infections by PET. Angew. Chem. Int. Ed Engl. 2014, 53 (51), 14096–14101. 3.

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P-1.3.11
Min Oscillations as Reporter of Bacterial Photodynamic Inactivation at the SingleCell Level
Ingrid V. Ortega1, Joaquim Torra1 and Cristina Flors1
1) IMDEA Nanociencia, C/ Faraday 9, Ciudad Universitaria de Cantoblanco, 28049, Madrid, Spain.
The Min protein system is a cell division regulator in the bacterium Escherichia coli1. Under normal growth
conditions, MinD is associated to the membrane and undergoes pole to pole oscillations. The period of theseoscillations has been previously proposed as a reporter for bacterial physiological state at the single-cell level. Thismethod has been used to monitor the effect of bacterial challenges from antibiotics, 2 temperature3 or mechanicalfatigue.4 Using real-time single-cell fluorescence imaging, we explore here the effect of antimicrobial photodynamic
treatment on MinD oscillations in E. coli. Irradiation of bacteria in the presence of the photosensitizer methylene
blue (MB) disrupts the Min oscillation pattern depending on the MB concentration. Different MB incubation times
and irradiation powers are also explored. In contrast to antibiotics, which slow down the oscillation, photodynamic
treatment results in an abrupt interruption of the oscillation and reflects divergent physiological mechanisms that leadto bacterial death. We propose that single-cell microscopy studies using Min oscillations as a reporter are a useful
tool to unravel the initial physiological response in antimicrobial photodynamic therapy.

References
1) De Boer, Piet A.J., Crossley, R. E. and Rothfield, L. I. A division inhibitor and a topological specificity
factor coded for by the minicell locus determine proper placement of the division septum in E. coli. Cell,
56(4), 641-649, 1989.
2) Downing, B. P., Rutenberg, A. D., Touhami, A., and Jericho, M. Subcellular Min oscillations as a singlecell reporter of the action of polycations, protamine, and gentamicin on Escherichia coli. PloS one, 4(9),e7285, 2009.
3) Touhami, A., Jericho, M., and Rutenberg, A. D. Temperature dependance of MinD oscillation in
Escherichia coli: running hot and fast. Journal of Bacteriology, 7661-7667, 2006
4) Del Valle, A., Torra, J., Bondia, P., Tone, C. M., Pedraz, P., Vadillo-Rodriguez, V. And Flors, C.
Mechanically induced bacterial death imaged in real time: a simultaneous nanoindentation andfluorescence microscopy study. ACS Appl. Mater. Interfaces, 12(28), 31235-31241, 2020.
Acknowledgements. This work was supported by grants PRE2018-084983 and PGC2018-094802-B-I00 from the
Spanish Ministerio Ciencia, Innovación y Universidades.

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19th Congress of the European Society
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.12

Water hardness shows strong impact on the photodynamic activity of cationicpyridylporphyrins against Legionella pneumophila and biofilm
Iva Ćavar1, Martina Mušković1, Ivana Gobin2, Nela Malatesti1
1) University of Rijeka, Department of biotechnology, Rijeka, Croatia;
2) University of Rijeka, Faculty of medicine, Department for microbiology and parasitology, Rijeka, Croatia
Legionella pneumophila, a gram negative, aerobic bacterium, is a member of the opportunistic premises plumbing
pathogens (OPPP) group. The main characteristics of this group of pathogens are their capacity to live in different
aquatic systems, the possibility of biofilm formation and their resistance to common disinfection methods , such as
chlorination and high temperatures1.
To date, only a small number of studies of photodynamic inactivation (PDI) of Legionella pneumophila have been
published. Our research group recently described PDI with cationic tetrapyrid-3-ylporphyrin and two cationictripyrid-3-ylporphyrins, one hydrophilic and the other conjugated with long alkyl chain (C18), on L. pneumophila.
Although all tested porphyrins showed a strong PDI effect on L. pneumophila, the amphiphilic tricationic porphyrinwith a long chain stood out with its strong anti-legionella activity already at nanomolar concentration (25 nM)2.
In this work, we study the effect of water samples of different hardness, taken from water wells, on the sameporphyrins and the PDI of L. pneumophila (Philadelphia strain). The source of irradiation is a LED based violet andblue light (395 nm with fluence rate 20 mW/cm2 for microbiology experiments, and 411 nm with 3.5-11 mW/cm2
fluence rate for studies of photophysical properties, respectively). In the case of tested porphyrins, their stability,
spectroscopic properties as well as singlet oxygen production by photodegradation of water soluble 9,10anthracenediyl-bis(methylene)dimalonic acid (ABMDMA) will be described, while the effect on L. pneumophila
will be investigated through MEK determination, photoinactivation tests and the destruction tests of the early andlate biofilms. We will discuss the observed strong impact of water hardness on the PDI of L. pneumophila
Philadelphia-1 and biofilm formation and growth.
References
1) J. O. Falkinhan, E. D. Hilborn, M. J. Arudino, A. Puden and M. A. Edwards, Epidemiology and Ecology ofOpportunistic Premise Plumbing Pathogens: Legionella pneumophila, Mycobacterium avium andPseudomonas aeruginosa. Enviromental Health Perspectives, August 2015.
2) A. Lesar, M. Mušković, G. Begić, M. Lončarić, D. Tomić Linšak, N. Malatesti and I. Gobin, Cationic
Porphyrins as Effective Agents in Photodynamic Inactivation of Opportunistic Plumbing Pathogen
Legionella pneumophila. International Journal of Molecular Sciences, July 2020.
Acknowledgements. This work was supported by the University of Rijeka grant uniri-prirod-18-173. The equipmentused in this work is made available thanks to the project “Research Infrastructure for Campus-based Laboratories at
the University of Rijeka”, financed by the European Regional Development Fund (ERDF) and the Ministry of
Science and Education of the Republic of Croatia (RC.2.2.06-0001).

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19th Congress of the European Society
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.13

Biofilm and planktonic cultures of Enterococcus sp. resensitized to antimicrobial withaPDI application
Beata Kruszewska1, Agata Woźniak1, Mariusz Grinholc1
University of Gdansk and Medical University of Gdansk, Intercollegiate Faculty of Biotechnology, Laboratory of
Molecular Diagnostics, 80-307 Gdansk

Enterococcus sp. is a human opportunistic pathogen that can cause infection of the urinary tract or persistentendodontic infections. In addition, the presence of multidrug resistant strains poses a serious clinicalchallenge. Besides, E. faecium is classified as a pathogen belonging to the ESKAPE group of microbes thatcaused hospital acquired infections. What is more, these bacteria can create biofilm which could be even
1000 more resistant to antimicrobials than planktonic cultures. It may result from the slow penetration ofsubstances through biofilm layers1. A chance for reuse of antibiotics is antimicrobial Photodynamic
Inactivation (aPDI) which displays huge potential in bacteria eradication and resensitization to antibiotics.
aPDI requires visible light which activates photosensitizer molecules (PS) accumulated or present in close
proximity to microorganisms leading to transfer energy or electron to oxygen molecules. This results in the
production of reactive oxygen species (ROS) which are toxic to cells and may lead to their damage.
The current study was aimed at investigation whether combined therapy employing clinically used
antibiotics and aPDI (involving exogenously administered PS – rose bengal and green light, λmax 515 nm)
demonstrates a synergy. Two clinical isolates of multidrug resistant strains were investigated: E. faecium
EU87 and E. faecalis EU92. Their drug resistance profile and the synergy between aPDI and antibiotics
were characterized with the application of different approaches, i.e. diffusion assays, MIC evaluation, ETEST, checkerboard assay, and postantibiotic effect. All experiments were performed in accordance to the
EUCAST standards. Also, biofilm was formed using approved methodology for flow-biofilm culturing in
a CDC reactor and treated with antibiotics and aPDI. The growth of biofilms and microbial survival after
treatment was visualized using confocal microscopy. To investigate the potential mechanisms of obtainedsynergy the fluorescent probes detecting ROS and membrane permeabilization were applied.
The obtained results confirmed that aPDI demonstrates a significant impact on Enterococcus sp. survival
rate. Synergy was detected for aPDI in combination with gentamycin (GEN), ciprofloxacin (CIP) anddaptomycin (DAP) against EU92. For EU87, synergy was observed with the application of GEN or CIP,
while for aPDI with vancomycin or DAP, the antagonism was observed. Postantibiotic effect test for EU87
demonstrated that this isolate exposed to aPDI in combination with GEN, streptomycin (STR), tigecycline,
doxycycline, or DAP exhibits delayed growth in comparison to monotherapies. Combined treatment with
STR and CIP also reduced bacterial load in biofilms by approx. 3-4 log10 CFU/cm2 what could be visualized
on microscopic images. In addition, the obtained results indicated that the combined treatment contributed
to the increased ROS in the case of CIP and aPDI. Induced increase in membrane permeabilization may
explain the observed synergy. All results confirm the effectiveness of aPDI in sensitizing Enterococcus sp.
to antimicrobials what has an amazing potential in treating infection caused by multidrug resistant strains.2
References
1) Cieplik, F.; Deng, D.; Crielaard, W.; Buchalla, W.; Hellwig, E.; Al-Ahmad, A.; Maisch, T. Antimicrobial
photodynamic therapy—What we know and what we don’t. Crit. Rev. Microbiol. 2018, 7828, 1–19.
2) Wozniak, A.; Kruszewska, B.; Pieranski, M.K.; Rychłowski, M.; Grinholc, M. Antimicrobial Photodynamic
Inactivation Affects the Antibiotic Susceptibility of Enterococcus spp. Clinical Isolates in Biofilm and
Planktonic Cultures. Biomolecules 2021, 11, 693.
Acknowledgements. This research was supported by National Science Centre 2015/19/B/NZ7/02487
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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.14

Biofilm and planktonic cultures of Enterococcus sp. resensitized to antimicrobial withaPDI application
Beata Kruszewska1, Agata Woźniak1, Mariusz Grinholc1
University of Gdansk and Medical University of Gdansk, Intercollegiate Faculty of Biotechnology, Laboratory of
Molecular Diagnostics, 80-307 Gdansk

Enterococcus sp. is a human opportunistic pathogen that can cause infection of the urinary tract or persistentendodontic infections. In addition, the presence of multidrug resistant strains poses a serious clinicalchallenge. Besides, E. faecium is classified as a pathogen belonging to the ESKAPE group of microbes thatcaused hospital acquired infections. What is more, these bacteria can create biofilm which could be even
1000 more resistant to antimicrobials than planktonic cultures. It may result from the slow penetration ofsubstances through biofilm layers1. A chance for reuse of antibiotics is antimicrobial Photodynamic
Inactivation (aPDI) which displays huge potential in bacteria eradication and resensitization to antibiotics.
aPDI requires visible light which activates photosensitizer molecules (PS) accumulated or present in close
proximity to microorganisms leading to transfer energy or electron to oxygen molecules. This results in the
production of reactive oxygen species (ROS) which are toxic to cells and may lead to their damage.
The current study was aimed at investigation whether combined therapy employing clinically used
antibiotics and aPDI (involving exogenously administered PS – rose bengal and green light, λmax 515 nm)
demonstrates a synergy. Two clinical isolates of multidrug resistant strains were investigated: E. faecium
EU87 and E. faecalis EU92. Their drug resistance profile and the synergy between aPDI and antibiotics
were characterized with the application of different approaches, i.e. diffusion assays, MIC evaluation, ETEST, checkerboard assay, and postantibiotic effect. All experiments were performed in accordance to the
EUCAST standards. Also, biofilm was formed using approved methodology for flow-biofilm culturing in
a CDC reactor and treated with antibiotics and aPDI. The growth of biofilms and microbial survival after
treatment was visualized using confocal microscopy. To investigate the potential mechanisms of obtainedsynergy the fluorescent probes detecting ROS and membrane permeabilization were applied.
The obtained results confirmed that aPDI demonstrates a significant impact on Enterococcus sp. survival
rate. Synergy was detected for aPDI in combination with gentamycin (GEN), ciprofloxacin (CIP) anddaptomycin (DAP) against EU92. For EU87, synergy was observed with the application of GEN or CIP,
while for aPDI with vancomycin or DAP, the antagonism was observed. Postantibiotic effect test for EU87
demonstrated that this isolate exposed to aPDI in combination with GEN, streptomycin (STR), tigecycline,
doxycycline, or DAP exhibits delayed growth in comparison to monotherapies. Combined treatment with
STR and CIP also reduced bacterial load in biofilms by approx. 3-4 log10 CFU/cm2 what could be visualized
on microscopic images. In addition, the obtained results indicated that the combined treatment contributed
to the increased ROS in the case of CIP and aPDI. Induced increase in membrane permeabilization may
explain the observed synergy. All results confirm the effectiveness of aPDI in sensitizing Enterococcus sp.
to antimicrobials what has an amazing potential in treating infection caused by multidrug resistant strains.2
References
1) Cieplik, F.; Deng, D.; Crielaard, W.; Buchalla, W.; Hellwig, E.; Al-Ahmad, A.; Maisch, T. Antimicrobial
photodynamic therapy—What we know and what we don’t. Crit. Rev. Microbiol. 2018, 7828, 1–19.
2) Wozniak, A.; Kruszewska, B.; Pieranski, M.K.; Rychłowski, M.; Grinholc, M. Antimicrobial Photodynamic
Inactivation Affects the Antibiotic Susceptibility of Enterococcus spp. Clinical Isolates in Biofilm and
Planktonic Cultures. Biomolecules 2021, 11, 693.
Acknowledgements. This research was supported by National Science Centre 2015/19/B/NZ7/02487
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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.15

Antimicrobial photodynamic inactivation against staphylococcal enterotoxin A – invitro and ex vivo studies
Patrycja Ogonowska1, Adrian Kobiela2, Danuta Gutowska-Owsiak2,3, Joanna Nakonieczna1
1

Laboratory of Molecular Diagnostics, Intercollegiate Faculty of Biotechnology of University of Gdansk andMedical University of Gdansk, Gdansk, Poland
2
Experimental and Translational Immunology Group, Intercollegiate Faculty of Biotechnology of University ofGdansk and Medical University of Gdansk, Gdansk, Poland
3
MRC Human Immunology Unit, Radcliffe Department of Medicine, MRC Weatherall Institute of Molecular
Medicine, University of Oxford, United Kingdom
Staphylococcal enterotoxin A (SEA) is a virulence factor produced by Staphylococcus aureus. SEA ismainly involved in staphylococcal food poisoning and, along with other staphylococcal enterotoxins (SEs), plays arole as an exacerbation factor for atopic dermatitis 1,2,3. In the treatment of atopic dermatitis eradication of S. aureus
and restoration of the natural bacterial diversity in the skin is crucial4. Moreover, SEs have superantigenic
properties5. All these features constitute a huge therapeutic challenge. Therefore, in our research we propose
antimicrobial photodynamic inactivation (aPDI) as the effective tool to combat both bacterial cells and virulence
factors.
The main objectives of the presented studies were (i) to assess the in vitro efficacy of aPDI against S.
aureus clinical isolates and (ii) to verify the impact of aPDI on the production of staphylococcal enterotoxin A at
the gene and protein levels (in vitro and ex vivo experimental models).
In the aPDI studies, a combination of rose bengal (RB) and green light (λmax=515 nm, irradiance 35
mW/cm2) was used for all clinical S. aureus strains tested. An MTT assay was performed on human keratinocytes(HaCaT cells) to verify the cyto- and phototoxic effects of aPDI on eukaryotic cells. The expression profile of sea
gene under sub-lethal aPDI conditions was examined using the qPCR technique. To confirm the observed resultson the protein level, the Western Blot technique and the proliferation assay were used. To verify our observations
from in vitro experiments, the analysis at the gene and protein levels was performed on a porcine skin model.
This study indicated high efficacy (5-6 log10 reduction in bacterial survival) of aPDI against planktonic S.
aureus clinical isolates, carrying sea gene. The aPDI conditions tested showed no cyto- and phototoxic effect oneukaryotic cells. The results obtained indicated decrease in the level of sea gene upon aPDI conditions (a 2.8 log2
unit decrease after 20 min of irradiation and 2.48 log2 unit decrease after 40 min). Western Blot analysis did notreveal any significant changes in the level of the protein. However, a proliferation assay proved that under aPDI
treatment SEA loses its biological function. Our preliminary ex vivo studies demonstrated the ability to monitor thepresence of SEA protein at various time points after aPDI.
The presented studies indicate that aPDI can effectively reduce the level of virulence factors and deprive
them of their biological functions.
References
1) Kadariya J., Smith T.C., Staphylococcus aureus and Staphylococcal Food-Borne Disease: An Ongoing
Challenge in Public Health. Biomed. Res. Int., 2014.
2) Bunikowski R., Mielke M., Evidence for a disease-promoting effect on Staphylococcus aureus-derived
exotoxins in atopic dermatitis. J. Allergy Clin. Immunol., 2000.
3) Taskapan M., Kumar P., Role of staphylococcal superantigens in atopic dermatitis: from colonization to
inflammation. Ann. Allergy Asthma Immunol., 2000.
4) Hendricks A.J., Mills B. Skin bacterial transplant in atopic dermatitis: knowns, unknowns and emerging
trends. J. Dermatol. Sco, 2019.
5) Spaulding A., Salgado-Pabón W., Staphylococcal and streptococcal superantigen exotoxins. Clin.
Microbiol. Rev., 2013.
Acknowledgements. This work was supported by National Science Centre grant no. 2017/27/B/NZ7/02323 (JN).
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.16

Protein based targeted delivery systems for antimicrobial PDT
Andrea Mussini1, Eleonora Uriati1,2, Pietro Delcanale1, Paolo Bianchini2,3, Alberto Diaspro2,3,4, Stefania
Abbruzzetti1, Cristiano Viappiani1, Cormac Hally5, Montserrat Agut5 and Santi Nonell51) Dipartimento di Scienze Matematiche, Fisiche e Informatiche, Università di Parma, Italy; 2) Nanoscopy, Istituto
Italiano di Tecnologia, Genoa, Italy; 3) Nikon Imaging Center, Istituto Italiano di Tecnologia, Genoa, Italy; 4)
Department of Physics, University of Genoa, Genoa, Italy; 5) Institut Químic de Sarrià, Universitat Ramon Llull,
Spain
Water soluble proteins are natural candidates as biocompatible delivery systems for photosensitizers. We report thedevelopment of a supramolecular structure endowed with photosensitizing properties and targeting capability for
antimicrobial photodynamic inactivation. Our synthetic strategy uses the tetrameric bacterial protein streptavidin,labelled with eosin, as the main building block. Biotinylated immunoglobulin G (IgG) from human serum, known
to associate with S. aureus protein A, was bound to the complex streptavidin-eosin. The supramolecular complex
was used to demonstrate photoinactivation of S. aureus suspensions. FCS and STED nanoscopy experiments
confirmed binding of the complex to S. aureus. Efficient photoinactivation is observed for S. aureus suspensions
treated with IgG-streptavidin-eosin at concentrations higher than 0.5 M and exposed to green light. The proposed
strategy offers a flexible platform for targeting a variety of molecular species.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.17
The use of curcumin to photoinactivate Trypanozoma cruzi.
T. Buenfil-Chi, D. Ortega-Zambrano and H. Mercado-Uribe
CINVESTAV-Monterrey, Parque PIIT, Apodaca, NL. 66000, Mexico.

Chagas disease is caused by the protozoan Trypanozoma cruzi. This is an endemic pathogen
of Latin America that has developed a significant resistance against the usual treating methods.
Therefore, it has become an important health problem. In this work, we applied the phodotodynamic inactivation procedure on cultures of CL Brener and Ninoa strains in their epimastigote
form, when the parasite is at its fastest growing stage. We investigated the join effect of curcumin and blue light at 9x10-6 M and 16 mW, respectively. We did not observe curcumin toxicity ineither strains. The viability of T. cruzi was immediately evaluated after irradiation at 89 J/cm2 (1hour). We found a 1 log10 reduction for CL Brener, and a little less for the Ninoa strain. However, we would like to remark that there was a delayed effect on the viability of the parasites. Indeed, after two days of the treatment, the mortality was 100%.
Acknowledgements. This work was supported by CONACyT, Mexico, Grant A1-S-8125.

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19th Congress of the European Society
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.18

Photodynamic Inactivation of fungal phytopathogens using Ce6 derivatives andassessment of their phytotoxicity.
Christoph Hamminger1, Michael Glueck1, Michael Fefer2, Jun Liu2, Raimund Tenhaken3, Kristjan Plaetzer1
1) University of Salzburg, Department of Biosciences, Laboratory of Photodynamic Inactivation of
Microorganisms, Salzburg, Austria; 2) Suncor AgroScience, Mississauga Ontario, Canada, 3) University ofSalzburg, Department of Biosciences, Plant Physiology, Salzburg, Austria
An increasing demand for organic and conventionally produced foodstuff necessitates a more sustainable,
biocompatible, and holistic approach to crop protection. This study aims towards adding Photodynamic Inactivation(PDI) as highly biocompatible strategy suitable for crop protection to the farmer’s toolbox. Since the antimicrobial
effectiveness of PDI is not diminished by classical resistance mechanisms of bacterial or fungal pathogens - which
trouble growers all over the globe - we believe this approach has the potential to benefit farmers as well as theecosystem. In this study Alternaria solani and Botrytis cinerea were used as model pathogens. These two fungi are
the cause for considerable crop and monetary shortfalls in the food production industry. Two photoactive compounds,
Sodium-Magnesium-Chlorophyllin (Chl, food additive E140) in combination with Na2EDTA and the chlorin e6
derivative B17-0024 holding cationic moieties were used as photosensitizers. Mycelial spheres with a diameter of 23 mm of A. solani and B. cinerea were incubated with the photosensitizers for one hundred minutes and illuminatedwith 106.6 J/cm² using a LED array with a wavelength of 395 nm. Samples not showing any growth on agar plates
after seven days were counted as dead. One hundred micromolar Chl with 5 mM Na2EDTA successfully
photoinactivated 91.7% of B. cinerea and 94.1% of A. solani. PDI based on B17-0024 thoroughly killed (100 %) A.
solani at concentrations of 10 and 100 µM. Ten micromolar of B17-0024 lead to 66.7 % and 100 µM to complete
eradication of B. cinerea. To screen for possible phytotoxic effects of the PDI treatment, a plant compatibility assay
based on the small woodland strawberry (Fragaria vesca) was performed. The strawberry plants at BBCH1 stage 14
were sprayed once – or three times on consecutive days - with 300 µL of the PS formulation used for PDI. Plant
health and growth was monitored for twenty-one days post PDI treatment. Although Na2EDTA induced minor
damage to the treated leaves, the assay demonstrated that the two photosensitizers do not cause significant damage
and have no adverse effect on plant development. In conclusion, we show that tissue-like fungal structures of A.solani
and B. cinerea can be successfully treated by PDI without causing harm to the host plants. Therefore our results
suggest that Chl / EDTA as well as B17-0024 are effective photofungizides.
References
1) U. Meier, H. Graf, H. Hack, M. Hess, W. Kennel, R. Klose (Eds.), Growth stages of mono-and dicotyledonous
plants, BBCH monograph, Strawberries (2th ed.), Federal Biological Research Centre for Agriculture and Forestry,
Berlin/Braunschweig, Germany (2001)
Acknowledgements. This work was supported by Suncor AgroScience.

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19th Congress of the European Society
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.19

A semi-theoretical action spectrum for antimicrobial photoinactivation in the lungs
Alfonso Dell’Accio1, Chiara Treghini1, Maria Micieli1, Eleonora Riccobono1, Felice Valzano2, Franco Fusi1, Gian
Maria Rossolini3, Lucia Pallecchi2, Giovanni Romano1, Light4Lungs group4
1
University of Florence, Department of Clinical and Experimental Biomedical Sciences “Mario Serio”, Florence,
Italy; 2 University of Siena, Department of Medical Biotechnologies, Siena, Italy;3 University of Florence,
Department of Experimental and Clinical Medicine, Florence, Italy; 4S. Nonell, Institut Quimic de Sarria,
Barcelona, Spain; G. Romano, University of Florence, Florence, Italy; N. Henry, Sorbonne Université, Paris,
France; E. Palomares, Fundacio Privada Institut Catala D’Investigacio Quimica, Tarragona, Spain; E. Reddi,
University of Padova, Padova, Italy; G. Taylor, Cardiff Scintigraphics Ltd, Cardiff, UK; A. Kadioglu, The
University of Liverpool, Liverpool, UK
In the framework of the search for innovative solutions to overcome antibiotic-resistance, the European project“Light4Lungs” aims at synthesizing and testing an inhalable light source to control lung infections associated with
biofilm-forming and multidrug-resistant bacteria such as P. aeruginosa and S. aureus, with notable applications in
cystic fibrosis (CF). In this context, the project proposes an antimicrobial photoinactivation (aPDI) approach, taking
advantage of the presence of endogenous photosensitizers (porphyrins) in the considered bacterial species.
The choice of the most suitable wavelength derives from the definition and calculation of the photo-killing action
spectrum in the lungs, considering the case of biofilm-associated infections caused by P. aeruginosa and S. aureus in
CF patients. This was obtained by applying a semi-theoretical modelling with Monte Carlo simulations to the infectedlung regions, according to optical theory [1] and previously published methodology related to stomach infection
phototherapy [2]. The variability of in vivo pathologic conditions [3] was accounted for by a careful literature
analysis, relative to the variation in the concentration of oxygen, bacteria and relative endogenous porphyrins, besides
the presence of other relevant absorbers/diffusers inside the biofilm/mucous layer. The obtained semi-theoreticalaction spectrum is peaked at 394 nm and mostly follows porphyrins extinction coefficient behaviour.
To confront these results with an experimental validation, we started by performing in vitro irradiation on preformed
biofilms grown on a standardized static model (MBEC Assay), considering one reference strain and one CF clinicalisolate for both P. aeruginosa and S. aureus. Biofilms were irradiated with representative wavelengths, considering
first LED sources centred in the violet region (415nm) and in the green (525nm). Light at 415nm showed a dosedependent and strain-dependent antimicrobial effect, with reduction of at least 2 log CFU at 60 J/cm2 in three of thefour tested strains. Light at 525 nm showed a scant or no effect, as expected. Additional wavelengths will beconsidered to explore the relative efficacy of the porphyrin secondary absorption peaks and merge data with themucous and biofilm absorption/scattering properties.
The obtained results can offer important indications for the synthesis of the aerosolized light source and definition ofits most effective emission spectrum, suggesting also a flexible platform for dosimetry calculation to be consideredin further applications.
References
1) Jacques, S. L., Simple optical theory for light dosimetry during PDT. Proc. SPIE 1645, 155–165, 1992.
2) Gnerucci, A., Faraoni, P. et al., Influence of stomach mucosa tissue on the efficacy of intragastric antibacterial
PDT. Photochem. Photobiol. Sci., 19, 34–39, 2020.
3) Cowley, E. S., Kopf, S. H. et al. Pediatric cystic fibrosis sputum can be chemically dynamic, anoxic, and extremely
reduced due to hydrogen sulfide formation. MBio, 6, 1–15, 2015.
Acknowledgements. This work was supported by the H2020 research and innovation programme under grant
agreement n.863102. This poster was displayed on behalf of the Light4Lungs Consortium.

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19th Congress of the European Society
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.20

Porphyrin-based light activated antimicrobial coatings prepared by spin coating
Melania Rogowska1, Martin Tran1, Ellen Bruzell2, Håkon Valen2, Ola Nilsen1
1) Centre for Materials Science and Nanotechnology (SMN), Department of Chemistry, University of Oslo, Oslo,
Norway; 2) Nordic Institute of Dental Materials, Oslo, Norway
Porphyrins are attractive functional materials commonly used as visible light absorbing photosensitizers inphotodynamic therapy (PDT). Furthermore, different transition metals can be introduced into the porphyrin ring tocreate metalloporphyrin complexes with various physiochemical properties.1 In this work, novel light-activatedantimicrobial thin films were prepared based on biocompatible zirconium (Zr) and tetrakis(4carboxyphenyl)porphyrin (TCPP) using spin coating deposition technique. The physicochemical properties of the
ZrTCPP films were characterized by UV-Vis and Fourier transform infrared (FTIR) spectroscopy, X-ray diffraction(XRD), atomic force microscopy (AFM) and contact angle measurements. The antibacterial activity of the films wasinvestigated using the gram-positive bacteria Enterococcus faecalis, both against newly adhered bacteria and after
biofilm formation on the surfaces. The photodynamic inactivation was induced after exposure of active films
combined with blue light (λ: 400 – 520 nm) using light doses of 5.7, 11.4 and 22.8 J/cm2. This study shows highinhibitory activity of the ZrTCPP complexes in the presence of light, however, the films were also active against
bacteria in the absence of light pointing to cytotoxicity of the Zr-metalloporphyrin complex.

Fig. 1 - Structure of tetrakis(4-carboxyphenyl)porphyrin (TCPP) used in in this work and UV-Vis absorption spectra of the
ZrTCPP thin film and the corresponding TCPP dissolved in ethanol.

References
1) Stojiljkovic, I., Evavold, B. D., Kumar, V. Antimicrobial properties of porphyrins. Exp. Opin. Invest. Drugs, 10(2),309-320 (2001).
Acknowledgements. This project has received funding from the European Union’s Horizon 2020 research andinnovation programme under the Marie Skłodowska-Curie grant agreement No. 765378.
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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.21
Decolonization of human skin by application of a photodynamically active hydrogel
Daniel B. Eckl1, Anja K. Hoffmann1, Nicole Landgraf1, Anja Eichner2, Harald Huber1, Wolfgang Bäumler2
1) Universität Regensburg, Lehrstuhl für Mikrobiologie, Regensburg, Germany; 2) Universitätsklinikum
Regensburg, Klinik und Poliklinik für Dermatologie, Regensburg, Germany,
Skin colonization with pathogenic, even multi-resistant bacteria is a potential source of infections, especially in
hospital-associated environments. It has been shown previously that photodynamic inactivation (PDI) of bacteria is
a promising, new approach for the decolonization of the human skin. In a preliminary study it was shown that PDI
experiments already provided a decontamination efficacy on human ex vivo skin of 99.9% using an aqueous
photosensitizer solution1. Furthermore, the application of aqueous photosensitizer solutions did not lead todifferences in mitochondrial survival compared to the untreated control. In a follow up study, we aimed to improvethe decolonization efficacy as well as the applicability in practice by applying a photodynamically active hydrogel.
UV-VIS spectroscopy was applied to investigate the chemical stability of photosensitizers in the hydrogel suspension.
Initial tests of the photodynamically active hydrogels were performed on inanimate surfaces with bacterial cells
suspended in different ionic solutions such as an artificial sweat solution in order to evaluate the basic parameters for
efficient photodynamic inactivation on human skin. Fresh ex vivo human skin was inoculated with bacterial
suspensions of methicillin-resistant Staphylococcus aureus (MRSA). After visible drying of bacteria, differenthydrogels containing different photosensitizers were added to the inoculated skin surface. Subsequently, the skinsurface was irradiated with visible blue light at different radiant exposures (J/cm²). Then, bacteria were recoveredfrom skin with a swap and plated via drop-plate method on Mueller Hinton Agar. Colony count unfold reduction
efficacy when referenced to internal controls. After recovery, skin biopsies were taken and nitro blue tetrazolium(NBT) staining was applied to check viability of skin cells via mitochondrial activity. TUNEL stain revealed thepresence of apoptotic skin cells.
Firstly, spectroscopy results suggest that the hydrogel did not affect the chemical structure of photosensitizers.
Artificial experiments on inanimate surfaces showed that the hydrogel is capable of highly efficient bacterial
reduction even in presence of an artificial sweat solution. The microbiology results of ex vivo human skin showed an
improved MRSA reduction on skin of around 99.99% as compared to internal references. NBT staining of the tissue
revealed no lack in mitochondrial activity of the human cells due to the application of the photodynamically activehydrogel. Lastly, TUNEL stained histological sections did not show noteworthy apoptotic effects after thephotodynamic treatment.
The data indicate that PDI represents an easy, quick and efficient method to decolonize human skin from MRSA.
The results of the study are remarkable, as substances on skin like inorganic ions or small organic molecules such as
histidine are capable to inhibit the photodynamic efficacy but seem to interfere less in a hydrogel environment. Since
PDI is known to eradicate a large set of pathogens independent of their antibiotic resistance profiles, this newtechnology should be tested in clinical studies in the near future.

References
1) M. Schreiner, W. Bäumler, D. Eckl, A. Späth, B. König, A. Eichner, MRSA decolonization of human skin via
photodynamic treatment, British Journal of Dermatology 179, 1358-1367, 2018
Acknowledgements. This work was supported by the German Research Foundation DFG (GEPRIS project number
415812443)

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19th Congress of the European Society
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.22

Antimicrobial photosensitizers and their formulations: A potential solution to
current world scenario
Bhavya Khurana1, 2, Tan Ouk1, Mathias O. Senge2, 3, Vincent Sol1
1.
Université de Limoges, Laboratoire PEIRENE.
2.
School of Chemistry, Trinity Biomedical Sciences Institute, Trinity College Dublin, The University of
Dublin.
3.
Medicinal Chemistry, Trinity Translational Medicine Institute, Trinity Centre for Health Sciences, Trinity
College Dublin, The University of Dublin
In order to provide a long-lasting solution to infections affecting the current world scenario, photodynamic therapy(PDT) offers a means to destroy pathogenic microbes via formation of reactive oxygen species, promoting thedamage of microbial targets such as nucleic acids (DNA or RNA), proteins, lipids, protein complexes, or by impeding
the biofilm matrix.1
Thus, the main aim of the study is to design and synthesize photoactive moieties based on porphyrin and chlorin
macrocycles and BODIPY dyes for antimicrobial photodynamic therapy (aPDT).2 Furthermore, incorporating thesephoto-moieties into biopolymeric hydrogelsfor a variety of biomedical applications are targeted.
Porphyrin based cationic photosensitizers (PSs) were synthesized and a chlorin based PS was extracted from
Spirulina maxima and modified to be included on a biopolymeric hyaluronic acid hydrogel platform. This platform
was characterized spectroscopically and evaluated for antimicrobial photoactivity via microbial evaluation ondifferent gram strains of bacterial species. Singlet oxygen production was determined as well to evaluate thephotoactivity of this polymeric hydrogel platform.
Furthermore, BODIPY dye-based PS species have been synthesized and modified for their activity as aPDT agents.
Several N-heterocyclic BODIPY-dyes have been positively charged or functionalized for incorporation into hydrogel
platforms. These dyes exhibit good water solubility. A library of such N-heterocyclic BODIPY dyes was preparedand characterized and will be evaluated for photoactivity against microbes.
References
1.
W.C. de Melo, P. Avci, M.N. de Oliveira, A. Gupta, D. Vecchio, M. Sadasivam, R. Chandran, Y.Y. Huang,
R. Yin, L.R. Perussi, G.P. Tegos, J.R. Perussi, T. Dai, M.R. Hamblin, Photodynamic inactivation of biofilm: taking
a lightly colored approach to stubborn infection, Expert Rev Anti Infect Ther , 2013, 11, 669-693.
2.
A. Wiehe, J.M. O'Brien, M.O. Senge, Trends and targets in antiviral phototherapy, Photochem. Photobiol.
Sci., 2019, 18, 2565-2612.
Acknowledgements. This work was supported by grant 42 from the Ministry of Truth.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.3.23

Photodynamic Antimicrobial Therapy: Chemical kinetic modeling to improvetreatment efficacy
Jeffrey C. Peterson1,2, Yu-Cherng Chang1,2, Irene Kochevar3, Fabrice Manns1,2, Jean-Marie Parel1,2,4
1) University of Miami Miller School of Medicine, Ophthalmic Biophysics Center, Department of Ophthalmology,
Bascom Palmer Eye Institute, Miami, FL, USA;
2) University of Miami College of Engineering, Department of Biomedical Engineering, University of Miami,
Coral Gables, FL, USA;
3) Harvard Medical School, Wellman Center for Photomedicine, Massachusetts General Hospital, Boston, MA,
USA;4) University of New South Wales, Brien Holden Vision Center, Sydney, NSW, Australia.
Purpose: Rose Bengal Photodynamic Antimicrobial Therapy (RB-PDAT) has been shown to effectively treat
antimicrobial resistant and atypical infectious keratitis.1–3 RB-PDAT works by generating antimicrobial singlet
oxygen (1O2) by exciting Rose Bengal (RB) photosensitizer with 525 nm light. The cumulative 1O2 dose
distribution generated during RB-PDAT is currently unknown, this project aims to create a predictive model tocalculate the 1O2 distribution under clinical conditions and will enable optimization of 1O2 dose by optimizing RBPDAT treatment parameters.
Methods: A chemical kinetics model previously developed (Wang et al., 2010) for 1O2 dose in tumors treated withphotodynamic therapy was used as the basis for this model.4 To describe ground-state oxygen (O2) supply specific tothe cornea, the oxygen supply term was adapted to include the model described by Larrea & Büchler and Castillo etal.5,6 RB distribution was modeled using Fick’s second law, and 525 nm excitation light distribution by timedependent application of Beer’s law. Rate constants for RB diffusion were derived from experimental data describing
RB corneal penetrance.7–9 Minimal lateral variation of all species was assumed to allow for one-dimensionalmodeling. The resulting system of partial differential equations was solved numerically in MATLAB. Cumulative
1
O2 dose was calculated for RB-PDAT during typical conditions (6 mW/cm2, 5.4 J/cm2, 0.1% RB), for pulsed andincreased light dose, and for addition of 90% supplemental O2.

Fig. 1 – (a) Calculated corneal O2 concentration as it varies throughout typical RB-PDAT application (0.1% RB applied for 30
min at 6 mW/cm2 for 15 min, 5.4 J/cm2). (b) Same graph as (a) zoomed in to 0-50 µm.

Results: Under typical RB-PDAT conditions, calculated O2 decreases to approximately zero beyond 10 μm depth in
the first 1 min of treatment. In this scenario, 1O2 dose distribution was calculated to be limited to <10 μm corneal
depth. Supplemental O2 increases 1O2 dose depth by approximately 5–10 μm, with similar results observed with 1s
on, 1s off pulsing with doubled fluence rate. Continuous wave, doubled fluence rate for half the treatment time (i.e.
6 mW/cm2 for 15 min vs. 12 mW/cm2 for 7.5 min) decreases calculated 1O2 depth for all treatment conditions.
Increasing RB concentration increases calculated cumulative 1O2 dose approximately linearly. Increasing light dosefrom 5.4 J/cm2 to 10 J/cm2 increases calculated 1O2 surface dose 1.4 to 1.7x.

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Fig. 2 – (a) Calculated cumulative 1O2 concentration with varying applied RB concentration, fluence rate, and continuous wave
light vs. 1s on 1s off pulsing (b) Calculated cumulative 1O2 concentration with varying total light energy, fluence rate, continuous
wave light vs. 1s on 1s off pulsing, or providing 90% supplemental O2.

Conclusions: We demonstrate a proof-of-concept predictive model for 1O2 dose generated during RB-PDAT,
capable of testing a range of experimental parameters, which can be used for optimizing RB-PDAT efficacy.
Follow-up studies, such as direct in vivo measurement of O2 during RB-PDAT, will be necessary to validate themodel predictions.
References
1) Naranjo A, Arboleda A, Martinez JD, et al. Rose Bengal Photodynamic Antimicrobial Therapy for patients with
progressive infectious keratitis: A pilot clinical study. American Journal of Ophthalmology. 2019;208:387-396.
2) Amescua G, Arboleda A, Nikpoor N, et al. Rose Bengal Photodynamic Antimicrobial Therapy: A novel
treatment for resistant Fusarium Keratitis. Cornea. 2017;36(9):1141-1144.
3) Arboleda A, Miller D, Cabot F, et al. Assessment of Rose Bengal versus riboflavin photodynamic therapy forinhibition of fungal keratitis isolates. American Journal of Ophthalmology. 2014;158(1):64-70.e2.
4) Wang KK-H, Finlay JC, Busch TM, Hahn SM, Zhu TC. Explicit dosimetry for photodynamic therapy:
macroscopic singlet oxygen modeling. Journal of biophotonics. 2010;3(5-6):304-318.
5) Larrea X, Büchler P. A transient diffusion model of the cornea for the assessment of oxygen diffusivity andconsumption. Investigative Ophthalmology & Visual Science. 2009;50(3):1076-1080.
6) Del Castillo LF, da Silva ARF, Hernández SI, et al. Diffusion and Monod kinetics model to determine in vivo
human corneal oxygen-consumption rate during soft contact lens wear. Journal of Optometry. 2015;8(1):12-18.
7) Naranjo A, Pelaez D, Arrieta E, et al. Cellular and molecular assessment of Rose Bengal photodynamic
antimicrobial therapy on keratocytes, corneal endothelium and limbal stem cell niche. Experimental Eye
Research. Published online September 2019:107808.
8) Peterson JC, Naranjo A, Martinez JD, et al. Rose Bengal depth in human donor cornea after Rose Bengal
Photodynamic Antimicrobial Therapy. Presented at the: American Society for Photobiology 2018 Biennial
Meeting;
May
12,
2018;
Tampa,
FL,
USA.
https://photobiology.org/2018minisite/ASP%.
202018%20Schedule.pdf
9) Wertheimer CM, Elhardt C, Kaminsky SM, et al. Enhancing Rose Bengal-photosensitized protein crosslinking
in the cornea. Investigative ophthalmology & visual science. 2019;60(6):1845-1852
Acknowledgements. This research was supported in part by the Robson Foundation, the Florida Lions Eye Bank
and Beauty of Sight Foundation; Gifts from Drs. HW Flynn Jr, KR Olsen, ME Hildebrandt, R Urs and A Furtado;
NIH Center Grant P30EY14801; unrestricted funds from Research to Prevent Blindness to the department ofOphthalmology; the Henri and Flore Lesieur Foundation (J.-M. Parel). Jeffrey Peterson was supported in part by NIH
Medical Scientist Training Program T32 GM112601-04. Provisional patent #39756179 filed 6/18/2020 for 1O2
measurement system for RB-PDAT, the University of Miami has all rights.
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P-1.3.24

Breaking the rebellion: Photodynamic Inactivation of Erwina amylovora resistant to
streptomycin
Annette Wimmer1, Michael Glueck1, Michael Fefer2, Jun Liu2, Kristjan Plaetzer1
1) University of Salzburg, Department of Biosciences, Laboratory of Photodynamic Inactivation of
Microorganisms, Salzburg, Austria; 2) Suncor AgroScience, Mississauga Ontario, Canada
Global crop production plays a key role in food security. The ever-increasing spread of plant pathogens causes highlosses in agriculture. Erwinia amylovora, the causal agent of the contagious fire blight disease, has already evolved
resistance to the antibiotic streptomycin (Sm) as conventional treatment strategy. Photodynamic Inactivation (PDI)
of microorganisms has recently been introduced as innovative method for plant protection. PDI is based on theprinciple that a per se non-toxic photosensitive substance is activated by visible light, by this forming reactive oxygen
species, which in turn directly induce phototoxicity in target cells. The aim of the study was to prove whether
E. amylovora resistant to Sm (E. amylovora SmR) can be killed by PDI. Considering the field application, naturalderived sodium magnesium chlorophyllin (Chl) combined with 1.2 % polyaspartic acid (PASA) or B17-0024, a
synthethic compound carrying cationic moieties, were used as eco-friendly photosensitizers (PS). In vitro
experiments were performed with the respective PS in concentrations of 1, 10 and 100 µM and five as well as
30 minutes incubation in the dark, followed by illumination at 395 nm with a radiant exposure of 26.6 J/cm2. To
investigate additional effects of Sm in media with and without this antibiotic were used. Efficiency testing wasevaluated by counting of colony forming units (CFU). Our results show that E. amylovora SmR can be successfully
treated with PDI. The highest inactivation of 7 log steps (equivalent to a 99.99999% reduction of viable bacteria)
was induced at 100 µM B17-0024 and 30 minutes incubation. Chlorophyllin in combination with 1.2 % PASA
reduced the number of bacteria by 6 log steps. Our results are in line with a previous study performed by Glueck etal.1, where the susceptible strain of E. amylovora was successfully photoinactivated. As conclusion this study proves
principle that PDI can be used to treat plant diseases even if the causing bacteria are resistant to conventional
treatment.
References
1) Glueck, M., C. Hamminger, M. Fefer, J. Liu and K. Plaetzer, Save the crop: Photodynamic Inactivation of plant
pathogens I: bacteria. Photochemical & Photobiological Sciences 18(7): 1700-1708, 2019.
Acknowledgements.
The authors are grateful to Suncor AgroScience, Mississauga Ontario, Canada for providing B17-0024.

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P-1.3.25

Targeted photodynamic approach: the case study of Ga3+ meso-PPIX and hemetransporters.
Michalska Klaudia1, Nakonieczna Joanna1
1) Intercollegiate Faculty of Biotechnology, University of Gdańsk and Medical University of Gdańsk, Departmentof Biotechnology, Laboratory of Molecular Diagnostics, Gdańsk, Poland
Introduction: The efficiency of antimicrobial photodynamic treatment (aPDI) may dependent on the uptake of thephotosenstizier (PS). Photosensitizers can be accumulated in various ways, depending on: the structure of bacterial
cell, environmental factors and the mechanism involved, e.g. active transport. The concept of a targeted
photodynamic approach is based on the uptake by molecular targets such as membrane receptors that recognize PS
as a natural ligand. One of the proposed compounds for the targeted PS are metal conjugated porphyrins(metalloporphyrin, MP). Gallium protoporphyrin IX conjugates such as Ga3+PPIX or meso-substituted Ga3+ mesoPPIX (GaMPPIX), mimic the binding between the heme structure (Fe3+protoporphyrin IX) and their acquisition
machinery. The heme can be acquired intracellularly under iron deficiency conditions by two systems oftransmembrane protein cascades: Isd (iron-regulated surface determinant) and Hts (heme transport system).
Paradoxically, heme might elaborate the toxicity at higher concentrations. The HrtAB (heme-regulated transporter)
two-component detoxification machinery senses and pumps an overdose of the compound out of the cell. The aim of
this study was to investigate the effect of heme or impairment of the heme acquisition and detoxification machineryon phototreatment with GaMPPIX.
Materials and methods: In this study, Staphyloccocus aureus ATCC 25923 cells were photodynamically treated invarious proportions of the mixture [heme and photosensitizer]. The conditions for aPDI were as follows: GaMPPIX
and subsequent green LED irradiation λmax= 522 nm, 10.6 mW/cm2. Bacterial viability was measured by serial
dilution of the cells and counting of the colonies plated on agar plates after treatment (CFU/mL). Similarly, S. aureus
Newman and its isogenic mutants (ΔHrtA, ΔIsdD and ΔHtsA) were subjected to aPDI under green light irradiationin the presence and absence of iron in the medium. Accordingly, accumulation of GaMPPIX was measured. Briefly,
overnight bacterial cultures were diluted and incubated with photosensitizer for 10 minutes at 37°C with shaking.
Then, washed twice with PBS buffer and suspended in 0.1M NaOH/1% SDS solution. After 24 hours of incubation,
the fluorescence of lysate was measured and the quantity of accumulated molecules of PS per cell were calculated.
Results: Post-treatment cell count analysis showed that the combined GaMPPIX-mediated aPDI and hemin action
decreased photodynamic efficiency with a competitive effect (10x and 5x higher hemin concentrations over
GaMPPIX resulted in an increase of S. aureus survival from 2 to 4 log10 units). This suggests that GaMPPIX is a
substrate for the studied heme transport system HrtA. The photoinactivation results were also confirmed by lower
intracellular accumulation of GaMPPIX in cells after addition of 10 µM of heme (calculated as an 18,3% reductionin accumulation of GaMPPIX). Impairment in HrtA ATPase resulted in the most sensitive phenotype in aPDI. The
sensitivity of ΔHrtA to aPDI was correlated with higher accumulation of GaMPPIX compared to the wild-type strain.
Acknowledgements. This work was supported by Small Grants Program for early career scientist funded by
University of Gdańsk – Ugrants-Start no. 533-0C30-GS31-21 (KM)

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P-1.3.26

Histopathological Study of the Zoonotic Anisakis Parasite Treated with aPDT as aControl Approach
Paula Ramos1, Ana Grade1, Maria G. P. S. Neves2, Adelaide Almeida3, Maria A. F Faustino2
1

Aquaculture, Valorisation and Bioprospective Division, Department of Sea and Marine Resources, Portuguese
Institute for Sea and Atmosphere, Av. Doutor Alfredo Magalhães Ramalho, nº6, 1495-165 Lisboa, Portugal;
2
Department of Chemistry and LAQV-REQUIMTE, University of Aveiro, 3810-193 Aveiro, Portugal; 3Departmentof Biology and CESAM, University of Aveiro, 3810-193 Aveiro, Portugal.
The presence of Anisakis larvae in fishery products jeopardizes their safety and quality, which is a potential healthrisk to consumers, being also a matter of concern for official authorities. The use of photodynamic action can beconsidered a potential approach to treat this zoonotic parasite in fresh sea products.1 Thus, several experiments werecarried out to assess the efficacy of antimicrobial photodynamic therapy (aPDT) to inactivate the Anisakis larvae
collected from Atlantic horse mackerel. Experiments with different photodynamic treatment time exposure (2, 8 or24 h), light irradiance (4.0, 14 and 67 mW/cm2) and photosensitizer concentrations (5.0, 25 and 50 µM of TMPyP)
took place in microcosm conditions. The viability of the Anisakis larvae was evaluated at 2 h and 4 h after treatment.
and in one case after 24 h. In order to evaluate tissue damage, histopathological study of the larvae was done at theend of the assays. Although not all parasites have been death during photodynamic treatment, larvae sections reveal
anatomopathological changes. In the first third of the larvae, the muscular oesophagus changes its characteristic
triradiate lumen. Additionally, on the posterior two thirds the intestine epithelium, the columnar cells present often
loss of their nuclei and of their shape. It was also observed that some sections of the parasite, in both areas, showed
irregular polymyarian muscle layer. This study aims to highlight the best conditions to inactivate Anisakis larvae andenvisage its application in food industry; and consequently, lead to the reduction of the biological hazard associatedwith their presence in fishery products. In this communication will be presented and discussed the results of thehistopathological study of the zoonotic Anisakis parasite treated with aPDT.
References
1) Preuß, A., Pfitzner, M., & Röder, B. Mosquito larvae control by photodynamic inactivation of their intestinal flora– a proof of principal study on Chaoborus sp. Photochem. Photobiol. Sci., 2019, 18, 2374-2380
Acknowledgements: Thanks are due to University of Aveiro and FCT/MCTES (Fundação para a Ciência eTecnologia/Ministério da Ciência e Tecnologia, Portugal) for the financial support to LAQV-REQUIMTE(UIDB/50006/2020) and Institute for Biomedicine – iBiMED (UID/BIM/04501/2013 and POCI-01-0145-FEDER007628) and also to supported by PREVINE project PTDC/ASP-PES/29576/2017 through national funds and, whenapplicable, co-financed by European Union, FEDER, QREN, and COMPETE within the PT2020 Partnership
Agreement. This work was also supported by I9+Proalga project MAR2020-16-01-03-FMP-0011 and Sanáqua
project MAR-02.05.01-FEAMP-0004.

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P-1.3.27

Synthesis and photophysical evaluation of new porphyrin-azabicyclo conjugates for
PDT applications
Federico Frendo1,2, Carlos J. P. Monteiro1, Ana S. Joaquinito1, Maria G. P. M. S. Neves1, Giorgia Miolo2, Maria A.
F. Faustino1
1) LAQV-REQUIMTE, Department of Chemistry, University of Aveiro, 3810-193 Aveiro, Portugal; 2) Departmentof Pharmaceutical and Pharmacological Sciences, University of Padova, Via Marzolo 5, 35131 Padova, Italy
Photodynamic therapy (PDT) is clinically approved treatment based on the interaction of a photosensitizing dye (PS),light and dioxygen and can find application in different areas, such treatment for oncologic and non-oncologic
diseases, tumour detection, viruses and bacteria inactivation.1 The mechanism of action involves the production,
cytotoxic reactive oxygen species (ROS) which trigger a cascade of reactions that lead to the destruction of targets.
The main advantages of PDT compared with traditional oncological therapies are due to its relatively non-invasivenature, lower systemic toxicity, relatively selective destruction of undesired cells, good cosmetic outcome and its
ability to stimulate the immune system, as well as its good tolerability profile. However, PDT has also some
limitations, and therefore there is significant room for improvement, especially in the search for new and more
efficient PS. Our research group has been exploring the synthetic versatility of 5,10,15,20tetrakis(pentafluorophenyl)porphyrin (TPFFP) that has revealed to be a multipurpose template to be efficiently andselectively derivatized by means of nucleophilic aromatic substitution reactions, devising a plethora of functionalgroups 2, generating libraries of photo-active compounds with biological interest.

In this communication, the functionalization by nucleophilic aromatic substitution of TPFFP, with different
azabicyclic amidines will be reported. The structural characterization, photophysical properties of new compounds
will be presented.
References
1) Hamblin, M.R. Photochem. Photobiol. 2020, 96, 506-516.
2) J. I. T. Costa, A. C. Tomé, M. G. P. M. S. Neves, J. A. S. Cavaleiro, J. Porphyrins Phthalocyanines 2011, 15 (1112), 1116-1133.
Acknowledgements. The authors thank University of Aveiro and FCT/MCT for the financial support provided to
QOPNA research Unit (FCT UID/QUI/00062/2019), the Associate Laboratory CESAM (FCTUID/AMB/50017/2019) and to Project PREVINE - FCT-PTDC/ASP-PES/29576/2017), through national funds (OE)and where applicable co-financed by the FEDER-Operational Thematic Program for Competitiveness andInternationalization-COMPETE 2020, within the PT2020 Partnership Agreement. Thanks are also due to thePortuguese NMR and Mass Networks. Federico Frendo thanks to the Erasmus mobility programme.

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P-1.3.28
Evaluation of Photosensitizer-Containing Superhydrophobic Surfaces for the
Antibacterial Treatment of Periodontal Biofilms
Caroline Coradi Tonon1, Shoaib Ashraf1, Alessandra Nara de Sousa Rastelli2, Goutam Ghosh, Tayyaba
Hasan1,3, QianFeng Xu4, Alexander Greer4,6,7 and Alan M. Lyons4,5,6
1) Wellman Center for Photomedicine, Massachusetts General Hospital and Harvard Medical School, 40 Blossom
St, Boston, MA 02114, United State;2) Department of Restorative Dentistry, School of Dentistry, Araraquara, São Paulo State University-UNESP, 1680
Humaitá St., Araraquara, SP 14801-385, Brazil;
3) Division of Health Sciences and Technology, Harvard University and Massachusetts Institute of Technology,
Cambridge, Massachusetts 02139, United States;
4) SingletO2 Therapeutics LLC, 75 Clinton Street, Staten Island, NY 10304, United States;
5) Department of Chemistry, College of Staten Island, City University of New York, Staten Island, New York 10314,
United States;
6) Ph.D. Program in Chemistry, The Graduate Center of the City University of New York, 365 Fifth Avenue, New
York, New York 10016, United States;
7) Department of Chemistry, Brooklyn College, City University of New York, Brooklyn, New York 11210, UnitedStates.
Antimicrobial photodynamic therapy (aPDT) has emerged as a promising approach to control biofilms involved inperiodontal diseases. A unique superhydrophobic surface (SH) that generates airborne singlet oxygen was fabricatedto potentially overcome the disadvantages of conventional aPDT. The aim of this in vitro study was to investigate
the efficacy of the SH surface against a multi-species periodontopathogenic biofilm model. Biofilms were cultured onhydroxyapatite discs and treated by a SH surface loaded with chlorin e6 in amounts ranging from 96 nmols/cm2 to1100 nmols/cm2 and illuminated with a 660 nm light-emitting diode. Most light incident on the SH surface wasabsorbed by the PS, the amount transmitted to the biofilm ranged from 18 J/cm2 to 89 J/cm2. The killing efficacy wasassessed by counting of colony forming units, biofilm metabolism by XTT and confocal microscopy. The SH-aPDT
showed a significant ability to inhibit both gram-positive and gram-negative bacteria in the multi-species biofilm
model compared to chlorohexidine (CHX) and other controls. The effect of the treatments was dependent on the lightdose and concentration of the embedded photosensitizer. The singlet oxygen (1O2) produced by the SH surface andtransported to the biofilm inhibited both Gram-positive and Gram-negative periodontal bacteria, showing >5-log
reduction of all three species: Porphyromonas gingivalis, Streptococcus mutans and Actinomyces naeslundii. Due tothe absence of physical contact of the PS with the bacteria themselves, the SH device avoids staining, unlike
conventional aPDT and CHX. Moreover, no drug-light interval is needed for SH device (pre-incubation time). The
lifetime of airborne 1O2 is 5000-fold greater than water-solvated 1O2, which enables transport of this ROS acrossmillimeter length-scales rather than difusion across nanometer lenght-scales for solvated PS systems. The results
demonstrate that this unique SH technology is a promising method to disinfect and treat periodontal disease.

Acknowledgements. This work was supported by grant 2R44DE026083-03 from the National Institutes of Health.

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P-1.3.29
The impact of photoinactivation with Ga3+ meso-PPIX on virulence factors of
Staphylococcus aureus and Pseudomonas aeruginosa
Michalska Klaudia1, Woźniak Agata1, Krupińska Martyna1, Moszyńska Agata 1, Nakonieczna Joanna 1, Grinholc
Mariusz 1
1) Laboratory of Molecular Diagnostics, Intercollegiate Faculty of Biotechnology University of Gdansk andMedical University of Gdansk, Gdansk, Poland.
Introduction: The ability of pathogenic bacteria to cause disease is dependent upon the production of agents termed‘virulence factors’, such as toxins and adhesion molecules, that actively cause damage to host tissues during infection.
The potential antimicrobial strategy might be also targeting virulence factors (e.g. inhibition of their production or
activity) which would be harmful to the pathogen and allow the host immune system a better chance of clearing theinfection. Further studies had revealed a significant impact of antimicrobial photodynamic inactivation (aPDI)
towards virulence factors produced by both Staphylococcus aureus and Pseudomonas aeruginosa 1,2,3. Ga3+ mesoPPIX (Ga 3+ MPPIX) is dual function photosensitizer, which not only could block iron metabolism, but also due tothe porphyrin structure acquire photodynamic properties.
Aim: The aim of this study was to determine of the influence of Ga3+ meso-PPIX-mediated aPDI on excreted
virulence factors produced by Gram-positive (S. aureus) and Gram-negative (P. aeruginosa) representatives.
Material and Methods: To investigate the effectiveness of aPDI on virulence factors production and activity, sublethal doses of photoinactivation with Ga 3+ MPPIX upon green LED light irradiation (λmax= 522 nm, 10.6 mW/cm2)
was determined for S. aureus clinical isolate and reference strain of P. aeruginosa ATCC 10145. The sub-lethal dose
was defined as a dose that reduces the survival rate of treated bacteria vs. untreated bacteria by 0.5 – 2 log10 CFU/ml.
Impact of sub-lethal doses of aPDI with Ga3+ MPPIX was determined on TSST-1 (Toxic Shock Syndrome Toxin 1)
S. aureus superantigen production and on two P. aeruginosa virulence factors: pyocyanin and elastase B. Evaluationof TSST-1 production after aPDI exposure was performed based on the gene expression measured with the qRTPCR method and the level of proteins produced by Western Blot immunoblotting. Moreover, changes in virulence
factors after aPDI exposure e.g., pyocyanin and elastase B activity was measured with Elastin Congo Red Assay andspectrophotometric quantification of pyocyanin concentration respectively, before and after exposure to sub-lethal
doses of aPDI.
Results: Photoinactivation with Ga 3+ MPPIX had revealed the highest reduction in the survival rate of clinicalisolate of S. aureus up to 6.08 log10 of CFU/mL. TSST-1 protein level decreased under the sub-lethal aPDI but there
was no significant effect of aPDI on the expression level of tsst-1 gene (fold change = -0.2 log2). Applied
photoinactivation reduced the bacterial viability of P. aeruginosa strain (>3 log10 of CFU/ml) and reduced
significantly pyocyanin concentration and elastase B activity after exposure P. aeruginosa to sub-lethal doses.
References
1) Bartolomeu, M.; Rocha, S.; Cunha, Â.; Neves, M.G.P.M.S.; Faustino, M.A.F.; Almeida, A. Effect of
Photodynamic Therapy on the Virulence Factors of Staphylococcus aureus. Frontiers in Microbiology, 2016
2) Fila, G.; Kawiak, A.; Grinholc, M.S. Blue light treatment of Pseudomonas aeruginosa: Strong bactericidal activity,
synergism with antibiotics and inactivation of virulence factors. Virulence, 20163) Ogonowska, P.; Nakonieczna J. Validation of stable reference genes in Staphylococcus aureus to study gene
expression under photodynamic treatment: a case study of SEB virulence factor analysis. Scientific Reports, 2021
Acknowledgements. This work was supported by grant from National Science Centre: 2018/30/Q/NZ7/00281 (MG)Book of Abstracts ������������������������������������������������������������������������������������������������ 56

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P-1.3.30
Development of new antimicrobial materials based on porphyrinoids
Tatevik Chilingaryan1,2, Dr. Céline Frochot,1 Dr. Philippe Arnoux,1 Dr. Benoit Habermeyer2
1) Université de Lorraine, Laboratoire Réactions et Génie des Procédés, Nancy, France; 2) PorphyChem SAS,
Dijon, France.
In France, each year more than 150.000 peoples suffer from infections caused by multi-resistant bacteria. Huge efforts
have been dedicated to find alternative treatment methods to conventional antibiotics and Photodynamic Inactivation(PDI)1,2 is considered as a promising one. PDI3 is an innovative technique that is highly efficient against both grampositive and gram-negative bacteria. Furthermore, it was demonstrated that bacteria are unbale to develop resistanceagainst this technique. PDI is the combination of a photosensitizer (PS) and light to generate cytotoxic reactive
oxygen species (ROS) that kill bacteria but also viruses, fungi and parasites.
In this project, we were interested in the development of new antimicrobial films, based on porphyrinoids(porphyrins, chlorins and phthalocyanines), for PDI applications mainly in hygiene and food safety. These films canbe used on all kind of surfaces in order to kill bacteria and to prevent their growth. These antimicrobial materials also
prevent surface contamination.

Fig. 1 – Perrin-Jablonski diagram for ROS generation via Type I and Type II.

For fabrication of antimicrobial films, two components are necessary: a photosensitizer that fills most of the criteria
required by PDI (stability of the molecule, high molar absorption coefficient, high quantum yield of singlet oxygen)
and a polymer film. Several candidates were studied as PSs and among them, water-soluble porphyrins. PSs wereincorporated onto polyethylene terephthalate (PET) film surface and singlet oxygen production was studied.
References
1. L. Jiang, C. R. R. Gan, J. Gao, X. J. Loh, Small 2016, 12, 3609-3644.
2. P. H. H. Duong, P-Y. Hong, V. Musteata, K. V. Peinemann, S. Nunes, J. Nanomaterials and Polymers, 2017, 2,
6612-6616.
3. B. Habermeyer, T. Chilingaryan, R. Guilard, Bacterial Efficiency of Porphyrin Systems, J. Porphyrins andPhthalocyanines, 2021, 25, 359-381.
Acknowledgements. This work was supported by grant ANRT.

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IL-1.4.1

Regulation of cyanobacterial photoprotection
Adjélé Wilson1, Jiao Zhan1, Fernando Muzzopappa1, Claudia Steglich2, Stanislaw Nizinski3, Elena Andreeva4 ,Jacques-Phillipe Colletier4, Michel Sliwa3, Wolfgang Hess2, Diana Kirilovsky1
1

Université Paris-Saclay, CEA, CNRS, Institute for Integrative Biology of the Cell (I2BC), Gif-sur
Yvette, France; 2University of Freiburg, Faculty of Biology, Institute of Biology III, Freiburg, Germany; 3
Univ. Lille, CNRS, UMR 8516, LASIRE, Lille, France ; 4 Univ. Grenoble Alpes, CEA, CNRS, Institut deBiologie Structurale, Grenoble, France
Photosynthetic organisms need to sense and respond to light changing environmental conditions in orderto maximize light absorption and photosynthetic efficiency while avoiding the formation of dangerous
reactive oxygen species. Cyanobacteria can adapt their photosynthetic apparatus to efficiently harvest
energy under low light intensities by increasing the synthesis and concentration of the phycobilisomes(PBSs), the cyanobacterial antenna, and avoid photodamage under high light conditions by increasingheat dissipation of excess absorbed excitation energy. This mechanism is induced by the modular
photoactive Orange Carotenoid Protein (OCP) by interacting with the PBSs. The amplitude ofphotoprotection depends on the concentration of OCP in the cell, the rate of photoactivation and thestrength of PBS-OCP interaction. The expression of most OCPs is constitutive but is increased understress conditions in which phycobiliprotein synthesis decreases. I will describe the recently discovery of apost-transcriptional regulatory mechanism linking the expression of PBS core proteins directly and in an
inverse fashion to the synthesis of OCP1. OCP synthesis is regulated by a small regulatory RNA (sRNA),
ApcZ, originating from the 3’ end of the apcABC operon. ApcZ inhibits OCP expression by base pairing
to the start codon, the following four codons and the region upstream of the start. The absence (ordecrease) of ApcZ largely increases OCP concentration under stress conditions. The OCP is formed by Nterminal (NTD) and C-terminal (CTD) domains which strongly interact in the orange inactive form
(OCPO) and shared a ketocarotenoid molecule. Upon strong illumination OCPO converts into the open
active red OCP (OCPR). Between the photoexcitation of the carotenoid and the final red open
conformation several red intermediary steps occur. The protein becomes red already in ps and the
carotenoid translocates into the NTD in several µs while the opening of the protein occurs slower. We
investigated several factors influencing these intermediary steps and the rates of photoactivation andrecovery including the carotenoid nature and the position of tags. Only the red open active OCPR is able
to interact with the PBS. Almost nothing is known about the OCP and PBS amino acids involved in thisinteraction. We constructed 23 OCP mutants in the different NTD surfaces to learn more about OCP-PBS
interaction. We demonstrated that only the NTD surface that in the orange OCP interacts with the CTD is
involved in OCP binding to PBS. Modification of these amino acids also influences OCP photoactivation
and/or recovery rates indicating that they are also involved in the translocation of the carotenoid.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.4.2
From light-harvesting to quenching in plant antenna complexes: a new perspectivefrom atomistic simulations
Benedetta Mennucci
Dipartimento di Chimica e Chimica Industriale, University of Pisa, via G. Moruzzi 13, 56124, Pisa, Italy
Light-harvesting complexes (LHCs) of plants can switch from an active, light-harvesting state to a quenched state
where excess energy is dissipated into heat however the mechanism of quenching and the identity of thequenched/active states are still subject to debate. Here we apply an integrated computational strategy1 which
combines molecular dynamics, enhanced sampling techniques, and multiscale methods to CP29, a minor LHC,
finding that the commonly accepted hypothesis of a quenching mechanism localized on a single pair of strongly
coupled pigments is not sufficient, but a dynamic network of pigments and interactions must be considered.
References
1) Cupellini, L., Bondanza, M., Nottoli, M. & Mennucci, B. Successes & challenges in the atomistic modeling oflight-harvesting and its photoregulation. BBA - Bioenergetics 1861, 148049 (2020).

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.4.3
Exploring Carotenoid-Mediated Photophysics in Plants with Ultrabroadband 2D
Electronic Spectroscopy
Minjung Son1, Alberta Pinnola2, Roberto Bassi3, Gabriela S. Schlau-Cohen1
1) Massachusetts Institute of Technology, Department of Chemistry, Cambridge, USA; 2) University of Pavia,
Department of Biology and Biotechnology, Pavia, Italy; 3) University of Verona, Department of Biotechnology,
Verona, Italy
Plants absorb sunlight across the visible region of the solar spectrum and collect the energy required forphotochemical reactions in lower-lying states. Previous 2D experiments to study plants were limited to these lowenergy states. We describe the development of ultrabroadband 2D electronic spectroscopy and its application to themajor antenna complex of plants.1 First, we identify a debated dark state on a single carotenoid, lutein 2, thatmediates relaxation.2 Second, we measure chlorophyll-to-carotenoid energy transfer, a hypothesized but previously
unobserved pathway to safely dissipate excess energy.3 We also investigate the molecular and environmental
parameters that can control the magnitude of this dissipative energy transfer.4
References
1) M. Son, S. Mosquera-Vasquez, G. S. Schlau-Cohen, “Ultrabroadband 2D Electronic Spectroscopy with HighSpeed, Shot-to-Shot Detection.” Optics Express, 25, 18950 (2017).
2) M. Son, A. Pinnola, R. Bassi, G.S. Schlau-Cohen, “The Electronic Structure of Lutein 2 is Optimized for Light
Harvesting in Plants.” Chem, 5, 575 (2019).
3) 10. M. Son, A. Pinnola, S.C. Gordon, R. Bassi, G.S. Schlau-Cohen. “Observation of Dissipative Chlorophyllto-Carotenoid Energy Transfer in Light-Harvesting Complex II in Membrane Nanodiscs.” Nature Communications
11, 1 (2020).
4) M. Son, A. Pinnola, G.S. Schlau-Cohen, “Zeaxanthin Independence of Photophysics in Light-Harvesting
Complex II in a Membrane Environment.” BBA-Bioenergetics 5-6, 1861 (2020).
Acknowledgements. This work was supported by the U.S. Department of Energy, Office of Science, Office ofBasic Energy Sciences, Division of Chemical Sciences, Geosciences, and Biosciencesunder Award #DESC0018097 to G.S.S.-C.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-1.4.4
The keto group in β2 of the carotenoid tunes the Orange Carotenoid Protein
photocycle kinetics
Volha Chukhutsina1 and Jasper van Thor1,
1) Department of Life Sciences, Imperial College London, London SW7 2AZ, United Kingdom
All vital biological processes, including
photosynthesis, rely on the function of various
15 µs
photosensors
andphotoreceptors.
In
13µs
CTD NTDDCTD NTDcyanobacteria
light-harvestingofthephotosynthetic machinery is controlled by a
ECH:
CAN:
uniquephotosensitive
protein,
Orange80 µs
17 µs
Carotenoid Protein (OCP). Being found only
among cyanobacteria, OCP is the only known
photoreceptor which uses a carotenoid for its
4.5 ms430 µslight activation. Understanding and, therefore,
NTE
NTE
gaining control over OCP light-activation has a
CTD
CTD
NT
NT
OCPR
D
broad range of applications from improvingDphotosynthesis productivity to potentiallyFig. 1 – Photocycle intermediates resolved for OCP constructs with revolutionizing the field of optogenetics.
OCP is a water-soluble protein
the two different carotenoids: ECH -echinenone, CAN-canthaxanthincomposed of the two domains: an all-helical Nterminal domain (NTD) and a C- terminal domain (CTD). Absorption of blue–green light by the carotenoid induces
conformational changes, converting stable inactive OCPO (the so-called orange form) into unstable but active OCPR
(red form), which are also followed by drastic changes in the OCP absorption. The OCP has been shown to bind andbe activated by various keto-carotenoids (3-hydroxyl-echinenone, hECN; echinenone, ECN; and canthaxanthin,
CAN). The conserved keto group of the β1-ring of the keto-carotenoid is hydrogen bonded to Tyr201 and Trp288 of
CTD. The other ring (β2) is nestled within a group of conserved aromatic residues (Trp41, Tyr44, Trp110) in NTD.
The structure of β2-ring is actually specific for different keto-carotenoids. For example, CAN contains a second ketogroup at the 4’ position, which is absent in ECH. The mechanism of OCP light activation is considered to be
independent on the keto-carotenoid bound to it and is initiated by hydrogen bonded rupture between the keto groupof the β1 ring and Tyr201 and Trp2881. Therefore, the photocycle intermediates resolved from OCPECH are considered
to be characteristic for OCP in general2.
In this study we for the first time to our knowledge study the effect of the keto group in the β2 ring of the ketocarotenoid on the OCP photocycle intermediates and activation energies. For that OCP constructs with ECH and
CAN in their structure were studied by performing us-ms time-resolved spectroscopy and Arrhenius temperaturedependence measurements. The results indicate different activation energies and photocycle intermediate rates for
the OCP with different carotenoids imbedded. The OCPCAN demonstrates lower activation energies both for OCPOOCPR and OCPR-OCPO reactions. As a result, the photocycle rates are also faster for OCPCAN. The results for the firsttime indicate that the presence of a keto group in β2 tunes the OCP photocycle.
NTE

NTE

OCPO

O

O

O

E

E

O

D

0.8 ms

6.5 ms

O

NT

NT

D
CT

O

NT

NTE

CT

NT

D

D

O

NTE

O

O

O

O

CTD

CTD

NTD

O

NTD

O

O

O

References
1) F. Muzzopappa & D. Kirilovsky, Changing Color for Photoprotection: The Orange Carotenoid Protein.
Trends in Plant Science (2020)2) P. E. Konold, et al. Photoactivation Mechanism, Timing of Protein Secondary Structure Dynamics andCarotenoid Translocation in the Orange Carotenoid Protein. J. Am. Chem. Soc. (2019).
Acknowledgements. This project was supported by the EMBO long-term fellowship (EMBO ALTF 244-2017) andhas received funding from the European Union Horizon 2020 research and innovation programme under the Marie
Sklodowska-Curie grant agreement No 839389) to V.C.

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IL-1.4.5

Hydrophobic mismatch as a possible trigger of NPQ
Alexander V. Ruban
School of Biological and Chemical Sciences, Queen Mary, University of London, UK
Non-photochemical quenching, NPQ, is a physiological process that protects the photosynthetic apparatus ofoxygenic photosystem II from harmful unused energy that is accumulated in excess in high light environment.
Despite a number of decades of intense NPQ research no clarity exists as to how exactly the proton gradient triggers
the quenching in LHCII antenna. Here, I present a novel view on this process proposing the onset of the hydrophobic
mismatch between proteins of antenna and photosynthetic membrane triggered by the gradient. This proposal is basedon the synthesis of various experimental and theoretically-obtained data available so far. Observation of thylakoid
membrane thinning and enhanced hydrophobicity following illumination of leaves and algal cells, effect of sterols,
cross-linkers and other agents promoting or mitigating the hydrophobic mismatch will be discussed. The involvementof hydrophobic mismatch in NPQ of plants is consistent with the fact that the minimum components of the quenching
are the proton gradient and the major LHCII complex, whilst zeaxanthin and PsbS protein are mere regulators of theextent of the mismatch and response of the protein.

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IL-1.4.6

Chlorophyll f site assignments in far-red light-acclimated photosystem I
Christopher J. Gisriel1, David A. Flesher2, Hao-Li Huang1, Krystle Reiss1, Victor S. Batista1, Jimin Wang2, Donald
A. Bryant3, and Gary W. Brudvig1,2
1) Yale University, Department of Chemistry, New Haven, USA; 2) Yale University, Department of Molecular
Biophysics and Biochemistry, New Haven, USA; 3) The Pennsylvania State University, Department of Biochemistry
and Molecular Biology, State College, USA

During far-red light (FRL) photoacclimation, chlorophyll (Chl) f molecules are bound to photosystem I(PSI) in addition to Chl a, allowing for far-red lightabsorption1. Two FRL-PSI structures have been reported
using cryo-EM from different cyanobacterial species2,3,
but they exhibited unexpected inconsistencies in their Chl
f site assignments4. We reprocessed one of these structures
to a higher resolution, and performed a quantitative
assessment of Chl substituents in the cryo-EM maps to
identify Chl f-binding sites in the two cryo-EM maps. The
two cryo-EM maps provide direct evidence for highoccupancy Chl f-binding at two and three sites,
respectively, and three more sites in each structure exhibit
strong indirect evidence for Chl f binding. Challenges that
accompany making Chl f assignments based on cryo-EM
maps, and common themes in Chl f binding are describedthat clarify the current understanding of the molecularbasis for FRL photoacclimation in photosystems.
Fig. 1 –Chl f sites in two FRL-PSI structures. Left and right panels show one monomer of the FRL-PSI structuresfrom H. hongdechloris and F. thermalis, respectively. Top panels show a stromal view and bottom panels show amembrane plane view from the center of the trimer. Only tetrapyrrole rings are shown for clarity. Pink glows areshown for Chl sites that exhibit strong direct and indirect evidence for being highly occupied Chl f (three sites in H.
hongdechloris and two sites in F. thermalis), and yellow glows are shown for Chl sites that exhibit strong indirect
evidence only for binding by Chl f at high occupancy (three sites in each structure).
References
1) Gan, F. et al. Extensive remodeling of a cyanobacterial photosynthetic apparatus in far-red light. Science 345,
1312–1317 (2014).
2) Gisriel, C. et al. The structure of Photosystem I acclimated to far-red light illuminates an ecologically important
acclimation process in photosynthesis. Sci. Adv. 6, eaay6415 (2020).
3) Kato, K. et al. Structural basis for the adaptation and function of chlorophyll f in photosystem I. Nat. Commun.
11, 238 (2020).
4) Gisriel, C. J., Wang, J., Brudvig, G. W. & Bryant, D. A. Opportunities and challenges for assigning cofactors incryo-EM density maps of chlorophyll-containing proteins. Commun. Biol. 3, 408 (2020).
Acknowledgements. This work was supported by Department of Energy, Office of Basic Energy Sciences, Divisionof Chemical Sciences grant DE-FG02-05ER15646 to G.W.B. and Grant DE-SC0001423 to V.S.B, National Science
Foundation grant MCB-1613022 to D.A.B., and the Forest B.H. and Elizabeth D.W. Brown Postdoctoral Fellowshipfor Plant Science to C.J.G.

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19th Congress of the European Society
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-1.4.7

BREAKING THE RED-LIMIT: DRIVING OXYGENIC PHOTOSYNTHESISWITH FAR-RED LIGHT
Roberta Croce
Vrije Univesiteit Amsterdam, Amsterdam, the Netherlands(*) r.croce@vu.nl
Photosynthetic organisms use sunlight energy to fix CO2 into carbohydrates, in this way sustaining
almost all life on our planet. The capacity of these organisms to harvest light is a crucial factor in
the photosynthetic process, especially in light-limited conditions, which occur in greenhouses andcanopies. However, plants and algae only use the visible part of the solar spectrum, discarding morethan 50% of the photons reaching the surface of the Earth. This is because their photosynthetic
proteins bind as main pigments chlorophyll (Chl) a and b, which have intense absorption in the red
and the blue regions of the electromagnetic spectrum but do not absorb above 700 nm. For a longtime, it was believed that cyanobacteria, the prokaryotic ancestors of plant chloroplasts, also could
only use visible light to drive photosynthesis. The discovery of species containing Chl d and Chl f,which absorb in the far-red region of the spectrum, has shown that this is not the case. However,
due to their different energetics, Chl d and f are expected to alter the excited state dynamics of thephotosynthetic units and, ultimately, their performances. How can thus cyanobacteria use far-red
light for efficient photochemistry?
To answer this question we use a combination of biochemistry and spectroscopic measurements on
intact cells and isolated complexes. We show that chlorophyll f insertion marginally affects the
charge separation efficiency of Photosystem I [1] but decreases significantly that of Photosystem
II [2]. The difference between the two photosystems and the possibility to introduce Chl d and f inplant photosynthetic complexes to extend the photosynthetic active radiation in crops will be
discussed.
References[1] Tros M, Mascoli V, Shen G, Ho MY, Bersanini L, Gisriel CJ, Bryant DA, Croce R, Chem, 7, 155-173(2021)[2] Mascoli V, Bersanini L, Croce R, Nature Plants, 6, 1044-1053 (2020)

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-1.4.8
Electrostatic Control of Reaction Centre Excitation in Photosystem II
Abhishek Sirohiwal1, Dimitrios A. Pantazis1
1) Max-Planck-Institute für Kohlenforschung, Department of Molecular Theory and Spectroscopy, Mülheim an derRuhr, Germany.
Photosystem II is a multi-subunit pigment-protein complex that utilizes sunlight to trigger charge-separation andcatalyse water oxidation. The charge separation cascade is initiated in the reaction centre (RC), which is composedof six pigments (four Chlorophyll a and two Pheophytin a) arranged symmetrically along the D1 and D2 core polypeptides (Fig. 1). Biological evolution favoured productive electron transfer only along the D1 side with the precise
nature of the initial excitation event(s) remaining under debate. In this work,1-2 we employ multiscale quantum
mechanics/molecular mechanics (QM/MM) coupled with high-level computations (full time-dependent density
functional theory with range-separated functionals benchmarked against coupled cluster theory) to investigate theexcited state profile of the RC (Fig. 1). Our results describe for the first time at a fundamental electronic structurelevel precisely how differential protein electrostatics create the observed excitation asymmetry within the RC. By
simultaneous quantum chemical treatment of multimeric pigment assemblies we identify the critical pairs of RC
pigments associated with low-lying charge-transfer states and we eventually propose a novel model to describe
excitation of Photosystem II reaction centre based on two parallel charge-separation pathways. Among others, our
new model explains the triggering of charge separation by direct absorption of far-red photons (700-800 nm), i.e.
beyond the known “red-limit” (680 nm) of oxygenic photosynthesis.

Fig. 1 – (left) Overview of symmetrically arranged pigments in the reaction centre along with key redox-active components of
Photosystem II and (right) molecular-mechanics based model of the lipid-bilayer bound Photosystem II.

References
1) A. Sirohiwal, F. Neese, D. A. Pantazis, Protein Matrix Control of Reaction Center Excitation in Photosystem II.
J. Am. Chem. Soc., 142, 42, 18174-18190, 2020.
2) A. Sirohiwal, F. Neese, D. A. Pantazis, How Can We Predict Accurate Electrochromic Shifts for
Biochromophores? A Case Study on the Photosynthetic Reaction Center. J. Chem. Theory Comput., 17, 3, 18581873, 2021.
Acknowledgements. The authors gratefully acknowledge support by the Max Planck Society.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.4.9

How pulse energy and wavelength affect ultrafast dynamics of Orange Carotenoid
Protein? Attempt to define safe regime of femtosecond excitation based experiments.
Stanislaw Nizinski1,2, Elena Andreeva3, Jacques-Philippe Colletier3, Adjele Wilson4, Diana Kirilovsky4, Gotard
Burdzinski1, Michel Sliwa21) Quantum Electronics Laboratory, Faculty of Physics, Adam Mickiewicz University in Poznan, Umultowska 85,
61-614 Poznan, Poland; 2) Univ. Lille, CNRS, UMR 8516 - LASIR - Laboratoire de Spectrochimie Infrarouge etRaman, F-59000 Lille, France; 3) Institut de Biologie Structurale (IBS), Univ. Grenoble Alpes, CEA, CNRS, 38044
Grenoble, France; 4) Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Universite Paris-Sud,
Universite Paris-Saclay, 91198 Gif-sur-Yvette, France
Orange Carotenoid Protein (OCP) is a water-soluble pigment protein responsible for dissipation of excited stateenergy harvested by cyanobacterial antenna complexes, phycobilisomes. OCP performs its photoprotectivefunction in a selective way: in the dark it occurs in inactive form (OCPO), but after illumination with a blue-green
light it undergoes photoconversion with a low quantum yield (< 1%) to quenching-capable form called OCP red(OCPR). The mechanism that controls the photoconversion of OCPO to OCPR has been recently discussed inliterature in a multiple papers. Upon absorption of a blue-green photon, the carotenoid embedded inside OCP
undergoes S0→S2 transition, forming two S1 / ICT coupled states and a distorted S* state within hundreds of fs.
These states decay to the ground state, generating redshifted forms of OCP with very small yield under 1%. These
forms are precursors of the final quenching-capable OCPR.
Femtosecond transient absorption spectroscopy is a powerful technique, which has been already widely employed
by a many groups to describe first steps of OCP photoactivation. However current state of literature does notprovide perfectly consistent picture of these steps, and detailed information about excitation pulse energy density is
not always provided by the authors. We have found that it is very hard to obtain always the same consistentdescription of OCP photodynamics when experiments are repeated, due to high complexity of mechanism, sample
unstability and heterogeneity. Therefore it is critical to perfectly control all important experimental conditions, andunderstand impact of each of them. We found, that when energy density of the pump pulse exceeds linear regime,
additional long living products are formed, which artificially boost calculated yield of the first red OCP
intermediate (P1). Obviously, observation of such products does not constructively contribute to the biologicalknowledge about the system, as they are artefacts of the technique which require high energy pulses, unreachable in
the natural environment. From the other side, observation of the low-yield products is required in OCP studies,
therefore one cannot discard high energy pulses as they help to maximize signal to noise ratio. It is important to go
for tradeoff and localize the maximum safe regime of excitation energy density. So paradoxically, it interesting toinvestigate formation of high pulse energy products, to find the pulse energy which forms them in negligible
quantity and does not significantly distorts the formation of biologically-relevant species.
Acknowledgements. This work was performed with financial support of the Polish National Science Centre(NCN), project 2018/31/N/ST4/03983, and French National Research Agency (ANR), project DynOCP (ANR-18CE11-0005).

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-1.4.10

Steady-state and time-resolved X-ray scattering and UV-vis spectroscopy
demonstrate that oligomerization processes limit photoactivation and recovery of the
cyanobacterial Orange Carotenoid Protein
Elena A. Andreeva1, Stanislaw Nizinski2, Adjélé Wilson3, Matteo Levantino4, Elke De Zitter1, Fernando
Muzzopappa3, Aurélien Thureau5, Ninon Zala1, Gotard Burdzinski2, Michel Sliwa6, Diana Kirilovsky3, Giorgio
Schirò1, Jacques-Philippe Colletier1
1) Institut de Biologie Structurale (IBS), Univ. Grenoble Alpes, CEA, CNRS, 38044 Grenoble, France; 2) Quantum
Electronics Laboratory, Faculty of Physics, Adam Mickiewicz University in Poznan, Poznan, Poland; 3) Institutefor Integrative Biology of the Cell (I2BC), CEA, CNRS, Université Paris-Sud, Université Paris- Saclay, Gif-surYvette, France; 4) European Synchrotron Radiation Facility (ESRF), BP 220, Grenoble, France; 5) Synchrotron
Soleil, Beamline SWING, Gif Sur Yvette Cedex, France; 6) Univ. Lille, CNRS, UMR 8516 - LASIR - Laboratoire deSpectrochimie Infrarouge et Raman, Lille, France.
Email: eandreeva@ibs.fr
The photoactive Orange Carotenoid Protein (OCP), involved in cyanobacterial photo-protection [1], is a twodomain protein which upon light absorption transitions from a compact inactive “dark” state (OCP O) to an extended
photoactive “light” state (OCPR) [2,3]. The molecular mechanism enabling photoactivation involves migration of
the carotenoid pigment from the interface between the two domains into the NTD, and subsequent domain
dissociation. It remains unclear what the exact structure of OCPR is, on which timescale it forms, and whether or
not oligomerization [2,4] is involved in the regulation of OCP function – either in the inactive or photo-activatedstate. Here, we used a combination of steady-state and time-resolved (TR) X-ray scattering and spectroscopy toaddress these three specific issues. Our results inform on the time scales associated to the dissociation andreassociation of domains in monomeric OCP, yielding OCPR and enabling recovery of OCPO, respectively. They
furthermore demonstrate that oligomerization occurs at the OCPO and OCPR level, limiting the photoactivation of
OCPO and the OCPO to OCPR thermal recovery, respectively.
References[1] Kirilovsky D. & Kerfeld C. A., (2016). Cyanobacterial photoprotection by the orange carotenoid protein. Nat.
Plants 2, 16180.
[2] Wilson A. et al., (2008). A photoactive carotenoid protein acting as light intensity sensor. PNAS, 105 (33):12075-12080[3] Haijun L. et al., (2016). Dramatic Domain Rearrangements of the Cyanobacterial Orange Carotenoid Protein
upon Photoactivation. Biochemistry, 55 (7): 1003–1009[4] Golub M. et al., (2019). Solution Structure and Conformational Flexibility in the Active State of the Orange
Carotenoid Protein: Part I. Small-Angle Scattering. J. Phys. Chem. B, 123, 9525−9535

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P-1.4.11

Blue light modulates seed germination in tomato through the interaction of thecryptochrome 1a with ABA and GA
Rogério Falleiros Carvalho1, Reginaldo Oliveira1
1) Faculty of Agronomic and Veterinary Science-FCAV-São Paulo State University-UNESP, Department of
Biology Applied to Agriculture, Jaboticabal, SP, Brazil
A multifaceted approach has been discussed regarding the role of blue light (BL) in the seed germination in
tomato1,2. However, from BL signaling, many points remain to be elucidated, including participation of the abscisicacid (ABA), gibberellin (GA), and BL photoreceptor cryptochrome (cry). We hypothesized for the first time thatcry1a of tomato interacts with ABA and GA in the seed germination from blue light BL signaling. The experiments
were carried out in growth chamber with continuous BL fluence rate (0 or dark; 1; 5; 10; 15 and 25 µmol m-2s-1).
We observed that tomato (cv. Moneymaker) seeds presented high germination rate (G%) and index of germination
speed (IGS) in the dark. However, all BL fluence partially inhibits the germination, increasing the time to reach
50% of final/maximum germination (T50). Regarding the function of cry1a, the loss of its function in the mutant(cry1a) reduced the G% and IGS, besides the fact that 5 µmol of gibberellic acid (GA3) did not recover thegermination, whereas 0,25 µmol of abscisic acid (ABA) amplified the negative effects of BL in the germination ofthis genotype. We also observed in dark and BL that the notabilis, which is ABA-deficient mutant, and procera,
which shows constitutive response to GA, increased the G% and IGS. These responses were partially related tochanges in the activity of endo-β-mannanase, β-mannosidase and α-galactosidase enzymes. Histochemical analysisof tomato cv. Micro-Tom transgenic lines harboring the gene reporter GUS fused to promoters of the RD29B andGA2OX, responsive to ABA and GA, respectively, revealed a temporary increase (36h) in seeds exposed to dark,
but a reduction in BL after 72h, confirming the fundamental role of BL on these hormones during seedgermination. Thus, in this study, we showed for the first time the influence of cry1a in the photoregulation of seedgermination in tomato, and the role of BL in the activity of hydrolytic enzymes related to the physical breaking of
the barrier imposed by seed endosperm.
References
1) Aneli, N. A. 1940. Inhibiting action of blue light upon germination of tomato seeds. Comptes Rendus De L
Academie Des Sciences De L Urss 28:267-69.
2) Omidvar, V., and M. Fellner. 2015. DNA methylation and transcriptomic changes in response to different lights
and stresses in 7B-1 male-sterile tomato. Plos One 10 (4).
Acknowledgements. This work was supported by grant number 2018/20748-3 from São Paulo
Research Foundation – FAPESP.

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PL-4
Dark CPDs and Genomic Sites Hypersensitive to UV
Douglas E. Brash1, Sanjay Premi1,3, Alejandro Garcia-Ruiz1, Vijay Menon1, Carlos Angele-Martinez1, and Leticia
C. P. Goncalves1,2
1) Yale University School of Medicine, Department of Therapeutic Radiology, New Haven, CT, USA; 2)
Universidade de São Paulo, Departamento de Química Fundamental, São Paulo, SP, Brazil; 3) Present address:
Moffitt Cancer Center, Department of Tumor Biology, Tampa, FL, USA
Cyclobutane pyrimidine dimers (CPD) in DNA are lethal and mutagenic, causing the UV-signature mutations seenin skin tumors. Two surprising features of CPDs have surfaced recently. First, although creating a CPD always
requires high energies capable of exciting an electron to a higher orbital, this energy needn't come from UV radiation.
It can be generated chemically, by a process termed "chemiexcitation". Second, specific individual dipyrimidines are
100-fold more sensitive to forming CPDs than the rest of the genome; we've termed these sites "CPD hyperhotspots".
Chemiexcitation in the skin begins when UVB or UVA activate enzymes that synthesize the radicals superoxide andnitric oxide, resulting in peroxynitrite. The latter oxidizes melanin or its precursors to a form that can be attacked by
O2 to form a cyclic peroxide, dioxetane. This four-member ring is unstable and decomposes to give a carbonyl
containing an electron excited to a triplet state; the energy transfers radiationlessly to nearby DNA to form a CPD.
Enzyme activation can persist for hours, allowing chemiexcitation to generate CPDs in the dark. We find that otherbiomolecules present in skin can also be chemiexcitation substrates.
CPD hyperhotspots were discovered by nicking DNA lesion sites with a repair enzyme, tagging nicks with linkers
detectable by NextGen sequencing, and analyzing the data with statistical methods having the resolution to quantify
rare lesions at single-base resolution across the genome. High-frequency outliers constituted a distinct category andfell into three classes. The strongest class occurred in gene promoters, particularly for RNA-binding proteins, at a
motif present at a) ETS family transcription factor binding sites, previously known to be UV targets and now shown
to be among the most sensitive in the genome, and b) sites of translation regulation. These CPD sites aligned precisely
with recurrent UV-signature mutations in gene promoters in melanomas. Another class occurred at simple PyPyPyruns and the reason for their UV sensitivity remains unknown. The third class was intriguing, occurring at A2–
15TTCTPy, developing “dark CPDs” long after UV exposure, and repairing CPDs slowly; in melanocytes this class
had accumulated CPDs prior to the experiment. The CPD hyperhotspots were originally discovered in melanocytes
and fibroblasts; we now find similar hyperhotspots in keratinocytes.
The high frequency of CPDs at these sites means that, at sunburn levels of UV exposure, every cell would have ahyperhotspot CPD in each of the ∼20 cell pathways in which the motifs occur, letting hyperhotspots act as epigenetic
marks. UV could drive tumor evolution by direct changes in cell physiology rather than acting solely through rare
mutations. The high CPD prevalence also favors co-occurring mutations, allowing tumor evolution to use genes thatare only weak phenotypic drivers.
Outlier DNA sequences extraordinarily sensitive to environmental agents can be sentinels for monitoring personalcarcinogen exposure, e.g., by measuring CPD levels. We are investigating their use as genomic dosimeters formeasuring past UV exposure and future skin cancer risk, so that UV-sensitive patients can be monitored to detect
skin cancers at a curable stage.
Acknowledgements. This work was supported by National Institutes of Health grants R01AR070851,
R01ES030562, and P50CA121974 and Department of Defense CDMRP grant CA093473.

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PL-5

Does light have a unique niche for enabling nanotechnology?
Tayyaba Hasan
Wellman Center for Photomedicine, Massachusetts General Hospital and Harvard Medical School, Boston, MA
02114; Division of Health Sciences and Technology, Harvard University and Massachusetts Institute ofTechnology, Cambridge, MA 02139
With a fairly routine use of omics, there is evolving a clearer understanding of disease mechanisms and theirregulation. It is now becoming increasingly clear for cancer or non-cancer pathologies, that disabling of single
possibly what are believed to be the dominant pathways or targets, will not provide effective long-term cures.
Inhibition of a given pathway leads the cancer cell or the pathogen to commandeer compensatory mechanisms for its
survival. A combination of therapeutics addressing multiple mechanisms has therefore emerged as a desirable modeof disease management, but pharmacokinetics and dosing now become complex and crucial. While the concept of
combination treatment is by no means new, there are some newer emerging approaches to administering it that are
exciting. These include the use of nanotechnology and light as a switch, which can provide cytotoxicity while at thesame time priming the microenvironment and helping deliver molecules at the right time to the right place. In thisregard Photodynamic Therapy (PDT) may have a special niche since it provides the additional level of selectivity
because of the requirement of the sensitizing molecules and photons being present together in adequate
concentrations to achieve the cytotoxic effect. And where these “reagents” are below the threshold for cytotoxicity
they create a priming effect on the microenvironment and cause remote physiologic changes such as immune effects
and permeability enhancement that enable other therapies to provide more efficient and tolerable treatments.
Recognition of these phenomenon explains the distant control of metastasis by a local treatment. Similarly, induced
or inherent resistance can be mitigated using PDT-activated nanotechnology due to the diversity of mechanisms thatcan be brought into play with the appropriately designed nano constructs. Photodynamic activation and PDTtriggered approaches therefore can have special attributes to make them unique and potentially of high impact. But
for this to be successful, there are a significant number of questions that still remain unanswered. Results from theliterature and our own studies in the context of the potential of PDT-inspired combination therapeutics andnanotechnology will be discussed.
Acknowledgements. This work was supported by grants: P01 CA084203 through the NIH/NCI, UH3 CA189901
through the NIH/NCI, R01 CA231606 through the NIH/NCI, 2R44 DE026083 through the NIH (with Singlet02
Therapeutics LLC), FA9550-20-1-0063 through the DoD/AFOSR, and R01 CA266855 through the NIH /NCI.

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IL-2.1.1

Photosensitized oxidation of proteins: is disulfide bond damage
important?
Michael J. Davies1
1) Department of Biomedical Sciences, Panum Institute, University of Copenhagen, Copenhagen 2200, Denmark.
Disulfide bonds are key determinants of protein structure and function, and are highly conserved across nearly all
proteomes. They are particularly abundant in many extracellular proteins, including those with critical structural,
ligand binding or receptor function. In the studies discussed here, we demonstrate that photo-sensitized oxidation ofprotein disulfides by singlet oxygen (and also a number of other oxidants) results in the formation of long-lived (halflife of a few hours), but reactive species, believed to be either a R-S+(OO-)-S-R’ species (i.e. a peroxidic adductformed between singlet oxygen and the disulfide bond) or a thiosulfinate R-S(O)-S-R’. Accumulating evidence isconsistent with further reaction of these species with another thiol, with this resulting in the formation of a thiol
adduct to the protein at the former disulfide bond and cleavage of the initial disulfide1,2.
In the case of glutathione (GSH), this results in glutathionylation of the protein3,4. The protein-GSH adducts
have been characterized by both mass spectrometry (MS; ions with m/z +306 and +712 assigned to the addition of 1
and 2 GSH respectively) and immunoblotting after SDS-PAGE separation, using anti-GSH antibodies. The extent of
GSH addition is increased by the use of D2O buffers consistent with the intermediacy of 1O2. Such adducts have been
detected on multiple proteins, demonstrating that this is a common pathway3,4. GSH adduction is reversed by both
non-enzymatic and enzymatic (glutaredoxin) reducing systems, but this does not repair the original disulfide bond.
Removal of the disulfide bond before oxidation prevents adduct formation.
For proteins with free Cys residues, these reactions result in the formation of novel protein-protein dimers
linked by a new disulfide5,6. These reactions require both a free thiol and the oxidized disulfide bond. The new interprotein disulfide can be reduced by added reducing agents such as DTT and TCEP. The new dimeric species havebeen characterized by immunological, intact protein MS, and LC-MS peptide mass mapping. The latter has allowed
unequivocable identification of the sites of the disulfides and also exact residues involved (e.g. between Cys25 on
beta-2-microglobulin and the active site Cys149 on GAPDH, and Cys36 of C-reactive protein and Cys-34 of humanserum albmin)5,6. These new cross-links affect enzymatic activity (e.g. of GAPDH) and are detected both withisolated proteins, and in fresh human plasma (e.g. cross-link between C-reactive protein and human serum albumin).
Overall these data provide evidence for a novel and facile process (‘oxidant-mediated thiol-disulfide
exchange’) that gives rise to glutathionylated and inter-protein crosslinks, as a result of initial oxidation of a disulfide
bond and subsequent rapid reaction with another thiol molecule, either low-molecular-mass (e.g. N-acetyl-Cys or
GSH), or on another protein. This pathway may contribute to the accumulation of modified proteins in diseased
tissues, in alterations to redox stasis, and altered cell signaling.
References
1
Karimi, M., Crossett, B., Cordwell, S. J., Pattison, D. I. & Davies, M. J. Characterization of disulfide (cystine)oxidation by HOCl in a model peptide: evidence for oxygen addition, disulfide bond cleavage and adductformation with thiols. Free Radic. Biol. Med. 154, 62-74 (2020).
2
Karimi, M. et al. Reactivity of disulfide bonds is markedly affected by structure and environment:
implications for protein modification and stability. Sci. Rep. 6, 38572 (2016).
3
Carroll, L. et al. Oxidant-induced glutathionylation at protein disulfide bonds Free Radic. Biol. Med. 160,
513-525 (2020).
4
Jiang, S., Carroll, L., Rogowska-Wrzesinska, A., Rasmussen, L. M. & Davies, M. J. Oxidation of disulfide
bonds in proteins by singlet oxygen gives rise to glutathionylated proteins. Redox Biol. 38, 101822 (2021).
5
Jiang, S., Carroll, L., Mariotti, M., Hägglund, P. & Davies, M. J. Formation of protein cross-links by singlet
oxygen-mediated disulfide oxidation. Redox Biol. 41, 101874 (2021).
6
Jiang, S., Hagglund, P., Carroll, L., Rasmussen, L. M. & Davies, M. J. Crosslinking of human plasma Creactive protein to human serum albumin via disulfide bond oxidation. Redox Biol 41, 101925, (2021).

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IL-2.1.2

Deciphering biomembrane photodamage: Alkylation of a type I sensitizer enhances
the photo-induced oxidation of phospholipid membranes.
Andrés H. Thomas1, Alexander Greer2, Mauricio S. Baptista3, Rosangela Itri4, Mariana Vignoni1
1) Universidad Nacional de La Plata-CONICET, Instituto de Investigaciones Fisicoquímicas Teoricas y Aplicadas,
La Plata, Argentina;2) City University of New York, Brooklyn, Brooklyn College, New York, United States; 3)
Universidade de São Paulo, Instituto de Química, São Paulo, Brazil; 4) Universidade de São Paulo, Institute ofPhysics, São Paulo, Brazil.
Pterins are endogenous photosensitizers present in human skin that accumulate in this tissue in patients sufferingfrom vitiligo, a skin disorder characterized by the acquired loss of constitutional pigmentation. Experiments using
model biological targets, such as nucleotides and amino acids, have demonstrated that mainly act through type I
mechanism. These processes are initiated by a dynamic reaction, controlled by diffusion, in which an electrontransfer from the substrate to the triplet excited state of the photosensitizer takes place. All unsaturated lipids in cell
membranes are well-known targets of oxidative damage, which can occur by type I and type II photosensitizedoxidation. In the case of vesicles dispersions, a hydrophilic photosensitizer will remain in the aqueous phase andthe photosensitized oxidation of a target molecule in the membrane will be a dynamic process. On the other hand, if
the photosensitizer is lipophilic, an association with a biomembrane is expected and, as the photosensitization is not
limited by diffusion, the oxidation might be much faster.
Pterin (Ptr), the parent and model compound of oxidized pterins, is hydrophilic, does not bind to phospholipid
membranes and, therefore, freely crosses biomembranes. Ptr photoinduces the oxidation of polyunsaturated fatty
acids (PUFAs) of phospholipids present in large unilamellar vesicles (LUVs), predominantly through type I
mechanism.1 Moreover, upon UVA irradiation in the presence of Ptr, the viability of HeLa cells decreases and thestructural integrity of the cell membrane are affected. In short, although Ptr remains in bulk water, it is able tophotoinduce membrane damage in simple model systems (LUVs), as well as in eukaryotic cells.
In the search of better compounds that retain the photosensitizing properties of pterins and, at the same time, areable to bind to biomembranes, a series of decyl-pterin derivatives were synthesized.2 Among them, due to its
photochemical properties (efficient intersystem crossing), 4-(decyloxy)pteridin-2-amine (O-decyl-Ptr) was chosen
for further studies using phospholipid membranes with various compositions. Conjugation of a decyl chain to thepterin moiety enables its facile intercalation in LUVs. In particular, O-decyl-Ptr is positioned right between thepolar head and the beginning of the fatty acid chains.3 Upon UVA irradiation lipid peroxidation photosensitized by
O-decyl-Ptr leads to the formation of hydroxyl derivatives, hydroperoxides and hydroxyhydroperoxides.4 These
photoproducts undergo a fast conversion into short-chain secondary products by cleavage of the fatty acid chains
most likely due to further photosensitized processes. These short-chain oxidized lipids are responsible fordestabilizing the phospholipid bilayer and promoting membrane leakage. The efficiency of photodamage, assessedin terms of oxidized products formation rate and membrane permeabilization, is much higher for O-decyl-Ptr thanfor free Ptr,4 which indicates that the intercalation of the alkyl-pterin to the membrane enhances the photosensitizedreactions. O-decyl-Ptr is also much more efficient than Ptr in the photodynamic activity on HeLa cells.3
References
1) A. H. Thomas, A. Catalá, M. Vignoni, Soybean phosphatidylcholine liposomes as model membranes to study
lipid peroxidation photoinduced by pterin, Biochim. Biophys. Acta 1858 (2016) 139–145.
2) M. Vignoni, N. Walalawela, S. M. Bonesi, A. Greer, A.H. Thomas, Lipophilic decyl chain-pterin conjugateswith sensitizer properties, Mol. Pharm. 15 (2018) 798-807.
3) A. Vignoni, C. Layana, H.C. Junqueira, A.H. Thomas, R. Itri, M.S. Baptista, M. Vignoni, Alkylation of ahydrophilic photosensitizer enhances the contact-dependent photo-induced oxidation of phospholipid membranes,
Dyes Pigm. 187 (2021) 109131.
4) M. Vignoni, M. N. Urrutia, H. C. Junqueira, A. Greer, A. Reis, M. S. Baptista, R. Itri, A .H. Thomas, PhotoOxidation of unilamellar vesicles by a lipophilic pterin: deciphering. biomembrane photodamage, Langmuir 34(2018) 15578-15586.

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IL-2.1.3

Heavy-Atom-Free Photosensitizers for the Treatment of Cancer Cells
Carlos E. Crespo-Hernández,
Case Western Reserve University, Department of Chemistry, Cleveland, Ohio, 44106, USA
Heavy-atom-free photosensitizers based on thionation of carbonyl groups of readily accessible organic compounds
are rapidly emerging as a versatile class of molecules1. A significant portion of our research group has focused onunderstanding how thionation regulates the electronic relaxation pathways in nucleic acid bases2 and other organic
chromophores3 from experimental and computational perspectives and the application of this fundamental
information for the development of all-organic photosensitizers as therapeutic drugs to treat cancer cells. In the firstpart of this presentation, I will show that fundamental physicochemical investigations can be used to develop
analogues of the canonical DNA and RNA nucleobases exhibiting more than 100 nm redshifted absorption spectraand nearly 100% greater photoreactivity, which, when applied in vitro with a low dose of light, substantially decrease
the proliferation of skin cancer cells1,4. In the second part, I will discuss the generalization of this chemistry for thedevelopment of heavy-atom-free photosensitizers absorbing near-infrared radiation and exhibiting high yields ofsinglet oxygen generation for the prospective treatment of cancer cells3,5.

Fig. 1 – Development of all-organic photosensitizers based on thionation of carbonyl functional groups. In the chemical
structures, yellow, red, blue, gray, and white, represent sulfur, oxygen, nitrogen, carbon, and hydrogen, respectively.

References
1) Ortiz-Rodríguez, L. A.; Crespo-Hernández, C. E., Thionated Organic Compounds as Emerging Heavy-Atom-Free
Photodynamic Therapy Agents, Chem. Sci. 11, 11113-11123 (2020).
2) Ashwood, B.; Pollum, M.; Crespo-Hernández, C. E., Photochemical and Photodynamical Properties of SulfurSubstituted Nucleic Acid Bases, Photochem. Photobiol. 95, 33-58 (2019).
3) Ortiz-Rodríguez, L. A.; Hoehn, S. J.; Loredo, A.; Wang, L.; Xiao, H.; Crespo-Hernández, C. E., Electronic
Relaxation Pathways in Heavy-Atom-Free Photosensitizers Absorbing Near-Infrared Radiation and Exhibiting High
Yields of Singlet Oxygen Generation, J. Am. Chem. Soc. 143, 7, 2676–2681 (2021).
4) Pollum, M.; Minh, L.; Jockusch, S.; Crespo-Hernández, C. E., Dithionated Nucleobases as Effective
Photodynamic Agent Against Human Epidermoid Carcinoma Cells, ChemMedChem 13, 1044-1050 (2018).

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5) J. Tang , L. Wang , A. Loredo , C. Cole and H. Xiao, Single-Atom Replacement as a General Approach towards
Visible-Light/Near-Infrared Heavy-Atom-Free Photosensitizers for Photodynamic Therapy, Chem. Sci. 11 , 67016708 (2020).
Acknowledgements. The author acknowledges the National Science Foundation (Grant No. CHE-1800052). This
work made use of the High-Performance Computing Resource in the Core Facility for Advanced Research
Computing at CWRU.

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OC-2.1.4
Mechanistic Organic Photochemistry: Dark Processes and Toxicity Priming
Shakeela Jabeen1,2, Andrés M. Durantini1-3, and Alexander Greer1,21) Brooklyn College, City University of New York, Department of Chemistry, Brooklyn, New York, United States
2) The Graduate Center of the City University of New York, Ph.D. Program in Chemistry, New York, New York,
United States
3) IDAS-CONICET, Universidad Nacional de Río Cuarto, Departamento de Química, Río Cuarto, Córdoba,
Argentina
Results will be described with the use of air/liquid interfaces and particles as a way to deepen our understanding ofreactive oxygen intermediates. One result in Figure 1 is an accomplishment in sorting out light/dark processes thatare intertwined. Deconvolution of competitive light and dark processes offer mechanistic insight into a singlet oxygen
priming mechanism1. New insight is being added to solve mechanistic puzzles in the production of reactive oxygenin primary light-dependent and secondary dark processes. Our work with multiphasic systems has provided insightto light-to-dark processes in the destabilization of biological membranes and surfaces of environmental particles2.

Fig. 1 – Schematic showing that complementary light and dark reactions arise in separable processes. We focus on lightdependent reaction of singlet oxygen, and light-independent reaction of hydroperoxides.

References
1) Jabeen, S.; Farag, M.; Malek, B.; Choudhury, R.; Greer, A. A Singlet Oxygen Priming Mechanism: Disentanglingof Photooxidative and Downstream Dark Effects. J. Org. Chem. 2020, 85, 12505-12513.
2) Durantini, A. M.; Greer, A. Interparticle Delivery and Detection of Volatile Singlet Oxygen at Air/solid Interfaces.
Environ. Sci. Technol. 2021, 55, 3559-3567.
Acknowledgements. This work was supported by the US National Science Foundation (CHE-1856765).

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IL-2.1.5

Photosensitization by tyrosine kinase inhibitors
Miguel A. Miranda,1,2 Ignacio Vayá,1,2 Inmaculada Andreu1,21) Departamento de Química/Instituto de Tecnología
Química UPV-CSIC, Universitat Politècnica deValència, Camino de Vera s/n, 46022-València, Spain.

2) Unidad Mixta de Investigación UPV-IIS La Fe,
Hospital Universitari i Politècnic La Fe, Fernando
Abril Martorell 106, 46026-Valencia, Spain.

The epidermal growth factor receptors (EGFRs) are transmembrane glycoproteins with tyrosine kinase
activity. They are able to regulate a number of signaling pathways within cells including cell proliferation,
migration, differentiation, tissue repair and wound healing. Mutations and overexpression of the tyrosine
kinase receptors may result in the appearance of different types of cancers and may promote solid tumor
growth. Therefore, EGFRs are major targets for the design of anticancer agents. In this regard, tyrosine
kinase inhibitors (TKIs) are of high interest due to their ability to block the kinase activity of these receptors.
Many drugs are known to absorb solar radiation, and can induce photosensitivity reactions, such as
phototoxicity or photoallergy, but also photoaging, weakening of the immune system and skin cancer. Theseside effects can be associated with damage to biomolecules mediated by radicals or reactive oxygen species
arising from excited singlet or triplet states. Interestingly, drugs containing the quinazoline moiety (thecommon chromophore of a family of TKIs) are known to produce skin photosensitivity.
In this context, as an extension of our previous work on Imatinib, we have now investigated the photobiologicalresponse of Lapatinib and Gefitinib. Thus, their phototoxic potential has been evaluated by means of the NRU assay,
while their photooxidation activity has been assessed towards HSA, the main transport protein in human serum.
Besides, fluorescence and transient absorption spectroscopies from the femtosecond to the microsecond time-scale
have been used to investigate the photobehavior of these TKIs in solution, as well as in the presence of HSA or incellular systems. As a result, it has been observed that the excited state properties of these drugs are strongly affectedby the environment: locally excited singlet states are mainly formed in organic non-polar solvents and within HSAor cells, while intramolecular charge transfer sates predominate in organic polar solvents. In the case of Gefitinib,
the triplet excited state has been identified and completely characterized for the first time, and its potential to generate
ROS has also been assessed. In general, a good correlation is established between the photophysical behavior

and the photobiological properties of the investigated TKIs, which provides a mechanistic basis for theobserved phototoxicity. All the above features are of key importance in connection with the photosensitizing

potential of this family of drugs.

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IL-2.1.6
Singlet molecular oxygen reaction with biomolecules: Mechanistic studies using
18
O-labeled oxygen, mass spectrometry, and light emission measurements
Paolo Di Mascio
Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP, Brazil
In mammalian tissues, ultraweak chemiluminescence arising from biomolecules oxidation has been attributed byseveral authors to the radiative deactivation of singlet molecular oxygen [1O2] and excited triplet carbonyl products
as dioxetane intermediates1,2. Singlet molecular oxygen has been shown to be generated in biological systems andhave been implicated in cell defense mechanisms and dark reactions. Singlet oxygen that may be produced among
various possibilities by myeloperoxidase, an enzyme implicated in inflammation processes, and by the UVA
component of solar radiation is also a major source of peroxidation of several key cellular components. Singlet
oxygen exhibits a substantial reactivity toward electron-rich organic biomolecules (Fig 1A) including protein, lipids,
RNA and DNA, leading to the formation of allylic hydroperoxides, dioxetanes or endoperoxides1.
It was also shown that biomolecules oxidation reactions which often involve 1O2-mediated peroxidation of nucleic
acids, unsaturated lipids and targeted amino acids3 are implicated in several diseases including advanced humanarteriosclerosis, arthritis, cataract and diabetes. The measurement of near-infrared light emission and the use of [18O]labeled 1O2 and hydroperoxides generated via a clean naphthalene derivative [18O]-labeled thermolabile
endoperoxide (DHPN18O2) as a source of [18O]-labeled 1O2 (Fig.1B), can be applied to study mechanistic aspects
related to 1O2 generation and reactions in biological systems4. A relevant major topic deals with the search for themolecular signature of the 1O2 formation in targeted biomolecules within cells. The use of [18O]-labeled 1O2 released
from thermolabile endoperoxides in association with sensitive and accurate LC-ESI-MS/MS analysis provides ahighly suitable way to gain relevant mechanistic insights into the formation and the decomposition pathways of
initially 1O2 generated peroxidic compounds and to search for characteristic biomarkers in cells.
16O
16O

R
R

R

Δ (37oC)+< 0 - 5oC

16[1O ]
2

R
R= -CH3, -CH2OH, -(CH2)2COO-,-CH2SO3-, -(CH2)2CONHCH2CHOHCH2OH
18O

18O

R1

R1
R1

H
N

R1 =

O

DHPN18O2
18O

18O

Δ

+

OH
OH

18[1O ]
2

R1
DHPN
R2

R2

R2
DPA18O2 or EAS18O2

R2
R2 = -C6H5 (DPA), -OSO3- (EAS)

Fig. 1 – (A) 1O2 is capable of efficiently reacting with cellular constituents1. (B) Naphthalene and anthracene derivatives4 forrelease and capture of 1O2.

References
1) Paolo Di Mascio et al., Chem. Rev., 119, 2043-2086, 2019. 2) Luiz D. Ramos et al., Photochem. Photobiol.,
97,327-334, 2021. 3) Stanley, C. P. et al. Nature, 566, 548-552, 2019. 4) Marilene S. Oliveira et al., Photochem.
Photobiol. Sci., 19, 1590-1602, 2020.
Acknowledgements This work was supported by grant from FAPESP (N°. 2012/12663-1, CEPID Redoxoma N°
2013/07937-8), CNPq (N° 302120/2018-1) and PRPUSP (NAP Redoxoma N° 2011.1.9352.1.8).

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IL-2.1.7
Metallodrug Photosensitizers for Light-Based Cancer Therapy
Sherri A. McFarland1, Colin G. Cameron1, John A. Roque III1,2, Houston D. Cole1, Ge Shi1
1) The University of Texas at Arlington, Department of Chemistry and Biochemistry, Arlington, TX United States;
2) The University of North Carolina at Greensboro, Department of Chemistry and Biochemistry, Greensboro, NC
United States
There has been an ongoing interest in the development of new photosensitizers (PSs) for photodynamic therapy(PDT). A related area of active investigation has been the design of PSs with novel mechanisms of action, including
oxygen-independent photoprocesses (known as photochemotherapy, or PCT) or the capacity to switch to such modes
at low oxygen tension. Since tumor hypoxia can present a real challenge for PDT in certain cases, PSs that operate
through PCT mechanisms offer the possibility of treating some of the most aggressive and drug-resistant tumors that
resist the traditional photodynamic reactions. Transition metal complexes have emerged as attractive PSs for both
PDT and PCT. Certain coordination complexes of ruthenium (Ru) are potent phototoxins toward a variety of in vitro
and in vivo cancer models. One example is our own TLD1433, which is currently in a Phase 2 clinical trial for treating
nonmuscle invasive bladder cancer with PDT. Part of the interest in Ru PSs stems from the ability to access a variety
of excited state electronic configurations with visible or near-infrared light by judicious choice of ligand
combinations around the metal center. These excited states, in turn, may participate in traditional type I/II
photodynamic reactions as well as oxygen independent pathways that form the basis of PCT. In this conference
presentation, we will discuss the design and development of transition metal complex PSs that exploit both PDT and
PCT effects. The emphasis will be on the structural features and photophysical models that give rise to excited triplet
states with characteristic reactivities.
Acknowledgements. This work was supported in part by the National Cancer Institute (NCI) of the National
Institutes of Health (NIH) (Award R01CA222227). The content in this work is solely the responsibility of
the authors and does not necessarily represent the official views of the National Institutes of Health.

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OC-2.1.8

Inhibition of 6-formylindolo[3,2-b]carbazole metabolism sensitizes keratinocytes to
UVA-induced apoptosis: Implications for drug-induced phototoxicity
Katharina M. Rolfes1, Natalie C. Sondermann1, Motoki Nakamura2, Christian Vogeley1, Jean Krutmann1 andThomas Haarmann-Stemmann1
1) IUF – Leibniz Research Institute for Environmental Medicine, Duesseldorf, Germany; 2) Department ofEnvironmental and Geriatric Dermatology, Nagoya City University, Nagoya, Japan
Ultraviolet (UV) radiation induced DNA damage and associated signalling pathways are mainly responsible for avariety of acute and chronic cutaneous adverse effects. In this context, energy-rich UVB rays are of particularimportance, as they are mainly absorbed by the DNA of epidermal keratinocytes. If not fixed by specialized DNA
repair systems or removed through the apoptotic demise of damaged cells, these lesions may cause mutations andthereby eventually initiate skin carcinogenesis. In addition, the aryl hydrocarbon receptor (AHR), a ligand-activatedtranscription factor has been identified as a key regulator of the UVB-induced stress response in epidermal
keratinocytes1. In UVB-irradiated skin, AHR is activated by the intracellular generated tryptophan photoproduct 6formylindolo[3,2b]carbazol (FICZ) and, amongst others, mediates immunosuppressive and anti-apoptotic effects.
Therefore, transient inhibition of AHR is widely regarded as a suitable strategy to protect skin against the damaging
effects of UVB radiation1. In contrast to UVB radiation, UVA-photons are barely absorbed by DNA, but rather
through endogenous and exogenous photosensitizers. Excitation of these photosensitizers results in ROS-induced
oxidative DNA damages and apoptosis. Interestingly, FICZ was identified as a potent, nanomolar UVA
photosensitizer in epidermal keratinocytes2. The fact that FICZ induces the expression of the AHR target gene
cytochrome P450 (CYP) 1A1, which is mainly responsible for the metabolism of this tryptophan photoproduct3,
implies that a modulation of AHR activity has a direct effect on the UVA-induced phototoxicity of FICZ. Indeed, we
confirmed that a modulation of AHR and/or CYP1A1 enzyme activity affected the intracellular FICZ level and thus
the UVA-induced phototoxicity of FICZ. AHR-dependent CYP1A1 induction decreased the phototoxicity of
FICZ/UVA-treatment, whereas inhibition of AHR had the opposite effect. Even though FICZ levels are quite low
and a transient inhibition of AHR and associated CYP1A1 inhibition may not result in an accumulation of UVAreactive quantities, chronic inhibition of AHR/CYP1A1 however might reach UVA-reactive levels of FICZ.
Therefore, this scenario might be of clinical relevance in the context of drug-induced phototoxicity. Interestingly,
one of the most prominent examples of phototoxic drugs, namely the BRAF protein kinase inhibitor vemurafenib isalso a potent inhibitor of AHR4. This drug is approved for the treatment of advanced melanoma with activating BRAF
V600 mutations. In addition, we found out that vemurafenib is not only an antagonist of AHR but also a substrate of
CYP1A1 and therefore influences the metabolic degradation of FICZ and associated UVA-induced phototoxicity. In
fact, the obtained results indicate that a modulation of FICZ metabolism might be a relevant molecular mechanism
responsible for the UVA-induced phototoxicity of drugs. Induction of epidermal CYP1A1 enzyme activity therefore
might be a suitable therapeutic approach to minimize phototoxicity in patients suffering from melanoma.
References:
1) Vogeley C, Esser C, Tüting T, Krutmann J, Haarmann-Stemmann T. Role of the aryl hydrocarbon receptor inenvironmentally induced skin aging and skin carcinogenesis. Int J Mol Sci 20:6005, 2019.
2) Park SL, Justiniano R, Williams JD, Cabello CM, Qiao S, Wondrak GT. The Tryptophan-Derived Endogenous
Aryl Hydrocarbon Receptor Ligand 6-Formylindolo[3,2-b]Carbazole Is a Nanomolar UVA Photosensitizer in
Epidermal Keratinocytes. J Invest Dermatol 135:1649-58, 2015
3) Bergander L, Wincent E, Rannug A, Foroozesh M, Alworth W, Rannug U. Metabolic fate of the Ah receptor
ligand 6-formylindolo[3,2-b]carbazole. Chem Biol Interact 149:151-64, 2004.
4) Hawerkamp HC, Kislat A, Gerber PA, Pollet M, Rolfes KM, Soshilov AA, Denison MS, Momin AA, Arold ST,
Datsi A, Braun SA, Oláh P, Lacouture ME, Krutmann J, Haarmann-Stemmann T, Homey B, Meller S. Vemurafenib
acts as an aryl hydrocarbon receptor antagonist: Implications for inflammatory cutaneous adverse events. Allergy
74:2437-48, 2019.
Acknowledgements: This work was supported by the Jürgen Manchot Stiftung.
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P-2.1.9

Cytotoxicity and redox profile of a novel transition-metal nitrosyl compound forphoto-controlled NO delivery in human fibroblasts
Hande Özbaşak1, Hani Mohamed Abdullah Elbeheiry2, Erika Csekes1, Martin Schulz2, Lucia Račková1
1) Centre of Experimental Medicine Slovak Academy of Sciences, Institute of Experimental Pharmacology and
Toxicology, Bratislava, Slovakia; 2) Friedrich Schiller University Jena, Institute of Physical Chemistry, Jena,
Germany
The discovery that nitric oxide (NO) plays important roles as a biological messenger in a wide range ofphysiological processes has stimulated great interests in the studies of its chemical and biochemical properties,
especially the investigation of NO-mediated events. Among NO releasing compounds, poly-pyridine ruthenium
complexes containing nitrosyl group are subject to increasing amounts of research in recent years1. As NO
concentration is crucial and decisive factor of desired effects, ruthenium-terpyridine is highly prized as it is possible
to tweak concentration of NO released with L2 ligand site. The derivative HE-10, a ruthenium (II) nitrosyl-complex
incorporating 4′-phenyl-terpyridine and o-benzoquinone diimine ([Ru(ptp)(o-bqdi)NO](PF6)3), is a newly
synthesized photoinducible NO releasing compound. In this study, it was aimed to investigate the cytotoxic andprooxidant effects likely related to NO release induced from HE-10 with white LED light stimulation in the VH10
fibroblast cell line.
After pre-treatment of the cells with HE-10, 1 hour light exposure and following overnight incubation, theeffects on cell viability under light and dark conditions were determined by MTT assay. Moreover, NO2- levels in
the medium were detected by Griess method and intracellular reactive species formations in VH10 cells were
determined by H2DCF-DA probe. Cell cycle analysis under light and dark conditions was performed by flow
cytometry. Protein nitration levels, measured as 3-nitrotyrosine, in VH10 cells were detected by western blot.
Our data show that cytotoxicity of HE-10 was enhanced by exposure to white LED light, as confirmed bylower IC50 value obtained from the HE-treated light-exposed cells compared to the unexposed ones. Cytotoxicity was
accompanied by increased intracellular reactive oxygen/nitrogen species (ROS/RNS) production, G2/M cell cycle
arrest and protein nitration, depending on both, light exposure and concentration. Moreover, light-exposureenhanced toxic effect of HE-10 on VH10 fibroblast cells was most likely to be NO-dependent as confirmed by the
induced NO2- release in the medium, intracellular oxidation of H2DCF-DA probe sensitive to ROS and RNS in
fibroblasts, and increased protein nitration levels. According to the cell cycle analysis, it was determined that HE-10
decreased cell proliferation and antiproliferative effect was enhanced with light exposure.
With this research, we successfully demonstrated that novel ruthenium−terpyridine compound HE-10 iscapable of selective cytotoxicity induced by light exposure through releasing NO to VH10 fibroblast cells. Our results
indicate that HE-10 is promising in terms of causing cell death in proliferative VH10 cells which could potentiallyapply to other cell lines which will be investigated further. Further experiments will be conducted to confirm DNA
damage potentially caused by ROS and investigate the involved molecular pathways. All in all, our results suggestthat the novel compound HE-10 might be a promising candidate in terms of photoinducible therapies, such as
ruthenium-nitrosyl-based cancer therapy2.
References
1) Xiang HJ, An L, et al. Photo-controlled targeted intracellular delivery of both nitric oxide and singlet oxygenusing a fluorescence-trackable ruthenium nitrosyl functional nanoplatform. Chem Commun. 51: 2555-8,
2015.
2) Orlowska E, Babak MV, et al. NO Releasing and Anticancer Properties of Octahedral Ruthenium-Nitrosyl
Complexes with Equatorial 1H-Indazole Ligands. Inorg Chem. 57: 10702-17, 2018.
Acknowledgements. This work was supported by the Slovak Research and Development Agency under the Contract
no. APVV-18-0336 and LogicLab ITN funded by the European Union’s Horizon 2020 research and innovation
program under the Marie Skłodowska-Curie [grant agreement No 813920].
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P-2.1.10

Assessment of the phototoxicity induced by talazoparib
Alejandro Mateos-Pujante1,2, M. Consuelo Jiménez1,2, Inmaculada Andreu1,22) Unidad Mixta de Investigación Universitat
1) Departamento de Química-Universitat Politècnicade València, Camino de Vera s/n, 46022, Valencia,
Politècnica de València -Instituto de Investigación
Spain.
Sanitaria (IIS) La Fe, Hospital Universitari i
Politècnic La Fe, Avenida de Fernando Abril
Martorell 106, 46026, Valencia, Spain.
Breast cancer is the most common cancer in women and the second leading cause of cancer death around the world.
It is estimated that 1 in 8 women are likely to develop breast cancer in their lifetime1. Therefore, there is a need fordeveloping different treatments to choose the most suitable treatment for each patient.
In this context, Poly(ADP-ribose) polymerase inhibitors (PARPi) have emerged as potent antitumoral drugs, as theirability to inhibit PARP activity. PARPs are a family of multifunctional enzymes that play essential roles in cellular
processes. Among them, PARP-1 and PARP-2 plays an important role in DNA repair catalyzing the repair ofendogenous single strand breaks (SSB) that occurs frequently in cells, which is essential for their survival. However,a non-efficient SSB repair can be converted to double strand breaks (DSB), that are toxic to cells and must be repairedby other mechanisms such as homologous recombination (HR)Given this background, talazoparib, the last PARPi approved by the FDA in 2018, exerts its cytotoxic effect byinhibiting PARP activity, provoking the formation and accumulation of DSB. Thus, cells with BRCA 1/2 mutations
are unable to repair this DNA damage by HR, which cause further double strand DNA breaks and subsequently, thecellular death. Therefore, talazoparib is used for the treatment of breast tumours with mutations in genes BRCA 1/2,
which induced apoptosis in those cancer cells that are unable to repair DNA damage by HR2,3.
Regarding the photosafety of talazoparib (TLP), although photosensitivity reactions have not been reported yet, inthe present work we have assessed the phototoxicity induced by TLP in cells through Neutral Red Uptake (NRU)
assay. Hence, it is necessary to perform photophysical studies to know the phototoxicity mechanism involved. As it
is shown in Fig. 1, TLP displays a triplet excited stated in PBS, which is quenched by oxygen. Moreover, to evaluate
if the phototoxicity to keratinocyte (HaCaT) cells obtained can be attributed to DNA photosensitized damage, cometassay was performed, indicating a photo(geno)toxicity damage around 50 %. These results seem very interesting toprevent photosensitivity reactions associated to TLP.

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N2
Air
O2

0.8
0.6
0.4
0.2
0.0

0

5

10

Time (
 s)

15

120

Cell viability (%)

 A normalized

1.0

Dark
UVA-light

80

40

0

0

1

2

3

log [TLP] ( M)Fig. 1 – From left to right: Chemical structure of TLP, LFP signal decay monitored at 400 nm in PBS and cell viability doseresponse curve of HaCaT cells treated with TLP.

References
1) Key T.J., Verkasalo P.K., Banks E., Epidemiology of breast cancer. Lancet Oncol. 2(3) 133-140, 2001.
2) Zheng F., Zhang Y., Chen S., Weng X., Rao Y., Fang H, Mechanism and current progress of Poly ADP-ribose
polymerase (PARP) inhibitors in the treatment of ovarian cancer. Biomed. Pharmacother. 123:109661, 2020.
3) Hurvitz S.A. et al., Talazoparib in patients with a germline BRCA-mutated advanced breast cancer: detailed safety
analyses from the phase III EMBRACA trial. Oncologist 0493, 2019.
Acknowledgements. This work was supported by Generalitat Valenciana (ACIF/2018/159), the Spanish Ministry of
Science and Innovation (PID2019-105391GB-C22, CTQ2016-78875-P, BEAGAL 18/00211) and Carlos III Instituteof Health (Grant PI16/01877).

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P-2.1.11

A study of the photophysical properties of adapalene.
Juan Antonio Soler, Miguel A. Miranda, Virginie Lhiaubet-Vallet
InstitutoUniversitario Mixto de Tecnología Química (UPV-CSIC), Universitat Politècnica de València, Consejo
Superior de Investigaciones Científicas, Avda de los Naranjos s/n, Valencia, Spain
Adapalene, a third-generation topical retinoid, is used as a treatment for acne vulgaris. It is currently one of the three
topical retinoids approved so far by the Food and Drug Administration (FDA), along with tazarotene and tretinoin.1
However, combination of adapalene and sun exposure might result in the appearance of adverse effects. They havebeen associated with different causes such as skin irritation that can upset the natural skin photoprotection andincrease ultraviolet rays hurt, or a decrease of the thickness of the stratum corneum reducing the natural photobarrier.2
Moreover, as shown in Figure 1, adapalene exhibits an important UVB/UVA absorption. Thus, like other topicaldrugs,3 it might act as a potential photosensitizer toward biological components and be responsible for different
processes related to phototoxicity or photoallergy.
7x104
6x104

e (M-1cm-1)

5x104
4x104
3x104
2x104
1x104
0

250

300

350

400

450

500

l (nm)

Fig. 1 – Left, structure of adapalene molecule. Right, UV-vis spectra of adapalene in acetonitrile at 10-5 M.

The aim of this study is to determine the photophysical properties of adapalene to investigate its potential
photoactivity toward biomolecules. For this, experiments combining UV-vis spectroscopy, fluorescence andphosphorescence (steady-state and time-resolved) and laser flash photolysis were carried out. An efficient
fluorescence emission centered at 425 nm and with a quantum yield of ca. 0,8 was evidenced together with aphosphorescence band at 520 nm. Moreover, laser flash photolysis experiments revealed a triplet-triplet transientabsorption with a lifetime of 40 s, which was efficiently quenched in the presence of oxygen and -carotene. A pH
dependent photobehavior was also observed.
References
1) Kolli, S. S., Pecone, D., Pona, A., Cline, A. and Feldman, S. R., Topical Retinoids in Acne Vulgaris: A Systematic
Review. Am J Clin Dermatol, 20, 345-365, 2019.
2) Zaenglein, A. L, Topical Retinoids in the Treatment of Acne Vulgaris. Semin. Cutan. Med. Surg., 27, 177-182,

2008.

3) Lhiaubet-Vallet, V., Miranda, M. A., Phototoxicity of Drugs, In CRC Handbook of Organic Photochemistry andPhotobiology, Eds. F. Ghetti, A. G. Griesbeck and M. Oelgemöller, CRC Press, Chap. 66, pp. 1541-1555. 2012.

Acknowledgements.
This work has been supported via the pre doctoral FPI fellowship program from the Spanish Government and a
Project of the Spanish Ministry of Science, Innovation and Universities.

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P-2.1.12

Photobehavior of the tyrosine-kinase inhibitor gefitinib in solution and within humanserum albumin
L. Tamarit1,2, M. El Ouardi1,2, I. Andreu1,2, I. Vayá1,2, M. A. Miranda1,2
1) Departamento de Química/Instituto de Tecnología Química UPV-CSIC, Universitat Politècnica de València,
Camino de Vera s/n, 46022 València (Spain); 2) Instituto de Investigación Sanitaria La Fe, Hospital Universitari i
Politècnic La Fe, Avenida de Fernando Abril Martorell 106, 46026 Valencia (Spain)
Gefitinib (GFT) is an orally active first-generation tyrosine-kinase inhibitor (TKI), which is currently used for thetreatment of patients with non-small cell lung cancer; the mode of action involves specific binding of GFT to the
ATP site of HER1 preventing autophosphorylation in tumor cells.1,2 Although the benefits of this drug are evident, it
can also induce adverse effects, which are normally associated to rash, diarrhea, dry skin, nausea and vomiting.3
Interestingly, GFT contains the quinazoline moiety, which is known to induce damage by photosensitization. In thisregard, it has been recently reported that lapatinib (LAP), a TKI used for the treatment of breast and lung cancer, caninduce photooxidation and phototoxicity.4–6 Therefore, interaction of GFT with UV light could also induce
photosensitivity disorders. Thus, a thorough spectroscopic study combining ultrafast transient absorption
spectroscopy, fluorescence and laser flash photolysis has been performed on GFT in solution and in biologicalmedium.
The photobehavior of GFT, which is summarized in Fig. 1, has been investigated in organic solvents of different
polarities and within human serum albumin (HSA), the most abundant transport protein in the blood-stream.7

Fig. 1 – Schematic representation of the main photophysical processes arising from irradiation of GFT in polar and non-polarorganic solvents and within HSA.

The results show that the photobehavior of GFT was strongly affected by the environment. In general, LE singlet
species was instantaneously formed. In organic non-polar solvents, they emit at short wavelengths (~380 nm) withmoderate quantum yields (~0.19) and short lifetimes (ca. 1.3 ns). This species can also cross to form the triplet
excited state of GFT (3GFT*), which takes place in about 10 ps in toluene. By contrast, in organic polar solvents, LE
states rapidly evolve (~1 ps) towards the formation of ICT states. These species emit at longer wavelengths anddisplay higher lifetimes (ca. 3.4 ns) than those of LE states; they are also able to populate 3GFT* in MeCN.
Surprisingly, this process was not observed in ethanol, since ICT states are deactivated in about 0.7 ns, andconsequently the triplet state of the drug cannot be formed. Interestingly, formation of 1O2 has been detected in bothorganic polar and non-polar solvents, which in principle might have significant implications in the photosensitizing
properties of GFT. However, it is worth to note that 3GFT* has not been detected within the protein cavities;
consequently, the photosensitizing potential of GFT could not be attributed to this species but more likely to a type I
reaction from LE singlet states, which are the only excited species observed in the biological environment.
References
1)
Cohen MH, Williams GA, Sridhara R, Chen G, Pazdur R. FDA Drug Approval Summary: Gefitinib(ZD1839) (Iressa ® ) Tablets . Oncologist. 2003;8(4):303-306. doi:10.1634/theoncologist.8-4-303
2)
Arora A, Scholar EM. Role of tyrosine kinase inhibitors in cancer therapy. J. Pharmacol. Exp. Ther.
2005;315(3):971-979. doi:10.1124/jpet.105.084145
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3)
4)
5)
6)
7)

Cersosimo RJ. Gefitinib: An adverse effects profile. Expert Opin. Drug Saf. 2006;5(3):469-479.
doi:10.1517/14740338.5.3.469
García-Lainez G, Vayá I, Marín MP, Miranda MA, Andreu I. In vitro assessment of the photo(geno)toxicity
associated with Lapatinib, a Tyrosine Kinase inhibitor. Arch. Toxicol. 95(1):169-178, 2021
Vayá I, Andreu I, Lence E, et al. Characterization of the locally excited and charge-transfer states of the
anticancer drug lapatinib by ultrafast spectroscopy and computational studies. Chem. Eur. J. 26: 1592215930, 2020
Andreu I, Lence E, González-Bello C, et al. Protein Binding of Lapatinib and Its N- and O-Dealkylated
Metabolites Interrogated by Fluorescence, Ultrafast Spectroscopy and Molecular Dynamics Simulations.
Front Pharmacol. 2020;11. doi:10.3389/fphar.2020.576495
Peters T. Ligand Binding by Albumin. All About Albumin. Published online 1995:76-132.
doi:10.1016/b978-012552110-9/50005-2

Acknowledgements. Financial support from the Spanish Government (RYC-2015-17737, CTQ2017-89416-R and
BEAGAL 18/00211), the Carlos III Institute of Health (Grant PI16/01877) and Conselleria d’Educació Cultura i
Esport (PROMETEO/2017/075) is gratefully acknowledged.

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P-2.1.13

Hybrid nanomaterials based on graphene quantum dots for photodynamic therapy
Cristina J. Dias1,2, Carla I. M. Santos1,3, Fátima Liliana Monteiro2, Laura Rodríguez-Pérez4, Gil Gonçalves5, MariaA. F. Faustino1, Sandra Vieira2, Luisa Helguero2, M. Ángeles Herranz4, Nazario Martin4, M. Graça P. M. S.
Neves1, José M. G. Martinho3, Ermelinda M. S. Maçôas3
1) LAQV-REQUIMTE, Department of Chemistry, University of Aveiro, Campus Universitário de Santiago, 3810193 Aveiro, Portugal; 2) iBiMED - Institute of Biomedicine, Department of Medical Sciences, University of Aveiro,Campus Universitário de Santiago, 3810-193 Aveiro, Portugal; 3) CQE, Centro de Química Estrutural, Instituto
Superior Técnico, Av. Rovisco Pais, 1049-001 Lisboa, Portugal; 4) Department of Organic Chemistry, Faculty ofChemistry, Universidad Complutense de Madrid, E-28040 Madrid, Spain; 5) TEMA-Nanotechnology ResearchGroup, Mechanical Engineering Department, University of Aveiro, Campus Universitario de Santiago, 3810-193
Aveiro, Portugal
Cancer appears as the second leading cause of death worldwide and, according to data from 2020, breast cancer was
the most incident1. Therefore, there is an urgent need to find effective therapies to achieve a better treatment
efficiency. Under this context, Photodynamic Therapy (PDT) has been extensively investigated in the treatment ofseveral types of cancer with positive outcomes2. Porphyrins and analogues are recognised to be promising
photosensitisers (absorption in the red region of the electromagnetic spectrum and good generation of 1O2) but their
low solubility in physiological medium can compromise their PDT efficiency. Therefore, the development ofphotoactive molecules coupled to drug delivery systems is of prominent interest2. Carbon nanomaterials, in particulargraphene quantum dots (GQDs) are promising candidates as vehicles to deliver photoactive molecules to cancer cells
in PDT due to their high photostability, water solubility, biocompatibility and excellent optical properties3.
In this study we discuss the biological evaluation of two biocompatible hybrids based on GQD covalently linked toa porphyrin bearing a 2-aminoethylamino chain. The photoefficiency of the hybrids nanomaterials and cellular
localization were studied in a T-47D breast cancer cell line.
References
1) Sung, H.; Ferlay, J.; Siegel, R.L.; Laversanne, M.; Soerjomataram, I.; Jemal, A.; Bray, F. Global Cancer Statistics
2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA. Cancer J.
Clin. 0, 1–41, 2021.
2) Mesquita, M.Q.; Dias, C.J.; Gamelas, S.; Fardilha, M.; Neves, M.G.P.M.S.; Faustino, M.A.F. An insight on therole of photosensitizer nanocarriers for Photodynamic Therapy. Ann. Brazilian Acad. Sci. 90, 1101–1130, 2018.
3) Santos, C. I. M.; Gonçalves, G.; Cicuéndez, M.; Mariz, I.; Silva, V. S.; Oliveira, H.; Campos, F.; Vieira, S. I.;
Marques, P. A. A. P.; Maçôas, E. M. S.; Neves, M. G. P. M. S.; Martinho, J. M. G. Biocompatible Hybrids Based on
Nanographene Oxide Covalently Linked to Glycolporphyrins: Synthesis, Characterization and Biological Evaluation.
Carbon N. Y. 135, 202–214, 2018.
Acknowledgements. Thanks are due the University of Aveiro, Instituto Superior Técnico (IST, Portugal) andFCT/MCTES (Fundação para a Ciência e Tecnologia/Ministério da Ciência e Tecnologia, Portugal) for the financial
support to LAQV-REQUIMTE (UIDB/50006/2020) and (UID/NAN/50024/2019) and Institute for Biomedicine –iBiMED (UID/BIM/04501/2020); the projects: (UID/EMS/00481/2013), (PTDC/FIS-NAN/4662/2014),(PTDC/NAN-MAT/29317/2017), (PTDC/QUI-QFI/29319/2017), (LISBOA-01-0145-FEDER-029319) and(UIDB/00100/2020) C. J. Dias and F. L. Monteiro thanks their grants (SFRH/BD/150676/2020) and(SFRH/BD/117818/2016), respectively, and C. I. M. Santos her research contract (REF.IST-ID/95/2018). Imageacquisition was performed in the LiM facility of iBiMED, a node of PPBI (Portuguese Platform of BioImaging):
POCI-01-0145-FEDER-022122. Financial support from the Spanish Ministry of Science and Innovation (MICINN)(projects CTQ2017-83531-R and CTQ2017-84327-P) is also acknowledged.

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P-2.1.14

SERS behaviour of corroles at the surface of colloidal metal particles
Joana F. B. Barata1, Sara Fateixa2, Helena I. S. Nogueira2, M. Graça P. M. S. Neves3, José A. S. Cavaleiro3 andTito Trindade2
1) CESAM and Department of Chemistry, University of Aveiro, 3810-193 Aveiro, Portugal; 2) CICECO and
Department of Chemistry, University of Aveiro, 3810-193 Aveiro, Portugal; 3) LAQV-REQUIMTE and Departmentof Chemistry, University of Aveiro, 3810-193 Aveiro, Portugal.
email: jbarata@ua.pt
The combination of porphyrinoids and metallic NPs is receiving significant attention from the scientific community
due to the relevance of the resulting nanomaterials for various applications, including catalysis, sensing, drugdelivery systems and photosensitization processes.[1] Following our interest in this field, new hybrids composed ofinorganic nanoparticles (NPs) and the contracted porphyrinoids corroles were prepared, and the vibrational andelectronic properties were analysed regarding their selection towards the reported applications.[2] Various factors
can affect the molecular adsorption of the macrocycles on metal surfaces, namely their structures and theinteraction mode between both components. Regarding this information, Surface-enhanced Raman Scattering(SERS) spectroscopy is a valuable tool since resonance SERS effect is expected when this type of macrocycles isattached or closely near to the surface of specific inorganic nanoparticles as gold or silver NPs.[3]In thiscommunication, we report the SERS behaviour of two corroles 5,10,15-trispentafluorophenylcorrole and 5-(4mercaptopropanethioloxy-2,3,5,6-tetrafluorophenyl)-10,15-bis(pentafluorophenyl)corrole on gold and silver NPs.
The intrinsic knowledge gathered from these studies will provide additional information, complementing ourcurrent studies on the development of corrole functionalized nanomaterials for photodynamic therapy.
References[1]E. J. Hong, D. G. Choi, M. S. Shim Acta Pharmaceutica Sinica B 2016, 6, 297[2] J. F. B. Barata, A. L. Daniel-da-Silva, M. G. P. M. S. Neves, J. A. S. Cavaleiro, T. Trindade, RSC Advances
2013, 3, 274. R. A. Pereira, T. Trindade, J. F. B. Barata, Magnetochemistry, 2018, 4, 37.
[3] I.Marcaida, M. Maguregui, H. Morillas, C. García-Florentino, V. Pintus, T. Aguayo, M. Campos-Vallette, J. M.
Madariaga Anal Bioanal Chem 2017, 409, 2221.
Acknowledgements. Thanks are due to University of Aveiro and FCT/MCTES for the financial support to
CESAM (UID/AMB/50017/2019), CICECO-Aveiro Institute of Materials (UIDB/50011/2020 &UIDP/50011/2020), LAQV-REQUIMTE (UIDB/50006/2020) through national funds and, where applicable, cofinanced by the FEDER, within the PT2020 Partnership Agreement, and to the Portuguese NMR Network, which
partially support the NMR spectrometers by Infrastructure Project Nº 022161 (co-financed by FEDER through
COMPETE 2020, POCI and PORL and FCT through PIDDAC). This work was financially supported by the
project Corlutna (POCI-01-0145-FEDER-031523) funded by FEDER, through COMPETE2020 - Programa
Operacional Competitividade e Internacionalização (POCI), and by national funds (OE), through FCT/MCTES. S.
Fateixa thanks FCT for his research contract (REF-069-88-ARH-2018) which is funded by national funds (OE).

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P-2.1.15

Corrole dimers – synthetic, spectroscopic and antimicrobial activity studies
Paula S. S. Lacerda1,2, Maria Bartolomeu3, Adelaide Almeida3, M. Amparo F. Faustino2, M. Graça P. M. S. Neves2,
Joana F. B. Barata1
1) CESAM, University of Aveiro, 3010-193 Aveiro, Portugal; 2) LAQV-Requimte and Department of Chemistry,
University of Aveiro, 3010-193 Aveiro, Portugal; 3) CESAM and Department of Biology, University of Aveiro,3010-193 Aveiro, Portugal
Corroles 1 (Fig. 1) are aromatic contracted tetrapyrrolic macrocycles with unique physicochemical features1. These
compounds possess key photophysical/photochemical properties as high excitation molar coefficients, fluorescencequantum yields, phosphorescence and photostability to be used in several therapeutic applications namely as
photosensitizers in PhotoDynamic Therapy (PDT)2. One of the main advantages of these macrocycles is their large
delocalized π-electronic conjugation and their stability under the PDT conditions. Additionally, conjugatedmacrocycles corrole-based 2 (Fig. 1) demonstrated that their absorption spectra fall within the therapeutic windowof 600–800 nm which make them promising candidates to be used as photosensitizers3.

Fig. 1 – The general structure of 5,10,15-triarylcorroles (1) and doubly linked corrole dimers 2a and 2b

In this context, it will be discussed a simple and peculiar alternative to the synthesis of corrole 2b (Fig.1), as well as
their structural and optical characterization. The antimicrobial activity studies, namely dark toxicity and photoactivity
under irradiation using the Gram-positive bacterium Staphylococcus aureus will also be presented.
References
1) J. F. B. Barata, M. G. P. M. S. Neves, et. al., Strategies for Corrole Functionalization. Chem. Rev., 117, 3192,
2017.
2) M. Wainwright, T. Maisch, et. al., Photoantimicrobials—are we afraid of the light? Lancet Infect. Dis., 17, e49,2017.
3) J. F. B. Barata, A. M. G. Silva, et al., β,β’–Corrole Dimers. Tetrahedron Lett., 47, 8171, 2006.
Acknowledgements. Thanks are due to University of Aveiro and FCT/MCTES for the financial support to CESAM(UIDP/50017/2020 + UIDB/50017/2020), and LAQV-REQUIMTE (UIDB/50006/2020) through national funds and,
where applicable, co-financed by the FEDER, within the PT2020 Partnership Agreement, and to the Portuguese
NMR Network, which partially support the NMR spectrometers by Infrastructure Project Nº 022161 (co-financed by
FEDER through COMPETE 2020, POCI and PORL and FCT through PIDDAC). This work was financially
supported by the project Corlutna (POCI-01-0145-FEDER-031523) funded by FEDER, through COMPETE2020 Programa Operacional Competitividade e Internacionalização (POCI), and by national funds (OE), throughFCT/MCTES.

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P-2.1.16

Toluidine blue derivatives and Human Serum albumin: Covalent conjugation andphotophysical studies.
Nory Marino-Ocampo1, Belinda Heyne2, and Denis Fuentealba1.
1) Pontificia Universidad Católica de Chile, Departamento de Química Física, Santiago, Chile; 2)Universityof Calgary, Department of Chemistry, Calgary, Alberta, Canada
Photodynamic therapy (PDT) is a therapeutic treatment for cancer. PDT involves the use of a photosensitizer (PS),light and oxygen. The PS is incorporated into the tumoral tissue, and it is excited by a light source leading toreactions with biological substrates and molecular oxygen, via Type I and II mechanism. These processes generate
reactive oxygen species (ROS). Singlet oxygen (1O2) is a ROS that can oxidize proteins, lipids, and DNA triggering
tumoral death. 1,2 Currently, several reports show strategies to improve the selectivity of the PS towards tumoral
tissues using different vehicles such as lipids, nanoparticles, macrocycles, proteins, etc. A useful protein to develop
PS carriers is human serum albumin (HSA). HSA has been shown to enhance the selectivity of different
chemotherapeutic drugs and PS. 3-5 The present work describes the derivatization of the PS Toluidine blue (TBO)with maleimide and pyridinium disulphide groups that could react with the only free thiol group of the HSA(Cysteine 34 -CYS34), potentially enhancing the selectivity of the PS towards tumoral cells. We have studied thephotophysical properties of the derivatives in reference to TBO, and the properties of the conjugated complex of
TBO derivatives with HSA using techniques like absorption and emission spectroscopy, time-resolved fluorescence
spectroscopy and time-resolved singlet oxygen phosphorescence emission.

Fig. 1 – Derivatives of Tolidine blue (TBO) that are conjugated with residue of CYS34 of HSA (PDB 1AO6).

References
1) R.R. Allison, K.Moghissi, Photodynamic Therapy (PDT): PDT Mechanisms. Clin Endosc, 2013,46,24-29.
2) M.S. Baptista, et al., Type I and Type II Photosensitized Oxidation Reactions: Guidelines and Mechanistic
Pathways. Photochem. Photobiol., 2017. 93(4), 912-919.
3) S. Absar, et al., Serum Albumin-protamine Conjugated for Biocompatible Platform for Targeted Deliveryof Therapeutic Macromolecules. J. Biomed. Mater. Res. Part A, 2014, 102, 2481-2490.
4) J. Caceres, et al., Photochemical behavior of biosupramolecular assemblies of photosensitizers,
cucurbit[n]urils and albumins. Phys. Chem. Chem. Phys., 2017, 19(3), 2574-2582.
5) S. Tayyab, S. R. Feroz., Advances in Protein Chemistry and Structural Biology, 2021, 123, 193-218.
Acknowledgements. This work was supported by grant ANID No.21210160, FONDECYT Nº1160443 and Global
Affairs Canada.
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IL-2.2.1

Responses of Cyanobacteria to UV Radiation
Rajeshwar P. Sinha
Laboratory of Photobiology and Molecular Microbiology, Centre of Advanced Study in Botany, Banaras Hindu
University, Varanasi, India
Cyanobacteria are Gram-negative prokaryotes and oxygenic phototrophs that originated around 3.5 billion years ago.
They are ubiquitous and diversified from unicellular to multicellular, coccoid to branched filaments, autotrophic toheterotrophic and established both as free-living as well as endosymbionts. A decrease in stratospheric ozone level
due to the release of chlorofluorocarbons and mononitrogen oxides results in an increased level of UV-B (280-315nm) radiation reaching the Earth’s surface. Nitrogen-fixing cyanobacteria are native of tropical agroclimaticconditions and most of them contribute to the carbon and nitrogen economy of soil. These cyanobacteria are mostly
susceptible to UV-B radiation. Pigmented proteins and DNA which have absorption maxima in the UV region are
the main targets of deleterious UV-B radiation in cyanobacteria. Results indicate that growth, survival, cellularmorphology, photosynthesis, N2 fixation, CO2 uptake and RuBISCO activity have been adversely affected by UV-B
radiation. However, cyanobacteria are not defenseless and have developed both enzymatic as well as non-enzymatic
defense mechanisms to counteract the damaging effects of UV-B radiation. Antioxidative enzymes such as
superoxide-dismutase (SOD), catalase (CAT), peroxidases (PODs) and ascorbate peroxidases (APXs) scavenge
reactive oxygen species generated in cyanobacteria due to UV-B radiation. Non-enzymatic mechanisms mediated by
the presence of photoprotectants such as mycosporine-like amino acids (MAAs) and scytonemin which absorb
mainly in the UV-B and UV-A region of the spectrum and help ecologically and economically importantcyanobacteria to grow and survive in habitats exposed to intense solar radiation. MAAs are colorless, water soluble
and low molecular weight compounds (<400 Da) with a maximum absorbance between 309 to 365 nm. Chemically,
these are composed of either aminocyclohexenone or an aminocyclohexinimine ring conjugated with the nitrogen
substituent of an amino acid or its amino alcohol. A number of MAAs such as shinorine, mycosporine-glycine,
palythine, palythinol, asterina-330 and porphyra-334 have been shown to protect cyanobacteria from UV-B radiation.
MAAs are among the strongest UV-A/UV-B-absorbing substances with high molar attenuation coefficients,
photostability and resistance against strong UVR, temperature, extreme pH and various solvents. Scytoneminconstitutes the second important class of UV-absorbing compounds in cyanobacteria that provides protection against
UV radiation due to its capacity to absorb significantly at 386, 300, 278 and 252 nm. It is a lipid-soluble yellowbrown dimeric pigment present in the extracellular polysaccharide sheath of some cyanobacteria which exists bothin oxidized (Mw 544 Da) and reduced (Mw 546 Da) forms. Any cyanobacteria endowed with the capacity to produce
photoprotective compounds will certainly be a good candidate to be used as antioxidant and natural sunscreen forhuman use against UV radiation and better biofertilizers to be used in rice paddy fields.

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IL-2.2.2
Algae and UV radiation: Examples from Antarctic ecosystems
Pirjo Huovinen1,2, Iván Gómez1,21) Universidad Austral de Chile, Instituto de Ciencias Marinas y Limnológicas, Valdivia, Chile;
2) Centro FONDAP de Investigación en Dinámica de Ecosistemas Marinos de Altas Latitudes (IDEAL), Valdivia,
Chile
Antarctic near-shore ecosystems host abundant macroalgal communities that thrive under permanently low
temperatures and extreme light seasonality (Fig. 1). Since the first surveys, the key role of these organisms in the
Antarctic costal systems was recognized, serving as a baseline for the revival of research efforts during the last
decade in the context of current environmental threats, such as the stratospheric ozone depletion and climatechange. In this presentation, an overview of our studies on the effects of UV radiation on macroalgae in the
Maritime Antarctic (King George Island) during the last decade is given, along with discussion of future scenarios.1
Antarctic macroalgae are strongly shade-adapted organisms, aphysiological feature that allow them colonizing significant depths(down to 40 m). Remarkably, a strong tolerance to high light (including
UV) stress has also been shown, which is a result of efficientphotosynthetic and bio-optical adaptations as well as constitutively highcontents of UV absorbing compounds. This paradox is well exemplified
by the large endemic brown algae, which are rich in phenolic
compounds (phlorotannins) that are related with several multifunctionalanti-stress mechanisms and are allocated in sensitive thallus parts, e.g.
reproductive tissues and growing zones. Overall, the impact of UV
radiation, e.g. DNA damage and inhibition of photosynthesis, is evident
in all life cycle stages, however, remarkable acclimation and tolerance
mechanisms are shown. For example, although early reproductive
stages, crucial for sustaining the populations, show higher susceptibility
to UV radiation (and temperature), they display efficient recovery
capacity resembling that reported in parental plants. In terms of UV
ecology, the responses of Antarctic macroalgae are defined by morphofunctional adaptations, origin and biogeographical affinity, which can beclearly recognized in their vertical zonation pattern. Shallower waters
Fig. 1 – View of coastal area of King
George Island, Maritime Antarctica.
are occupied by annual species that grow fast, intertidal zone being
dominated by ephemeral, widely distributed green algae with delicate
morphology. The large endemic brown algae (with leathery morphology) that dominate below 10 m provide
understory for also perennial coarsely branched rhodophytes. This configuration of functional groups provides theecosystem with high resilience to physical perturbations and key bio-engineering functions under changing
environment. Although recent reports point to a recovering tendency of the stratospheric ozone layer over
Antarctica, these ecosystems are currently threatened by other anthropogenic-driven perturbations, such aspollution, acidification and local decreases in salinity. Thus, the effects of UV radiation on macroalgae should be
examined considering the multiple environmental alterations that are being experienced by vast regions around theAntarctic. For example, warming temperatures (reported for West Antarctic Peninsula) pose a threat throughenhanced melting of snow and ice, which may modify the substrate availability and particularly the opticalproperties and transparency of water column, with far-reaching consequences for UV exposure of organisms.
References
1) Gómez I, Huovinen P (Eds), Antarctic Seaweeds: Diversity, Adaptation and Ecosystem Services. Springer.
2020.
Acknowledgements. This work was supported by CONICYT (Project Anillo ART110; FONDECYT 1161129 and1201053; FONDAP 15150003) and by Instituto Antártico Chileno (INACH; Grant T-20-09).
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IL-2.2.3

UV-B resistance strategies of green macroalgae
Frauke Pescheck1, Wolfgang Bilger1
1) Christian-Albrechts-University, Botanical Institute, Kiel, Germany
Green macroalgae belong to the morphologically and cytologically diversified class of Ulvophyceae. These algae
inhabit the intertidal of marine and brackish coasts as well as freshwater habitats worldwide. Although some species
grow submerged down to several meters water depth, many species thrive at the uppermost fringe of the water toland and are regularly exposed to unattenuated solar ultraviolet-B (UV-B) irradiation.
Notably, it appears that the majority of species of green macroalgae displays no apparent in vivo UV-B screening,with the exception of a few orders, e.g. the Cladophorales1. Nevertheless, many species of the Ulvophyceae arereported to be highly UV-B resistant2. For example, even over a prolonged time of exposure to unfiltered sunlight,
no strong accumulation of UV-B specific DNA damage was observed in floating tubular thalli of Ulva intestinalis,which is a representative of the group of non-UV screening green macroalgae3. In three species from this group,
different cellular UV-B tolerance mechanisms like photosystem II recovery and DNA repair were found to be fasteras compared to Cladophora sp., a green macroalga with efficient UV-B screening. Thus, cellular tolerance seems to
be capable to compensate for the absent or low UV-B screening in some green algae4. However, this was only
detected in field acclimated thalli and not in laboratory grown algae, revealing that UV-B tolerance could be downregulated under low light conditions.
Similarly, the UV-B screening capacity in some Cladophorales was observed to increase with growth irradianceunder controlled laboratory conditions. In contrast, in a freshwater Cladophora cultured outdoors apparent UV-B
screening diminished during summer indicating that other environmental factors than irradiance probably can
influence the acclimation of UV-B screening. Interestingly, growth rate was relatively high during times of low
screening. Hypothetically, this might be interpreted as a shift from an UV-B resistant to a more opportunistic lifestylein Cladophora. Since the identity of the substances that provide UV-B screening in the Cladophorales is still
unknown, knowledge about the chemical and physical characteristics is very limited. In addition to spectral properties
derived from chlorophyll fluorescence excitation spectra, physiological measures of light-dependent reactions were
employed to analyse the spectral effects of UV screening in vivo. The influence of UV-A and UV-B screening on
DNA damage induction and its reversion via photoreactivation was investigated. From the results, it could be deduced
that screening of UV-A radiation is affecting the rate of photoreactivation in Cladophora5. A discussion of UV-B
resistance in a pair of a non-UV screening and a UV screening species under natural sunlight will close the talk.
References
1) Pescheck, F., Bischof, K., Bilger, W., Screening of ultraviolet-A and ultraviolet-B radiation in marine green
macroalgae (Chlorophyta). Journal of Phycology, 46, 444-455. 2010.
2) Bischof, K., Gómez, I., Molis, M., Hanelt, D., Karsten, U., Lüder, U., Roleda, M., Zacher, K., Wiencke, C.,
Ultraviolet radiation shapes seaweed communities. Reviews in Environmental Science and Bio/Technology, 5,141-166, 2006.
3) Pescheck, F., Campen, H., Nichelmann, L., Bilger W., Relative sensitivity of DNA and photosystem II in Ulva
intestinalis under natural solar irradiation. Marine Ecology Progress Series, 555, 95-107, 2016.
4) Pescheck, F., Bilger, W., Compensation of lack of UV screening by cellular tolerance in green macroalgae(Ulvophyceae) from the upper eulittoral. Marine Biology, 165, 132, 2018.
5) Pescheck, F. UV-A screening in Cladophora sp. lowers internal UV-A availability and photoreactivation ascompared to non-UV screening in Ulva intestinalis. Photochemical & Photobiological Sciences, 18, 413-423, 2019.
Acknowledgements. This work was supported by a scholarship from the Christian-Albrechts-University of Kiel.

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OC-2.2.4
Chemical decoration and biochemical incorporation of photoactive molecules indiatoms for photonics and electronic applications
Danilo Vona1, Cesar Vicente Garcìa1, Stefania Roberta Cicco2, Roberta Ragni1, Gabriella Leone1, Gianluca Maria
Farinola1
1 Department of Chemistry, University of Bari, Via Orabona 4, 70125, Bari, Italy; 2 CNR-ICCOM Institute oforganometallic chemistry Via Orabona 4, 70125, Bari, Italy
Biomineralization is a biochemical process which converts inorganic salts into complex hybrid materials. Diatom
microalgae couple the photosynthetic energy with the production of mesoporous silica shells, called frustules, after
the biomineralization of uptaken silicate. These shells constitute a source of silica which exhibit high surface area,transparency, and mechanical resistance, and also an easy aptitude to be chemically functionalized, paving the way
to their use in photonics, sensing, optoelectronics and biomedicine [1]. In this context we present a plethora ofchemical or in vivo decorations for directly giving specific properties to silica from diatoms microalgae [2]. We
recently demonstrated the in vivo uptake of organometallic emitters containing Iridium, Rhutenium and Aluminum
metal cores, into Phaeodactylum tricornutum diatom specie,[3] adherent cells which self-populate and propagate ontransparent conductive ITO glasses while bearing luminescent complexes. These experiments successfully aim to
produce simple devices in principle suitable for producing photocurrent under light. Furthermore we considered thein vivo incorporation of a positively charged Iridium Complex into Coscinodiscus spp. and Thalassiosira weissflogii
diatoms frustules, till producing new silica-based luminescent micromaterials and luminescent nanoparticles forbiomedicine and drug delivery.[4] Also, organic dyes can be chosen for improving biomass and oxygen response ofliving microalgae directly exploiting some spectroscopic interactions with the photosynthetic apparata.[5] These lastresults exhibit an intrinsic aspect of technological applicability in the field of biomass production from algae, forcosmetics, pharmacology, and carbon dioxide or pollutants bioremediation.

Acknowledgements
This work was financially supported by the European Commission through EU project 800926-HyPhOE(Hybrid Electronics based on Photosynthetic Organisms).
References
1) Ragni, R., Cicco, S.R., Vona, D. and Farinola, G.M. Multiple Routes to Smart Nanostructured Materials from
Diatom Microalgae: A Chemical Perspective. Adv. Mater. 1-23, 1704289. 20172) Ragni, R., Scotognella, F., Vona, D., Moretti, L., Altamura, E., Ceccone, G., Mehn, D., Cicco, S.R., Palumbo, F.,
Lanzani, G. and G. M. Farinola. Hybrid Photonic Nanostructures by In Vivo Incorporation of an Organic Fluorophore
into Diatom Algae. Adv. Funct. Mater. 28 (24), 1706214-1706223. 2018
3) Vona, D., Ragni, R., Altamura, E., Albanese, P., Giangregorio, M.M., Cicco, S.R. and Farinola, G.M. LightEmitting Biosilica by In Vivo Functionalization of Phaeodactylum tricornutum Diatom Microalgae with
Organometallic Complexes. Appl. Sci. 11(8), doi.org/10.3390/app11083327. 2021
4) Della Rosa, G., Vona, D., Aloisi, A., Ragni, R., Di Corato, R., Lo Presti, M., Cicco, S.R., Altamura, E., Taurino,
A., Catalano, M., Farinola, G.M. and Rinaldi, R. Luminescent Silica-Based Nanostructures from in Vivo IridiumDoped Diatoms Microalgae. ACS Sust. Chem. & Eng., 7(2), 2207-2215. 20185) Leone, G., De la Cruz Valbuena, G., Cicco, S.R., Vona, D., Altamura, E., Ragni, R., Molotokaite, E., Cecchin,M., Cazzaniga, S., Ballottari, M., D’Andrea, C., Lanzani, G. and Farinola, G.M. Incorporating a molecular antenna
in diatom microalgae cells enhances photosynthesis. Sci. Rep. 11, doi.org/10.1038/s41598-021-84690-z. 2021

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IL-2.2.5

Do secondary compounds protect lichens against UV radiation?
Knut Asbjørn Solhaug
Norwegian University of Life Sciences, Faculty of Environmental Sciences and Natural Resource Management, Ås,
NorwayLichens are symbiotic associations between a mycobiont and a green algae and/or cyanobacterium photobiont The

lichen cortex often contain high amounts of secondary compounds synthesized by the fungal partner.
These compounds are situated as crystals on the surface of the fungal hypha. Most of these secondary
compounds absorb UV-B and a few visible light. They may screen visible and/or UV by absorption or byreflection when they are in crystal form. The ecological functions of the secondary compounds have beenstudied in a series of experiments using the acetone-rinsing method. Lichen compounds may be removedby acetone-rinsing without any harmful effect on the lichens. The susceptibility of the lichens against
various stresses can then be tested with or without secondary compounds. One main conclusion fromthese experiments is that the secondary compounds are not essential for UV screening. However, UV may
act as a signal for synthesis of the compounds, and their main function seem to be e.g. visible lightscreening and to deter herbivores.

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IL-2.2.6
Ultraviolet photoprotection in bryophytes - a polar perspective
Sharon A. Robinson1,2, Melinda J. Waterman1 , Johanna D. Turnbull1
1) University of Wollongong, School of Earth Atmosphere and Life Sciences & Global Challenges Program,
Wollongong, Australia 2) University of Wollongong, Securing Antarctica’s Environmental Future,
Wollongong, Australia
We discuss the effects of ultraviolet (UV) -B radiation on polar bryophytes. Multiple studies have measured polar
bryophyte responses to decreases in UV-B radiation under screens, fluctuations in UV-B irradiance in naturalenvironments as well as increases in UV-B radiation supplied from fluorescent UV lamps3. Exposure to UV-B
radiation increases the concentrations of UV-B absorbing compounds2-5 and increases DNA damage6,7. Bryophyteslack epidermal tissues and may sequester UV screening compounds preferentially in their cell walls as a method toenhance UV photoprotection1,4,8,9. We consider if these bryophytes exhibit unique photoprotective mechanisms toenable them to survive in these incredibly inhospitable environments.
References
1) Clarke, L. J., Robinson, S. A. Cell wall-bound ultraviolet-screening compounds explain the high ultraviolet
tolerance of the Antarctic moss, Ceratodon purpureus. New Phytologist 179: 776-783. 20082) Dunn J. L., Robinson S. A. Ultraviolet-B screening potential is higher in two cosmopolitan moss species
than in a co-occurring Antarctic endemic moss – implications of continuing ozone depletion. Global Change
Biology 12: 2282-2296. 2006
3) Newsham, K. K., Robinson, S. A. Responses of plants in polar regions to UVB exposure: a meta-analysis.
Global Change Biology 15: 2574-2589. 2009
4) Robinson, S. A., Waterman, M. J. Sunsafe bryophytes: photoprotection from excess and damaging solar
radiation. In D. Hanson & S. Rice (eds.), Photosynthesis in bryophytes and early land plants, pp. 113-130.
Springer, Dordrecht, The Netherlands. 2014
5) Snell K. R. S., Kokubun T., Griffiths H., Convey P., Hodgson D. A., Newsham K. K. Quantifying themetabolic cost to an Antarctic liverwort of responding to UV-B radiation exposure. Global Change Biology
15: 2563-2573 20096) Turnbull, J. D., Robinson, S. A. Accumulation of DNA damage in Antarctic mosses: correlations withultraviolet-B radiation, temperature and turf water content vary among species. Global Change Biology 15:319-329. 20097) Turnbull, J. D., Leslie, S. J., Robinson, S. A. Desiccation protects two Antarctic mosses from ultraviolet-B
induced DNA damage. Functional Plant Biology 36: 214-221. 20098) Waterman, M. J., Bramley-Alves, J., Miller, R. E., Keller, P. A., Robinson, S. A. Photoprotection enhanced
by red cell wall pigments in three East Antarctic mosses. Biological Research 51: 49, 20189) Waterman, M. J., Nugraha, A. S., Hendra, R., Ball, G. E., Robinson, S. A., Keller, P. A. Antarctic Moss
Biflavonoids Show High Antioxidant and Ultraviolet-Screening Activity. Journal of Natural Products 80:
2224-2231. 2017
Acknowledgements. This work was supported by grant DP200100223 from the Australian Research Council.

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IL-2.2.7
Responses of non‐flowering plants to UV radiation: an overview
Javier Martínez-Abaigar1, Encarnación Núñez-Olivera1
1) University of La Rioja, Faculty of Science and Technology, Logroño (La Rioja), Spain
Ultraviolet (UV) radiation is a minor component of the whole solar spectrum reaching the Earth’s surface, but it
constitutes an important environmental factor regulating life processes. Considering the term “plants” in its widest
sense, we can construct an integrative structural, physiological, ecological, and evolutionary portrait of the
responses of non-flowering plants (NFP, cryptogams) to UV radiation. In this wide sense, NFP would include
cyanobacteria, eukaryotic algae, lichens, bryophytes (mosses, liverworts and hornworts), and pteridophytes.
Although the research effort devoted to each group of organisms has been notably asymmetric, much knowledge is
now available on the UV effects on NFP, including both common and specific responses. Nevertheless, theexperimental conditions applied in the different studies have been so diverse that it is difficult to generalize theresults obtained and to compare the UV tolerance of the different organisms. With the aim to synthesize ourcomparative knowledge on the UV tolerance of NFP, and focusing on UV-B radiation, we applied the same
treatment to 107 species (26 algae, 6 lichens, 64 bryophytes, and 11 pteridophytes), in the (apparently) mostextensive study to date. UV-B tolerance was assessed using chlorophyll fluorescence variables. Our results showthat UV-B tolerance of NFP depends on the species considered, and is greatly influenced by the taxonomic groupand (except lichens) by the concomitant level of structural complexity. Overall, the scale of UV-B tolerance was:(pteridophytes and lichens) > mosses > (liverworts and hornworts) > algae. UV-B tolerance was significantly
correlated with the content of potentially protective UV-absorbing compounds, and with the sclerophylly of thevegetative bodies. These results may have ecological and evolutionary implications, and can help understand therole of UV radiation in the water-to-land transition of photosynthetic organisms and their subsequent conquest ofland.
Acknowledgements. This work was supported by the Spanish Ministry of Science, Innovation and Universities
and European Regional Development Fund (ERDF), through the project PGC2018-093824-B-C42.

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P-2.2.8

UV-induced changes in phenolic and antioxidant profiles of Nicotiana tabacum leaves
outdoors and in a growth chamber
Kristóf Csepregi, Arnold Rácz, Gyula Czégény, Éva Hideg
Department of Plant Biology, Faculty of Sciences, University of Pécs, Hungary
Phenolic compounds (flavonoids, phenolic acids) are synthesized via the phenylpropanoid pathway. These
compounds are localized in various tissues and documentedly play a protective role against environmental stresses,
such as high light intensity or UV radiation as antioxidants and screening pigments. The complex effect of sunlightoutdoors and the effect of UV single factor in controlled experiments indoors are distinct. Understanding the latter isimportant for exploiting UV as modifier of special metabolites. In this study we examined phenolic and antioxidant
profiles of tobacco (Nicotiana tabacum) leaves grown either in growth chamber or under natural sunlight using UVfilters to separate different UV ranges. Outdoor experiments were carried out in June-July 2020 in Pécs (Hungary,
46.07° N, 18.23° E), and plants were grown from seeds to 5-weeks old under the three different light conditions. In
the indoor experiments tobacco plants were grown for four weeks under visible light only, then exposed tosupplementary UV radiation for 5 days (Q-panel UVB-313EL, 6.9 kJ m-2 d-1 biologically effective dose). There were
three treatment groups both indoors and outdoors: visible light only, visible plus UV-A, visible plus UV-A and UVB. Photosynthesis measurements demonstrated that neither conditions were stressful but resulted in acclimative
responses.
The aims were to (i) identify UV-responsive main phenolic compounds with HPLC, and (ii) find a relationshipbetween antioxidant capacities and phenolic levels. We identified the same phenolic compounds in both experiments,
although in different amounts and relative ratios. The main phenolic acids were chlorogenic acid and it’s two isomers
neochlorogenic acid and cryptochlorogenic acid, major flavonoids included quercetin-3-O-rutinoside andkaempferol-3-O-rutinoside. Total antioxidant capacities were measured as TEAC, FRAP and FC in order to provide
a multi-angle analysis1, and identified higher capacities in leaves grown outdoors. Interestingly, although full sunlightand UV-supplemented growth light were very different, some components (neochlorogenic and cryptochlorogenic
acids, and quercetin-3-O-rutinoside) were present at the same amounts in leaves under these two conditions. Other
compounds (chlorogenic acid and other flavonol glucosides) were present at 1.5-4-times higher amounts in sunlightthan indoors. The presence of UV had a larger positive effect on phenolic components and antioxidants indoors thanoutdoors, due to differences in the ratio of photosyntetically active radiation to UV in these two experiments.
Correlations between UV absorbing and antioxidant capacities of leaf extracts and amounts with phenolic contents
reflected different properties2 of individual compounds.
References
1) Csepregi K, Neugart S, Schreiner M, Hideg É., Comparative evaluation of total antioxidant capacities of plant
polyphenols. Molecules. 21, 208, 2016
2) Csepregi K, Hideg É., Phenolic compound diversity explored in the context of photo-oxidative stress protection.
Phytochemical analysis. 29, 2, 129-136, 8, 2018
Acknowledgements. This work was supported by the National Research, Development and Innovation Office (grants
NN-128806 to KCs and PD-131625 to GyCz).

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IL-2.3.1

Germicidal Ultraviolet UV-C:
Rediscovery of an Old Method to Reduce Infection Risk During COVID-19
David H. Sliney, Ph.D.1
1) Johns Hopkins University Bloomberg School of Public Health, Department of Environmental Health andEngineering, Baltimore, USA
Historically, germicidal ultraviolet germicidal irradiation (UVGI) dates back more than a century andwas widely used in hospitals and public places to reduce infections by inactivating airborne parthenogens in
the 1930s – 1950s.1 Studies from those times demonstrated efficacy,2 but UVGI use later dropped out of favor
in the 1960s after vaccines against a number of childhood diseases, such as polio and measles had been
virtually eliminated. Of infectious diseases of significant severity in the developed world only tuberculosis
has been without a vaccine and for that reason UVGI has remained in TB clinics – particularly in thosecountries where TB continues to be a major problem.3 In these countries, some expertise has been retained.
Today with the COVID-19 pandemic, the lessons learned from TB control allow for a ‘rediscovery’ of thistechnology for use in the current pandemic.3-4 Furthermore, the pandemic has greatly accelerated development
of UV-C LEDs and other lamp types such as the krypton-chloride (222-nm) lamp to augment the traditionaluse of low-pressure mercury (254 nm) lamp.5 Sadly, misconceptions about UVGI, such as a perceived skincancer risk remain and a lack of understanding of proper safety precautions continue to slow the wide
acceptance of UVGI.6-8
References
1) Reed, N. G. The history of ultraviolet germicidal irradiation for air disinfection. Pub Health Rep. 125, 15–
24 (2010).
2) Wells, W. F., Wilder, T. S. The environmental control of epidemic contagion: I. An epidemiologic studyof radiant disinfection of air in day schools. Am. J. Hyg. 35, 97–121 (1942).
3) Volchenkov, G. Experience with UV-C air disinfection in some Russian hospitals. Photochem.
Photobiol., 97: 549–551 (2021).
4) Nardell, E. A. Air disinfection for airborne infection control with a focus on COVID-19: Why germicidal
UV is essential. Photochem. Photobiol., 97: 493–497 (2021).
5) Bergman, R. S. Germicidal UV sources and systems. Photochem. Photobiol., 97: 466–470 (2021).
6) Bergman, R., Brenner, D., et al. Air disinfection with germicidal ultraviolet: For this pandemic and thenext. Photochem. Photobiol., 97: 464–465 (2021).
7) Sliney, D.H., Stuck, B.E. A need to revise human exposure limits for ultraviolet UV-C radiation.
Photochem. Photobiol., 97: 485–492 (2021).
8) Claus, H. Claus, H. (2021). Ozone generation by ultraviolet lamps. Photochem. Photobiol. 97: 471–476(2021).

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IL-2.3.2

Photoinduced inactivation of SARS-CoV-2
Mauricio S. Baptista
Universidade de São Paulo, Chemistry Institute, Department of Biochemistry.
baptista@iq.usp.br
The COVID-19 pandemic has accelerated the need for the implementation of effective light-based technologies totarget the causative agent SARS-CoV-2. Light of different wavelength can act in the virus and in the hostindividuals by different mechanisms and used as environmental disinfectant with distinct efficiency, as well as, canbe used to easing the consequences of the infection in individuals. While UV radiation target mainly the nucleic
acids of the virus, visible light in the presence of proper a photosensitizer can target other compartments such as thevirus envelop, by the so-called contact-dependent photosensitized oxidation reactions. I will comment on thebenefits, challenges, and pitfalls of repurposing light-based antimicrobial strategies in the COVID-19 pandemic.
References
CP Sabino, et al. Light-based technologies for management of COVID-19 pandemic crisis. Journal of
Photochemistry and Photobiology B: Biology 2020, 111999.
LVM de Assis et al. How does the skin sense sun light? An integrative view of light sensing molecules. Journal of
Photochemistry and Photobiology C: Photochemistry Reviews 2021, 100403.
IOL Bacellar et al. Photosensitized membrane permeabilization requires contact-dependent reactions between
photosensitizer and lipids. Journal of the American Chemical Society 2021, 140 (30), 9606.

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IL-2.3.3

Is there a therapeutic role for PDT in Covid 19 SARS infection?
K. Moghissi
The Yorkshire Laser Centre, Goole, UK
Background: In December 2019 a new virus; Corona Virus-2 causing Severe Acute Respiratory Syndrome (SARS)
appeared in Wuhan (China). The virus was baptized COVID 19 and took the world by surprise because of its rapid
spread, its highly lethal characteristics and its lack of response to all available antimicrobial/antiviral drugs.
COVID 19 started as an epidemic in Wuhan and quickly went out of control to become a pandemic killing - andcontinuing to kill - millions of people globally.
Ever since its emergence, COVID 19 has presented the world with a serious public health issue, has brought thehospitals’ function to a halt and has diverted all resources to symptomatic (palliative) treatment of patients and theattempt to stop the spread of the infection.
Lockdown, social distancing, intensive care unit admission and the treating of SARS by rich oxygen ventilation
support to combat hypoxia and, at the last resort, the use of ECMO have been employed. They all are preventive,
symptomatic treatments.
Additionally, there has been international cooperation to produce suitable vaccines which have been successful toimmunize the recipients.
In short there has been:
1) Shielding and immunization as preventive measures.
2) Symptomatic treatment.
However, no report of specific treatment to kill the virus in vivo has been proposed.
Aims and Objectives:
The aims and objective of the presentation is to suggest that there is a place for Photodynamic Therapy (PDT) in thetreatment of SARS COVID 19, based on Photodynamic Reaction (PDR/Photodynamische Wirkung) to destroy thevirus and investigate how it can, potentially, be clinically applied.
The presentation comprises of:
•
•
•
•
•
•

A short reminder of PDT mechanisms emphasizing that it is basically a Local Therapy.
Briefly reviewing relevant anti-microbial/antiviral effects of PDT.
Discussing the problems in using PDT in the particular case of COVID 19 and its lethal SARSIn this discussion I will highlight the relevant structure COVID 19 and its defense mechanisms.
Explain the difficulties of finding a suitable Photosensitiser which could reach the virus and break its defense
mechanism.
How PDT can be applied clinically in order to neutralize COVID 19 which is the causative agent of SARS
which in turn afflicts and kills the patient.

Basically, therefore, PDT has to target the initial injury and the virus colony within the airway; this seems to be thepath the virus takes to invade and create the lethal pulmonary injury, respiratory failure and gas exchange crash.
Finally, the currently existing planned protocol of study and effort to use PDT as a treatment of SARS COVID 19
will be explored and exposed.

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OC-2.3.4
Correlation of the ultraviolet index (UVI) and the temperature with incidence andseverity of Covid-19 in Spain
Yolanda Gilaberte1, Natalia Martín 2, Juan B. Pérez-Gilaberte3, Alba Navarro-Bielsa1, M. Victoria de Galbe4, José
Aguilera4.
1) Miguel Servet University Hospital, Department of Dermatology, Zaragoza, Spain; 2) University of Zaragoza;3)Miguel Servet University Hospital, Department of Internal Medicine, Zaragoza, Spain; 4)University of Malaga, ,Department of Dermatolology, Málaga, Spain.
Introduction: severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) In the present study, the causal
agent of coronavirus disease 2019 (COVID-19), has become pandemic on March 11. Different waves have occurredin the different countries, being modified the incidence by the social distancing measures taken. Low levels of vitamin
D have been associated with the higher severity of the disease. Considering that the most important source ofvitamin D in ultraviolet radiation (UVR) exposure, the aim was to study the correlation between ultraviolet index(UVI) and temperature with the incidence and the severity of COVID-19 in different latitudes in Spain.
Material and Methods: an observational prospective study was performed analyzing the daily UVI and maximum
and minimum temperatures with the incidence of cases of COVID-19, hospitalized patients, admitted in intensive
care units (ICU), and deaths in 5 Spanish cities located in different latitudes: San Sebastian (N43.25), Zaragoza(N41.65), Madrid (40.41), Málaga (N36.72) and Tenerife (N28.48) from February 2020 to January 2021. Pearson
correlation test and linear regression were used. P-value less than 0.005 was considered statistically significant.
Results: An inverse correlation, statistically significant, was found between UVI and the incidence of cases per
100.000 inhabitants in all the studied cities. Those cities located by the sea had the highest correlation coefficients,
one in the north (San Sebastian r=-0.426, p=0.001) and 2 in the South (Málaga (r=-0.46, p<0.001) and Tenerife (r=0.569, p<0.001)). Mortality was significantly inversely correlated with UVI also in these 3 cities. Regarding
temperatures, the most relevant inverse correlations were found between the maximum temperature and the mortality
in the cities of Tenerife (r=-0.528, p<0.001) and Málaga (r=-0.410, p=0,001). According to the linear regression
analysis, in Zaragoza 43% of the incidence of COVID-19 can be explained by the UVI and the temperature, whereas
in Madrid only 19% can be explained by them.
Conclusion: UVI inversely influences the incidence of COVID-19 independently of the latitude, and the highest
correlations are found in cities with sea. Although the correlation of these meteorological variables with severity
could not be established, mortality was inversely correlated with UVI and maximum temperatures in cities of theSouth.

.

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IL-2.3.5

Broad-spectrum Photodynamic Disinfection in the Treatment of SARS-CoV-2
N. Loebel1, C. Romo1, C. Cross1, J. Hickok1, R. Andersen1
1) Ondine Biomedical, Inc., Vancouver, BC, Canada
Direct-acting antiviral (DAA) therapies generally target proteins mediating specific stages in the viral lifecycle
such as proteases, polymerases, nonstructural proteins and other viral constituents. These DAA approaches canprovide high viral eradication rates, in some cases exceeding 90%.1 DAAs offer the potential for treatment in cases
where vaccine access is lacking or contraindicated; in deficient host immune response secondary to viral
downregulation of major histocompatibility class I molecules2; or in the evolution of newly-infectious viral variants
known as quasispecies3 resulting from high viral replication and error rates in unvaccinated or
immunocompromised populations. In a particularly threatening recent finding, SARS-CoV-2 has been
demonstrated to engage in cell-to-cell transmission in a mode known as virus cell-surfing,4 an actin filamentmediated intercellular propagation technique efficiently utilized by HIV, poxvirus and other DNA and RNA
viruses. Cell-to-cell transmission enhances immune evasion and results in reduced clearance by neutralizing
antibodies.5
Photodynamic disinfection is a broad-spectrum,6 non-resistance forming,7 rapid,8 and safe9 approach to eliminationof pathogens from epithelial surfaces in vivo. The technique has been clinically deployed for many years as a potent
preoperative decolonization method in the anterior nares,10 minimizing or eliminating the use of mupirocin andother resistance-prone agents. In this work, we report on the use of the technique as a topical DAA against SARSCoV-2 in laboratory settings, followed by a summary of field-use reports by industrial teams in meat-packing
plants, where high potential exists for worker-to-worker and worker-to-package transmission of SARS-CoV-2.

References
1) M McConnell, JK Lim, et al, Hepatitis C Vaccine Development in the Era of Direct‐Acting Antivirals. Clin
Liver Dis (Hoboken), 2018.
2) S Ishido, C Wang, et al, Downregulation of Major Histocompatibility Complex Class I Molecules by Kaposi's
Sarcoma-Associated Herpesvirus K3 and K5 Proteins. J Virol, 2000.
3) C Trucchi, A Orsi, et al, State of the Art, Unresolved Issues, and Future Research Directions in the Fightagainst Hepatitis C Virus: Perspectives for Screening, Diagnostics of Resistances, and Immunization. J
Immunol Res, 2016.
4) LA Caldas, FA Carneiro, et al, Ultrastructural analysis of SARS-CoV-2 interactions with the host cell via highresolution scanning electron microscopy. Scientific Reports, 2020.
5) J Dufloo, T Bruel, et al, HIV-1 cell-to-cell transmission and broadly neutralizing antibodies. Retrovirology,
2018.
6) M Wainwright, Photodynamic antimicrobial chemotherapy (PACT). J Antimicrob Chemother, 1998.
7) LA Pedigo, AJ Gibbs, et al, Absence of bacterial resistance following repeat exposure to photodynamic
therapy. SPIE 7380 Photodynamic Therapy: Back to the Future, 2009.
8) F Cieplik, D Deng, et al, Antimicrobial photodynamic therapy – what we know and what we don’t. Crit. Rev.
Microbiol, 2018.
9) D Meller, J Hickok, et al, Photodisinfection Therapy: Essential Technology for Infection Control.
InfectionControl.tips, 2020.
10) E Bryce, T Wong, et al, Nasal photodisinfection and chlorhexidine wipes decrease surgical site infections: ahistorical control study and propensity analysis. J Hosp Infect, 2014.
Acknowledgements. All authors are employees of Ondine Biomedical, Inc., Vancouver, BC, Canada.

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IL-2.3.6
Photodynamic therapy for Covid-19 and other infectious diseases
Luis G. Arnaut1, Mariette M. Pereira1, Zoe A. Arnaut1, Rafael T. Aroso1, Sara M. A. Pinto1, Ana Catarina S. Lobo2,
Paulo Rodrigues-Santos3
1) CQC, Chemistry Department, University of Coimbra, 3004-535 Coimbra, Portugal; 2) LaserLeap Technologies,
R. Col. Veiga Simão, 3025-307 Coimbra, Portugal; 3) Immunology Institute, Faculty of Science, University of
Coimbra, 3004-504 Coimbra, Portugal
The Covid-19 pandemic raised the global awareness on the fragilities of medical responses to infectious diseases.
Repositioning of existing antiviral medications gave, at best, controversial results and no clear clinical benefit. Drugdevelopment targeting directly the SARS-CoV-2 virus could not offer a timely response to the pandemic. Vaccines
developed and approved with unprecedented rapidity offer a medium/long term protection against the pandemic, butalso illustrate the shocking asymmetry of medical care, where the clinical solutions are not placed where they are
most needed and bounce back with variants that have higher infection potentials. The need for a new approach tocontrol infectious diseases became increasingly evident with the Covid-19 pandemic.
Photodynamic disinfection (PDDI) is based on the generation of reactive oxygen species when an electronically
excited photosensitizer molecule encounters molecular oxygen and transfers its energy or an electron, to produce
singlet oxygen or superoxide ion, respectively. The local oxidative stress generated by PDDI can damage proteins,
lipids or nucleic acids. Enveloped viruses, such as the coronavirus, offer various accessible targets to PDDI. A mostappealing property of PDDI is that it can work equally well for all enveloped virus, independently of mutations,which is a valuable tool for the armamentarium of measures against infectious diseases, known at present or emerging
in the future.
We developed a new photosensitizer to photoinactive enveloped virus and show how it works against clinical samplesof SARS-CoV-2. Virus collected from patients were transported alive to a proper biosafety lab and photoinactivatedwith the new photosensitizer. A 150 nM concentration and 5 J/cm2 at 644 nm inactivated >99.99% of the virus in 5
minutes, i.e., the amount of virus in the clinical sample became undetectable by RT-qPCT. Under the same drug andlight doses, the viability of HaCat (human epidermal keratinocyte) cell remained 100%. It is possible to selectively
inactivate SARS-CoV-2 without toxicity to human cells. The mechanisms of inactivation are discussed with reference
to results obtained with genetically engineered lentivirus that express the spike protein at their surface. Comparison
with the combined drug and light doses of PDDI using methylene blue with SARS-CoV-2 shows that the newphotosensitizer is >300 times more potent.
Acknowledgements. This work was supported by the Portuguese Science Foundation (grants UIDB/00313/2020,
PTDC/QUI-OUT/27996/2017-DUALPI and PD/BD/143123/2019) and by P2020 (Covid-19, project no. 49708)

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IL-2.3.7

Ultraviolet C Exposure Testing of Materials; An Important Aspect of Surface
Sterilization
Matt McGreer1
1) Atlas Material Testing Technology, Ametek Measurement, Communications and Testing, Mount Prospect,
Illinois, United States of America
UVC radiation (between 200-280 nanometers) has been used for sterilization purposes for over 40 years. This
activity has increased tremendously because of the COVID-19 pandemic, expanding from hospital operating rooms
to retail and other public spaces, transportation, and electronic devices. In most cases, the polymers and coatings
used in these applications have not been engineered or tested to withstand repeated applications of UVC. Very few
studies have been conducted on a limited number of polymers.
UVC lamps with a peak output of 254 nanometers have recently been employed in modified testing devices thathave been traditionally used with standard fluorescent UV lamps. Modifications to these devices include special
features to protect users from stray light, sensors that can measure and control irradiance of the UVC lamps, andmore robust materials that can withstand the higher energy from these lamps.
This presentation will show results from UVC exposures of a variety of materials to (a) compare material propertychange from UVC exposure to that of standard UVA-340 and UVB-313 exposures, (b) analyze the impact ofdifferent irradiance levels, black panel temperatures, and light/dark cycling on these materials, and (c) evaluate
standard performance characteristics such as repeatability and uniformity of these devices. Finally, an “Exposure
Dosage Calculator” will be presented to assist users with determination of appropriate exposure levels based on
selected use cases.

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OC-2.3.8
Characterization of 222nm UVC-based Cleaning Impacts on Vegetative and Biofilm
Cells
Jeffrey Busick1, Julie Zwiesler-Vollick1, and Janet Price2
1) Lawrence Technological University, Department of Natural Sciences, Southfield, MI, USA; 2) JustLight PBC,
Ann Arbor, MI, USA
Illnesses spread through indirect means, such as common colds, rhinovirus and the flu, pose significant risk tocommunities and drain collective resources. The flu alone affects ~29 million people annually, costing $90 billion
in medical costs and resulting in 38,000 deaths1, 2. Reduction of pathogens on a surface by 99% reduces incidence
of infection after contact to less than 1 in a million chance 3. UVC light is quickly becoming a mainstream methodof cleaning air, surfaces, and food due to its germicidal properties and shallow penetration depth. JustLight isleveraging these properties of UVC light in order to develop a light-based hand cleaner, Violet. To support this, we
assessed low doses of 222nm UVC light for efficacy against common indirectly transmitted pathogens in vitro andin the context of a biofilm. Monocultured pathogens inoculated on a surface displayed concentration and dosedependent reduction after UVC exposure. From this, we were able to calculate a density of pathogens on a surface
that was sufficient to block the effect of the UVC light from penetrating to underlying cells. This result was echoedin the biofilm context, as transient pathogens on the surface of hands were reduced at a higher rate than theunderlying biofilm cells. These results support Violet and 222nm UVC as an effective method for reducing
transient pathogens without damaging the skin microbiome or underlying tissue.
References
1) Centers for Disease Control, Disease Burden of Influenza. CDC.gov, 2020.
2) Roger Dobson, Flu costs the US $90bn a year, report shows. BMJ, 2007.
3) Michael Ryan, Charles Haas, Application of quantitative microbial risk assessment for selection of microbial
reduction targets for hard surface disinfectants, Am J Infect Control, 2014.

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P-2.3.9

Characterising ‘far-UVC’ KrCl excimer lamps for safety and inactivation of viruses
Ewan Eadie1, Paul O’Mahoney2, Louise Finlayson3, Sally H. Ibbotson2, Kenneth Wood4
1) NHS Tayside, Photobiology Unit, Ninewells Hospital, Dundee, United Kingdom; 2) University of Dundee,
Photobiology Unit, Ninewells Hospital, Dundee, United Kingdom; 3) University of St Andrews, SUPA School ofPhysics and Astronomy, St Andrews, United Kingdom
Krypton-Chloride (KrCl) excimer lamps have a primary emission wavelength of 222 nm, with residual shorter andlonger wavelengths throughout the ultraviolet (UV) spectrum. It has been demonstrated in laboratory research thatthese lamps can effectively inactivate a wide range of pathogens, including SARS-CoV-21-3. The peak emission
wavelength at 222 nm appears to be strongly absorbed in the skin’s stratum corneum and in the tear layer of theeye, thus not inducing the acute reactions that are characteristic of other UVC and UVB wavelengths 4-6. However,residual longer wavelength UV radiation from unfiltered KrCl lamps does appear to penetrate beyond the stratum
corneum, inducing both erythema and cyclobutane pyrimidine dimers (CPD) in the deeper layers of epidermis,7,8. It
is therefore important to understand the emission characteristics of a source to accurately determine the hazard it
may present.
We investigated the spectral irradiance and the change-in-irradiance-with-distance, from 13 commercially available
and research prototype sources with peak emission wavelength at 222 nm. Spectral irradiance was measured with adouble-grating spectroradiometer (IDR300, Bentham Instruments Ltd, Reading, UK) with cosine corrected input
optics and change-in-irradiance-with-distance was measured with a broadband radiometer (IL1400 SEL220
QNDS2 W, International Light Technologies, Massachusetts, USA). Spectral irradiance measurements were used
to determine S(lamp), which is the total effective spectral hazard of a lamp as defined by Buonanno et al.8 S(lamp)
was determined for the current International Commission on Non-Ionizing Radiation Protection (ICNIRP) hazard
weighting function (S(lampI)) as well as the proposed hazard weighting functions from the American Conferenceof Governmental Industrial Hygienists (ACGIH) for the eye (S(lampAE)) and skin (S(lampAS)). The spectral angular
distribution from one of the sources was also measured both with and without a diffusing material incorporated into
the device. The results of these measurements were incorporated into a Monte Carlo radiative transfer model todetermine the depth penetration of each source in skin and the resulting modelled CPD. A comparison withcomputer modelled sun exposure was also performed.

Fig. 1 – Normalised spectral irradiance measurements from three of the 13 commercial sources measured. The normalised
spectral irradiance from Woods et al.7 is plotted for comparison.

Variation in the measured spectral irradiance was observed from the 13 commercial sources (Fig. 1). The total
effective spectral hazard of each lamp, S(lampI), varied from 0.126 to 0.166. The difference between lamps wasBook of Abstracts ����������������������������������������������������������������������������������������������� 106

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more marked when considering the proposed new ACGIH limits - S(lampAE) from 0.020 to 0.072 and S(lampAS)
from 0.007 to 0.031. Angular spectral measurements showed that the spectral distribution from one of the sources
varied with viewing angle, a phenomenon well known with certain types of optical filter. The introduction of adiffusing material into the lamp housing removed the angular spectral mismatch and improved the emission
distribution (Fig. 2). Computer modelling demonstrated vastly less CPD from any of the KrCl sources whencompared to 10 minutes of sun exposure in both temperate and Mediterranean climates.

Fig. 2 –Spectral distribution as a function of viewing angle for source without diffuser (left) and with diffuser (right)

In conclusion, there is variation in the spectral distribution of currently available commercial KrCl sources, which
necessitates full spectrum assessment against the exposure limits and not just assessment against the 222 nm
exposure limit. Full spectrum assessment will become even more important if new ACGIH-proposed threshold
limit values are adopted. With real-world installations, if there is overlapping emission from fixtures, it should not
be assumed that the greatest hazard is located directly underneath a fixture.
References
1) Kitagawa, H., Nomura, T., et al. Effectiveness of 222-nm ultraviolet light on disinfecting SARS-CoV-2
surface contamination. American Journal of Infection Control, 2020.
2) Narita, K., Asano, K., et al. Ultraviolet C light with wavelength of 222 nm inactivates a wide spectrum of
microbial pathogens. Journal of Hospital Infection, 2020.
3) Welch, D., Buonanno, M., et al. Far-UVC light: A new tool to control the spread of airborne-mediatedmicrobial diseases. Scientific Reports, 2018.
4) Hickerson, R., Conneely, M., et al. Minimal, superficial DNA damage in human skin from filtered farultraviolet-C (UV-C). British Journal of Dermatology, 2021.
5) Eadie, E., Barnard, I.R.M., et al. Extreme Exposure to Filtered Far-UVC: A Case Study. Photochemistry
and Photobiology, 2021.
6) Kaidzu, S., Sugihara, K., et al. Re-Evaluation of Rat Corneal Damage by Short Wavelength UV Revealed
Extremely Less Hazardous Property of Far-UV-C. Photochemistry and Photobiology, 2021.
7) Woods, J., Evans, A., et al. The effect of 222-nm UVC phototesting on healthy volunteer skin: a pilot
study. Photodermatology, Photoimmunology and Photomedicine, 2015.
8) Buonanno, M., Welch, D. and Brenner, D. Exposure of Human Skin Models to KrCl Excimer Lamps: The
Impact of Optical Filtering. Photochemistry and Photobiology, 2021.
Acknowledgements. We would like to thank all the commercial enterprises who donated or loaned their KrCl
sources used in this research. No commercial enterprise had an influence in this research. Dr Ewan Eadie
acknowledges financial support from NHS Research Scotland Career Researcher Fellowship. Dr Paul O’Mahoneyis funded by Medi-lase (registered charity SC037390) and the Alfred Stewart Trust. Louise Finlayson
acknowledges financial support from EPSRC Industrial Doctorate Centre Scheme (2262922) and the Laser
Research and Therapy Fund (registered charity SC030850).

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IL-2.4.1

Advantages of NIR light-triggered cancer therapies
Mladen Korbelik
BC Cancer, Integrative Oncology Department, Vancouver BC, Canada
Recent years have seen an increased interest in research and development of phototherapies performed in nearinfrared (NIR) sections of light spectrum, particularly photodynamic therapy (PDT) and photothermal therapy(PTT) for treatment of cancer. These efforts are motivated by several advantageous attributes:






High penetration depths in biological tissues;
Less toxic consequences for normal cells and tissues;
Lower light irradiation doses involved;
More effective inhibition of cancer cell proliferation through cellular DNA damage;Opportunity for concomitant enhanced nanodrug delivery to targeted sites.

This presentation will particularly elaborate how both PDT and PTT function as photoimmunotherapy (PIT) that isextremely efficacious in engaging the patient’s immune system through the induction of immunogenic cell death(ICD), release of damage-associated molecular patterns (DAMPs), and greatly increased availability of highly
immunogenic tumor antigens. The resultant advantages are highlighted by systemic tumor immune rejection
favorable for treatment of metastatic cancer, and immunological memory generation preventing disease recurrence.

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IL-2.4.2
Dye Doped Silica Nanoparticles as Photoactive Organized Systems for Nanomedicine
Luca Prodi1, Matteo Cingolani, Damiano Genovese, Liviana Mummolo, Enrico Rampazzo, Nelsi Zaccheroni
Dipartimento di Chimica “Giacomo Ciamician” – Università di Bologna – Bologna – Italy
Silica nanoparticles are versatile platforms with many intrinsic features, including a low toxicity. Their proper designand derivatization yield particularly stable bright nano-systems displaying multiple functions,1 which can be used for
either optical and photoacoustic imaging2 and for photoluminescence (PL) and electrochemiluminescence (ECL)
sensing.3 In addition, silica nanoparticles can also be used as platforms for photodynamic and photothermal
therapies.2 For these reasons, silica nanoparticles already offer unique opportunities, and further improvement andoptimization can substantially expand their possible applications in fields of high impact, such as medical diagnostics
and therapy, environmental analysis, and security.
In this context, we have developed a direct micelle assisted strategy based on the use of Pluronic F127 as highmolecular weight surfactants. The one-pot synthesis yields PEGylated silica nanoparticles endowed with very highmonodispersity, colloidal stability and core-shell structure. These Pluronic Silica NanoParticles (PluS NPs), with asilica core of about 10 nm and an overall hydrodynamic diameter of about 25 nm, can be tailored for optimization ofprocesses such as directional energy transfer, providing systems with extremely valuable functions: high lightharvesting capability, signal-to-noise maximization, multiplex output, and signal amplification. In-vivo experimentproved the absence of toxic effects on mice even after three months from injection.4 A library of functionalized
Pluronic surfactants allowed for a straightforward and modular method for surface derivatization, a fundamental stepsince we found that their possible applications can be controlled by nanoparticle functionalization; the control ofsurface charge (density and localization) allows, for example, for sentinel lymph node targeting.4 Furthermore, we
demonstrated that the drug loading ability can be tuned with a suitable choice of the silica precursor. Finally, weinvestigated the interaction with fluorogenic hyaluronan, a synthetic nanogel that improves cell internalization of
PluS NPs.6
References
1) M. Montalti, L. Prodi, E. Rampazzo, N. Zaccheroni, “Dye Doped Silica Nanoparticles as Luminescent Organized
Systems for Nanomedicine”, Chem. Soc. Rev., 2014, 43, 4243.
2) S. Biffi, L. Petrizza, C. Garrovo, E. Rampazzo, L. Andolfi, P. Giustetto, I. Nicolov, G. Kurdi, M. Danailov, G.
Zauli, P. Secchiero, L. Prodi, “Multimodal NIR-Emitting PluS Silica Nanoparticles with Fluorescent,
Photoacoustic and Photothermal Capabilities”, Int. J. Nanomedicine, 2016, 11, 4865.
3) A. Zanut, F. Palomba, M. Rossi Scota, S. Rebeccani, M. Marcaccio, E. Rampazzo, G. Valenti, F. Paolucci, Luca
Prodi, “Dye‐Doped Silica Nanoparticles for Enhanced ECL‐Based Immunoassay Analytical Performance”,
Angew. Chem. Int. Ed., 2020, 59, 21858.
4) M. Helle, E. Rampazzo, M. Monchanin, F. Marchal, F. Guillemin, S. Bonacchi, F. Salis, L. Prodi, L. Bezdetnaya,“The Surface Chemistry Architecture of Silica Nanoparticles Determine the Efficiency of in Vivo Fluorescence
Lymph Node Mapping”, ACS Nano, 2013, 7, 8645.
5) F. Palomba, D. Genovese, E. Rampazzo, N. Zaccheroni, L. Prodi, L. Morbidelli, “PluS Nanoparticles loaded withSorafenib: synthetic approach and their effects on endothelial cells”, ACS Omega, 2019, 4, 13962.
6) F. Palomba, E. Rampazzo, N. Zaccheroni, M. Malferrari, S. Rapino, V. Greco, C. Satriano, D. Genovese, and L.
Prodi, ACS Appl. Mater. Interfaces, 2020, 12, 6, 6806–6813
Acknowledgements. This work was supported by Italian Ministero dell’Istruzione, Università e Ricerca (PRIN
project number 2017EKCS3)

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IL-2.4.3
Drug Delivery Systems Targeting Tumour Microenvironment for Near-Infrared
Photodynamic Therapy
Takahiro Nomoto1
1) Tokyo Institute of Technology, Institute of Innovative Research, Yokohama, Japan
Photodynamic therapy (PDT) combines delivery of photosensitizers (PSs) to a target site and subsequent
photoirradiation to excite the PSs. The key to extending PDT application is the development of PSs that canaccumulate selectively within a target site without retaining in normal tissues. For this purpose, we developed two
kinds of drug delivery systems (DDSs) targeting acidic tumour environment and tumour-associated blood vessels.
The DDS targeting acidic tumour environment was developed by utilizing the polymer that exerts
hydrophilic-to-hydrophobic phase transition. Generally, hydrophilic character of drugs prevents unfavourable
interaction with biological components, but it also compromises interaction with target cancer cells, decreasing
therapeutic efficiency. Meanwhile, the hydrophobic character facilitates the cellular uptake, but it causes untoward
distribution to normal tissues due to nonspecific interaction. We thus expected that the switching ofhydrophilic/hydrophobic characters in response to tumour acidic environment may permit selective accumulationof PSs. To realize this concept, we developed poly(N-isopropylacrylamide/2-aminoisopropylacrylamide)(P(NIPAAm/AIPAAm)) whose AIPAAm units were further modified with hydrophilic acid-labile 2-propionic-3methylmaleic (PMM) amide1. This polymer, termed P(NIPAAm/AIPAAm-PMM), exhibits hydrophilic character at
pH 7.4; however, at pH 6.8, PMM moieties were cleaved from the polymer and P(NIPAAm/AIPAAm) exerts
hydrophobicity due to its lower critical solution temperature (~30°C). Owing to this phase transition,
P(NIPAAm/AIPAAm-PMM) accomplished efficient cellular uptake in cultured cancer cells in response to mild
acidic pH. We then conjugated hydrophilic phthalocyanine-based PS, IRDye 700DX, with the polymer andexamined the potential in PDT2. The IRDye 700DX-conjugated polymer importantly accomplished enhanced
accumulation within a subcutaneous tumor in a mouse after intravenous injection and ultimately improved the PDT
efficiency of IRDye 700DX, while the unfavorable photochemical damage to the skin was considerably prevented
by avoiding the penetration to the normal tissue due to the hydrophilic character in the bloodstream.
Although acidic pH of tumours has been widely reported by many researchers, some tumours do not show
homogeneous acidic environment because of the tumour heterogenicity. In such cases, it is difficult to deliver PSs
with the aforementioned polymer homogeneously to the tumour tissue, which may lead to the recurrence after PDT.
In this regard, vascular targeted PDT may offer great potential because obstruction of tumor-associated vasculature
will block the flow of oxygen and nutrient, thereby killing tumor cells in a deep region where PSs cannot bedelivered. For this purpose, we synthesized poly(ethylene glycol)-poly(L-glutamate) whose side chains weremodified with multiple cyclic RGD peptides, which have the strong affinity with avb3 integrins overexpressed on
tumour-associated blood vessels as well as many cancer cells3. The polymer conjugated with IRDye 700DX
heterogeneously distributed in the tumour tissue and preferentially accumulated within the tumour-associatedvasculature, thereby accomplishing significant PDT effect in a subcutaneous tumour model. Our approaches using
these DDSs may have potential to extend the PDT application.
References
1) Muttaqien, S. E., Nomoto, T. et al. Poly(N-isopropylacrylamide)-based polymer-inducing isothermalhydrophilic-to-hydrophobic phase transition via detachment of hydrophilic acid-labile moiety.
Biomacromolecules 20, 1493-1504 (2019).
2) Muttaqien, S. E., Nomoto, T. et al. Photodynamic therapy using LCST polymers exerting pH-responsive
isothermal phase transition. Journal of Controlled Release 328, 608-616 (2020).
3) Dou, X., Nomoto, T. et al. Effect of multiple cyclic RGD peptides on tumor accumulation and intratumoral
distribution of IRDye 700DX-conjugated polymers. Scientific Reports 8, 8126 (2018).
Acknowledgements. This work was supported by Center of Innovation (COI) program (JPMJCE1305) from Japan
Science and Technology Agency (JST), JSPS KAKENHI (JP26882022 and JP20K20196) and Kobayashi
International Scholarship Foundation.
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OC-2.4.4
The impact of atropisomerim on Redaporfin photodynamic therapy efficacy
Claire Donohoe1,2, Fábio Scharbele1,Mariette M. Pareira1, Mathias O. Senge2, Luís G. Arnaut1, Lígia C. Gomes-daSilva1
1) Department of Chemistry, University of Coimbra, Rua Larga, 3004-535 Coimbra, Portugal
2) Medicinal Chemistry, Trinity Translational Medicine Institute, Trinity Centre for Health Sciences, St. James’s
Hospital, Trinity College Dublin, The University of Dublin, Dublin 8, Ireland
Atropisomerism presents an intriguing, often neglected, source of structural variety in drug development. Suchdiversity in tetrapyrrole structures may enhance photosensitizer development for photodynamic therapy (PDT) andrelated drug developments. The present work demonstrates that the photosensitizer Redaporfin (and related precursor porphyrin molecules) atropisomers can be separated, do not interconvert at room temperature and exert
different biological effects. Redaporfin, a pre-clinical synthetic sulfonamide fluorinated bacteriochlorin, presents
ideal properties for studying PDT including enhanced photostability and strong absorption at 750 nm. Hinderedrotation of the Cm-aryl-macrocyclic bonds of the bacteriochlorin and porphyrin structures results in different spatial
orientations of the sulfonamide groups in the meta positions. They are defined as: α 4 when all of the sulfonamide
substituents of the phenyl groups are on the same side of the macrocycle plane; α 3β when three of the sulfonamides
are on the same side of the plane and one is on the opposite side; α2β2 when two sulfonamide groups are on each side
and adjacent to each other and finally, αβαβ when two sulfonamides are on each side but alternate in the positions
with respect to the macrocycle. Although the photo- and physicochemical properties of the four atropisomers are
similar, their therapeutic efficacies are dramatically different. In vitro studies have demonstrated significant
variability of the atropisomer phototoxicity and cellular internalization levels. In particular, the α4 atropisomer has
displayed the highest levels of uptake and phototoxicity. Atropisomers have presented similar mechanisms of cellular
uptake, primarily passive diffusion with subcellular localization in the endoplasmic-reticulum-Golgi complex.
Heightened α4 internalization by cells of the tumor microenvironment has been observed in vivo. Atropisomertherapeutic efficacy has been demonstrated to vary in vivo when sufficient time for photosensitizer tumor cellinternalization occurs. A better understanding of how atropisomerism impacts therapeutic effects may contribute to
the establishment of enhanced drug development strategies for atropisomer drug mixtures.
References
1) L.G. Arnaut, M.M. Pereira, J.M. Dabrowski, E.F. Silva, F.A. Schaberle, A.R. Abreu, L.B. Rocha, M.M. Barsan,
K. Urbanska, G. Stochel, C.M. Brett, Photodynamic Therapy Efficacy Enhanced by Dynamics: The Role of Charge
Transfer and Photostability in the Selection of Photosensitizers, Chemistry, 2014, 20, 5346-5357.
2) L.B. Rocha, L.C. Gomes-da-Silva, J.M. Dabrowski, L.G. Arnaut, L.B. Rocha, L.C. Gomes-da-Silva, J.M.
Dabrowski, L.G. Arnaut, Elimination of primary tumours and control of metastasis with rationally designed
bacteriochlorin photodynamic therapy regimens. Eur J Cancer, 2015, 51, 1822-1830. Eur J Cancer, 2015, 51, 18221830.
Acknowledgements. This project has received funding from the European Union’s Horizon 2020 research andinnovation programme under the Marie Sklodowska-Curie grant agreement number 764837 (Polythea—How light
can save lives)

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IL-2.4.5

Ru(II) and Os(II) complexes as Hypoxia-Active Photosensitizer Classes for PDT: thecontribution of computational studies
Marta Erminia Alberto
1) Università della Calabria, Dipartimento di Chimica e Tecnologie Chimiche, Arcavacata di Rende (CS), Italy
Due to their appealing physicochemical properties, Ru(II) compounds have found wide applications in biological andmedical fields. Complexes of its analogue group 8 counterpart Osmium instead, are relatively unexplored in
medicinal chemistry. Despite this, they have an advantage for photobiological applications due to their extended
absorption windows, and both Ru(II) and Os(II) are now attracting an increasing interest as potential candidates for
Photodynamic Therapy (PDT) and Photochemotherapy (PCT) approaches.
To shed light on the complex photochemistry of these molecules, the information that can be gained fromcomputational studies is very useful, as it has the potential to increase the understanding of the entire photochemical
pathways involved, providing a rationale for the observed photobiology along with useful guidelines for the theorydriven design of next-generation agents.
The important contribution of computational studies in this field is reviewed presenting the outcomes of joint
experimental and theoretical studies on several Os(II)- and Ru(II)-polypiridyl based scaffold combined with theimidazo[4,5-f ][1,10]-phenanthroline-oligothiophene (IP-nT) ligand motif (n=0-4).1,2 (Fig.1)
Those compounds show both powerful 1O2 sensitization and unprecedented PIs for hypoxic photoactivity (1% O2),
among the largest reported to date, making of our Os(II)-complexes, the first hypoxia-active Os-based
photosensitizers ever proposed, demonstrating the utility of such metal for phototherapy applications.
DFT results reveal the pivotal role of the switch in the nature of lowest-lying triplet excited state from a metal-toligand charge transfer (3MLCT) to intraligand charge transfer one (3ILCT) at n=3, with a lower energy and longer
lifetime for n=4, for both Ru(II) and Os(II) families. Moreover, it provides fundamental insights into the structureactivity relationships along the series of related complexes from n=0 to 4 with respect to ground and excited stategeometries, photophysical properties, excited state lifetimes and decay mechanisms.

Fig. 1 – Molecular Structure of a representative of the Ru(II)- and Os(II)-polypiridyl IP-nT complexes proposed as HypoxiaActive Photosensitizers

References
1) C.G. Cameron, M.E. Alberto, S.A. McFarland et al. Breaking the Barrier: An Osmium Photosensitizer with
Unprecedented Hypoxic Phototoxicity for Real World Photodynamic Therapy, Chem. Sci., 11, 9784-9806, 2020;
2) C.G. Cameron, M.E. Alberto, S.A. McFarland et al. “Os(II) Oligothienyl Complexes as a Hypoxia-Active
Photosensitizer Class for Photodynamic Therapy, Inorg Chem, 59, 16341−16360, 2020;3) M.E. Alberto, J. Pirillo, N. Russo, C. Adamo, A theoretical exploration of TypeI/TypeII dual photoreactivity of
new promising Ru(II)-dyads for PDT approach, Inorg Chem, 55, 11185–11192, 2016;
Acknowledgements. We acknowledge the CINECA award under the ISCRA initiative, for the availability of highperformance computing resources and support (IsC85_Hypo-PDT)
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IL-2.4.6

Light-driven coordination compounds and hybrid assemblies as multimodal
bioimaging agents and ROS-photosensitizers: Design, synthesis and implementation
Cristian A. Strassert
Institut für Anorganische und Analytische Chemie & Cells in Motion Interfaculty Centre – Westfälische WilhelmsUniversität Münster, Münster, Germany
Diverse approaches have allowed us to fully control the aggregation of planar coordination compounds and to tune
their photophysical properties for phototherapy, functional microscopy and multimodal bioimaging.
We have designed a series of (na)phthalocyanine-based photosensitizers able to target, to label and to photoinactivate
pathogenic and antibiotic resistant bacteria upon irradiation with red light. For this purpose, it was necessary to avoid
stacking by diverse (supra)molecular strategies. Currently, we are extending these concepts to targeted, fully watersoluble and biodegradable platforms, a prerequisite for biomedical applications. These approaches include the use of
dextrin conjugates and cyclodextrin vesicles that selectively photoinactivate Gram-positive strains. On the other
hand, axially decorated dicationic Si(IV) phthalocyanines can kill both Gram-positive and Gram-negative bacteria,
despite showing antibiotic resistance. We have also implemented light-driven arrays for spatiotemporally resolved
functional microscopy to monitor in situ the response towards ROS of eukaryotic as well as prokaryotic cells andbiofilms. Insertion of open-shell transition metal cations and tuning of the macrocycles’ substitution pattern yielded
NIR-absorbing sonophores for in vivo photoacoustic imaging.
Acknowledgements. This work was supported by the Deutsche Forschungsgemeinschaft (Cluster of Excellence
Cells in Motion EXC1003).

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IL-2.4.7

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OC-2.4.8, P-2.4.10

Clinical use of a near-infrared fluorescence imaging system in photodynamic therapy
using liposomal indocyanine green: a case series study
Hiromi Muranishi1-3, Yasuo Komura1-3, Atsushi Hori3, Shinichi Hori3
1) Rinku Medical Clinic, Osaka, Japan; 2) Kyoto Oike Medical clinic, Kyoto, Japan; 3) IGT clinic, Osaka, Japan(Background) Photodynamic therapy (PDT) is an alternative minimally invasive therapeutic procedure for malignant
diseases currently used in clinical practice. Indocyanine green (ICG) is considered as a good photosensitizer for PDT.
ICG has been encapsulated in an inner aqueous compartment of liposome (liposomal ICG), improving its stability
and efficacy. However, because of the lack of clinical evidence, the biodistribution of liposomal ICG and the optimal
clinical strategy for PDT using liposomal ICG is unclear. We reported a series of cases using a near-infrared
fluorescence (NIR) imaging system to evaluate the biodistribution of liposomal ICG in patients with breast cancer
undergoing PDT.
(Purpose) This case-series study aimed to evaluate the biodistribution of liposomal ICG in patients with breast cancer
undergoing PDT.
(Method and Result) Four patients with breast cancer underwent PDT with liposomal ICG in addition to TACE from
August 2020 to October 2020. Patients ranged from 41 to 55 years of age. Three patients had metastatic breast cancer,
but daily activity was preserved. The patients had either finished or rejected standard therapy. Patients wereadministered 300 mg liposomal ICG (180 mg intravenously and 120 mg intratumorally via the feeding artery) 24
hours before PDT during a transcatheter arterial chemoembolization (TACE) procedure. We used near-infrared
fluorescence imaging system (LIGHTVISION®; Shimadzu Corporation) to detect the biodistribution of liposomal
ICG 3, 6, and 24 hours before the PDT procedure. The peak intertumoral liposomal ICG uptake was shown 24 hours
after liposomal ICG administration in three patients. Only one patient had peak uptake at 6 hours, with no uptake at
24 hours.
(Discussion) Although several preclinical studies have shown the efficacy and safety of PDT with liposomal ICG for
several malignant diseases, few have reported using PDT with liposomal ICG in the clinical setting 1-3. However,some clinical studies have already reported that the optimal time for PDT with ICG is 24 - 48 hours after ICG
administration. In a similar fashion we found that the optimal time for PDT procedure was 24 hours after liposomal
ICG administration, except in one case. Compared with other photosensitizers, a strength for using ICG is that
detection by NIR-imaging systems might be easier. Our study was a case-series study, and further studies are needed
to clarify the optimal clinical strategy for PDT with liposomal ICG.
(Conclusion) NIR-imaging system may be and adjuvant in evaluation of liposomal ICG biodistribution in patientswith breast cancer and assisting in the decision-making for the use of PDT with liposomal ICG.

Fig. 1 - Liposomal indocyanine green in scanning electron microscope

Fig. 2 - Findings of NIR-imaging system

References
1) Lv Chen, Lulu Zhou, et al: Tumor-Targeted Drug and CpG Delivery System for Phototherapy and DocetaxelEnhanced Immunotherapy with Polarization toward M1-Type Macrophages on Triple Negative Breast Cancers. Adv
Mater. 2019 Dec;31(52):e1904997
2) María Julia Lamberti, Natalia Belén Rumie Vittar, et al; Breast cancer as photodynamic therapy target: Enhanced
therapeutic efficiency by overview of tumor complexity. World J Clin Oncol 2014; 5(5): 901-907
3) ElżbietaOstańska, DavidAebisher, et al; The potential of photodynamic therapy in current breast cancer treatment
methodologies. Biomed Pharmacother. 2021 May;137:111302
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OC-2.4.9

A low molecular weight carboxamide halogenated bacteriochlorin for the treatmentof highly aggressive tumors
Bárbara Lima1, Helder Tão1, Mariette Pereira1, Lígia C. Gomes-da-Silva1, Luis G. Arnaut1
1) CQC, Coimbra Center of Chemistry, Coimbra, Portugal
Photodynamic therapy (PDT) is a clinically approved therapy that combines the use of a photosensitizer (PS) andlight, which in the presence of molecular oxygen generates reactive oxygen species. The generated oxidative stressleads to cell death of cancer cells and other cells of the tumour microenvironment. Additionaly, PDT is being
perceived as a new form of photoimmunotherapy owing to its ability to trigger anti-tumor immunity.
Despite promising, PDT (as other oncological therapies) has a small effect on very aggressive and poorly
immunogenic tumors such as, the 4T1 orthotopic mouse model. This model closely mimics the immunogenicity,
metastatic properties and growth characteristics of stage IV of human breast cancer. It is very challenging to treat
4T1 orthotopic tumors with a variety of photosensitizers namely, chlorin-e6, verteporfin, talaporfin sodium andredaporfin. The poor vascularization of 4T1 tumors and the high accumulation of immunosuppressive cells contributeto the poor responses to various therapies, including PDT. Additional challenges rely on its orthotopic localization at
the mammary gland that significantly increases the risk of lethality upon tumor illumination.
A new family of bacteriochlorins with relatively smaller molecular weight and moderate lipophilicity was recently
developed by our group with the expectation that these properties will favor cellular internalization. In vitro
screenings pointed out LUZ51b as a promising compound. LUZ51b is a small lipophilic halogenated carboxamide
bacteriochlorin (MM= 595.59 g/mol; log Pow = 2.9) that is characterized by an elevated ROS quantum yield andstrong absorption at 734 nm.
Preliminary results using phosphatidylcholine liposomes demonstrated that LUZ51b interacts more with the lipid
membranes than redaporfin, a bacteriochlorin of 1135 g/mol and log Pow = 1.9. In accordance, after a short time ofincubation, such as 4 h, high levels of cellular internalization of LUZ51b are observed, which is in contrast to
redaporfin that only peaks at 24h. LUZ51b mainly accumulates at the endoplasmic reticulum and Golgi
compartments (being a potential inducer of immunogenic cell death) and remarkably, it kills cancer cells atconcentration in the range of nM. Pre-clinical studies confirmed the high efficacy of photo-activated LUZ51b incancer mouse models of different aggressiveness: low (subcutaneous CT26 tumors), moderate (subcutaneous B16F10
tumors) and high (orthotopic 4T1 tumors). Remarkably, an optimized protocol consisting in the intravenous
administration of 0.25 mg/kg of LUZ51 followed by the deliver of 20 J/cm2 enabled the cure of approximately 60%(3 out of 5) Balb/c mice bearing 4T1 tumors. Further rechallenge of these mice with 4T1 cells revealed delay oftumor growth which might indicate the presence of immunological memory.
Overall, LUZ51b combine unique properties that turn it in a very promising photosensitizer for the treatment of
aggressive and advanced tumors. Further studies will focus on its potential for the treatment of pancreatic cancer,
alone or in the combination with immunotherapy. This might constitute a major breakthrough in oncology as thistype of tumors lack effective therapeutic options.
Acknowledgements. This project has received funding from the European Union’s Horizon 2020 research andinnovation programme under the Marie Sklodowska-Curie grant agreement n°764837 and Coimbra Chemistry Center
under the project UID/QUI/00313/2019).

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P-2.4.11

Metal-based photosensitizers: approaches to enhance the efficiency of photodynamic
therapy in cancer treatment
Ge Shi, John Roque III, Houston D. Cole, Colin G. Cameron, Sherri A. McFarland
Department of Chemistry and Biochemistry, The University of Texas at Arlington, Arlington, Texas, 76019 USA
Photodynamic therapy (PDT) has become an approved medical intervention to treat certain types of cancer. It has
been utilized to complement and in certain cases to replace traditional anticancer treatment such as surgery,chemotherapy or radiotherapy. In PDT, a photosensitizer that is generally organic compound is administrated
locally or systemically at a nontoxic dose and then activated with red light in the PDT window to produce a potent
PDT response with minimal dark toxicity. The photosensitizers that have been designed and developed in our lab
are metal-based coordination complexes, which we believe are promising PDT agents dues to their special
photophysical and photobiological properties. Ruthenium- and osmium-polypyridyl complexes serve as examples,with our own TLD1433 currently undergoing a phase II clinical trial for bladder cancer. This presentation will
share insights regarding the development of some of our latest photosensitizers for PDT.

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PL-6.1
Radiotherapeutic effects of radioluminescent nanomaterials
Anne-Laure Bulin
Synchrotron Radiation for Biomedicine, Grenoble Alpes University, INSERM UA7, Grenoble, France.
More than 50% of cancer patients undergo radiation therapy in the course of their treatment. However, because of alack of specificity for tumor tissues, delivering therapeutically effective doses of X-rays with tolerable toxicity on
healthy surrounding tissues remains a challenge. In order to enhance the therapeutic window of radiotherapy for
difficult-to-treat cancers, innovative nanoscintillators may induce various radiotherapeutic effects. Nanoscintillators,
also known as radioluminescent nanoparticles, down-convert ionizing radiations into visible light. Thus, they can act
as internal light sources for X-ray activated PDT 1. When accumulated in the tumor prior to radiotherapy,
nanoscintillators can induce various effects that can synergize. First, when conjugated to photosensitizers,
nanoscintillators can induce deep-tissue photodynamic therapy (PDT). This strategy would overcome the shallow
penetration of light in tissues, one of the major limitations of PDT 2. Second, when nanoscintillators emit in theultraviolet (UV)-range, and more specifically in the UV-C, direct DNA-damage can be induced 3,4. Finally, because
nanoscintillators are made of high-Z element, their accumulation in the tumor prior to radiotherapy will create apurely physical effect of radiation dose-enhancement 5. This effect is initiated by a higher photoelectric absorptionof orthovoltage X-rays by high-Z elements compared to soft tissues that leads to a higher production of photo- andAuger electrons that enhance the damage to cancer tissues.
While proof-of-concept studies of radiotherapeutic effects of nanoscintillators have already been demonstrated by us
and others, our goal is now to provide a comprehensive description of the mechanisms involved during radiotherapy.
We leverage a multidisciplinary approach ranging from physics to biology, to preclinical trials. Our approach
includes Monte Carlo simulations to guide the design of optimized nanoconjugates, measurements with reactive
oxygen species-sensitive fluorescent probes, in vitro experiments performed on 3D models of glioblastoma andpancreatic cultures and preclinical tests. In order to investigate the particular role of the X-ray energy in the overall
treatment efficacy, radiotherapy is delivered by monochromatic tunable synchrotron radiation.
References
1. Dujardin, C. et al. Needs, Trends, and Advances in Inorganic Scintillators. IEEE T Nucl Sci 65, 1977–1997(2018).
2. Sun, W., Zhou, Z., Pratx, G., Chen, X. & Chen, H. Nanoscintillator-Mediated X-Ray Induced Photodynamic
Therapy for Deep-Seated Tumors: From Concept to Biomedical Applications. Theranostics 10, 1296–1318(2020).
3. Squillante, M. R. et al. Fabrication and characterization of UV-emitting nanoparticles as novel radiation
sensitizers targeting hypoxic tumor cells. Opt Mater 80, 197–202 (2018).
4. Müller, M. et al. UV scintillating particles as radiosensitizer enhance cell killing after X-ray excitation.
Radiother Oncol 129, 589–594 (2018).
5. Bulin, A.-L. et al. Radiation Dose-Enhancement Is a Potent Radiotherapeutic Effect of Rare-Earth Composite
Nanoscintillators in Preclinical Models of Glioblastoma. Advanced Science 7, 2001675 (2020).
Acknowledgements. My group is supported by grants from the French Medical Research Foundation (Fondation
pour la Recherche Médicale, ARF20170938739), the Auvergne Rhône-Alpes Region (NEPTUNE project), INSERM
Cancer, the French Ligue Contre le Cancer Foundation, the LABEX PRIMES of the Université of Lyon (ANR-11LABX-0063), the “Rotary Club” Association through the program “Jetons Cancer” and the INSERM.

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PL-6.2A NanoBioengineering Frontier for Next-Generation Optical Devices
Ardemis A. Boghossian1
1) Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
The vast expansion of available synthetic biology tools has led to explosive developments in the field of materials
science. No longer confined to engineering just synthetic materials, the increased accessibility of these tools has
pushed the frontier of materials science into the field of engineering biological and even living materials. By
coupling the tunability of nanomaterials with the prospect of re-programming living devices, one can re-purpose
biology to fulfill needs that are otherwise intractable using traditional engineering approaches.
Optical technologies in particular could benefit from capitalizing on untapped potential in coupling the opticalproperties of nanomaterials with the specificity and scalability of biological materials. This presentation highlights
specific applications in optical sensing and light-harvesting energy technologies that exploit the synergistic
coupling of nanobio-hybrid materials. We discuss the development of bio-conjugated single-walled carbon
nanotubes (SWCNTs) for near-infrared fluorescence sensing and the application of these nano-bioptic sensors forcontinuous measurements in living cells and organisms. We further explore the development living photovoltaics
based on bioengineered, photosynthetic organisms with augmented capabilities.
Acknowledgements. This work was supported by the Swiss National Science Foundation (SNSF), European
Research Council (ERC) Starting Grant, CODEV seed funding, and Swiss Competence Center (SCCER) for
Hydrogen Production & Storage.

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IL-3.1.1

Type I interferons enhance repair of ultraviolet radiation induced DNA damage andregulate cutaneous immune suppression
Mohammad Asif Sherwani1, Israr Ahmad1, Ahmed Abdelgawad1, Kevin Yang1, Chander Raman1, Craig A
Elmets1,2,3, Nabiha Yusuf1,2,3
1) Department of Dermatology, 2) O’Neal Comprehensive Cancer Center, University of Alabama at Birmingham,
AL, USA; 3) Veteran Affairs Medical Center, Birmingham, AL, USA.
Type I interferons (IFNs) are important enhancers of immune responses which are downregulated in humancancers, including skin cancer. Solar ultraviolet (UV) B radiation is a proven environmental carcinogen, and its
exposure contributes to the high prevalence of skin cancer. The carcinogenic effects of UV light can be attributed
to the formation of cyclobutane pyrimidine dimers (CPD) and errors in repair and replication of DNA. Treatment
with a single dose of UVB (100 mJ/cm2) upregulated IFNα and IFNβ in the skin of C57BL/6 mice. They werepredominantly produced by CD11b+ cells. Mice lacking the type I IFN receptor 1 (IFNAR1) had decreased repairof CPD following cutaneous exposure to a single dose of UVB (100 mJ/cm2). UVB induced expression of the DNA
repair gene xeroderma pigmentosum A (XPA) in wild type (WT) mice. In contrast such treatment in IFNAR1(IFNAR1-/-) mice downregulated XPA. A local UVB regimen consisting of UVB radiation (150 mJ/cm2) for 4 days
followed by sensitization with the hapten 2,4, dinitrofluorobenzene (DNFB) resulted in significant suppression of
immune responses in both WT and IFNAR1-/- mice. However, there were significantly higher
CD4+CD25+Foxp3+ regulatory T-cells and CD11b+Gr1+ myeloid cells in draining lymph nodes of IFNAR1-/mice in comparison to WT mice. Overall, our studies reveal a previously unknown action of type I IFNs in repair ofphotodamage. Strategies that enhance type I IFN production may be useful for prevention of pre-malignant skinlesions before they develop into skin cancer.
Acknowledgements. This work was supported by 1R01AR071157-01A1 from National Institute of Arthritis andMusculoskeletal and Skin Diseases (NY), American Skin Association (KY and NY), and I01BX003395 from
Veterans Administration (CAE).

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IL-3.1.2
Modulation of B cell responses to TLR7 activation following narrowband UVBphototherapy in early multiple sclerosis

Stephanie Trend1,2, Jonatan Leffler1, Matthew N Cooper1, Scott N Byrne3,4, Allan G Kermode2,5, Martyn A French6,
Prue H Hart1
1) Telethon Kids Institute, University of Western Australia, Perth, WA, Australia;2) Centre for Neuromuscular and
Neurological Disorders, Perron Institute for Neurological and Translational Science, University of Western
Australia, Perth, WA, Australia;3) School of Medical Sciences, Faculty of Medicine and Health, The University ofSydney, Sydney, NSW, Australia;4) Centre for Immunology and Allergy Research, Westmead Institute for Medical
Research, Westmead, NSW, Australia;5) Institute for Immunology and Infectious Disease, Murdoch University,
Perth, WA, Australia;6) UWA Medical School and School of Biomedical Sciences, University of Western Australia,
Perth, WA, Australia.
Multiple sclerosis (MS) is an immune-mediated disease that involves episodes of inflammation in the central nervous
system, and is the most common degenerative neurological condition affecting young adults. Low UV radiation
exposure is associated with an increased risk of developing MS. To investigate whether UV radiation would be
beneficial for people with the earliest form of MS, (Clinically Isolated Syndrome [CIS]), the PhoCIS trial wasestablished, and participants received either sub-erythemal narrowband UVB (311-312 nm) phototherapy three times
per week for 8 weeks (n=7) or no phototherapy (controls; n=6). Peripheral blood mononuclear cells (PBMC) werecollected at baseline and at 8 weeks, following completion of therapy, and flow cytometry was used to investigate
the frequencies of B cell subsets. In addition, the expression of TNF and IL-10 in B cells cultured with or without a
TLR7/8 agonist (R848) was investigated in each individual at 8 weeks, adjusted for responses at baseline. Significant
changes in the relative frequencies of naive, marginal zone (MZ)-like and class switched memory B cells wereobserved after the treatment period in individuals receiving phototherapy. Phototherapy had no detectable effect on
IL-10 expression in B cells. However, the expression of TNF following culture with R848 was significantly decreasedin B cells, particularly the MZ-like cell population, after phototherapy. These results suggest that phototherapyassociated priming effects were detected upon subsequent polyclonal B-cell activation in B cells.
Acknowledgements. This work was supported by grants from Multiple Sclerosis WA. The trial recruiting the CIS
phototherapy patients was supported by the National Health and Medical Research Council of Australia (ID1067209).

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IL-3.1.3

Exposure to systemic immunosuppressive solar simulated ultraviolet radiation alters
T cell recirculation through sphingosine 1 phosphate
Scott N. Byrne*1,2, Benita C.Y. Tse1, Rachael Ireland1,2, Jun Yup Lee1, Felix Marsh-Wakefield1, Lai Fong Kok1 andAnthony Don3,4
1) The University of Sydney, School of Medical Sciences, Faculty of Medicine and Health, Sydney, New South
Wales, Australia; 2) Westmead Institute for Medical Research, Centre for Immunology and Allergy Research,
Westmead, New South Wales, Australia; 3) Centenary Institute, The University of Sydney, New South Wales,
Australia; 4) The University of Sydney, NHMRC Clinical Trials Centre, Faculty of Medicine and Health, NewSouth Wales, Australia.
*Presenting author: scott.byrne@sydney.edu.au
Systemic suppression of adaptive immune responses is a major way through which ultraviolet (UV) radiation
contributes to skin cancer development. Immune suppression is also likely to explain how UV protects from someautoimmune diseases such as multiple sclerosis. However, the mechanisms underlying UV-mediated systemic
immune suppression are not well understood. Exposure of mice to doses of solar simulated UV known to suppress
systemic autoimmunity led to the accumulation of cells within the skin-draining lymph nodes and away from nonskin-draining lymph nodes. Transfer of CD45.1+ cells from non-irradiated donors into CD45.2+ UV-irradiatedrecipients resulted in preferential accumulation of donor naïve T cells and a decrease in activated T cells withinskin-draining lymph nodes. A single dose of immune suppressive solar-simulated UV was all that was required tocause a redistribution of naïve and central memory T cells from peripheral blood to the skin-draining lymph nodes.
Specifically, CD69-independent increases in S1P receptor 1 (S1P1)-negative naïve and central memory T cells
occurred in these lymph nodes. Mass spectrometry analysis showed UV-mediated activation of sphingosine
kinase 1 activity, resulting in an increase in S1P levels within the lymph nodes. Topical application of asphingosine kinase inhibitor on the skin prior to UV-irradiation eliminated the UV-induced increase in S1P. Thus,
exposure to immunosuppressive UV disrupts T cell recirculation by manipulating the S1P pathway.

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OC-3.1.4
Local and systemic effects of narrowband UVB irradiation in mice
Rachael Ireland1,2, Benita Tse1, Anthony Don3, Scott Byrne1,2
1) The University of Sydney, Cellular Photoimmunology Group, Sydney, Australia; 2) The Westmead Institute for
Medical Research, Centre for Immunology and Allergy Research, Sydney, Australia; 3) The University of Sydney,
Centenary Institute, Sydney, Australia
Ultraviolet radiation (UVR) stimulates local and systemic immunoregulatory effects in a wavelength dependantmanner. Narrowband UVB (NBUVB) phototherapy is a common and effective treatment for inflammatory skindiseases including psoriasis, but its mechanism of action is not fully understood. This study aims to characterise local
and systemic changes in mice following NBUVB irradiation. In a model of contact hypersensitivity, a single exposure
to 3 J/cm2 NBUVB suppressed the response to an irritant applied at an unirradiated site. Like exposure to solar
simulated UV (ssUV), this immune suppressive dose of NBUVB resulted in Langerhans cell depletion and dermal
neutrophil infiltration. Mast cells are a key mediator of ssUV induced immune suppression, but their dermalfrequency was not altered by single or repeated exposure to NBUVB. NBUVB irradiation also generated fewerchanges in both the plasma lipidome and lymphocyte recirculation compared to ssUV. While NBUVB is a potent
local immunomodulator, these findings suggest it has relatively low capacity to induce systemic effects, which has
implications for the clinical use of NBUVB in the treatment of a wider range of diseases.

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IL-3.1.5
Oestrogen-related factors and incidence of melanoma in the U.S. Radiologic
Technologists study
Jim Z. Mai1, Rui Zhang1, Michael R. Sargen1, Mark Little1, Bruce H. Alexander3, Margaret A. Tucker1, Cari
Kitahara1, Elizabeth K. Cahoon1
1) National Cancer Institute, Division of Cancer Epidemiology and Genetics, Bethesda, United States; 2) Universityof Minnesota, School of Public Health, Minneapolis, United States
While the photosensitizing effects of oestrogens may increase the impact of ultraviolet (UV) radiation on melanoma
risk, data from large cohort studies with information on personal sun sensitivity and UV radiation exposure arelacking. Few studies have examined effect modification interaction between UVR, oestrogens-related factors andrisk of melanoma. We examined the association between reproductive factors, exogenous hormone use, and firstprimary invasive melanoma of the skin while accounting for UV radiation exposure across different life periods,
personal sun sensitivity, and lifestyle factors for geographically dispersed non-Hispanic white women exposed to awide range of ambient UV radiation, using data from the U.S. Radiologic Technologists (USRT) Study.
The USRT is a large nationwide prospective cohort of 146,022 radiologic technologist. The study population included
white female participants who completed both the second (baseline; 1994-1998; mean age at entry =46.8 years) andthird questionnaires (2003-2005), and did not report having cancer (except for keratinocyte carcinoma, KC) atbaseline. Participants (N=46,544) were followed from their age at completion of the baseline questionnaire until theearlier of first primary cancer diagnosis, including invasive melanoma of the skin, or completion of (either the thirdor fourth) questionnaire. To examine the association of reproductive factors, exogenous hormone use, and firstprimary invasive melanoma of the skin, we used Poisson regression to calculate rate ratios (RRs) and 95% likelihoodbased confidence intervals (CIs), adjusting for attained age, birth cohort (<1941, 1941-1945, 1946-1950, 1951-1955,
1956+), lifetime average annual ambient UV radiation (quartile 1-4), contraceptives and menopausal hormone
therapy use. To address the effect modification of ambient UV radiation exposure and personal sun sensitivities onmelanoma risk, we conducted likelihood-ratio tests for multiplicative interaction.
Over a median follow-up time of 17.1 years, 0.95% of eligible participants had an incident first primary melanoma(n=444). Higher melanoma risks incidence rates were observed in participants with older attained age, blue/green
eye colour, blonde/red/auburn natural hair colour at age 15, fair skin complexion, and higher UV radiation. We foundan increased risk of melanoma in women who experienced menarche at an early age (13 and ≤12 years versus 14+
years: RR=1.33, 95% CI=1.03-1.75; 1.27, 0.93-1.73; P-value for trend=0.17), and in women with older age at firstbirth (25-29 and ≥30 years versus <25 years; 1.09, 0.86-1.39; 1.48, 1.12-1.95; P-value for trend<0.001). However,no significant association was observed for other reproductive factors, and for all exogenous hormone use. The
associations of melanoma incidence for most reproductive factors and exogenous hormone use were not modified byambient UVR, eye colour, natural hair colour at age 15 and skin complexion. The exception was that natural haircolour at age 15 significantly modified the associations of melanoma for age at menarche (P-value for interaction =0.002) and age at first birth among parous women (0.028). In participants with blonde/red/auburn natural hair colour
at age 15, we found significantly increased risk of melanoma among women who experienced menarche at an earlyage (13 and ≤12 years vs 14+ years) (HR = 3.02, 95% CI = 1.74-5.80; 2.67, 1.42-5.39, respectively; P-value for trend= 0.011). The positive association between age at menarche and melanoma was null in participants with brown naturalhair colour at age 15.
Early age at menarche and late age at first birth were associated with higher risks of melanoma in this large,
geographically dispersed cohort of non-Hispanic white female radiologic technologists. These findings strengthen
the evidence that oestrogen-related factors alone were not associated with melanoma. However, women with late age
at first birth and early age at menarche, especially those with blonde/red/auburn natural hair colour at age 15, or
residing in areas with elevated levels of ambient UV radiation may constitute an additional high-risk group in needof more frequent skin cancer examinations. Future larger studies could collect more detailed information on
photosensitizing medication use and breastfeeding, and ideally, measurements of genetically predicted levels ofBook of Abstracts ������������������������������������������������������������������������������������������������ 124

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oestrogen, particularly for childhood. Identifying susceptible periods of exposure or factors that modify UVRsusceptibility may aid in guiding more targeted guidelines for melanoma prevention.
References
1) Siegel, R.L., et al., Cancer Statistics, 2021. CA: A Cancer Journal for Clinicians, 2021.
2) Bray, F., et al., Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for
36 cancers in 185 countries. CA: A Cancer Journal for Clinicians, 2018.
3) Olsen, C.M., et al., Evaluation of Sex-Specific Incidence of Melanoma. JAMA Dermatology, 2020.
4) Sun, Q., et al., Effects of Exogenous Hormones and Reproductive Factors on Female Melanoma: A Meta-Analysis.
Clinical Epidemiology, 2020.
5) Gandini, S., et al., Hormonal and reproductive factors in relation to melanoma in women: current review and metaanalysis. European Journal of Cancer, 2011.
6) Donley, G.M., et al., Reproductive factors, exogenous hormone use and incidence of melanoma among women in
the United States. British Journal of Cancer, 2019.
7) National Aeronautics Space Administration. Total ozone mapping spectrometer data product: erythemal UV
exposure, 2004.
8) Fuhrman, B.J., et al., Association of the Age at Menarche with Site-Specific Cancer Risks in Pooled Data from
Nine Cohorts. Cancer Research, 2021.
9) Olsen, C., et al., Reproductive factors, hormone use and melanoma risk: an Australian prospective cohort study.
British Journal of Dermatology, 2021.
10) Armstrong, B.K., C.M. Vajdic, and A.E. Cust, Melanoma, in Cancer Epidemiology and Prevention, M. Thun, etal., Editors. 2017.

Acknowledgements. We thank the radiologic technologists who participated in the study, Dr Jerry Reid of theAmerican Registry of Radiologic Technologists for continued support, and Diane Kampa and Allison Iwan of the
University of Minnesota for data management and collection. This research was supported by the Intramural Research
Program, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health,
U.S. Department of Health and Human Services.

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IL-3.1.6
Photoprotection by vitamin D compounds: uncovering markers that predict efficacy
in reducing photocarcinogenesis.
Furkan A. Ince1, Artur Shariev1, Mark S. Rybchyn1, W.G. Manori De Silva1, B.Y. McCarthy1, C. Yang1, Myriam
Abboud2, Rebecca S. Mason1 and Katie M. Dixon1
1) School of Medical Sciences, Faculty of Medicine and Health, The University of Sydney, Sydney NSW 2006
Australia. 2) College of Natural and Health Sciences, Zayed University, United Arab Emirates.
The key ingredients for development of ultraviolet radiation (UV)-induced skin tumours are UV-induced DNA
damage, some of which is inadequately repaired, resulting in mutations, and UV-induced immune suppression, which
results in developing skin tumours not being recognised and eliminated by immune surveillance. The vitamin D
hormone, 1,25-dihydroxyvitamin D (1,25(OH)2D3), applied immediately after UV exposure, reduces DNA damage,
immune suppression and skin tumours, as do the vitamin D-like compounds 1,25-dihydroxylumisterol andtetrahydrocurcumin1,2. Yet several of the vitamin D-like compounds and other potentially photoprotective agents wehave examined in acute and chronic UV studies have reduced UV-generated DNA damage in skin cells and in skin,and reduced UV-induced immune suppression in mice1, but have not reduced photocarcinogenesis in this well
accepted murine model. There is thus a need to find readily measurable markers which can identify agents that will
reduce skin tumours in mice exposed to chronic UV, since reductions in UV-induced DNA damage and immune
suppression alone are not necessarily predictive.
Both phosphatase and tensin homolog (PTEN) and N-myc downstream regulated gene-1 (NDRG1) are protein
markers which are lost or suppressed during carcinogenesis and metastasis. We demonstrated that expression of boththese proteins is significantly reduced 24 h after UV in primary melanocytes and in Skh:hr1 mouse skin, but
significantly increased with 1,25(OH)2D3 treatment. Phosphorylation of Cyclic AMP response element binding
protein (CREB) is known to increase in skin cells after UV and is linked to carcinogenic potential3. Our preliminary
data indicates that 1,25(OH)2D3 also reduces pCREB after UV. Interleukin-6 (IL-6) is a marker of inflammation,
increased after UV. IL-10 also increases following UV and contributes to immune suppression. We havedemonstrated reductions in both of these cytokines with 1,25(OH)2D3. Using vitamin D and vitamin D-likecompounds previously tested for their effects on DNA damage, immune suppression and photocarcinogenesis in bothhuman in vitro and murine in vivo models, we performed further studies in primary human skin cells, Skh:hr1 miceand ex vivo human skin, with the aim of developing a matrix to determine which of these markers or combination ofmarkers best predict ability of the agent to reduce UV-induced skin tumours.
Testing for efficacy against UV-induced skin tumours, while essential to the development of biological enhancers tosimple filtration of UV by sunscreens, is a long (40 week protocol), tedious and very expensive process. If there were
early markers for likely efficacy and to identify those agents which were unlikely to reduce tumours, this would
considerably streamline the process for identifying suitable candidates for this lengthy testing.
References
1) Dixon, K.M., et al., 1alpha,25(OH)2-Vitamin D and a Nongenomic Vitamin D Analogue Inhibit Ultraviolet
Radiation-Induced Skin Carcinogenesis. Cancer Prevention Research, 2011. 4(9): p. 1485-94.
2) Tongkao-On, W., Role of Vitamin D and Other Compounds in the Protection of Skin Cells from UV, in
Physiology, School of Medical Sciences, University of Sydney, 2015 [PhD Thesis].
3) Rybchyn, M.S., et al., Enhanced Repair of UV-Induced DNA Damage by 1,25-Dihydroxyvitamin D3 in Skin
Is Linked to Pathways that Control Cellular Energy. J Invest Dermatol, 2018. 138(5): p. 1146-1156.

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IL-3.1.7
UV-induced growth and migration of melanoma cells in a spheroid model.
Linda Vainikka, Petra Wäster, Linda Rößler and Karin Öllinger1
1) Linköping University, Department of biomedicine and Clinical Sciences, Linköping, Sweden

Cutaneous melanoma is a highly aggressive cancer with a propensity for metastasis to various organs.
The most important risk factor to develop the disease is exposure to UV radiation. UVB wavelengths has
ability to cause tumor-initiating mutations while UVA exposure generates free radicals and may act
immunosuppressive. How the microenvironmental effects of UV radiation influence melanoma
pathogenesis and its ability to metastasize remains to be elucidated. In a previous report we have shownthat UVA exposure triggers increased secretion of extracellular vesicles. UVA cause plasma membrane
damage which is promptly repaired by transportation of lysosomes to the damaged membrane to form a
resealing patch. Subsequently, extracellular vesicles are released outside the cell (see Figure 1). The
vesicles promote signaling event when taken up by non-irradiated cells and increased growth andmigration is noted1.
Figure 1. UVA-radiation induces plasma
membrane damage resulting in calcium influx,
which trigger transport of lysosomes to thedamaged site. The lysosomal membranes forms
a resealing patch and lysosomal content isreleased outside the cell. Subsequently, excess
membrane is shredded as extracellular vesicles.

In this study results from UV exposure of malignant melanoma cells will be presented and the possible
signaling pathways induce by UV will be discussed. We generated melanoma tumor spheroids from four
different cell lines and exposed to sub-lethal doses of UVA- and UVB-radiation. We found spontaneous
secretion of extracellular vesicles by all melanoma cell lines, but after UVA, increased number of large
size vesicles were found. Following radiation, the spheroids were embedded in Matrigel and observed
microscopically to follow cell migration and invasion. Cell migration out from the spheroid was higher in
spheroids exposed to UVA compared to control and UVB exposed samples. A transcriptome analysis was
performed 24 h after irradiation of the spheroids, and revealed significantly increased mRNA levels of
matrix metalloprotease-1 (MMP1; +3.3-fold, p=0.0009) after UVA exposure. MMP-1 is a collagenase
involved in extracellular matrix degradation and the protease is secreted and activated outside the cell.
We found elevated protein level of MMP1 and secretion to the medium in three out of four tested
melanoma spheroids after UVA exposure and in one of four after UVB. We conclude that melanoma
migration and invasion is elevated more efficiently after UVA than UVB radiation and MMP1 is acandidate protein to promote this effect.
References
1. Wäster P, Eriksson I, Öllinger K. Intercellular cross-talk mediated by extracellular vesicles promotes survival andproliferation via miR-21-signaling. Pigment Cell and Melanoma Research 2020; 33(4):542
Acknowledgements. This work was supported by grant from Swedish Cancer Society and Welander Finsten Skin
Foundation.

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OC-3.1.8

Characterization of Photo damaged skin using 3D Line-field optical coherence
tomography and histopathological correlation
J Pérez-Anker1, B Alejo1, P Iglesias2, V. Berot3, C Cano4, E Cinotti5, V Del Marmol4, M Fontaine4, M Miyamoto4, J
Monnier6, JL Perrot3, P Rubegni5, L Tognetti5, M Suppa4,7, Ann’Laure Demessant - Flavigny8, Caroline Le Floc’h8,
Leonor Prieto9, J Malvehy1,2, S Puig1,2
1. Dermatology Department, Melanoma Unit, Hospital Clínic de Barcelona, IDIBAPS, Universitat de Barcelona,
Barcelona, Spain, 2. Centro de Investigación Biomédica en Red de Enfermedades Raras, CIBERER, nstituto de Salud
Carlos III, Barcelona, Spain, 3. Dermatology Department, University Hospital of Saint-Etienne, Saint-Etienne,
France, 4. Dermatology Department, Hôpital Erasme, Université Libre de Bruxelles, Brussels, Belgium, 5.
Dermatology Unit, Department of Medical, Surgical and Neurological Sciences, University of Siena, Siena, Italy, 6.
Dermatology Department, AP-HM, Aix-Marseille University, Marseille, France, 7. Dermatology Department,
Institut Jules Bordet, Universite Libre de Bruxelles, Brussels, Belgium, 8 International Scientific Direction,
Laboratoire Dermatologique La Roche-Posay, France, 9. Scientific Direction, Laboratoire Dermatologique La
Roche-Posay, L’ORÉAL Cosmética Activa, Madrid, Spain
Chronic sun exposure is responsible of skin photoaging. Noninvasive imaging techniques are able tomorphologically characterize photoaging and generate qualitative analysis in photodamaged skin. Optical coherence
tomography (OCT) is one of the non-invasive available technologies that allows a deep laser penetration in thesurface, reaching a good quality image of dermis visualization. It creates in vivo cross-sectional (vertical) and en face(horizontal) images of the skin with an isotropic resolution of 1.3 µm and a penetration depth of 500 µm. A new
three-dimensional technology, 3D lineal confocal OCT (LC-OCT) (DAMAE Medical, Paris) has been introduced,
allowing a vertical and horizontal reconstruction of the complete skin with cellular resolution.
Our objectives were to observe in an in vivo three-dimensional mode, the morphological and cytological changes ofphotodamaged skin, in order to better understand the changes of the different layers of the skin, induced by chronic
sun exposure and responsible for chronic sun-damage.
In this study, we conducted a descriptive and observational investigation, after the approval of the Ethics Committeeat Hospital Clínic de Barcelona, in order to determine the 3D LC-OCT characteristics of sun damaged skin. In June
and July of 2020, nine phototype II-III patients in their fourth to sixth decade of life were prospectively included.
Dermoscopy and LC-OCT was performed on two different zones: the sun-damaged area on dorsal forearm (L01) andthe non-sun-damaged area on ventral side of the same forearm (L03). Skin biopsies were obtained of both observed
areas.
Quantification of the structural morphology of the skin`s upper layers was obtained: The thickening of the Stratum
Corneum (SC), of the stratum spinosum, and the flattening of the Dermal Epidermal Junction (DEJ) were evaluated.
The number and density of keratinocyte nuclei were also quantified. All metrics were compared as paired data on 9
patients. No statistically significant differences were observed for the SC thickness and the DEJ undulation. By image
analysis, we showed that the epidermis thickness was increased (mean: 5.39 µm, p= 0.0078) and that the number ofkeratinocytes was higher (mean=0.997; p= 0.002) in exposed area (L01) in comparison with non-exposed one (L03).
Under LC OCT analysis, the volume of the different layers of keratinocytes seemed to be larger for the more sunexposed area (mean 0-20%: 4.61 µm3, mean 40-60%: 26.22 µm3, mean 80-100%: 55.34 µm3).
Concerning the histology comparison between both areas as paired data, we showed an increased thickness of theepidermis (mean:11.07µm, p=0.011) on exposed areas. Significant differences were also observed for the number ofkeratinocytes layers that was higher for the chronic photodamaged skin (mean:0.77, p=0.029). Those results are wellcorrelated with LC-OCT one’s.
We present for the first time a study of photodamaged skin in 3D, with quantitative and objective measures comparing
normal and chronic photodamaged skin. We demonstrated that the epidermis was significantly thicker on the exposedarea and that the number of keratinocytes layers was higher in the same area with good correlation withhistopathological evaluation.

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IL-3.2.1

NIR biphotonic chromophores in the service of biology
Chantal Andraud1
1) Laboratory of Chemistry, Ecole Normale Supérieure de Lyon, CNRS, Lyon, FranceThis presentation will illustrate our different recent approaches in order to fulfill main requirements for in vivo
biophotonics. We will present our results based on : (1) molecular engineering approaches for enhancing organic
based chromophores biphotonic properties and further spectrocopic requirements in the NIR1 ; (2) methods ofhydrosolubilisation and biocompatibility for these biphotonic chromophores2 . Intravital fluorescence imaging willbe then discussed as a function of chromophores characteristics (Fig. 1) and perspectives for theranostic or bimodal
probes will be presented3.

Fig. 1 – Example of fluorophore with a 3D image of the functional cerebral vasculature in the motor cortex of mice using twophoton laser scanning microscopy.

References
1) C. Monnereau, ,C. Andraud et al. A water soluble probe with near infra-red two-photon absorption and polarityinduced fluorescence for cerebral vascular imaging Chem. Sci., 2013; C. Monnereau, C. Andraud et al. Fluorescent
Polymer Probe with High Selectivity toward Vascular Endothelial Cells for and beyond Noninvasive Two-Photon
Intravital Imaging of Brain Vasculature, ACS Appl. Mater. Interfaces 2016, 8, 17047 ; C. Andraud, O. Maury et al. ,Keto-polymethines: a versatile class of dyes with outstanding spectroscopic properties for in cellulo and in vivo twophoton microscopy imaging, Chem. Sci. 2017, 8, 381; Y. Bretonnière, C. Andraud et al. Near-Infrared Solid-State
Emitters Based on Isophorone: Synthesis, Crystal Structure and Spectroscopic Properties Chem. Mat. 2011, 23, 862;
Y. Bretonnière, C. Andraud et al. , Solid state red biphotonic excited emission from small dipolar fluorophores,, J.
Mater. Chem. C, 2016,4, 766 ; C. Andraud, Y. Bretonnière et al. , Tuning the solid-state emission of small push-pull
dipolar dyes to the far-red through variation of the electron-acceptor group, Dyes and Pigments 2018, 156, 116 ; C.
Andraud, Y. Bretonnière et al. , 4,5,5-Trimethyl-2,5-dihydrofuran-Based Electron-Withdrawing Groups for NIREmitting Push–Pull Dipolar Fluorophores, J. Org. Chem. 2019, 84, 9965
2) C. Andraud, Y. Bretonnière et al., Photostable far-red emitting pluronic silicate nanoparticles: perfect blood pool
fluorophores for biphotonic in-vivo imaging of the leaky tumour vasculature, RSC Advanced 2016, 6, 94200; C.
Andraud, Y. Bretonniere, J. Bernard et al General and Scalable Approach to Bright, Stable and Functional AIE
Fluorogen Colloidal Nanocrystals for in Vivo Imaging ACS Appl. Mater. Interfaces 2018, 10, 25154 ; Y.
Bretonnière, C. Andraud, X. Cattoën et al., Ultrabright Silica-Coated Organic Nanocrystals for Two-Photon InVivo
Imaging Ultrabright Silica-Coated Organic Nanocrystals for Two-Photon InVivo Imaging ACS Appl.Nano Mater.
2020, 3, 11933
3) C. Andraud, C. Monnereau, O. Maury et al., A “Multi-Heavy-Atom” Approach Toward Biphotonic
Photosensitizers With Improved Singlet Oxygen Generation Properties Chemistry - A European J. 2019, 25, 9026; Y.
Bretonnière, C. Andraud, S. Parola et al., Hybrid multimodal contrast agent for multiscale in vivo investigation ofneuroinflammation Nanoscale, 2021,13, 3767
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-3.2.3

Monitoring of Wound Healing by Using Label-free Multiphoton Microscopy and the3D Printed Live-cell Imaging Chamber
Haijiang Zhang1#, Che-Wei Chang1#, Rafael Camacho1, Julia Fernandez-Rodriguez1*
1) Centre of Cellular Imaging, Core Facilities, the Sahlgrenska Academy, University of Gothenburg, Sweden#Co-first authors*Correspondence should be addressed to Julia Fernandez-Rodriguez; email address:
julia.fernandez-rodriguez@cci.sahlgrenska.gu.se
Current skin research typically relies on classical histopathological examination, either by studying the abnormalityof the surface or vertical sections across the tissue. The drawbacks of this type of end-point experiment for studying
wound healing are its destructive nature and that the outcome can only be rationalized retrospectively. To reliably
observe the skin biology directly, the optimal experiment would require monitoring live tissue in real time with animaging system capable of recording cellular responses a few hundred μm under the skin surface. One-photon
microscopy, although capable of achieving sub-micron resolution, generally can only image the surface of the skintissue, due to the nature of the one-photon absorption/emission, and more importantly, the imaging limitations
caused by the tightly packed keratinocyte cells. This study demonstrates that multiphoton laser scanningmicroscopy is a good imaging technique for monitoring wound healing in ex vivo skin models, while using a 3D
printed microfluidics chamber for keeping the tissue viable. Multiphoton microscopy is capable of deep tissue
imaging down to ~300 μm and is able to monitor the cellular proliferation and collagen regeneration without
adding external fluorescent markers, which could potentially interfere with native cellular functions. Further, the3D printed microfluidics chamber we designed can be integrated into commercial microscopy incubation systems
to create an artificial growth minienvironment for keeping tissue alive for prolonged periods of time. In this study,
real-time monitoring of wound healing was achieved at the depth of ~300 μm during 3-7 days.

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OC-3.2.4

A
novel
CathepsinBdegradable
nanoparticle
intraoperative NIR imaging and treatment of pancreatic cancer

platformfor

Fabiola Sciscione1, Shiv Patel1, Hamzah Masood1, Marym Mohammed Hadi1, Bala S. Ramesh1, John F. Callan2,
Alexander MacRobert1, Anthony McHale2 and Nikolitsa Nomikou1*1) University College London, Division of Surgery & Interventional Science, , Royal Free Hospital, London, UK;
2) Ulster University, Biomedical Sciences Research Institute, Coleraine, UK

Pancreatic cancer is ranked amongst the most lethal forms of cancer. Currently, surgical resection is the only practical
approach for patients with locally advanced disease. Extension of the limits of tumour resectability can be achieved
with image-guided surgery, which can help accurately delineate the borders of the tumour and facilitate surgicalresection. This, in combination with near-infrared (NIR) laser treatment of the resection site immediately after tumour
removal, can help destroy residual microscopic tumour fragments and prevent their progression after surgery. In thisregard, we have developed a novel multi stimulus-responsive nanoparticle formulation containing indocyanine green(ICG), a clinically approved NIR fluorescent agent with photothermal properties. Previously, poly-l-glutamic acid
(PGA) was used as the carrier polymer.1 In this work PGA was modified by partial conjugation of the carboxylate
side groups with a lipid molecule, which resulted in the formation of nanoparticles with increased ICG-loading yield.
The new nanoparticles proved to be digested by the proteolytic enzyme Cathepsin B (CB) in phosphate buffer at pH
6.4 and 7.4 and release ICG, as confirmed by fluorescence spectroscopy studies. CB is overexpressed in advanced
pancreatic cancer and its production and activity are particularly enhanced at the tumour acidic pH. To demonstrate
the feasibility of the novel formulation for efficient NIR-guided imaging/delineation of highly aggressive tumours
with elevated proteolytic enzyme activity, we developed a three-dimensional (3D) cell culture system in which thepancreatic BxPC-3 cells were embedded in the extracellular matrix-resembling 3D structures. These systems wereincubated with the nanoparticle formulation at either pH 7.4 or 6.4, in the absence or in the presence of CB. An NIR
excitation diode and a high sensitivity CMOS camera were used to generate an NIR image of the 3D cell systems.
An increase of ICG fluorescence signal was observed after 24 hours in CB-rich systems, with the highest intensity at
pH 6.4. Following these positive preliminary results, we are currently carrying out 2D and 3D in-vitro NIR laser
treatments to establish the potential and efficacy of the formulation in eliminating residual microscopic disease
following combined NIR-imaging and treatment.

References
1) S. Tarassoli, S. Martinez de Pinillos, H. Pye, C.A. Mosse; J.F. Callan, S. Macrobert, A.P. McHale; N. Nomikou,
Cathepsin B-degradable, NIR-responsive nanoparticulate platform for target-specific cancer therapy.
Nanotechnology 2017, 28(5),055101.
Acknowledgments. The work is supported by the Engineering and Physical Sciences Research Council UK . EPSRCDTP (Ref: EP/N509577/1) and EPSRC Standard Grant (Ref: EP/R035008/1).

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IL-3.2.5

Imaging of stress- and damage- induced metabolic adapations in skin
Florian Gruber1,2,3, Arvand Haschemi4, Anne Miller4, Marie-Sophie Narzt1,3, Christopher Kremslehner1,2,3
1) Medical University of Vienna, Department of Dermatology,Vienna, Austria; 2) Christian Doppler Laboratory
for Skin Multimodal Analytical Imaging of Aging and Senescence – SKINMAGINE- and 3) - Christian Doppler
Laboratory for Biotechnology of Skin Aging, Vienna, Austria; 4) Medical University of Vienna, Department ofLaboratory Medicine, Vienna, Austria
In the differentiation of epidermal keratinocytes their function, chemical composition, physical properties,
metabolism and secretion profoundly change. Extrinsic stressor, like ultraviolet (UV) radiation and intrinsic factorsthat are associated with aging thus may differently affect the epidermal keratinocytes, depending on differentiation
stage. Exposure to UV elicits various responses including the DNA damage response and activation of pathways
which detoxify or repair damage or induce cell death when the damage was irreparable. Recently, rapid diversionof glucose flux into the pentose phosphate pathway (PPP) was discovered as additional mechanism by which cells
instantly generate reduction equivalents and precursors for nucleotides – both being in demand after UV damage.
There is however little known about the correlation of such metabolic activity with differentiation state, cell
damage and tissue localization of epidermal cells. We developed a method to correlate the activity of G6PD, thefirst and rate-limiting enzyme of this metabolic UV response, at cellular resolution to cell type, differentiation state,
and cell damage in human skin and in organotypic reconstructed epidermis. We thereby could verify rapid
activation of G6PD as an immediate UVB response not only in basal but also in differentiating epidermal
keratinocytes and found increased activity in cells which initiated DNA damage responses. We also found that theanti-diabetic and senomorphic drug metformin strongly induced G6PD activity throughout reconstructed epidermis.
This is of relevance as cellular senescence of skin cells affects their metabolic profile. Activation of the protective
pentose phosphate pathway may be useful to modulate the skin's antioxidant defense systems and DNA damage
repair capacity on demand.
References
1) Kremslehner, et al. “Imaging of metabolic activity adaptations to UV stress, drugs and differentiation at cellular
resolution in skin and skin equivalents – Implications for oxidative UV damage.”, Redox Biology, 2020
Acknowledgements. The work of the CDLs is funded by the Federal Ministry for Digital and Economic Affairs(BMWFW) of Austria and the National Foundation for Research, Technology, and Development of Austria and by
CHANEL Parfums et Beauté and the Herzfelder'sche Familienstiftung, Austria

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IL-3.2.6

Imaging Mass Spectrometry – Benefits, Challenges, Potentials
Martina Marchetti-Deschmann
TU Wien, Institute of Chemical Technologies and Analytics, Vienna, Austria
A single modality is very often not good enough to understand all functional, structural, temporal and chemical
relations underlying certain biological conditions. Imaging a specimen with two or more complementary modalities
creates an informative, composite view of a sample that spans all relevant resolution ranges.
Here we shortly introduce the concept of MALDI Mass Spectrometry based imaging (MALDI-MSI) and its
potential to localize molecules in a tissue without a priori knowledge. Briefly the advantages, limitations andpotentials for future applications will be highlighted. The successful combination of different mass spectrometricapproaches will be shown but also the combination of MSI with vibrational spectroscopy and μXRF.
Acknowledgements. Parts of the present work were supported by COMULIS (COST CA 17121), SKINMAGINE(Christian Doppler Society) or MEIBio (TUWien).

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IL-3.2.7
Hyperspectral imaging extended to circularly polarized light
Guillaume Raffy1, André Del Guerzo1, Dario M. Bassani1
1) Institut des Sciences Moléculaires, CNRS UMR5211 and University of Bordeaux, Talence, France
Even when using light from a perfectly unpolarized source, reflections and interactions with absorbing media can
induce polarization. Furthermore, because electronic transitions in molecules are aligned with the electronictransition dipole moments, both absorption and emission of light occurs preferentially along specific molecularaxes that may differ between absorption and emission processes. This has been recognized and exploited innumerous experimental setups designed to probe the relative orientation of chromophores, their motion orinteraction in solution. In solid matter microscopy, linear polarization allows probing of the local order and the lossof excitation-induced polarization can provide information on exciton diffusion in solids and of solid-state
photoinduced reactions.1 Compared to the use of linear polarization, circular polarization has received much lessattention. It is, however, becoming increasingly recognized in importance since becoming the focus of newmaterials for CPL in material science.2 In Nature, some examples of CPL active materials exist, although it is not
yet clear whether this property plays a role in the evolution of the species. In the study of CPL-active new materialsof biological or synthetic origin, resolution at the sub-micron scale can provide information on the heterogeneity
which in turn can be used to understand function and optimization. However, collecting circular polarization
represents a formidable challenge due to the inherently small ∆I that need to be sensed. Indeed, glum values formany organic chromophores are below 10–3, which renders detection challenging. Furthermore, the numerous
optical components in the detection setup need to be carefully considered to minimize losses and scrambling of thepolarization components in a complex way of the sample's luminescence.3 Our recent progress towards attaining
sub-micron resolution with CPL detection will be presented.

Wide-field CPL microscope image of two enantiomeric Eu3+ complexes exhibiting opposite CPL. Polarplot of blue and red regions are shown
below

References
1) Guillaume Raffy, Debdas Ray, Cheng-Che Chu, André Del Guerzo, Dario M. Bassani"Controlling the emission polarization from single crystals using light: Towards photopolic materials"
Angew. Chem. Int. Ed. 50, 9584 – 9588, 2011.
2), Tonghan Zhao, Jianlei Han, Pengfei Duan, Minghua Liu, New Perspectives to Trigger and Modulate Circularly
Polarized Luminescence of Complex and Aggregated Systems: Energy Transfer, Photon Upconversion, Charge
Transfer, and Organic Radical, Acc. Chem. Res., 53, 1279-1292, 2020.
3) P. Torok, P. D. Higdon, T.Wilson, On the general properties of polarised light conventional and confocal
microscopes, Optics Commun., 148, 300-315, 1998.

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OC-3.2.8

Analysis of UV-exposed skin sections via MALDI Imaging Mass Spectrometry
Samuele Zoratto1,2, Michaela Sochorova2,3, Christopher Kremslehner2,3, Markus Schosserer2,4, Florian Gruber2,3,5,
Martina Marchetti-Deschmann1,2,5,6
1) Institute of Chemical Technologies and Analytics, TU Wien, Vienna, Austria; 2) Christian Doppler Laboratory
for Skin Multimodal Imaging of Aging and Senescence, Vienna, Austria; 3) Department of Dermatology, Medical
University of Vienna, Vienna, Austria; 4) University of Natural Resources and Life Sciences, Vienna, Austria;
5) Vienna Center for Engineering in Medicine; 6) Austrian Cluster for Tissue Regeneration, Vienna, Austria.
Our skin is constantly exposed to solar radiation, high oxygen levels, and environmental pollutants. These areaccelerant stress factors for premature skin aging, tissue inflammation, and photocarcinogenesis. In skin aging, theevident and well-known clinical effect is the alteration of the skin appearance. In addition, metabolism and cellcommunication are impaired, and an increase of senescent cells is evident. Cellular senescence is a phenomenon
characterized by the arrest of cell division and is commonly linked to the aging phenotype. Nevertheless, it plays anessential role in tissue healing and tumor suppression. Although the shortening of the telomeres is usually the leading
promoter of senescence; also oxidative stress or oncogenes activation can trigger this cellular response as sources of
DNA damage. To counteract the oxidative stress produced by the stress-inducing agents, the cells activate several
epidermal and dermal lipoxygenases. Indeed, oxidized lipids can act as danger-associated molecular patterns(DAMPs) messengers and are involved in the senescence-associated secretory phenotype (SASP).1
In previous work from Gruber et al2, several oxidized phosphatidylcholines (OxPCs) were characterized in asemitargeted lipidomic approach. The method involved the exposure of human dermal fibroblasts at defined UVA
fluences, followed by liquid-liquid extraction, and lastly, mass spectrometric analysis by means of reversed-phase
HPLC coupled to electrospray ionization MS/MS (HPLC-ESI-MS/MS).
In this work, artificially aged skin samples were provided by the Medical University of Vienna (MUW). Skin
specimens were irradiated in specific areas with either UVA/B radiation at different fluencies before embedding andcryo-sectioning in the same fashion as the aforementioned publication. Our study aims to evaluate the epilipidomic
effects via an untargeted multimodal approach focused on OxPCs species by high-resolution mass spectrometry
analysis. Indeed, we employed high-resolution matrix-assisted laser desorption/ionization Fourier-transform ion
cyclotron resonance mass spectrometry (MALDI-FT-ICR MS) to generate MS images (MSI) of skin section after
autofluorescence light-microscopy analysis. The high mass accuracy of FT-ICR MS instrumentation, combined withthe analytes’ spatial localization and relative abundance, allow us to locate and putatively identify several OxPCs
with great confidence. Lastly, the integration with high-definition images from light-microscopy yields a detailed
and comprehensive result of the effects of UV-induced alteration.
In conclusion, the untargeted MS-based methodology allows us to expand our analysis to known and unknown
analytes. The multimodal approach unifies the results from different techniques and helps us to discover the crucial
factors of scientific importance.
The authors declare no conflict of interest.
References:
1) Chunya Ni et al, Autophagy deficient melanocytes display a senescence associated secretory phenotype that
includes oxidized lipid mediators. Int J Biochem Cell Biol. 81(Pt B):375-382, 2016.
2) Gruber et al, A simplified procedure for semi-targeted lipidomic analysis of oxidized phosphatidylcholines induced
by UVA irradiation. J Lipid Res. 53(6): 1232–1242, 2012
Acknowledgments:
We acknowledge financial support of the Federal Ministry Republic of Austria for Education, Science and Research
and TU Wien (HRSM 2016), the Christian Doppler Research Association and CHANEL Parfums et Beauté forfunding SKINMAGINE. This work is supported by COMULIS (CA17121) and EpiLipiNet (CA 19105).

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P-3.2.9

Application of UVA irradiation for the determination of the redox status in excised
skin by EPR spectroscopy
Lohan SB1, Ivanov D1, Schüler N2, Berger B2, Lademann J1, Meinke MC1
1) Charité – Universitätsmedizin Berlin, Corporate member of Freie Universität Berlin, Humboldt-Universität zu
Berlin, and Berlin Institute of Health, Department of Dermatology, Venerology and Allergology, Berlin, Germany;
2) Freiberg Instruments GmbH, Freiberg, Germany
Free radicals in the body are essential for the metabolism, signal transport and are part of our immune response.
Under physiological conditions, a dynamic equilibrium between the process of radical production and detoxification
is given. But when the stress level is too high, oxidative stress occurs, and an imbalance between the body's
antioxidant defense system and reactive metabolites is existing. If oxidative stress occurs, first reactive oxygen
species (ROS) are formed which can be well controlled by the antioxidant system. With increasing stress, C-centered(CCR) or alkoxy radicals (lipid oxygen species, LOS) are produced, which can be harmful to metabolic processes.
Concerning the skin physiology, they are significantly involved in the development of inflammatory skin diseases,
immunosuppression, skin cancer, and premature skin aging. The partitioning of the different types of radicals could
give information about the stress level.
With a share of 46% a large amount of radicals is generated by sunlight irradiation in the UVA region with amaximum at ~ 360nm1,2.
In this study, radical production in excised porcine skin was characterized and quantified by electron paramagnetic
resonance (EPR) spectroscopy using UVA irradiation as stress source. The aim was to determine the dose at which
the switch between eustress and distress occurs.
The study revealed that during UVA radiation the ratio of ROS and LOS turned with increasing dose. The region ofturn appears at half of the minimal erythema dose of UVA. Important is a sufficient irradiance of the UVA light,
because at low irradiance adaptation processes inhibit the revers effect. This method could detect heat induced stressin the skin of pre-stressed pigs in the summer time due to temperatures above 35°C during husbandry or transport.
In these ears, the ratio was already reversed, more LOS were produced right from the start of irradiation. Thisillustrates that the ratio of ROS and LOS produced by irradiation in skin could be used as a stress marker3.
References

1) L. Zastrow, N. Groth, F. Klein, D. Kockott, J. Lademann, R. Renneberg, L. Ferrero, The missing link-light-induced (280-1,600 nm) free radical formation in human skin, Skin Pharmacol Physiol, 22, 31-44(2009).
2) S.B. Lohan, R. Muller, S. Albrecht, K. Mink, K. Tscherch, F. Ismaeel, J. Lademann, S. Rohn, M.C.
Meinke, Free radicals induced by sunlight in different spectral regions - in vivo versus ex vivo study,
Exp Dermatol, 25, 380-385 (2016).
3) S.B. Lohan, D. Ivanov, N. Schuler, B. Berger, L. Zastrow, J. Lademann, M.C. Meinke, Switching fromhealthy to unhealthy oxidative stress - does the radical type can be used as an indicator?, Free Radical
Bio Med, 162; 401-411 (2021).
Acknowledgements. The work is funded by the German Federal Ministry of Education and Research BMBF (FKZ
03ZZ0140A) within the program “Zwanzig20 – Partnerschaft für Innovation” and carried out within the consortium
Advanced UV for Life.

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P-3.2.10

Using infrared beam to track the cell fate of transgenic flatworms.
Jakub Wudarski1, Yasuhiro Kamei2, Kiyokazu Agata 1
1) National Institute for Basic Biology, Laboratory of Biological Diversity, Okazaki, Japan;
2) National Institute for Basic Biology, Spectrography and Bioimaging Facility, Okazaki, Japan
Flatworms are considered champions of regeneration.
The key to this process lies in a population of adultstem cells called neoblasts. So far live imaging of thesestem cells was impossible because of the absence oftransgenic flatworms. I have developed a protocol toestablish transgenic lines of the flatworm Macrostomum
lignano1 and used heat shock promoter to drive theexpression of fluorescent proteins2. Now, together with
Dr. Yasuhiro Kamei, we are combining our hsp linewith the infrared laser evoked gene operator (IRLEGO)3 technology to track the cell development inlive flatworms (Fig. 1). Using this approach, we will
also be able to selectively overexpress genes of choicein any part of the worm and track the results.
In my presentation I will show our progress in adapting
the IR-LEGO method to the transgenic flatworm
system of M. lignano.

Fig. 1 – Cells expressing mScarlet under Hsp20
promoter 24h after activation using IR-LEGO.

References
1.
2.
3.

Wudarski, J. et al. Efficient transgenesis and annotated genome sequence of the regenerative flatworm model Macrostomum lignano.
Nat. Commun. 8, 2120 (2017).
Wudarski, J., Ustyantsev, K., Glazenburg, L. & Berezikov, E. Influence of temperature on development , reproduction andregeneration in the flatworm model organism , Macrostomum lignano. Zool. Lett. 1–10 (2019) doi:10.1186/s40851-019-0122-6.
Kamei, Y. et al. Infrared laser-mediated gene induction in targeted single cells in vivo. Nat. Methods 6, 79–81 (2009).

Acknowledgements. This work is supported by grant 16H06376 from the JSPS.

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P-3.2.11

C. aggregans LOV domain: a framework for engineering of fluorescent reporters
Ivan Gushchin1, Anna Yudenko1, Anastasia Smolentseva1, Vera V. Nazarenko1, Alina Remeeva1, Oleg Semenov1
1) Moscow Institute of Physics and Technology, Research Center for Molecular Mechanisms of Aging and AgeRelated Diseases, Dolgoprudny, Russia
LOV domains, ubiquitous photoreceptors of plants, fungi, bacteria and archaea, have found recently a number ofapplications in molecular biology. They have been used as genetically encoded fluorescent reporters, generators ofreactive oxygen species, and optogenetic tools. Yet, there are still many possibilities for improvement such as
generation of colour-shifted variants or engineering of LOV-based sensors of molecular signals.
Recently, we developed a flavin-based fluorescent protein CagFbFP based on the LOV domain of a hybrid
histidine kinase from Chloroflexus aggregans1. The protein can be used for imaging both in bacteria andeukaryotes (Fig. 1). High thermal stability of CagFbFP and its amenability to high-resolution structural studies
allowed us to engineer a number of variants for different applications.
First, we tested whether CagFbFP fluorescence excitation and emission maxima can be shifted using mutations of aconserved glutamine amino acid adjacent to the flavin chromophore. Effects of the mutations were modest, andnone of the variants exhibited a red-shift2. Crystallographic structures revealed the reason for this: potentiallycharged amino acids didn’t interact with flavin, and in some cases the active site was unlatched and the side chainswere oriented away from the chromophore2. Consequently, based on the data for iLOV, we generated a new variant
where the positively charged lysine side chain was stabilized near the flavin by a second mutation; this variant
could be used for two-color imaging of bacterial cells3.
We also used structural information to engineer split-CagFbFP pairs that produced fluorescence while being fused
to interacting proteins of interest, but were not photoactive otherwise4. Moreover, as a proof of concept, we wereable to engineer a calcium-responsive pair inspired by the calcium sensor FGCaMP74. Thus, our data show that
CagFbFP is a promising framework for engineering of fluorescent reporters for different applications.

Fig. 1 – Fluorescence microscopy images of CagFbFP expressed in E. coli (left), mitochondria (center) and lysosomes (right)of mammalian cells.

References
1) Nazarenko et al., A thermostable flavin-based fluorescent protein from Chloroflexus aggregans: a framework forultra-high resolution structural studies. Photochemical & Photobiological Sciences, 2019.
2) Remeeva et al., Insights into the mechanisms of light-oxygen-voltage domain color tuning from a set of highresolution X-ray structures. Proteins: Structure, Function, and Bioinformatics, 2021.
3) Röllen et al., The molecular basis of spectral tuning in blue- and red-shifted flavin-binding fluorescent proteins.
Journal of Biological Chemistry, 2021.
4) Yudenko et al., Rational Design of a Split Flavin-Based Fluorescent Reporter. ACS Synthetic Biology, 2020.
Acknowledgements. The work is supported by the Russian Foundation for Basic Research, grant 20-34-70109.

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P-3.2.12
Engineering cellulose acetate fabrics loaded with photosensitizer and graphene
oxide endowed with bactericidal activity
Ilse G. J. Manet1, Rania Morsi1, Barbara Ventura1 , Denis Gentili2 , Franco Corticelli3, Francesca Bonvicini41) ISOF, National Research Council, Via Gobetti 101, 40129 Bologna, Italy;2) ISMN, National Research Council, Via Gobetti 101, 40129 Bologna, Italy;3) IMM, National Research Council, Via Gobetti 101, 40129 Bologna, Italy;4) Dpt. of Pharmacy and Biotechnology, Univ. of Bologna, Via Massarenti 9, 40138 Bologna, Italy

Composite materials based on cellulose acetate matrices and porphyrinic photosensitizer (PS) dyes wereprepared, characterized and tested as devices with antibacterial activity. Scope of the work is to use light toenable phototoxic bactericidal activity mediated by the production of reactive oxygen species, triggeringoxidative damage to pathogens. The materials were fabricated as porous membranes by phase inversion
process. Photosensitizers were loaded into the cellulose acetate fabrics as pre-loading, dissolving the dye into
the polymer solution before membrane preparation. Different PS loading amounts were explored. We further
prepared the fabrics also in the presence of a surfactant or graphene oxide (GO), the latter added to impartmechanical robustness to the final material.
We will present the results of the morphological study of the prepared fabrics investigated by scanning electron
microscopy and atomic force microscopy. We will further discuss the detailed photophysical characterizationof the PS on the fabrics with and without GO performed for the composite materials by means of bulk andspatially resolved techniques. In particular absorption and time-resolved fluorescence data were collected
evidencing high PS concentrations caused fluorescence lifetime shortening. Similar effects were observed in
the presence of the surfactant. Confocal time-resolved fluorescence lifetime imaging (Figure 1) of the PSloaded fabrics confirmed aggregation phenomena at the basis of the short lifetimes. The most promising fabrics
obtained loading the PS in different amounts were used to study the bactericidal effect of the fabrics on S.
Aureus bacterial population illuminated with red light.
FittingofthePS
fluorescence
decay
converged
withtwolifetimes of 2,2 ns and 8,6
ns. The image has beenobtained plotting the pixel
intenisty of the two lifetimes(red for the long lifetime
component and green for the
short lifetime component).

Fig. 1 – Fluorescence lifetime image of the cellulose acetate matrix and 0,2 % porphyrinic photosensitizer

Acknowledgements. This work was supported by H2020 MSCA-IF Grant ID: 800317 Enhanced-MUMS.

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P-3.2.13

Porphyrin-loaded polymeric microcapsules for Photodiagnosis of cancer

Please leave this line empty as spacer - delete this text. Font is Times New Roman 11
Mariana C. S. Vallejo1, Sofia G. Serra 2, Vanda V. Serra2, Nuno M. M. Moura1, M. Graça P. M. S. Neves1
Font is Times New Roman 11. Affiliation number superscript. Presenting author underlined
1) LAQV-REQUIMTE, Department of Chemistry, University of Aveiro, 3810-193 Aveiro, Portugal;
2) Centro de Química Estrutural, Instituto Superior Técnico, Universidade de Lisboa, 1049-001 Lisboa, Portugal
Please leave this line empty as spacer - delete this text. Font is Times New Roman 11
Tetrapyrrolic compounds, such as porphyrins, are of great interest for the scientific community due to their
physico- and photochemical features, allowing them to be used in a variety of research fields, including biomedical
sciences [1,2]. One particular interest of these molecules is related with their mode of action as light emitters: in thepresence of light, they can release energy in the form of fluorescence, being excellent candidates to be used in
Photodiagnosis of cancer [3]. Polymeric particles are a smart and controllable way to deliver these molecules to thetarget site. These particles, which are usually highly stable, biocompatible and stimuli-responsive, can also be
loaded with hydrophobic and hydrophilic cargos, such as porphyrins [4]. In this work, it is proposed the controlled
loading of cationic porphyrins polymeric particles for Photodiagnosis of cancerous cells, through local delivery andreal-time fluorescence imaging.

Fig. 1 – Synthesized cationic porphyrins to be incorporated in the polymeric particles (A); Scanning Electron Microscopy ofthe particles (B) and Fluorescence Lifetime Imaging Microscopy of the dopped particles (C).

References
1) Cerqueira, A. F. R.; Moura, N. M. M.; Serra, V. V.; Faustino, M. A. F.; Tomé, A. C.; Cavaleiro, J. A. S.;
Neves, M. G. P. M. S. Molecules, 22, 1269, 2017.
2) Kadish, K. M.; Smith, K. M.; Guilard, R. Handbook of Porphyrin Science, World Scientific Publishing
Company Co, Singapore, Vol 10-12, 2010.
3) Lim, E.-K.; Kim, T.; Paik, S.; Haam, S.; Huh, Y.-M.; Lee, K. Chem. Rev., 115, 327, 2015.
4) Parveen, S.; Sahoo, S. k. Journal of Drug Targeting, 16, 108, 2008.
Acknowledgements. We thank the University of Aveiro and FCT/MCT for the financial support for QOPNA
research Unit (FCT UID/QUI/00062/2019), REQUIMTE (UIDB/50006/2020) and the FCT project THERMIC(PTDC/QUI-COL/29379/2017), through national founds and were co-financed by the FEDER, within the PT2020
Partnership Agreement, and to the Portuguese NMR Network. The research grants of M.C.S. Vallejo (REF: BIUI51-8642-2019) and S.G. Serra are supported by the project THERMIC (PTDC/QUI-COL/29379/2017), and theresearch contracts of N.M.M. Moura (REF.-048-88-ARH/2018) and Vanda Vaz Serra are funded by national funds(OE), through FCT.

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IL-3.3.1

Photoreceptors from the plant symbiont Methylobacterium radiotolerans
Aba Losi1, Wolfgang Gärtner 2
1) University of Parma, Department of Mathematical, Physical and Computer Sciences, Parma, Italy1; 2)
University of Leipzig, a.Institute for Analytical Chemistry, Leipzig, Germany 2
Methylobacteria are pink-pigmented facultative methylotrophic (PPFMs) alpha-proteobacteria, most often living in
the phyllosphere as plant symbionts. They are able to feed on C1 organic compounds produced by plants, and in turnprovide their hosts with hormones and other metabolites that promote plant fitness and growth. As such,
Methylobacteria are considered as biofertilisers and plant probiotics representing a potential resource for sustainableagriculture.1 Methylobacteria are extremely rich in genes encoding for photoreceptors, mostly belonging to red/far
red light (RL/FRL) sensing bacterial phytochromes (BphP), and to the blue-light (BL) sensing LOV (Light, Oxygen
and Voltage) families.2 Here we present the molecular characterization of two such photoreceptors fromMethylobacterium radiotolerans, respectively MrBphP1 and Mr4511. MrBphP1 shows the expected
photochromicity between a red and far red absorbing form (Pr and Pfr), but with higher quantum yield (0.2 for bothroutes) than other BphPs. The LOV protein Mr4511 also shows the expected photochemical reactions, with BLdriven formation of a flavin-cysteine covalent adduct, but with an unusually slow recovery kinetics and an astonishingresistance to denaturation with urea. Upon substitution of the reactive Cys71 (C71S or C71G), Mr4511 becomes anefficient photosensitizer for singlet oxygen (SO), with ca. 20% efficiency.3 This unusual feature is due the absenceof a tryptophan residue at position 112, otherwise conserved in the majority of LOV domains. With a series of point
mutations at different distances from the flavin chromophore (between 0.6 and 1.7 nm), it was possible to map the
efficiency by which Tyr and Trp residues are able to quench the excited states of the flavin chromophore by electrontransfer (Fig.1). Substitution Y116F (0.9 nm from the flavin chromophore) results in a prolonged lifetime of theflavin triplet state, thereby largely increasing the efficiency of SO production. Instead, mutation F130W (0.6 nm fromthe flavin) results in complete quenching of the chromophore singlet state, also preventing adduct formation when
the reactive cysteine is present. The most efficient quencher for the flavin chromophore excited states is Trp, followedby Tyr and His, in agreement with the standard redox potentials of aromatic amino acids.

K57

~11 Å

~17 Å
~11 Å

~9 Å

~6 Å

Y129

F130

Q112
Y116

Fig. 1 – Structural model of Mr4511-LOV with positions mutated in this works

References
1) M. Kumar, D. Kour, A. N. Yadav, R. Saxena, P. K. Rai, A. Jyoti and R. S. Tomar, Biodiversity of
methylotrophic microbial communities and their potential role in mitigation of abiotic stresses in plants. Biologia(Bratisl). 74, 287–308, 20192) E. Consiglieri, Q. Xu, K.-H. Zhao, W. Gärtner and A. Losi, First molecular characterisation of blue- and redlight photoreceptors from Methylobacterium radiotolerans, Phys. Chem. Chem. Phys. 22, 12434–12446, 2020.
3) E. Consiglieri, Q. Xu, M. Bregnhøj, M. Westberg, P. R. Ogilby and A. Losi, Single mutation in a novel bacterial
LOV protein yields a singlet oxygen generator, Photochem. Photobiol. Sci. 18, 2657–2660, 2019.

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IL-3.3.2
Ralstonia solanacearum virulence and bacterial physiology are modulate by light andLOV photoreceptor.
María Josefina Tano1,2*, María Belén Ripa1,2*, Analía Carrau1,2, María Victoria Rodriguez2, Silvana Petrocelli2,
Elena G. Orellano1,2. *These authors contributed equally to the work.
1) Instituto de Biología Molecular y Celular de Rosario, Universidad Nacional de Rosario, Santa Fe, Argentina, 2)Área Biología Molecular, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario,
Santa Fe, Argentina, 3) Área Biología Vegetal, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad
Nacional de Rosario, Santa Fe, Argentina.

Microorganisms have developed metabolic and physiological capacities to adapt to a wide variety of
external stimuli. The light signals in the environment are perceived by photoreceptor proteins, which trigger
the signal transduction cascades that regulate light-biochemical responses. LOV (Light, Oxygen or Voltage)
photoreceptors are flavin-binding proteins using flavin mononucleotide (FMN) as chromophore. Ralstonia
solanacearum is the bacterium responsible for "bacterial wilt", one of the most destructive plant diseases
in the world. Currently, Ralstonia genus is considered a complex of species that can be differentiated basedon the attacking hosts, their geographic distribution, aggressiveness, and physiological properties. For thisreason, Ralstonia pseudosolanacearum (Rpso) GMI1000 is considered a model organism for the plantpathogen interaction. Rpso GMI1000 genome presents a Rsp0254 gene encoding a putative LOV domain
photoreceptor. To evaluate whether light would have a role in bacterial physiology and in the infectivity
capacity in tomato plants, mediated by this photoreceptor, it was first decided to analyze the light effect on
different bacterial virulence factors. Rpso exhibited lower adhesion capacity, motilities and biofilm
formation under white light. Once it was evidenced that light regulates different pathogenicity factors, themutant strain of Rpso in the Rpso0254 gene called Rpso lov was constructed to study whether this regulationis mediated by the LOV-type photoreceptor. The lov gene was analyzed by qRT-PCR under white light anddark treatments revealing that this gene is expressed. The absence of lov gene modified physiologicalparameters involved in the virulence of Rpso such as motility, biofilm and adhesion compared to wild typestrain. Finally, the infection capacity of tomato host plants (Solanum lycopersicum var. Minitomato)
mediated by both strains grown in different lighting conditions was studied, showing differences in bacterialcolonization depending on the light treatment applied. Altogether these results suggest a light-dependent
regulation of Rpso virulence during the host plant colonization.
Acknowledgements. This work was supported by grant from the ANPCyT (PICT 2017-2242), UNR (PID-1BIO432)
and CONICET (PIP 0451).

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IL-3.3.3"Effects of light on the non-photosynthetic plant growth promoting rhizobacteria A.
brasilense Az39”
Molina Romina1, Nievas Sofía1, Coniglio Anahí1, Mora Verónica1, Rosas Susana2, Cassán Fabricio1
1) Laboratorio de Fisiología Vegetal y de la Interacción Planta microorganismo, Instituto de Agrobiotecnología(INIAB), Universidad Nacional de Río Cuarto, Ruta 36 Km 601, Río Cuarto, Córdoba, Argentina; 2) Instituto de
Biotecnología Ambiental y Salud (INBIAS), Universidad Nacional de Río Cuarto, Ruta 36 Km 601, Río Cuarto,
Córdoba, Argentina.
Light is an essential environmental factor for the growth and development of photosynthetic microorganisms;
however, it can be a threat to those who are not 1.With the discovery of photoreceptors in prokaryotes, progress wasmade in the understanding of their functionality in nonphotosynthetic bacteria2. A. brasilense Az39 is anonphotosynthetic rhizobacterium used as a biofertilizer in South America during the last 40 years. In our laboratory,
we analyzed the Az39 genome, generated mutants at the level of its photoreceptors, and studied its physiological andmicrobiological responses to white, blue and red light (PAR38) under different experimental systems. In acomplementary way, we analyzed its physiological behavior in soybean leaves inoculated with this bacterium andexposed to different light conditions. Our results allowed us to understand that the tolerance of Az39 to light depends
on the culture conditions. In particularly, white and blue light were found to be lethal for this bacterium underconditions of gregarious growth (culture on solid medium); however, positive effects were observed under liquid
culture conditions, at level of the phytohormones production and lifestyle characters. The survival of these bacteria
on soybean leaves was drastically affected by the exposure to all light sources after inoculation, but mostly by whitelight on bacterial phytochrome expression deficient strain. Summarizing, A. brasilense Az39 is differentially affectedby light conditions in both positive and negative ways, and this capacity is dependent on the bacterial growth
conditions.
References
1) Elías-Arnanz, M., Padmanabhan, S., and Murillo, F. J. Light-dependent gene regulation in nonphototrophic
bacteria. Current opinion in microbiology, 14(2), 128-135, 2011.
2) Davis, S.J., Vener, A.V., and Vierstra, R.D. Bacteriophytochromes: phytochrome-like photoreceptors from
nonphotosynthetic eubacteria. Science, 286(5449), 2517-2520, 1999.
Acknowledgements. We thank Universidad Nacional de Río Cuarto, Consejo Nacional de Investigaciones
Científicas y Tecnológicas (CONICET) and FONCyT for the finantial support to this research.

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OC-3.3.4

New Insights from the Exciting Photobiological World of Mushrooms
Bianka Siewert1, Fabian Hammerle1, Johannes Fiala1 Lesley Huymann1,2, Gabrijela Curak1, Anna Hannecker1,
Ursula Peintner2, Ilse Kranner3, Harald Schöbel4, Michaele Lackner5, Christoph Schinagl2, Pamela Vrabl2
1) University of Innsbruck, Pharmacy/Pharmacognosy, Innsbruck, Austria; 2) University of Innsbruck,
Microbiology, Innsbruck, Austria; 3) University of Innsbruck, Botany, Innsbruck, Austria; 4) MCI Management
Center Innsbruck, Innsbruck, Austria; 4) Medical University Innsbruck, Innsbruck, Austria
Whenever insects nibble on fruiting bodies of the green Cortinarius austrovenetus the bite site turns reddish. The
chemistry behind it is as easy as it is fascinating: Hypericin is formed by oxidation from the fungal pigment
austrovenetin as soon as the fungal tissue is damaged. Inspired by this striking observation, we started tosystematically explore the photobiology of fruiting bodies.
Via an established workflow2, species of the colorful webcaps (Cortinarius) –especially of the subgenus Dermocybe–
were identified as a promising source for new photosensitizers. The photoactive principles were studied by the meansof molecular-network analysis and obtained via photo-activity guided isolation. The isolated pigments were photochemically as well as photo-pharmaceutically characterized; The most-promising fungal PS for PDT applications
showed a singlet oxygen photo-yield of ΦΔ = 20% (d4-MeOH, λirr = 450 nm, 15mW) and was highly photo-cytotoxic(EC50, A549 = 64 nM (468 nm, 9.3 J/cm², Selectivity Index Dark/Light > 61)3. In addition, we explored thephotoantimicrobial potential of these fungal PSs by submitting them to our new photoantimicrobial HTS-assay4.
While most extracts and fungal PSs were highly active against gram-positive bacteria, the photoactivity against
several Candida species –for example against the resistant Candida auris or C. tropicalis– was of special interest.
Furthermore, we studied the distribution of the photoactivity in the different organs (cap, lamellas, and stem) of the
fruiting bodies to test the hypothesis that mushrooms produce PSs to protect their spores. Our HPLC-DAD analysisof over 200 extracts clearly showed that the photoactivity is concentrated in the lamellas. In sum, here we will present
our newest insights from the photobiological world of fungi, highlighting Dermocybes as a promising and fascinating
source for new photosensitizers, photoantimicrobials, and phototoxic defense effectors.

Figure 1. Schematic representation of dermocyboid Cortinarii producing singlet oxygen under light irradiation. The
photobiological action is based on monomeric and dimeric anthraquinones and can be used to combat malignant cancer cells
and resistant microbes, as we will show here.

References
2) Siewert, B.; Vrabl, P; A convenient workflow to spot photosensitizers revealed photo-activity in basidiomycetes;
RSC Advances; 20193) Hammerle, F.; I. Bingger; Targeted Isolation of Photoactive Pigments from Mushrooms Yielded a Highly Potent
New Photosensitizer: 7,7’-Biphyscion; ChemRxiv; 2021
4) Fiala, J.; Schöbel, H.; A new High-Throughput-Screening-assay for Photoantimicrobials Based on EUCAST
Revealed Photoantimicrobials in Cortinariaceae; BioRxiv, 2021
Acknowledgements. This work was supported by the Austrian Science Fund (FWF, P319105) and the Tyrolian
Science Fund.
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IL-3.3.5
Photoperception in plant- and rock-associated black fungi (Ascomycota)
Julia Schumacher1,2, Anna A. Gorbushina1,21) Bundesanstalt für Materialforschung und -prüfung (BAM), 4 – Materials and the Environment, Berlin,
Germany; 2) Freie Universität Berlin, Department of Biology, Chemistry, Pharmacy, Berlin, GermanyPRs(photoreceptors)

3 PHYs2 OPs
Botrytis cinerea

Chromatin modifiersConserved signaltransduction(MAPKs, cAMP, Ca2+)

Fungal specific signal transduction(Velvet proteins)

LIGHT SIGNALING MACHINERY

LTFs(light-responsive
transcription factors)

PROTECTION

MORPHOGENESISTROPISMENTRAINMENT

pigmentation/sunscreens
enzymatic systems

conidiation,
apothecial development

conidiophores, apothecia,
conidial germ tubes

of a FREQUENCY-based
circadian clock

TRANSDUCTION

4 LOVsOUTPUT

X 2 CRYs

INPUT

Fungi that share light-flooded habitats with phototrophs may profit from their excess photosynthetic products.
Sunlight-associated stresses are however multiple: high temperatures, UV radiation with associated DNA damage,accumulation of reactive oxygen species (ROS), desiccation and osmotic stresses. Ascomycota dominating lightflooded habitats accurately sense and respond to changes in light using it as a cue to coordinate growth, stressresponses as well as to establish pathogenic or symbiotic relationships. Two species from two light-flooded habitats– phyllosphere and sun-exposed solid surfaces – were analysed for their photoreceptor distribution.
In both habitats phototroph-associated and black [dihydroxynaphthalene (DHN) melanin-containing] fungi areprevalent. This diversity was sampled with the plant-associated fungus Botrytis cinerea (Leotiomycetes), while
Knufia petricola (Eurotiomycetes) was included as a typical biofilm-former on sun-exposed solid surfaces e.g. rocks,
building facades, roofs, and solar panels. The analysis has shown that genomes of black fungi contain more
photoreceptors than animal pathogens and saprophytes such as Aspergillus nidulans and Neurospora crassa1,2.
B. cinerea that causes the grey mould disease by infecting the above-ground parts of more than 200 dicots has ahighly sophisticated photosensory and signalling system that helps to avoid light and to locate susceptible hosts1.

Fig. 1 – B. cinerea uses signals from at least 11 photoreceptors to coordinate stress responses, development, and virulence.

Rock-inhabiting Dothideomycetes and Eurotiomycetes including Knufia petricola possess equal numbers ofphotoreceptors along with the same set of protective metabolites i.e. melanin, carotenoids and mycosporines2. Thissimilarity between black fungi from plant and rock surfaces suggests that photoperception and -regulation areimportant for sun-stressed fungi that receive nutrients through cooperation with phototrophs. CRISPR/Cas9-based
genetic tools for manipulating K. petricola were established3 and are currently used for elucidating the functions of
the different photoreceptors in the biology of rock-inhabiting fungi.
References
1) Schumacher J. How light affects the life of Botrytis. Fungal Genetics and Biology 106: 26-41, 2017.
2) Schumacher J, Gorbushina AA. Light sensing in plant- and rock-associated black fungi. Fungal Biology 124:407417, 2020.
3) Voigt O, Knabe N, Nitsche S, Erdmann EA, Schumacher J, Gorbushina AA. An advanced genetic toolkit for
exploring the biology of the rock-inhabiting black fungus Knufia petricola. Scientific Reports 10(1), 2020.
Acknowledgements. This work was supported by the grant SCHU 2833/4-1 from the German Research Foundation(DFG) and internal funds of the BAM.
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IL-3.3.6
Analysis of the light response in Alternaria alternata – mitochondria as novel
phytochrome assembly stations
Reinhard Fischer, Kai Leister, Christian Streng
Karlsruhe Institute of Technology, Department of Microbiology, Karlsruhe, Germany
Mitochondria are essential organelles because of their function in energy conservation. Here, we show an
involvement of mitochondria in phytochrome-dependent light sensing in fungi. Phytochrome photoreceptors arefound in plants, bacteria and fungi and contain a linear, heme-derived tetrapyrrole as chromophore. Linearization ofheme requires heme oxygenases (HOs) which reside inside chloroplasts in planta. Despite the poor degree ofconservation of HOs, we identified two candidates in the fungus Alternaria alternata. Deletion of either one
phenocopied phytochrome-deletion. The two enzymes had a cooperative effect and physically interacted withphytochrome, suggesting metabolon formation. The metabolon was attached to the surface of mitochondria with aC-terminal anchor sequence in HoxA. The affinity of phytochrome apoprotein to HoxA was 57,000-fold higher
than the affinity of the holoprotein, suggesting a “kiss-and go” mechanism for chromophore loading and a functionof mitochondria as assembly platforms for functional phytochrome. Hence, two alternative approaches forchromophore biosynthesis and insertion into phytochrome evolved in plants and in fungi.

Fig. 1 – Phytochrome apoprotein is produced in thecytoplasm and travels to mitochondria, where a two heme
oxygenases provide the linear tetrapyrrole biliverdin andinsert it into the phytochrome protein. Functional
phytochrome interacts with the phophoptransfer protein
YpdA to induce the MAP kinase pathway. A fraction ofholo-phytochrome is imported into nuclei, where it isinvolved in chromatin remodeling.

References
Igbalajobi, O., Yu, Z. and Fischer, R. (2019). Red- and blue-light sensing in the plant pathogen Alternaria
alternata depends on phytochrome and the white-collar protein LreA. MBio 10, e00371-19.
Igbalajobi, O., Gao, J. and Fischer, R. (2020). The HOG pathway plays different roles in conidia and hyphaeduring virulence of Alternaria alternata. Mol Plant Microbe Interact 33, 1405-1410.
Streng, C., Hartmann, J., Leister, K., Krauß, N., Lamparter, T., Frankenberg-Dinkel, N., Weth, F.,
Bastmeyer, M., Yu, Z. and Fischer, R. (2021) A novel role for mitochondria in light sensing. EMBO J., inrevision.
Yu, Z., Armant, O. and Fischer, R. (2016). Fungi use the SakA (HogA) pathway for phytochrome-dependent
light signaling. Nat. Microbiol. 1, 16019.
Yu, Z. and Fischer, R. (2019). Light sensing and responses in fungi. Nat. Rev. Microbiol. 17, 25-36.
Acknowledgements. This work was supported by the Deutsche Forschungsgemeinschaft.

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IL-3.3.7
Light regulates the degradation of VE-1, a component of the regulatory velvet
complex in the fungus Neurospora crassa
Luis M. Corrochano1, M. del Mar Gil-Sánchez1, Sara Cea-Sánchez1, María Corrochano-Luque1, Alejandro
Miralles-Durán1, Cristina De Andrés-Gil1, Eva M. Luque1, Guilherme TP Brancini1, Gabriel Gutiérrez1 and David
Cánovas1
1) University of Seville, Department of Genetics, Seville, Spain
Conidiating mycelia of species of the fungus Neurospora colonize the surface of burned vegetation and facilitate
the recycling and restoration of forests after fires. The velvet complex is a protein complex that serves as a key
element in the coordination of fungal development, secondary metabolism, and the perception of environmental
signals, such as light. Most fungi use proteins similar to WC-1 and WC-2 from Neurospora crassa for sensing blue
light. In N. crassa and other fungi these two proteins form a photoreceptor and transcription factor complex (WCC)that binds to the promoters of light-regulated genes to activate transcription. Several other photoreceptors havebeen identified in fungi, including phytochromes, opsins and cryptochromes, but they play a secondary role in thephotobiology of N. crassa.
The velvet regulators are members of a family of proteins with a conserved domain that help to coordinate growth,differentiation and secondary metabolism in fungi. There are four genes containing a velvet domain in the genomeof N. crassa (ve-1, ve-2, ve-3 and vos-1). Mutants in genes ve-1 and ve-2 show defective aerial hyphae growth,
increased conidiation, and reduced carotenoid accumulation. VE-1, VE-2 and the methyltransferase LAE-1 areobserved in vegetative mycelia and during conidiation. These three proteins interact to form the velvet complex
that, presumably, regulates transcription during conidiation and the biosynthesis of secondary metabolites such as
carotenoid protective pigments. During the characterization of the abundance of the components of the velvet
complex during conidiation we noted that VE-1 was absent in aerial hyphae grown in the dark despite the presenceof ve-1 mRNA. The other components of the velvet complex, VE-2 and LAE-1, accumulated in similar amounts invegetative and conidiating mycelia. We have shown that the absence of VE-1 in aerial hyphae in the dark is due toits degradation through the proteasome with a key role for the adaptor protein FWD-1. The degradation of VE-1 isreduced by light in a process that requires the blue-light photoreceptor WC-1. We propose that the light-dependent
regulation of VE-1 stability modifies the components of the velvet complex resulting in changes in thetranscriptome during conidiation.
Acknowledgements. Supported by European funds (European Regional Development Fund) and the Ministry of
Science, Innovation and Universities, Spain (RTI2018-098636-B-I00) to LMC and DC. The work of Guilherme TP
Brancini at the University of Seville was supported by a BEPE short-term fellowship from FAPESP (2018/003557), Brazil.

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OC-3.3.8

Effects of LOV photoreceptor deletion in the Pseudomonas syringae-tomato system
Daniela Ceresini1, Francesca Degola1, Aba Losi2, Ada Ricci1
1) University of Parma, Department of Chemistry, Life Sciences and Environmental Sustainability, Parco Area delle
Scienze 11/A, 43124, Parma, Italia; 2) University of Parma, Department of Mathematical, Physical and Computer
Sciences, Parco Area delle Scienze 7/A, 43124, Parma, Italia.
Light is an important environmental signal and its perception plays a crucial role in many physiological processes,
even in non-phototrophic organisms associated with the phyllosphere. These organisms possess photosensory
proteins capable of perceiving light and transducing light signal to evoke a response in the bacterial cell.
Pseudomonas syringae pv. tomato DC3000 (Pst), the etiological agent of bacterial speck in tomato plants, represents
one of the most important models for studying plant-pathogen interaction. It causes large losses of crops and thiscalls for more research on the physiological basis of its infectiveness and virulence. The interest in this microorganism
is linked to the presence in its genome of various gene sequences coding for photoreceptors. Pst is indeed equippedwith two red/far-red (R/FR) light sensing bacteriophytochromes (BphP), binding biliverdin as chromophore and with
one blue light sensing LOV (Light, Oxygen and Voltage) protein binding flavins as chromophore, mimicking thephotosensing ability of host plants.
Here, a major target is to determine if and how Pst LOV photoreceptor interfere with plant defense system. We report
the study of the effects of infection on tomato plants with LOV mutant strain deleted in the LOV gene. We alsostudied the effects of salicylic acid (SA), an important endogenous plant defense signal of attack by pathogens, on
the growth of LOV mutant strain. Furthermore, we have studied the plant's response to infection by quantifying
callose production.
The mutant strain lacking the photosensory protein LOV is more virulent and invasive towards tomato plants than
Pst-WT. Moreover, we found that SA concentrations of 1mM and 5mM inhibit the growth of both the mutant LOV
strain and the Pst-WT strain in vitro, while growth stimulation is surprisingly observed at 0.1mM concentration. As
far as the plant production of callose is concerned, a higher production of callose after 24h in plants infected with the
mutant LOV strain is reported compared to those infected with the Pst-WT strain.
Further studies are needed to better understand the role of this photoreceptor in the Pseudomonas syringae-tomato
system.
References
1) Ricci, A., Dramis, L., Shah, R., Gärtner, W. and Losi, A. (2015), Bacterial photoreceptors in plant–pathogen
interaction. Environmental Microbiology Reports, 7: 795-802.
2) Shah, R., Pathak, G., Drepper, T. and Gärtner, W. (2016), Selective Photoreceptor Gene Knock-out Reveals a
Regulatory Role for the Growth Behavior of Pseudomonas syringae. Photochem Photobiol, 92: 571-578.

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OC-3.4.1

Highly sensitive detection of 2-photon photosensitized singlet oxygen using a novel
optofluidic system
Sergio Adan Bermudeza, Jochen Arltb and Anita C. JonesaaSchool of Chemistry, The University of Edinburgh, Edinburgh EH9 3FJ, UKb
School of Physics and Astronomy, The University of Edinburgh, Edinburgh EH9 3FD, UK
Photosensitized production of singlet oxygen (1O2) is used in the treatment of cancer by photodynamic therapy (PDT),
in which a photosensitizer is administered and excited locally in the tumour tissue, generating 1O2 to destroy,
selectively, malignant cells. PDT is also used in the treatment of age-related macular degeneration, atherosclerosis,
psoriasis and acne. The use of two-photon excitation (TPE) in PDT has significant advantages in achieving enhanced
spatial selectivity and greater depth of penetration through tissue. This is driving development of new
photosensitizers with high 2-photon cross-sections, but progress is inhibited by the difficulty of determining, in vitro,
the efficiency of 1O2 production in response to TPE by these new pro-drugs.
In conventional experiments, the extremely high photon intensity needed to achieve two-photon absorption is createdby focusing a pulsed laser beam into a spot of about 1 μm in diameter. This miniscule (femtolitre) excitation volume
makes the detection of two-photon-induced 1O2 generation extremely challenging. We will describe a new approachto the detection of 2-photon photosensitized 1O2, which exploits the unique optofluidic properties of hollow-core
photonic crystal fibre (HC-PCF).1 In HC-PCF, light is trapped in the hollow core by the surrounding microstructured
fused-silica cladding. This allows the infiltration of a sample solution into the hollow core, which is typically 10’s ofm in diameter, while maintaining the high optical transmission efficiency of the fibre. As we have shown
previously, the confinement of both laser beam and sample solution within the fibre core permits two-photon
excitation to be sustained over a path-length of more than 10 cm.2
We will report the development a HC-PCF-based optofluidic platform for the detection of 2-photon photosensitizedsinglet oxygen, to enable the in vitro screening and quantitative characterisation of two-photon photosensitizers,
under mechanistically relevant conditions (in aqueous solution and using low pulse-energy excitation). Highly
sensitive detection is achieved using a 1O2-specific fluorescent probe, employing a two-colour pump-probe
methodology, with an 800-nm pump beam (TPE of photosensitizer) and a 488-nm probe beam (excitation offluorescent probe) co-coupled into the hollow core of the fibre. Using this approach, we are able to detect
quantitatively singlet oxygen produced by TPE of sub-micromolar concentrations of photosensitizer in aqueous
solution.
References
1. A. M. Cubillas, S. Unterkofler, T. G. Euser, B. J. Etzold, A. C. Jones, P. J. Sadler, P. Wasserscheid, P. S.
Russell Chem. Soc. Rev. 2013, 42, 8629-8648.
2. G. O. S. Williams, T. G. Euser, J. Arlt, P. S. J. Russell, A. C. Jones ACS Photonics 2014, 1, 790-793.

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OC-3.4.2

Photoprotection, energy quenching and PSBS, is it really important?
Christo Schiphorst1, Koen Staring1, Emilie Wientjes1
1) Wageningen University, Biophysics, Wageningen, Netherlands
Photosynthesis is crucial for life on earth, but it is not very efficient. In most conditions, less than 1% of the incoming
light-energy is converted into biomass, which is far below the theoretical maximum. One of the major losses happensin the form of energy quenching (qE), a process where light energy is converted into heat. In plants this is activatedby the protein PSBS upon a decrease in the pH of the thylakoid lumen. We are investigating when and how PSBS
becomes important for photoprotection, water usage and crop-yield in various light and temperature conditions.

Figure 1 Leaf area of WT, npq4 in continuous fluctuating light conditions. Left inset 1h 1000µmol*m 2*s-1 & 0.5h 100µmol*m2*s-1. Right
inset: 1min 1000µmol*m2*s-1 & 5min 100µmol*m2*s-1.

However, despite the general believe that PSBS is essential for photoprotection and optimal growth, finding
conditions in which there is a difference in growth between WT and a mutant without PSBS (npq4 mutant) is not as
simple, see Fig. 1. Therefore, we are combining growth experiments with more direct measurements like CO2
assimilation and chlorophyll fluorescence to assess in real time what is happening during the light fluctuations, seeFig. 2.

Figure 2 CO2 assimilation measurements and Chlorophyll fluorescence of WT and npq4 plants in different fluctuating light conditions.

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OC-3.4.3
Oxybenzone solar filter as a photoremovable protecting group for carbonylcompounds of biological interest
Mauricio Lineros-Rosa, Miguel A. Miranda and Virginie Lhiaubet-Vallet
Instituto Universitario Mixto de Tecnología Química UPV-CSIC, Universitat Politècnica de València, Consejo
Superior de Investigaciones Científicas, Avda de los Naranjos s/n, 46022 Valencia, Spain
Nowadays, there is a growing number of cases dealing with photosensitization due to the increasing use of
xenobiotics (such as cosmetics, fragrances and drugs) in combination with exposure to sunlight. These harmful
effects are closely related to the development of skin cancer. Therefore, the design of new photoprotection strategies
to counteract these negative consequences is of utmost importance.1 In this context, a new photoremovable carbonylprotecting group based on the oxybenzone (OB) solar filter is presented.2 This system has demonstrated the ability
to photorelease aromatic ketones along with the aforementioned UV filter in a simultaneous and controlled manner.
Thus, the OB protects the carbonyl derivative against the deleterious effect of UV light (Figure 1A). In the case ofaromatic ketones, this photoprotection is of special interest in view of their well-known photoreactivity. Among thepossible aromatic ketone substrates for this new PPG, ketoprofen (KP) can be highlighted given that it is a worldwide
employed nonsteroidal anti-inflammatory drug of topical use and responsible for severe photosensitizing effects.3
(A)
R1
O

O

R2

O

hν

OH

O

O
Oxybenzone
solar filter

Oxybenzone-based
systems

(B)

O

O

O

O

O

O
OB-CBP

R1

R2

Carbonylcompounds

OH

O

O
OB-BP

O
+

O

O

O
OB-AP

O

HO

O

O

O

O
OB-TL

O
OB-KP

Figure 1. Photorelease of oxybenzone and carbonyl derivatives compounds from the oxybenzone-based systems (A) and thestructures of the studied systems (B).

In this work, the photophysical and photochemical properties of the OB-BP and OB-CBP systems (Figure 1B) havebeen thoroughly studied through experimental techniques such as laser flash photolysis, HPLC, UV-Vis spectroscopy
and mass spectrometry. Based on these results, a photorelease mechanism involving a first d-hydrogen abstraction
followed by photoionization has been proposed. Additionally, this system has also been applied for the photoreleaseof other aromatic ketones, i.e. a-tetralone (TL), acetophenone (AP) and ketoprofen (KP).
References
1) (a) I. Aparici-Espert, M. C. Cuquerella, C. Paris, V. Lhiaubet-Vallet and M. A. Miranda, Chem. Commun., 2016,
52, 14215-14218. (b) M. Lineros-Rosa, M. C. Cuquerella, A. Francés-Monerris, A. Monari, M. A. Miranda, V.
Lhiaubet-Vallet. Org. Biomol. Chem., 2021, 19, 1752-1759.
2) M. Lineros-Rosa, M. A. Miranda, V. Lhiaubet-Vallet. Chem. Eur. J., 2020, 26, 7205-7211.
3) V. Lhiaubet-Vallet and M. A. Miranda, CRC Handbook of Organic Photochemistry and Photobiology, CRC Press,
Boca Raton, 3rd edn, 2012, ch. 66, vol. 2, p. 1541.
Acknowledgements. This work was supported by the Universitat Politècnica de València (FPI grant) and by the
Spanish government (project PGC2018-096684-B-I00).
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OC-3.4.4

Photostability of Ipilimumab and Nivolumab in the formulation and sterile saline or
glucose solutions for parenteral administration.
Benedetta Fongaro, Valentina Cian, Giorgia Miolo and Patrizia Polverino de Laureto
University of Padova, Department of Pharmaceutical and Pharmacological Sciences, Padova, Italy
Background. Monoclonal antibodies are complex protein molecules, and their structural integrity influences their
biological and pharmacological activity. Ipilimumab and Nivolumab, targeting the molecules CTLA-4, PD-1,
respectively, have shown efficacy against several types of cancer and have demonstrated the superiority of the
combination strategy compared to the single agent therapy.1
Aim. We investigated the impact of some stress factors, i.e., shaking, vibrations and light exposure, on two
formulated monoclonal antibodies (mAbs) Ipilimumab and Nivolumab, to mimic their routine handling once
released from the pharma industry, shipped to the hospital, diluted for the parenteral administration, and finallyadministered to the patients.
Methods. Protein stability analyses, including aggregation and excipient effects on the protein structure, werecarried out by spectroscopic methods (Uv-Vis, Fluorescence, and Circular dichroism), native and denaturing
electrophoresis, and size exclusion chromatography analysis. Suntest CPS+ was used to simulate the conditions of
one day or 3 weeks of sunlight exposure.
Results. Among the selected critical stress factors to simulate the real-life handling of mAbs during transport,
pneumatic systems, reconstitution, and administration to patients, only light exposure was able to affect the protein
drug products. The two mAbs demonstrated a general instability to light as intact products and after dilution insaline and glucose solutions (both homemade and commercial sterile solution). The highest instability under light
exposure was found in diluted glucose sterile solutions, suggesting the role of glucose degradation products on thephoto-modification process of the two mAbs.
Conclusion. Nivolumab and Ipilimumab are stable molecules under vigorous shaking and vibrations. However,they undergo structural change, mostly aggregation, upon exposure to artificial sunlight. The instability of mAbs
upon light exposure could have a potential impact on the safety and efficacy of these very active drugs inanticancer therapy.
1) Marchetti P. Botticelli A., Agnostic evaluation of ipilimumab and nivolumab association: a metanalysis.
J Transl Med, 2020.
2) Le Basle Y., Chennel P., Physicochemical Stability of Monoclonal Antibodies: A Review. Journal of
Pharmaceutical Sciences, 2020.

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OC-3.4.5
Chromatophores efficiently promote light-driven ATP synthesis and DNA
transcription inside hybrid multicompartment artificial cells
Emiliano Altamura,1 Paola Albanese,1 Roberto Marotta,2 Francesco Milano,3 Michele Fiore,4 Massimo Trotta,5Pasquale Stano6 and Fabio Mavelli1,71Department of Chemistry, University of Bari “Aldo Moro”, Bari, Italy.
Electron Microscopy Facility, Italian Institute of Technology (IIT), Genova, Italy.
3
CNR-ISPA, Institute of Sciences of Food Production, Lecce Unit, Lecce, Italy.
4
Institut de Chimie et Biochimie Moléculaires et Supramoléculaires, Université de Lyon “Claude Bernard Lyon
1”, Lyon, France.
5
CNR-IPCF, Institute for Physical and Chemical Processes, Bari, Italy.
6
Department of Biological and Environmental Sciences and Technologies (DiSTeBA), University of Salento,
Lecce, Italy.
7
CNR NANOTEC, Istituto di Nanotecnologia, Bari, Italy.
2

In the emerging field of bottom-up synthetic biology, scientists challenge to transfer cellular capabilities to simplified
artificial compartments1-7. Considering that every cellular active function need energy to be maintained, a crucial
requirement for an artificial cell is to be energetically autonomous.
Our research is aimed to overcome this criticality by constructing synthetic cells able to assimilate energy from lightand support an internal metabolism. We called these systems: Artificial Simplified-Autotroph Protocells (ASAPs)8.
Here we show a hybrid multi-compartment approach to build ASAPs in an effective manner. Chromatophores
obtained from Rhodobacter sphaeroides (Fig 1a and 1b) accomplish the photophosphorylation of ADP to ATP
functioning as nanosized photosynthetic organellae when encapsulated inside artificial giant phospholipid vesicles
prepared with the droplet transfer method. Under continuous illumination chromatophores produce ATP that in turnsustains the transcription of a DNA gene by T7 RNA polymerase inside ASAPs (Fig 1c).

Figure 1. Cartoon representation of chromatophore-containing ASAPs (a) Rhodobacter sphaeroides R-26 cell in section. The
membrane exhibits invaginations, large spherical vesicles (average diameter 80 nm), and several stratified flattened vesicles.
(b) Chromatophores are ∼80-nm closed vesicles derived from the lysis of R. sphaeroides R-26 cells, typically via Frenchpressing. Their membrane contains the entire photosynthetic system: Light Harvesting complex I (dark red), Reaction Center(blue), ubiquinol oxidase (or bc1, in red) and ATP synthase (yellow) in inside-out orientation when compared with theBook of Abstracts ������������������������������������������������������������������������������������������������ 153

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cytoplasmic membrane (ATP produced in the outer solution). (c) Chromatophores encapsulated inside giant phospholipid
vesicles act as ATP-producing organellae when illuminated by an 860-nm light. Co-encapsulated ADP, Pi, GTP, CTP, UTP, T7
RNA polymerase (in purple), and a DNA template give rise to an out-of-equilibrium system of coupled reactions: namely,
photophosphorylation and DNA transcription. The biosynthesized mRNA is revealed by Acridine Orange (green), which binds
it, forming a green-fluorescent complex. Image credit Filippo Trazi.

Cryo-EM and time-resolved spectroscopy were used for characterizing the chromatophore morphology and theorientation of the photophosphorylation proteins, which allow high ATP production rates (up to ~100 ATP/s per ATP
synthase). mRNA biosynthesis inside individual vesicles has been determined by confocal microscopy8. The hybrid
multi-compartment approach here proposed appears at the same time convenient and effective, and thus very
promising for the construction of full-fledged artificial protocells.

References
1) P. L. Luisi, Toward the engineering of minimal living cells. Anat. Rec. 268, 208–214 (2002).
2) Walde P. (2010) Building artificial cells and protocell models: experimental approaches with lipid vesicles
Bioassays Vol. 32, 296-303.
3) J. C. Blain, J. W. Szostak, Progress toward synthetic cells. Annu. Rev. Biochem. 83, 615–640 (2014).
4) B. C. Buddingh’, J. C. M. van Hest, Artificial cells: Synthetic compartments with life-like functionality andadaptivity. Acc. Chem. Res. 50, 769–777 (2017).
5) P. Schwille et al., MaxSynBio: Avenues towards creating cells from the bottom up. Angew. Chem. Int. Ed. Engl.
57, 13382–13392 (2018).
6) J. W. Hindley et al., Building a synthetic mechanosensitive signaling pathway in compartmentalized artificial
cells. Proc. Natl. Acad. Sci. U.S.A. 116, 16711–16716 (2019).
7) Y. Elani, Interfacing Living and Synthetic Cells as an Emerging Frontier in Synthetic Biology. Angew. Chem.
60(11): 5602-11.
8) E. Altamura, P. Albanese, et al. Chromatophores efficiently promote light-driven ATP synthesis and DNA
transcription inside hybrid multicompartment artificial cells. Proc. Natl. Acad. Sci. USA (2021), Vol. 118(7)Book of Abstracts ������������������������������������������������������������������������������������������������ 154

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OC-3.4.6

Hypericin against SARS-CoV-2: from binding to antiviral efficacy
Pietro Delcanale1, Andrea Mussini1, Eleonora Uriati1,2, Matteo Mariangeli1,2, Paolo Bianchini2,3, Alberto
Diaspro2,3,4, Stefania Abbruzzetti1, Davide Lelli5, Ana Moreno5, Cristiano Viappiani1
1) Department of Mathematical Physical and Computer Sciences, University of Parma, Parma, Italy; 2)Nanoscopy, Istituto Italiano di Tecnologia, Genoa, Italy; 3) Nikon Imaging Center, Istituto Italiano di Tecnologia,
Genoa, Italy; 4) Department of Physics, University of Genoa, Genoa, Italy; 5) Istituto Zooprofilattico Sperimentaledella Lombardia e dell’Emilia-Romagna, Brescia, Italy
Hypericin is photosensitizing drug with significant antiviral activity against membrane-enveloped viruses andtherefore constitutes a promising candidate for the treatment of SARS-CoV-2 1. The binding affinity of Hypericin toviral components and the distribution of Hypericin molecules within a population of viral particles are fundamental
parameters determining its ensemble efficacy 2. Hence, quantitative measurements addressing molecular binding anddistribution of Hypericin on SARS-CoV-2 virions can provide a rationale to guide the improvement of treatments
based on this photosensitizing drug, beyond the mere observation of a reduced infectivity.
In this work, we use a quantitative approach to follow the interaction of Hypericin with SARS-CoV-2, at the basis of
the efficacy of this drug. First, we exploited the fluorescence emission of Hypericin to visualize its binding onto viralparticles (see Fig.1). Then, using a combination of spectroscopy techniques, we identified the membrane envelope
as the main viral component targeted by the photosensitizer and we measured binding affinity. In addition, we usedfluorescence microscopy to investigate the loading of Hypericin molecules onto individual SARS-CoV-2 virions.
Finally, the results are correlated to the ensemble antiviral efficacy of Hypericin, both photoinduced and in the dark,
observed on infected Vero E6 cells. These findings demonstrate that Hypericin has a significant antiviral activity
against SARS-CoV-2 and provide a quantitative characterization of binding, which is key to understand themechanisms of action of Hypericin and to drive the rational development of antiviral treatments based on
photosensitization.

Fig. 1 – A: Representation of Hypericin binding to a SARS-CoV-2 virion: while unbound Hypericin is found in non-emissive
aggregates, virus-bound Hypericin molecules recover fluorescence emission, enabling selective detection. B: Confocal image of
SARS-CoV-2 virions exposed to Hypericin. Emission from virus-bound Hypericin is shown in red while emission from Hoechst,
labelling the viral RNA, is in green. Co-localized red and green emissions look yellow. Scale bar 1 μm.

References
1) Kubin A., Wierrani F., Hypericin -The facts about a controversial agent. Current Pharmaceutical Design, 2005.
2) Wiehe A., O'Brien J.M., Trends and targets in antiviral phototherapy. Photochemical & Photobiological Sciences,
2019.

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OC-3.4.7

An update on Photodynamic Decontamination to prevent foodborne disease
Michael Glueck1, Christoph Hamminger1, Markus Hoerl1, Barbara Schamberger1, Kristjan Plaetzer1
1) University of Salzburg, Department of Biosciences, Laboratory of Photodynamic Inactivation of
Microorganisms, Salzburg, Austria
Foodborne disease occurs regularly around the globe and often leads to severe illness and even death. To minimize
this threat and to ensure microbiological safety, the food industry applies different thermal and non-thermal
sterilisation technologies. For vegetables and fruits, which are often eaten raw, thermal treatment is impracticableand non-thermal technologies, such as treatment with chlorine gas or gamma irradiation, are deprecated byconsumers. Photodynamic Decontamination (PDc) represents a non-thermal alternative and has been proven to beeffective in killing of bacteria on food surfaces.1,2 Relevant bacterial genii in food hygiene are Listeria sp. (Gram+)and Escherichia sp. (Gram-). The aim of this study was therefore to test PDc based on two natural photosensitizers(PS), Na-chlorophyllin (Chl) and curcumin, against these bacteria.
Listeria innocua as model system for Listeria monocytogenes was grown in liquid culture and subsequently
PDc-treated using Chl (1 µM, 10 µM or 100 µM, 5 min incubation, 15.3 J/cm2 at 395 nm) or NovaSOL®- Curcumin(NoCu), a water soluble micellar formulation of curcumin (0.1 µM to 50 µM, 15 min incubation, 20.4 J/cm2 at
435 nm). The minimal radiant exposure for achievement of an antibacterial effect was investigated using NoCu at
5 µM against L. innocua after 15 min incubation. As example of use, PDc efficiency was also examined on fish skin(trout) using NoCu and on fenugreek seeds using Chl in combination with polyaspartic acid (PA). The foodstuff wascontaminated with L. innocua and subsequently phototreated. Evaluation was done by counting of colony forming
units (CFU). To study the influence of the target geometry, slices of cucumbers as flat, mung beans as round andgerminated mung beans as complex objects were purposely contaminated with E. coli, incubated with 10 µM, 50 µM
or 100 µM of SACUR-3 (a cationic derivative of curcumin) and illuminated at 435 nm (33.8 J/cm2).
The treatment of L. innocua in liquid culture was highly effective for both photoactive compounds. Concentrationsof 1 µM and above for NoCu and 10 µM and higher for Chl led to a complete kill of bacteria. At 5 µM NoCu a very
low radiant exposure of ca. 1.4 J/cm2 induced cell death in all bacteria of the sample. The PDc of L. innocua on fishskin and on fenugreek seeds showed phototoxicity achieving nearly 3 log10 reduction. On flat surfaces as cucumbers,
E. coli was killed with a relative inactivation of 3.4E4 using 100 µM SACUR-3. On mung beans, the relative
inactivation reached 5.0E3, but only 5.1E0 on germinated mung beans.
Photodynamic Decontamination is a promising approach to improve microbial food safety. The use of PSs approved
as food additives guarantees biocompatibility. Phototreatment based on Chlorophyllin and Curcumin has highpotential to contribute to the prevention of foodborne diseases. Jet there are limitations, which can partially beovercome by the development of 3D illumination devices.
References
1) Buchovec I., Lukseviciute V., Marsalka A., Reklaitis I. and Luksiene Z., Effective photosensitization-based
inactivation of Gram (-) food pathogens and molds using the chlorophyllin-chitosan complex: towards photoactive
edible coatings to preserve strawberries. Photochemical & Photobiological Sciences, 20162) Glueck M., Schamberger B., Eckl P. and Plaetzer K., New horizons in microbiological food safety:
Photodynamic Decontamination based on a curcumin derivative. Photochemical & Photobiological
Sciences, 2017
Acknowledgements. We thank Suncor AgroScience for providing support.

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OC-3.4.8

Photoactivatable metabolic warheads enable precise and safe ablation of target cells invivo
Sam Benson1, Fabio de Moliner1, Antonio Fernandez1, Erkin Kuru2, Nicholas L. Asiimwe3, Jun-Seok Lee4, Lloyd
Hamilton5, Dirk Seiger5, Isabel R.Bravo1, Abigail M. Elliot1, Yi Feng1 & Marc Vendrell1.
1) Centre for Inflamamtion Research, QMRI, Theuniversity of Edinburgh, Edinburgh, UK
2) Department of Genetics, Harvard Medical School, Boston, MA, USA.
3) Molecular Recognition Research Center, KIST, Seoul, South Korea
4) Departmetn of Pharmcaology, Korea University College of Medicine,Seoul, South Korea
5) The Centre for Discovery Brain Sciences, The University of Edinburgh, Edinburgh, UK
Photoactivatable molecules enable ablation of malignant cells under the control of light, yet current agents can beineffective at early stages of disease when target cells are similar to healthy surrounding tissues. We have created achemical platform based on amino substituted benzoselenadiazoles to build photoactivatable probes that mimic
native metabolites as indicators of disease onset and progression. Through a series of synthetic derivatives, we haveidentified the key chemical groups in the benzoselenadiazole scaffold responsible for its photodynamic activity,and subsequently designed photosensitive metabolic warheads to target cells associated with various diseases,
including bacterial infections and cancer. We demonstrate that versatile benzoselenadiazole metabolites canselectively kill pathogenic cells but not healthy cells with high precision after exposure to non-toxic visible light,
reducing any potential side effects in vivo. This chemical platform provides powerful tools to exploit cellular
metabolic signatures for safer therapeutic and surgical approaches.1

References
1) Benson, S.; de Moliner, F.; Fernandez, A.; Kuru, E.; Asiimwe, N. L.; Lee, J.-S.; Hamilton, L.; Sieger, D.;
Bravo, I. R.; Elliot, A. M.; Feng, Y.; Vendrell, M. Photoactivatable Metabolic Warheads Enable Precise
and Safe Ablation of Target Cells in Vivo. Nat. Commun. 2021, 12, 2369. https://doi.org/10.1038/s41467021-22578-2.
Acknowledgements. This work was supported by Medical Research Scotland (S.B.:879-2015) and the ERC
consolidator grant (M.V.:771443).

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OC-3.4.9

A portable NIR spectrometer directly quantifies singlet oxygen generated bynanostructures for Photodynamic Therapy in deep tissues
Orsi Davide1, Vaccari Marco1, Baraldi Andrea1, Cristofolini Luigi1
1) Università di Parma, Dipartimento di Scienze Matematiche, Fisiche e Informatiche, Parma (IT)We report on the setting up and calibration of a portable NIR spectrometer specifically developed for quantitative
direct detection of the highly reactive singlet oxygen (1O2) chemical specie, of great importance in Photodynamic
therapies. This quantification relies on the measurement of fluorescence emission of 1O2, which is peaked in the nearinfrared (NIR) at 𝜆𝜆 = 1270 𝑛𝑛𝑛𝑛.
In recent years, several nanostructures capable of generating reactive oxygen species (ROS) when activated bypenetrating radiation (X-rays, NIR light) have been developed to apply Photodynamic Therapy (PDT) to tumours indeep tissue, where visible light cannot penetrate1-3; these nanostructures combine a scintillating nanostructure and a
PDT photosensitizer. A bottleneck in their characterization is the lack of a fast and reliable technique to quantitatively
assess their performances in generating ROS, and in particular 1O2. For instance, the widely used PDT “Singlet
Oxygen Sensor Green” kit suffers from self-activation under X-ray irradiation.
To solve this difficulty, we propose here direct detection of 1O2 by a spectroscopic technique, thanks to recently
developed thermoelectrically cooled InGaAs single photon avalanche photodiodes (SPAD).
We couple an InGaAs SPAD to a custom-made integrating sphere. By calibrations on standard photosensitizers, thedetection threshold for our apparatus was determined: it turns to be ∼ 4 ⋅ 109 1O2 in realistic experimental condition
and for measurements extending to 1 minute of integration4.
Finally, we report on preliminary results of 1O2 generated by nanostructures for PDT activated by 6MeV X-rays
generated by a Radiotherapy Linear Accelerator of clinical use.

Fig. 1 – A nanostructure combining scintillating nanoparticles and a PDT photosensitizer generates 1O2 when irradiated byhigh-energy X-rays. 1O2 is detected by measuring its near-infrared emission at 1270nm using a specifically developed portable
NIR spectrometer.

References
1) T. Rimoldi et al. J. Mater. Sci. Mater. Med. 27, 159, 20162) D. Orsi et al. Nanomedicine. 13 2311–2326, 2018
3) F. Rossi et al. Sci. Rep. 5 1-6, 20154) D. Orsi et al, submitted, 2021
Acknowledgements. This work was supported by the programme “FIL-Quota Incentivante” of University of Parma,co-sponsored by Fondazione Cariparma.

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OC-3.4.10

Uncovering the potency of PSI-ALA-Hex as a fluorescence-guided surgery tool forbreast cancer
Martin Kiening, Norbert Lange
School of Pharmaceutical Sciences, Institute of Pharmaceutical Sciences of Western Switzerland, University ofGeneva, Switzerland
In 2018, breast cancer (BC) was the most commonly diagnosed cancer in women with more than 2 million new cases
and the first cause of female cancer death (>625,000) 1. Mammography being the only effective screening method,
researches started to extend photodiagnosis (PD) to breast cancer.
Protoporphyrin IX (PpIX) is a natural molecule whose production has been widely browsed for photodynamic
therapy (PDT) and PD applications due to its photosensitizing potential. Synthesis of this ultimate heme precursor in
mitochondria can be boosted following the intake of 5-aminolevulinic acid (5-ALA), a former molecule in the heme
biosynthesis pathway. However, the original hydrophilic 5-ALA toughly cross plasma membranes and showed apoor pharmacokinetic profile.
Designing more lipophilic derivatives such as methyl- and hexyl-5-ALA esters countered the 5-ALA uptake issue.
PSI-ALA-Hex, a new phosphatase-sensitive 5-ALA ester performed even better by its reduced acute toxicity andhigher stability2.

Fig. 1 – 5-Aminolevulinate (5-ALA), 5-ALA hexyl ester (HAL), phosphatase-sensitive prodrug of HAL (PSI-ALA-Hex)

In this study, we investigated in vitro PpIX production levels in BC spheroids treated with such 5-ALA derivatives.
Four BC cell lines were chosen to represent main BC tumour subtypes: MCF7 (luminal A), BT-474 (luminal B),
SKBR3 (HER2+) and MDA-MB-231 (Triple Negative (TN)). Cells were seeded in ultra-low attachment 96-wellplates at 5,000 cells/well in 3.5% matrigel and spheroids settled for 4 days. They were subsequently treated with
HAL or PSI-ALA-Hex at 0.033 / 0.1 / 0.33 or 1mM. Red PpIX fluorescence level was monitored over 3 days in an
Incucyte S3 live-cell imager and analysed with the Incucyte spheroid module.
In MDA-MB-231, PSI-ALA-Hex showed similar PpIX levels but a slightly faster elimination compared to HAL.
Fluorescence production was dose-dependent. MCF7 profiles of both molecules were highly similar, showing nodose-dependency. Skbr3 spheroids displayed an unusual profile with a fluorescence production phase in two times.
A break was observed at 6h, after which the fluorescence production rate lessend until reaching the peak. The lowest
concentrations 0.033 and 0.1 mM performed respectively the same with both molecules. However, PSI-ALA-Hexinduced fluorescence did not compete with HAL at 1mM.
Our results display huge discrepancies from one cell line to another that may be due to differences in enzyme
expression of the heme biosynthesis pathway. In order to balance a proper fluorescence intensity for tumour resection
and a rapid elimination of the photosensitive compound in most tumour types, an intermediate concentration of
0.1mM is proposed. PSI-ALA-Hex, a stable derivative of the well-studied and already marketed HAL is unveiling a
potential for systemic administration and wider tumour detection.

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References
1) Bray F, Ferlay J, Soerjomataram I, et al. Global cancer statistics 2018: GLOBOCAN estimates of incidence
and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin. 2018;68(6):394-424.
2) Herceg V, Lange N, et al. Activity of phosphatase-sensitive 5-aminolevulinic acid prodrugs in cancer celllines. J Photochem Photobiol B. 2017;171:34-42.
Acknowledgements. This work was supported by a grant of the Swiss National Science Foundation.

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OC-3.4.11

New porphyrin conjugates for the treatment of TNBC.
Miryam Chiara Malacarne1, Paolo Dognini2, Enrico Caruso1, Marzia Bruna Gariboldi1 and Francesca Giuntini2.
1) Department of Biotechnology and Life Sciences (DBSV), University of Insubria, Italy; 2) School of Pharmacy and
Biomolecular Science, Liverpool John Moores University, United Kingdom.
Triple Negative Breast Cancer (TNBC), an aggressive variant of breast cancer, is characterized by lack of expressionof the oestrogen (ER) and progesterone receptors (PRs) and the human epidermal growth factor receptor (EGFR) thatare commonly observed in other breast cancer subtypes. Treatment options are limited and for this reason TNBC is
an important area of research1. The association of photosensitizers (PSs) with peptides has been extensively exploited
to improve the performance of porphyrin-based anticancer photosensitizing agents2. Specific and successful targetingof a PS to TNBC would allow developing a more selective therapeutic approach based on photodynamic therapy(PDT).
This project aims at generating porphyrin-peptide conjugates targeting TNBC cells via a novel approach relying on
the aromatic displacement of a suitable leaving group on the porphyrin with a thiol group present on the peptide.

5,15-bis(4-carbomethoxyphenyl)porphyrin was synthetized via acid-catalyzed mixed condensation. This compound
was subsequently modified in the meso positions to insert a nitro group. A dodecapeptide that binds specifically to
EGFR was used as a cancer cell targeting peptide3. We used two other dodecapeptide sequences provided with one
cysteine residue at the N-terminus to obtain the desired conjugate. The conjugation of the peptides to the porphyrins
was carried out by aromatic nucleophilic substitution of nitro groups. Thiol groups are excellent nucleophiles, andtheir reactivity is known to take place under mild conditions compatible with the integrity of the delicate peptide
moiety. Before proceeding with the formation of the peptide-porphyrin conjugate, preliminary tests were carried out
to evaluate the reactivity of the nitro groups present in the meso position. These tests were performed using Nacetylcysteine. LC-MS analyses have shown the formation of the desired conjugate already after one-hour reaction.
This result provides positive information on the subsequent step for the formation of peptide-porphyrin conjugate, a
new class of compounds with high potential as PSs and as light-responsive materials.
The effects of the peptide-porphyrin conjugates on cell viability, along with the percentage of apoptotic and necrotic
cells, the production of ROS and 1O2 and, the cellular uptake, were evaluated on 4 breast cancer cell lines (MCF-7,
T74D and the TNBC lines MDA-MB-231 and MDA-MB-453), following 24h treatment with the PSs, exposure for
2h to a white light irradiation and 24h incubation in drug-free medium.
References
1) C.L. Griffiths, J.L. Olin, Triple Negative Breast Cancer: A Brief Review of its Characteristics and Treatment
Options. Journal of Pharmacy Practice, 2012.
2) F. Biscaglia, M. Gobbo, Porphyrin–peptide conjugates in biomedical applications. Peptide Science, 2018.
3) H. Hossein-Nejad-Ariani, E. Althagafi, K. Kaur, Small Peptide Ligands for Targeting EGFR in Triple Negative
Breast Cancer Cells. Scientific Reports, 2019.

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OC-3.4.12
Interrogating the physical and therapeutic attributes of NIR active moleculartargeted photonanomedicines in solid tumorsa)emedium EGFR

no EGFRb)high EGFR

Lorem ipsum

Lorem ipsum

mt-PNM specificity
0

1

2

3

Relative selectivity
or specifcity (/9L tumor)

Please leave this line empty as spacer - delete this text. Font is Times New Roman 11
Girgis Obaid,1,2 Kimberly Samkoe,3 Shazia Bano,2 Kenneth Tichauer,4 Brian Pogue3 and Tayyaba Hasan2,5Presenting author underlined
1) Department of Bioengineering, University of Texas at Dallas, Richardson, TX 75080, United States; 2) Wellman
Center for Photomedicine, Massachusetts General Hospital, Boston, MA 02114, United States; 3) Thayer School ofEngineering, Dartmouth College, Hanover, NH 03755, United States; 4) Illinois Institute of Technology, Chicago,
IL 60616, United States; 5) Division of Health Sciences and Technology, Harvard University and Massachusetts
Institute of Technology, Cambridge, MA 02139, United States
Please leave this line empty as spacer - delete this text. Font is Times New Roman 11
Molecular targeted photonanomedicines (mt-PNMs) are receptor-specific nanoplatforms that facilitate photodynamic
therapy-based treatment of cancer. Targeting solid tumors with mt-PNMs is particularly complex due to highlyvariable in vivo pharmacokinetics coupled with significant variations in the functionality of entrapped photosensitizer
molecules.1 This work interrogates the molecular specificity (true receptor recognition) of NIR activable mt-PNMs
in EGFR-overexpressing solid tumors using quantitative molecular imaging.2 Results reveal that molecularspecificity of mt-PNMs (as measured by molecular imaging) is distinct from tumor selective uptake, which has
traditionally been the conventional approach to predict mt-PNM success (Fig. 1). 1,2 Furthermore, we demonstratethat molecular specificity of mt-PNMs is entirely responsible for photodynamic therapy-based tumor destruction,
reversal of desmoplasia in pancreatic tumor xenografts and improvements in treatment tolerability.3 Emerging multispecific mt-PNMs also have demonstrate the capacity to address tumor receptor heterogeneity: the Achille’s Heel ofmolecular targeted therapies.4 The approaches presented here provide a rational, informed tumor biology-driven
approach to mt-PNM nanoengineering to increase their chances of ultimately increasing treatment efficacy andtolerability in cancer patients.
specificity
selectivity

1069.8% **

5

94.2%
0

0

2 104

4 104

EGFR expression (receptors per cell)Fig. 1. a) Specificity imaging
of EGFR-targeted mt-PNMs
in 9L (no EGFR), U87(medium EGFR) and U251
(high EGFR) tumors. b)
traditional
measuresofselectivity
grossly
underrepresent
EGFR
specificity of mt-PNM.

References
1) Bhandari, C., Guirguis, M., Savan, N.A., Shrivastava, N., Oliveira, S., Hasan, T. and Obaid, G. What NIR photodynamic
activation offers molecular targeted nanomedicines: Perspectives into the conundrum of tumor specificity and selectivity.
Nano Today, 2020, 36, p.101052.
2) Obaid, G.*, Samkoe, K.*, Tichauer, K., Bano, S., Park, Y., Silber, Z., Hodge, S., Callaghan, S., Guirguis, M., Mallidi,
S. and Pogue, B. and Hasan, T. Is tumor cell specificity distinct from tumor selectivity in vivo? A quantitative NIR
molecular imaging analysis of nanoliposome targeting. Nano Research, 2020, pp.1-11.
3) Obaid, G., Bano, S., Mallidi, S., Broekgaarden, M., Kuriakose, J., Silber, Z., Bulin, A.L., Wang, Y., Mai, Z., Jin, W.
and Simeone, D. Impacting Pancreatic Cancer Therapy in Heterotypic in Vitro Organoids and in Vivo Tumors with
Specificity-Tuned, NIR-Activable Photoimmunonanoconjugates: Towards Conquering Desmoplasia? Nano Letters, 2019.
19 (11), pp.7573-7587
4) Bano, S.*, Obaid, G.*, Swain, J.W., Yamada, M., Pogue, B.W., Wang, K. and Hasan, T. NIR Photodynamic Destructionof PDAC and HNSCC Nodules Using Triple-Receptor-Targeted Photoimmuno-Nanoconjugates: Targeting Heterogeneity
in Cancer. Journal of clinical medicine, 2020, 9(8), p.2390.
Acknowledgements. This work was supported by the National Institutes of Health (K99CA215301 and
R00CA215301 to G.O.; R37CA212187 to K.S.; and P01CA084203, R01CA156177, R01CA160998, R21CA220143
to T.H.).
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IL-4.1.1

Photochemistry of DNA damages
Virginie Lhiaubet-Vallet
Instituto Universitario Mixto de Tecnología Química (UPV-CSIC), Universitat Politècnica de València, Consejo
Superior de Investigaciones Científicas, Avenida de los Naranjos s/n, 46022 Valencia, Spain.
The natural DNA bases dissipate more than 90% of their excitation energy through efficient nonradiative channels
leading to the ground state, which explains their high resistance to UV excitation. Nevertheless, small structuralchanges can result in the lengthening of excited states lifetime and/or in the increase of intersystem crossing
efficiency and thus, can modify drastically the photochemical properties of DNA.
In this context, the photochemistry of lesions is of utmost importance as some of them are able to absorb in the UVAUVB region and behave as a potential intrinsic photosensitizer. Here, we will discuss the photophysical andphotochemical properties of damages such as the (6-4) photoproducts1-3 or 5-formylpyrimidine derivatives5-6 (seeFig. 1) to evaluate if their fulfil the basic requirements of a good DNA photosensitizer: (i) to absorb in the UVAUVB region, (ii) to populate efficiently their triplet excited state and (iii) to be able to interact with DNA components
through a Type I or II process and/or a triplet-triplet energy transfer.

Fig. 1 – Structures of the photolesions considered for their DNA photosensitizing effects.

References
1) V. Vendrell-Criado, G. M. Rodríguez-Muñiz, V. Lhiaubet-Vallet, M. C. Cuquerella, M. A. Miranda, The (6–4)
dimeric las a DNA photosensitizer. ChemPhysChem, 17, 1979-1982, 2016.
2) V. Vendrell-Criado, G. M. Rodriguez-Muniz, M. C. Cuquerella, V. Lhiaubet-Vallet, M. A. Miranda,
Photosensitization of DNA by 5-methyl-2-pyrimidone deoxyribonucleoside: (6-4) photoproduct as a possible Trojan
horse. Angew. Chem. Int. Ed., 52, 6476–6479, 2013.
3) I. Aparici-Espert, G. Garcia-Lainez, I. Andreu, M. A. Miranda, V. Lhiaubet-Vallet, Oxidatively generated lesions
as internal photosensitizers for pyrimidine dimerization in DNA. ACS Chem. Biol. 13, 542–547, 2018.
4) A. Frances-Monerris, C. Hognon, M. A. Miranda, V. Lhiaubet-Vallet, A. Monari, Triplet photosensitization
mechanism of thymine by an oxidized nucleobase: from a dimeric model to DNA environment. Phys. Chem. Chem.
Phys. 20, 25666–25675, 2018.
5) A. Francés-Monerris, M. Lineros-Rosa, M. A. Miranda, V. Lhiaubet-Vallet, A. Monari, Photoinduced intersystemcrossing in DNA oxidative lesions and epigenetic intermediates. Chem. Commun., 56, 4404–4407, 2020.
6) M. Lineros-Rosa, A. Frances-Monerris, A. Monari, M. A. Miranda, M.A.; Lhiaubet-Vallet, Experimental andtheoretical studies on thymine photodimerization mediated by oxidatively generated DNA lesions and epigenetic
intermediates. Phys. Chem. Chem. Phys. 22, 25661–25668, 2020.
Acknowledgements. This work was supported by the Spanish government (project PGC2018-096684-B-I00).
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IL-4.1.2

DNA damage and molecular modelling: Mechanistic aspects revealed by multiscale
simulations
Antonio Francés-Monerris1
1) Universitat de València, Departament de Química Física, 46100 Burjassot (València), Spain
DNA damage is a fundamental process of interest in chemistry, biology, and medicine. Among the different agentsthat can harm DNA/RNA, light represents a powerful aggressor that delivers the damage either by populating reactive
excited states localized in the nucleic acids or by producing reactive oxygen species (ROS) in the surroundings that
ultimately oxidize the DNA structure. Apart from the fundamental interest of DNA damage as the molecular basisof diseases such as skin cancer, and its role in the understanding of the natural DNA photostability, these processes
also represent an opportunity of treatment. In this context, photodynamic therapy (PDT) is a technique that uses light
to selectively kill ill tissues, in most cases by inducing DNA damage mechanisms activated by irradiation.
The scientific community uses a plethora of experimental techniques to study DNA damage. The underlying
processes have a wide variety of spatial and temporal scales: from the very first events that take place after lightabsorption, tracked with ultrafast transient spectroscopy, to the quantification of cellular damage, studied withcellular assays, the scientific community is progressively revealing DNA damage mechanisms in greater detail. In
this scenario, molecular modelling offers sophisticated methodologies that allow the full description of DNA damage
events from chemical, photochemical, and photobiological perspectives. In the present talk, I will highlight recent
advances in classical molecular dynamics (MD), quantum-mechanical (QM) methods and hybrid QM/MM schemesthat have opened the door to unprecedented descriptions of DNA damage and repair.1–3 I will illustrate the advanceof the field with recent examples, including the discovery of multiple inter-strand proton transfers induced by light,4the formation of photoinduced DNA lesions in bacteria,5 the phototoxic potential of oxidized DNA nucleobases6–8
and the mode of action of non-conventional PDT agents.9
References(1)
Francés-Monerris, A.; Gillet, N.; Dumont, E.; Monari, A. DNA Photodamage and Repair: Computational Photobiology in
Action. In QM/MM Studies of Light-responsive Biological Systems; Springer, 2021; pp 293–332.
(2)
Francés-Monerris, A.; Marazzi, M.; Besancenot, V.; Grandemange, S.; Assfeld, X.; Monari, A. Computational Spectroscopy
and Photophysics in Complex Biological Systems: Towards an In Silico Photobiology. In Computational Techniques forAnalytical Chemistry and Bioanalysis; The Royal Society of Chemistry, 2021; pp 202–246.
(3)
Roca-Sanjuán, D.; Carmona-García, J.; Ding, B.-W.; Navarrete-Miguel, M.; Francés-Monerris, A.; Giussani, A.; SegarraMartí, J.; Monari, A.; Liu, Y.-J. Photochemistry, Chemiluminescence and Dark Photochemistry: Computational Advances(2018–2019). In Photochemistry: Volume 48; The Royal Society of Chemistry, 2021; Vol. 48, pp 24–70.
(4)Francés-Monerris, A.; Gattuso, H.; Roca-Sanjuán, D.; Tuñón, I.; Marazzi, M.; Dumont, E.; Monari, A. Dynamics of the
Excited-State Hydrogen Transfer in a (DG)·(DC) Homopolymer: Intrinsic Photostability of DNA. Chem. Sci. 9, 7902–7911,
2018.
(5)
Francés-Monerris, A.; Hognon, C.; Douki, T.; Monari, A. Photoinduced DNA Lesions in Dormant Bacteria. The Peculiar RouteLeading to Spore Photoproduct Characterized by Multiscale Molecular Dynamics. Chem. Eur. J. 26, 14236–14241, 2020.
(6)Francés-Monerris, A.; Hognon, C.; Miranda, M. A.; Lhiaubet-Vallet, V.; Monari, A. Triplet Photosensitization Mechanism ofThymine by an Oxidized Nucleobase: From a Dimeric Model to DNA Environment. Phys. Chem. Chem. Phys. 20 (40), 25666–
25675, 2018.
(7)
Francés-Monerris, A.; Lineros-Rosa, M.; Miranda, M. A. A.; Lhiaubet-Vallet, V.; Monari, A. Photoinduced Intersystem
Crossing in DNA Oxidative Lesions and Epigenetic Intermediates. Chem. Commun. 56, 4404–4407, 2020.
(8)
Lineros-Rosa, M.; Francés-Monerris, A.; Monari, A.; Miranda, M. A.; Lhiaubet-Vallet, V. Experimental and Theoretical
Studies on Thymine Photodimerization Mediated by Oxidatively Generated DNA Lesions and Epigenetic Intermediates. Phys.
Chem. Chem. Phys. 22, 25661-25668, 2020.
Francés-Monerris, A.; Tuñón, I.; Monari, A. Hypoxia-Selective Dissociation Mechanism of a Nitroimidazole Nucleoside in a
(9)
DNA Environment. J. Phys. Chem. Lett. 10, 6750–6754, 2019.

Acknowledgements. This work was supported by the postdoctoral contract APOSTD/2019/149 and the project
GV/2020/226 from the Generalitat Valenciana and the European Social Fund.
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IL-4.1.3

Photoactivated behavior of Guanine-rich DNA Quadruple Helices
James Green1, Haritha Asha1, Lara Martinez-Fernandez2, Dimitra Markovitsi,3 Roberto Improta1
1) Istituto Biostrutture e Bioimmagini- Consiglio Nazionale delle Ricerche, Napoli, Italy 2) Departamento deQuímica,Universidad Autónoma de Madrid, Madrid, Spain 3) LIDYL, CEA, CNRS, Université Paris-Saclay,
FranceGuanine-rich DNA sequences can adopt a peculiar fold, a Quadruple helix, (G4) where four Guanine (G) bases arearranged in planar structures, the tetrads, stabilized by Hoogsteen-type hydrogen bonds,1 involved in key cellular
processes and have emerged as promising therapeutic targets. G4 exhibits peculiar absorption, emission andcircular dichroism spectra,2 and we show that Quantum Mechanical calculations can provide spectra in fairagreement with the experiments and crucial information for their assignment and interpretation.3-7 Both the staticand dynamical properties of G4 excited states are ruled by their topology. In general, after light absorption fromexcited states delocalized over multiple bases, the most important decay pathways involve localization of theexcitation over a single base or on two stacked guanines, excimers with different degrees of charge transfer
character. The two main photochemical reactions involve the photodimerization of two stacked guanine bases onthe ‘neutral’ excimer path and the ionization of guanine.4,6 Finally, we shall describe a general and flexible
approach, based on fragment diabatization, which incorporates charge transfer states and significantly increases thereliability of excitonic Hamiltonians for systems where the chromophores are very close. This model (FrDEx) is
used to compute the electronic circular dichroism and absorption spectra of several prototype G4, allowing
significant improvements with respect to “standard” excitonic Hamiltonians and giving access to interesting
insights into the chemical–physical effects modulating the spectral signals.
References
1) R. Hänsel-Hertsch et al Nature Rev. Mol. Cell Biol. 2017, 18, 279.
2) P. Changenet-Barret, et al. Electronic Excitations in Guanine Quadruplexes in Photoinduced Phenomena in
Nucleic Acids II: Springer International Publishing: Cham, 2015; 183.
3) R. Improta Chem. Eur. J. 2014, 20, 8106.
4) A. Banyasz,et al. J.Am. Chem. Soc 2017, 139, 10561.
5) L. Martínez-Fernández et al. Chem. Eur. J. 2018, 24, 15185.
6) L Martinez-Fernandez, et al J. Phys. Chem. Lett. 2019, 10, 6873
7) L. Martínez-Fernández, et al Photochem. Photobiol. Sci. 2020, 19, 436-444
8) J. Green, et al. J. Chem. Theory Comput. 2021, 17, 1, 405.
Acknowledgements. This work has received funding from the European Union\’s Horizon 2020 research andinnovation programme MSCA-ITN under grant agreement No. 765266 (LightDyNAmics)

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OC-4.1.4

Transfection of keratinocytes with in vitro synthesized CPD-specific photolyaseencoding mRNA is a model system to study the CPD-dependent cellular effects of
UVB
Csaba Hegedűs1, Eszter Fidrus1, Gábor Boros2, Eszter Anna Janka1, Gabriella Emri1, Katalin Karikó2, Éva
Remenyik1
1. University of Debrecen, Department of Dermatology, Debrecen, Hungary

2. BioNTech, Mainz, Germany

Cyclobutane pyrimidine dimer (CPD)-specific photolyase removes UVB-induced CPDs from DNA using the energyof visible light in a process called photoreactivation. However, this repair mechanism disappeared from placental
mammals during evolution. Previously we established a model system based on transient transfection of humankeratinocytes with codon-optimized in vitro transcribed mRNA containing 1-methylpseudouridine modifications that
encodes CPD-specific photolyase (CPD-PL mRNA)1. The RNA composition is similar to that used for BioNTech
COVID-19 vaccine. Following UVB irradiation HaCaT and human primary keratinocytes were either kept in thedark or exposed to photoreactivating light. Different cellular events were studied in a time-dependent manner.
Our results show that keratinocytes express functional photolyase upon transfection of CPD-PL mRNA. The encoded
protein starts to accumulate in the nucleus 5-8 hours post-transfection. CPDs were effectively removed byphotoreactivation immediately as well as 6 hour after CPD-PL mRNA transfection resulted in relieving the negative
effects of UVB on cell viability. Activation of photolyase prevented the loss of cell proliferation and G2/M cell cycle
block after UVB irradiation. The functional photolyase also diminished other UVB-mediated effects, including
enhanced IL-6 mRNA expression. Microarray analyses demonstrated that gene expression pattern is altered by UVB
induced CPDs. Using our model system, it has been demonstrated that CPDs are responsible for production of
mitochondrial reactive oxygen species followed by the activation of several energy sensor enzymes, andcompensatory metabolic changes in keratinocytes exposed to UVB. Interestingly, CPDs could be removed not only
from nuclear genome but from mitochondrial genome as well and restored mitochondrial DNA copy number
suggesting that damage to mitochondria can also be prevented by photolyase activation. Finally, UVB-induced
mutagenesis was completely abrogated by photoreactivation emphasizing the key role of CPDs in mediating DNA
damage and carcinogenesis.
These results suggest that activation of a non-human photolyase encoded by nucleoside-modified mRNA is able toprevent the UVB-induced cellular damages and introduction of photolyase mRNA into the skin may have a future
therapeutic potential.
References
1) Boros G, Miko E, Muramatsu H, Weissman D, Emri E, Rózsa D, Nagy G, Juhász A, Juhász I, van der Horst G,
Horkay I, Remenyik É, Karikó K, Emri G. Transfection of pseudouridine-modified mRNA encoding CPDphotolyase leads to repair of DNA damage in human keratinocytes: a new approach with future therapeuticpotential. J Photochem Photobiol B. 2013 Dec 5;129:93-9.
Acknowledgements. This work was supported by the European Union and the European Regional Development
Fund GINOP-2.3.2-15-2016-00005 and Hungarian Scientific Research Fund NKFIH K120206.

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IL-4.1.5

The triplet state as the precursor of the thietane intermediate in the formation of the
DNA 6-4 photoadductCarlos E. Crespo-Hernández,
Case Western Reserve University, Department of Chemistry, Cleveland, Ohio, 44106, USA
Notwithstanding the central biological role of the (6-4) photoadduct in the induction of skin cancer by sunlight,
crucial mechanistic details about its formation have evaded characterization despite efforts spanning more than halfa century1. 4-Thiothymidine (4tT) has been widely used as an important model system to study its mechanism of
formation2, but the excited-state precursor, the intermediate species, and the time scale leading to the formation ofthe (6-4) photoadduct have remained elusive. In this talk, I will present experimental and computational evidence
elucidating the excited state leading to the formation of the thietane intermediate, its rate, and the formation of the(6-4) photoadduct in the 5’-TT(4tT)T(4tT)TT-3’ single-stranded DNA oligonucleotide. It will be demonstrated that
efficient, sub-1 ps intersystem crossing leads to the population of a triplet minimum of the thietane intermediate inas short as 3 ps, which intersystem crosses to its ground state and rearranges to form the (6-4) photoadduct1.

Fig. 1 – The colored arrows represent electronic transitions. A 4tT base is highlighted with light orange in the single-stranded
DNA sequence depicted to the left to imply that 4tT bases are selectively photoactivated by ultraviolet-A radiation. 4tT and thethymidine 5′ relative to the 4tT are represented as tube in the single-stranded DNA sequence, while the other nucleobases are
depicted as wireframe to highlight the importance of having a thymine 5′ relative to the 4tT for the reaction to occur. In thechemical structures, yellow, red, blue, gray, and white, represent sulfur, oxygen, nitrogen, carbon, and hydrogen, respectively.

References
1) Ortiz-Rodríguez, L. A.; Reichardt, C.; Hoehn, S. J.; Jockusch, S.; Crespo-Hernández, C. E., Detection of thethietane precursor in the UVA formation of the DNA 6-4 photoadduct, Nature Commun. 11, 3599 (2020).
2) Clivio, P., Fourrey, J. L., Gasche, J. & Favre, A. DNA photodamage mechanistic studies: characterization of athietane intermediate in a model reaction relevant to “6-4 lesions”. J. Am. Chem. Soc. 113, 5481–5483 (1991).
3) Liu, J. & Taylor, J.-S. Remarkable photoreversal of a thio analog of the Dewar valence isomer of the (6-4)
photoproduct of DNA to the parent nucleotides. J. Am. Chem. Soc. 118, 3287–3288 (1996).

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4) Warren, M. A., Murray, J. B. & Connolly, B. A. Synthesis and characterization of oligodeoxynucleotides
containing thio analogues of (6-4) pyrimidine-pyrimidinone photo-dimers. J. Mol. Biol. 279, 89–100 (1998).
5) Reelfs, O. et al. Identification of potentially cytotoxic lesions induced by UVA photoactivation of DNA 4thiothymidine in human cells. Nucleic Acids Res. 39, 9620 (2011).
Acknowledgements. The authors acknowledge the National Science Foundation (Grant No. CHE-1800052). This
work made use of the High-Performance Computing Resource in the Core Facility for Advanced Research
Computing at CWRU.

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IL-4.1.6
In silico insight on the photophysics of non-canonical nucleobases. Implications in
DNA photostability and damage
Inés Corral1
1) Universidad Autónoma de Madrid, Departamento de Química, Cantoblanco, Madrid, Spain
Unnatural nucleobases, or heterocycles structurally differing from their five natural counterparts in different extents,
have captured the interest of the scientific community in recent years for their potential implications as prebiotic
ancestors of the current genetic letters1 or in biological or biotechnological applications, by conceding chemical andstructural diversity to DNA and RNA strands.2
Albeit minor, these structural modifications can dramatically condition the intra-, inter-, and macromolecular
structure or the photophysics of the genetic polymers. Thus, the self-assembly ability, and the chemical and lightstability of these systems can be considered as key selection pressures determining their viability in contemporary
experimental and primordial biopolymers.
The aim of this contribution is to provide an overview of the main photophysical properties of a selected group of
nucleobase derivatives, based on state-of-the-art quantum mechanical and molecular dynamics simulations including
full dimensionality. In particular, the impact in the optical properties and topographical features of the potential
energy landscapes of the position and the number of oxo and amino substituents, or of the replacement of carbonylby thiocarbonyl functions3 along the purine and pyrimidine cores will be discussed.
These results will help establishing the functionalization-photoactivity relationships in purine4 and pyrimidine5 cores,
and will contribute to determining the electronic and structural factors that established the superiority of the fiveDNA and RNA nucleobases against other organic heterocycles and to the rational design of unnatural biopolymers
allowing the coding flexibility necessary to synthetize new proteins.
References
1) A. C. Rios and Y. Tor, Israel Journal of Chemistry, 53, 469, 2013.
2) "Expansion of the Genetic Alphabet: Unnatural Nucleobases and Their Applications", Special Issue, Journal of
Nucleic Acids, 2012.
3) S. Arslancan, L. Martínez-Fernández, I. Corral, Molecules, 22, 998, 2017.
4) C. E. Crespo-Hernández, L. Martínez-Fernández, C. Rauer, C. Reichardt, S. Mai, M. Pollum, P. Marquetand, L.
González, I. Corral, The Journal of the American Chemical Society, 137, 4368, 2015.
5) E. M. Arpa, M. M. Brister, S. J. Hoehn, C. E. Crespo-Hernández, I. Corral, The Journal of Physical Chemistry
Letters, 11, 5156, 2020.
Acknowledgements. This work has been supported by the Ramón y Cajal program of the Ministerio de Economía
Industria y Competitividad of Spain, and the PGC2018-094644-B-C21 project and FPU and FPI grants from the
Ministerio de Ciencia, Innovación y Universidades of Spain. The Red Española de Supercomputación, and the Centro
de Computación Científica of the UAM (CCC-UAM) are also acknowledged for the generous allocation of computer
time and for their continued technical support.

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OC-4.1.7
Photoinduced azetidine decomposition reaction by photo-oxidation and photoreduction: Inverting the aza-Paternò-Büchi reaction
Miriam Navarrete Miguel1, Antonio Francés-Monerris2, Daniel Roca-Sanjuán1
1) Instituto de Ciencia Molecular, Universitat de València, P.O. Box 22085, ES-46071 València, Spain; 2)
Departamento de Química Física, Universitat de València, 46100 València, Spain
DNA in living beings is constantly damaged by both exogenous and endogenous agents, such as UV radiation. The
photoinduced DNA lesions can be initiated by Paternò–Büchi, or aza-Paternò-Büchi, photocycloaddition between
two adjacent pyrimidine bases, forming an oxetane, or an azetidine ring.
Then, the repair mechanism of these lesions can be represented with the inversion of the Paternò-Büchi reaction.
Some lesions can be repaired by photolyases also in presence of light and a photosensitizer via a photoprocess
whose molecular mechanism is not completely understood yet. In this contribution, we have focused on the azaversion of the Paternò-Büchi reaction. Then, two azabipyrimidinic azetidines have been studied as modelcompounds of the elusive azetidine intermediates present during the (6-4) photoproducts repair. The first one is
obtained by photocycloaddition between 6-azauracil and thymine (AZT-Thy), and the second one arises from thephotocycloaddition between 6-azauracil and cyclohexene (AZT-CH). DFT calculations have allowed to interpretthe experimental observations. In addition, they also help us to elucidate the ring-opening mechanisms during
photoreduction and photo-oxidation processes. On the one hand, regarding the AZT-Thy model, energy barriers
lower than 13 kcal mol-1 are obtained for both the photoreduction and photo-oxidation processes1. On the other
hand, regarding the AZT-CH system, energy barriers lower than 14 and 9 kcal mol-1 for the cis- and trans- isomers
are obtained for photoreduction, while values of about 36 kcal mol-1 are obtained for both isomers in the photooxidation process2.
References
1) Ana B. Fraga-Timiraos, Antonio Francés-Monerris, Gemma M. Rodríguez-Muñiz, Miriam Navarrete-Miguel,Miguel Ángel Miranda, Daniel Roca-Sanjuán, Virginie Lhiaubet-Vallet, Chem. - A Eur. J. 2018, 24, 15346–15354.
2) Miriam Navarrete-Miguel, Antonio Francés-Monerris, Miguel Ángel Miranda, Virginie Lhiaubet-Vallet, Daniel
Roca-Sanjuán, Molecules 2021, 26, 2911.
Acknowledgements. This work was supported by the D. R.-S. “Ramon y Cajal” grant from the Spanish“Ministerio de Ciencia e Innovación (MICINN)” (ref. RYC-2015-19234). A.F.-M. is grateful to the Région GrandEst government (France) and the Université de Lorraine for their financial support. This work has also been fundedby the Generalitat Valenciana and the European Social Fund through the postdoctoral contract APOSTD/2019/149
and the project GV/2020/226. It also was funded by the MICINN project CTQ2017-87054-C2-2-P, and a 2019
Leonardo Grant for Researchers and Cultural Creators, BBVA Foundation. M.N.-M. acknowledges the Generalitat
Valenciana for the predoctoral grant (Ref. ACIF/2020/075). Calculations were performed on the local QCEXVAL
cluster of the University of Valencia and on the LluisVives and Tirant III clusters of the Servei d’Informàtica
(University of Valencia).

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OC-4.1.8
An insight into etheno adducts optical properties.
Paloma Lizondo-Aranda1, Thomas Gustavsson2, Miguel Ángel Miranda1, Virginie Lhiaubet-Vallet1.
1) Instituto Universitario Mixto de Tecnología Química (UPV-CSIC), Universitat Politècnica de València, Avda delos Naranjos, s/n., 46022, Valencia, Spain.; 2) Laboratoire Interactions, Dynamiques et Lasers (LIDYL) CEA/Saclay, 91191 Gif-sur-Yvette, France.
The study of DNA damages has increased during the last decades in order to get more insight into theirinvolvement in the appearance of cancer.1 Among the large number of DNA lesions, etheno adducts have been thematter of interest because of their presence in chronically inflamed human tissues, making their quantification
useful as potential biomarkers for cancer of colon, prostate, lung, etc. Moreover, these lesions exhibit highly
mutagenic properties and induce base transitions or transversion in mammal cells.2 Etheno adducts are mainly
formed endogenously as a result of lipid peroxidation. This biochemical process produces reactive aldehydes suchas malondialdehyde (MDA), which can combine with DNA bases creating the exocyclic ring.

ETHENO ADDUCTS(BASE-MODIFIED NUCLEOSIDES )Norm. Fluorescence Intensity

1,0

m1dG
edC
edA
edG

0,8

0,6

0,4

0,2

0,0
250

300

350

400

450

500

550

Wavelength (nm)

600

650

700

0,07

t= 0.74
t= 0.60t= 1.80t= 1.40t= 1.00
t= 0.84

0,06

DOD

0,05

e

e

0,04
0,03
0,02
0,01

e

0,00
300 350 400 450 500 550 600 650 700 750 800 850
Wavelength (nm)Figure 1. Etheno adduct structures and some spectra from the present work.
Photochemically, the etheno derivatives are of great interest since they present an extended - conjugated systemthat might confer them optical properties different from those of the canonical bases and open the way to aparticular photoreactivity of the damaged DNA. This work presents a complete spectroscopic study of the ethenodamages combining steady-state and time-resolved fluorescence experiments together with transient absorption
studies. The obtained results demonstrate an ultrafast deactivation of their excited states, being however somewhat
slower than that of their respective canonical base.
References
1) J. Kaya, E. Thadhania, L. Samsona, B. Engelwarda, Inflammation-induced DNA damage, mutations and cancer.
DNA repair, 2019.
2) B. Hwa Yun, J. Guo, M. Bellamri, R.J. Turesky, DNA adducts: Formation, biological effects, and new
biospecimens for mass spectrometric measurements in humans. Mass spectrometry reviews, 2018.
Acknowledgements. This work was supported by Spanish Government (PGC2018-096684-B-I00 project, grantBES-2016-077517)

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SP-4.1.9, P-4.1.9

Risk assessment of irradiating skin models at 233 nm using far-UVC LEDs foreradiacation of MRSA and MSSA
Johannes Schleusener1, Paula Zwicker2, Silke B. Lohan1, Loris Busch1,3, Claudia Sicher2, Sven Einfeldt4, Michael
Kneissl4,5, Anja A. Kühl6, Cornelia M. Keck3, Christian Witzel7, Axel Kramer2 and Martina C. Meinke1
1) Charité – Universitätsmedizin Berlin, Department of Dermatology, Venerology and Allergology, Berlin,
Germany; 2) University Medicine Greifswald, Institute of Hygiene and Environmental Medicine, Greifswald,
Germany; 3) Philipps-Universität Marburg, Department of Pharmaceutics and Biopharmaceutics, Marburg,
Germany; 4) Ferdinand-Braun-Institut gGmbH, Leibniz-Institut für Höchstfrequenztechnik, Berlin, Germany; 5)Technische Universität Berlin, Institute of Solid State Physics, Berlin, Germany; 6) Charité – Universitätsmedizin
Berlin, iPATH.Berlin-Immunopathology for Experimental Models, Berlin, Germany; 7) Charité –Universitätsmedizin Berlin, Division of Plastic and Reconstructive Surgery, Berlin, Germany
Surgical Site Infections are mainly caused by the colonization of MRSA and MSSA and result in increased levelsof surgical morbidity and mortality. The eradication of these resistant pathogens is a continuing clinical problem, as
they cannot be reached by antiseptics in the depth of the skin.
UVC irradiation has been considered as a possible inactivation strategy in order to prevent infections during
surgery. Broad band UV sources with emission wavelengths between 200 and 400 nm are effectively bactericidal[1]. However, they can cannot be used in vivo, as they cause DNA damage in the skin. Recently, far-UVC-lamps at
222 nm could successfully be used for inactivation of bacteria. Due to the strong absorption of wavelengths below
240 nm, the radiation is mostly absorbed in the stratum corneum and does not reach the viable epidermis.
Therefore, DNA damage can effectively be eliminated.
Here, we present a study using a newly developed 233 nm far-UVC LED source [2] for the eradication of MRSA
and MSSA. Cell viability, DNA damage and radical production were assessed on different excised human skin andreconstructed human epidermis skin models in comparison to irradiation by conventional near-UVC (254 nm),
UVB (280–400 nm, positive control) and a conventional far-UVC radiation source at 222 nm.
At an effective bactericidal dose of 40 mJ/cm², where bacteria were reduced by 5 log10 levels, the investigated skinmodels showed no reduction in cell viability. The resulting DNA damage, which only occurred in the superficial
layer of the viable epidermis was below those caused by 0.1 minimal erythema dose (MED) UVB, which isregarded safe. This low damage disappeared after 24 h and irradiation on four consecutive days showed no
epidermal cells with DNA damage at this dose. We found a low radical formation load, which was far lower thanfor a dose equivalent to 20 min outdoor visible light, which can be compensated by the antioxidant defence system.
References
1) A. Kramer, M. Meinke, A. Patzelt, M.B. Stolpe, J. Lademann, K. Jacobs, Perspektiven physikalischer Verfahrender Antiseptik und Desinfektion, Hygiene und Medizin, 45;3/2020 D39.
2) M. C. Meinke, A Kramer, S Einfeldt, M. Kneissl, Krankenhauskeime mit UVC-Leuchtdioden bekämpfen,
Management und Krankenhaus 39; 9/2020, S.20
Acknowledgements. The work is funded by the German Federal Ministry of Education and Research BMBF (FKZ
03ZZ0146B) within the program “Zwanzig20 – Partnerschaft für Innovation” and carried out within the consortium
Advanced UV for Life

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SP-4.1.10, P-4.1.10
Photoprotection from UV light-induced telomere shortening and DNA damage by abroad-spectrum sunscreen productAna Guío-Carrión1, Almudena Martín2, José Aguilera3, María Matabuena2, Diego Megias4, Anne Laure DemessantFlavigny5, Isabel Castillejo6, Françoise Bernerd7, Leonor Prieto6, Maria A. Blasco1,*
1)Telomeres and Telomerase Group, Molecular Oncology Program, Spanish National Cancer Centre (CNIO),
Madrid, Spain; 2) Invitrotecnia Laboratory S.L., Alcobendas, Madrid, Spain; 3) Department of Medicine andDermatology, Faculty of Medicine, University of Málaga, Spain; 4) Microscopy Unit, Biotechnology Program,
Spanish National Cancer Research Centre (CNIO), Madrid, Spain; 5) International Scientific Direction, Laboratoire
Dermatologique La Roche-Posay, France; 6) Scientific Direction, Laboratoire Dermatologique La Roche-Posay,
L’ORÉAL Cosmética Activa, Madrid, Spain; 7)L’ORÉAL Research & Innovation, France
UV light is considered a class I carcinogen; it produces DNA damage, leading to photoaging, and in some cases
carcinogenesis. Upon UV-light exposure, the formation of photoproducts and reactive oxygen species highly toxic
for the cell induces the activation of DNA damage response (DDR) pathways that alter the metabolism of the cell,
ultimately inducing apoptosis or senescence. Telomeres are tandem repeats of the hexamer 5´-TTAGGG that cap andprotect the end of chromosomes. Telomeres are a prime target for UV-light induced damage due to their sequence
highly enriched in pyrimidines. Their length shortens upon UV irradiation in-vitro, compromising the regenerative
capacity and function of tissues, and telomeres in sun-exposed skin areas are shorter compared to telomeres innonexposed ones. Given the importance of telomere preservation in genome stability, we aimed to study whether abroad-spectrum sunscreen product prevents UV-induced DNA damage and telomere shortening.
We used a solar simulator lamp to provide 10J/cm2 SSUV to in-vitro cultured human keratinocytes and fibroblasts
and to a 3D skin model, and compared at 24h upon exposure among three conditions – nonirradiated control,
irradiated, and irradiated in the presence of a broad-spectrum sunscreen product, with a very high UVB and UVA
protection (SPF 50+ / PF UVA 46). We performed HT Q-FISH or FISH on skin slides to measure telomere length,gH2AX immunofluorescence as a readout for DNA damage, DAPI nuclei staining for cell viability, and hematoxylin& eosin staining of skin sections to examine skin structure.
Upon SSUV exposure, in-vitro cultured human keratinocytes and fibroblasts suffer from UV-induced damage, as
seen by reduced cell viability (41.6% and 79% respectively) and increased gH2AX and 53BP1, whereas, in thepresence of the sunscreen product, cell viability, gH2AX, and 53BP1 are comparable to controls. Significantly,
telomeres are shortened upon UV-light (26.8% and 13.5% of telomeric length loss respectively), while in the presenceof sunscreen the telomere length is partially preserved (shortening rate of 13.7% and 8.3% respectively) and the %of critically short telomeres is reduced. On the 3D skin model, SSUV exposure disrupts the skin structure with lossof skin layers stratification, loss of spatial orientation of basal keratinocytes, presence of immature cells and apoptosis
in the upper stratum of skin, and overall cellularity reduced to 77.4%. Exposure to SSUV in the presence of sunscreen
shows H&E staining and cellularity similar to controls. Importantly, there is a dramatic increase in gH2AX positive
cells in UV-exposed skin that is completely prevented in the presence of sunscreen. Strikingly, telomeres of cells in
the 3D skin environment, as opposed to isolated cells in in-vitro cultures, show similar telomeric length to control,
and this length is even increased when exposure occurs in the presence of the broad-spectrum sunscreen product,
together with a decrease in the % of short telomeres.
In conclusion, the broad-spectrum sunscreen product with a very high UVB and UVA protection (SPF 50+ / PF UVA46) preserves cell viability, reduces DNA damage in UV-exposed in-vitro cells or 3D skin model, and protects theintegrity of the stratified structure of the skin, crucial for its barrier function and natural protection against UV.
Importantly, this photoprotection also occurs at the telomere level since UV-induced telomere shortening is greatly
reduced in the presence of the sunscreen in in-vitro cultures. Furthermore, telomere length is even increased in SSUV
exposed 3D skin in the presence of sunscreen. These findings help in the understanding of the mechanisms ofphotoprotection by a sunscreen product against UV-induced damage and are relevant in the fields of photobiology
and skin carcinogenesis.
Acknowledgements. This work was supported by grant from La Roche Posay
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P-4.1.11

Molecular Basis of Dark Photochemistry
Juliana Cuéllar-Zuquin,1 Fernand Louisnard,1 Angelo Giussani1 and Daniel Roca-Sanjuán1
1) Instituto de Ciencia Molecular, Universitat de Valencia, Valencia, Spain.
Dark photochemistry is responsible for the production of melanomas when there is no UV radiation, occurring theprocess via chemiexcitation. This chemiexcitation is originated by a dioxetane derivate of melanin, which induces
the formation of lesions in the DNA pyrimidine bases1. In this work, we have carried out a theoretical study todetermine the molecular basis of the mentioned process.
Firstly, we have employed the 1,2-dioxetane molecule as a model of the dioxetane derivate of melanin, and an
ethylene dimer as a model of two stacked pyrimidine nucleobases. With these models, we have studied themechanism of the decomposition of dioxetane, which goes through a diradical intermediate that allows to populate
the triplet excited state of the formaldehyde2. We have also studied the mechanism of [2+2] cycloaddition of
ethylene on the triplet manifold. In this case, the triplet excited state evolves up to a diradical intermediate, where
the ground and excited state cross. Here, an inversion of multiplicity takes place and the systems evolves to theformation of the cyclobutane.
Secondly, we have carried out an energetic study to verify if the cycloaddition reaction induced by energy transferfrom the chemiexcited dioxetane is plausible. In the case of the models, we have obtained that the process cannot
take place, but in the case of the real molecules (melanin dioxetane and two stacked thymine nucleobases) theprocess is favourable, thus allowing to explain the phenomenon of melanoma produced via dark photochemistry.
At last, we have studied different factors which can be involved in the process such as the ionization potential of
the nucleobases, the formation of hydrogen bonds and the presence or absence of π-stacking interactions. All of theabove studies had been carried on over different dimers of the dioxetane derivative of the melanin and thenucleobases.
References
1) Premi, S.; Wallisch, S. Chemiexcitation of melanin derivatives induces DNA photoproducts long after UV
exposure. Science 2015.
2) Farahani, P.; Roca-Sanjuán, D. Revisiting the nonadiabatic process in 1,2-dioxetane. J. Chem. Theory
Comput. 2013.

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P-4.1.12
Developing a tool for capturing genomic regions of high-density skin clonal
mutations
Megan E Fitzgerald1,2, Sydney Grant1,2, Andrew D Patentreger1,2, Nico Shary2,3, Allison K McCarthy1,2, Barbara A
Foster4, Lei Wei5, Wendy J Huss1,2, Gyorgy Paragh1,2
1) Roswell Park Comprehensive Cancer Center, Department of Cell Stress Biology, Buffalo, NY, USA; 2) Roswell
Park Comprehensive Cancer Center, Department of Dermatology, Buffalo, NY, USA; 3) University at Buffalo,
Jacobs School of Medicine and Biological Sciences, Buffalo, NY, USA; 4) Roswell Park Comprehensive Cancer
Center, Department of Pharmacology and Therapeutics, Buffalo, NY, USA; 5) Roswell Park Comprehensive
Cancer Center, Department of Biostatistics and Bioinformatics, Buffalo, NY, USA
Nonmelanoma skin cancers (NMSCs) greatly exceed the number of all other human malignancies in the UnitedStates and show increasing incidence worldwide. Chronic long-term ultraviolet (UV) exposure is the primary causeof skin carcinogenesis and leads to the formation of clonogenic mutations (CMs). CMs are clinically andhistologically normal, the first steps during photocarcinogenesis, and the first known manifestations of fieldcancerization. It has been established that UV-exposed clinically normal-presenting areas of skin contain highlevels of CMs1, however the comparison of CMs and mutations in NMSC have yet to be systematically studied. To
understand how mutation profiles of clinically normal presenting skin correspond to mutations in NMSC, wedeveloped two analytic pipelines for comparing the genome of normal skin to mutational hotspots in cutaneous
squamous cell carcinoma (cSCC), a type of NMSC with metastatic potential. The mutation frequency binningpipeline takes each mutation in the dataset and identifies neighboring mutations within an assigned range of basepairs. If a mutation is found, the algorithm will search for additional mutations surrounding this one using this samerange. This repeats until no additional mutations are found within the designated range, outputting the total
concatenated base pair region covering all identified mutations. This pipeline may then identify the regions with thehighest quantity of mutations, by filtering out regions containing less than an assigned cutoff value. Our second
pipeline takes these defined regions from two different datasets and calculates the number of base pairs that overlap
between the two. Using these two pipelines with a genomic target of 100 base pairs and a cutoff value of fivemutations per genomic area, we identified a significant (p=0.001194) overlap between mutations found in normal
sun-exposed skin and cSCC. We also identified significant (p<0.00001) enrichment of individual point mutations
and identified numerous CM enriched areas with low mutation frequency in cSCC. The identified pattern ofmutation overlap will help to design optimal sequencing library targets for potentially cancer-causing CMs in sunexposed normal human skin.
References
1) Lei Wei, Sean R Christensen, Megan E Fitzgerald, Ultradeep sequencing differentiates patterns of skin clonal
mutations associated with sun-exposure status and skin cancer burden. Science Advances, 2021.

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P-4.1.13

Metalloporphyrins: promising G-quadruplex stabilizing ligands
Catarina IV Ramos,1 Ana Rita Monteiro, 1,2 Nuno MM Moura,1 Tito Trindade2, Maria AF Faustino,1 Maria Graça
PMS Neves1
1) LAQV-REQUIMTE, Department of Chemistry of University of Aveiro, Aveiro, Portugal; 2) CICECO,
Department of Chemistry of University of Aveiro, Aveiro, Portugal
G-quadruplexes (GQ) are DNA secondary structures found in several genome regions of biological significance,
namely in telomeres.1 The stabilization of these GQ DNA structures by ligands is a promising strategy for the
inhibition of telomerase, a reverse transcriptase enzyme expressed in a range of cancer cells, that allows continual
cell division without telomere shortening.2,3 So, the inhibition of telomerase by recurring to adequate GQ-stabilizing
ligands can be considered a promising therapeutic approach for cancer.
To assess the potential of a compound as a telomerase inhibitor, selectivity for quadruplex over duplex DNA is afundamental attribute, as the drug must be able to recognize quadruplex DNA in the presence of a large amount of
duplex DNA, in the cellular nucleus.4
Several studies have shown that porphyrins and analogues, in special the tetracationic 5,10,15,20-tetrakis(1methylpyridinium-4-yl)porphyrin (TMPyP), exhibit high affinity to GQ and thus are potential telomerase inhibitors.
However, it is also recognized that TMPyP has poor selectivity for GQ over duplex DNA structures.4
By using spectroscopic techniques and ensuring the same experimental conditions, the efficacy of the AgII, CuII, CoIII,
NiII, PdII and ZnII metal complexes of TMPyP (M-TMPyP, Fig. 1) as GQ stabilizing agents were evaluated andcompared. Formation of stable porphyrin-DNA adducts was observed for all the ligands with both DNA structures.
The analysis of the hypochromic and bathochromic effects in the ligand UV-Vis spectra due to the presence of the
DNA allowed to obtain binding constants and to predict that, in general, the interaction of the ligands occurs by endstacking in the GQ structure. The AgII ligand showed interesting and promising properties as G-quadruplex stabilizing
agent.

Fig. 1 – G-quadruplex-M-TMPyP adduct formation and telomerase inhibition.

References
1) a) D Sen, W Gilbert. Nature, 334, 364, 1988; b) EH Blackburn. Cell. 77, 621, 1994
2) a) A Moye, et al. Nature Commun. 6, 7643, 2015; b) J Nandakumar et al. Nat. Rev. Mol. Cell Biol. 14, 69, 2013;
3) a) L Oganesian, et al. BioEssay. 29, 155, 2007; b) Q Wang, et al. Nucleic Acids Res. 39, 6229,2011; c) J Bidzinska,
et al. Molecules. 18, 12368, 2013.
4) a) AR Monteiro, et al. ACS Omega. 2, 11184, 2018; b) CIV Ramos, et al. Molecules. 24, 733, 2019
Acknowledgements. The authors are grateful to the University of Aveiro and to the Fundação para a Ciência e a
Tecnologia (FCT) for the financial support to LAQV-REQUIMTE (Ref.UIDB/50006/2020), and, where applicable,
co-financed by the FEDER, within the PT2020 Partnership Agreement. Catarina IV Ramos (REF.-047-88ARH/2018) and Nuno MM Moura (REF.-048-88-ARH/2018) acknowledges the costs of their research contracts.
Ana R. Monteiro thanks FCT for the PhD grant SFRH/BD/137356/2018.
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P-4.1.14

Metalized Phthalocyanines as potential photosensitizer in Photodynamic Therapy
Miguel León1, Franklin Vargas1
1)Venezuelan Institute for Scientific Research (I.V.I.C), Organic Photochemistry Laboratory, Chemistry Center,
Caracas, Venezuela.
Phthalocyanines are organic compounds whose origin is related to porphyrin structure. They have many
applications in different fields, including biomedical, photobiological and chemical research. From a chemical
standpoint, phthalocyanines are macrocyclic tetraaza compounds, which are composed mainly of isoindolic groups
that confer aromaticity and planarity to the chemical structure1. Similar to other types of porphyrin compounds, thestructure of phthalocyanine is capable of chelating many metals, which more often contribute to its wide variety offunctions, including singlet oxygen and free radicals generation, fluorescence, absorption spectra, among others.
All of these properties are desirable for biomedical and pharmaceutical research. ROS generation is the main themein Photodynamic Therapy and phototoxic processes, as well as in antibacterial, antiviral and anticancer properties.
Therefore, the synthesis of phthalocyanines and in vitro testing are a relevant scientific topic2, 3, 4, 5. There are many
synthesis methods and mechanisms of action, each one of those have a different level of difficulty 5. In our present
experimental development, we expect to test the photosensitizer potential and Pbr322 plasmid DNA cleavage
activity of Zn, Cd and Hg metalized phthalocyanines by means of agarose gel electrophoresis andchemiluminescence. The excellent photochemical and pharmaceutical properties of phthalocyanines will be valued,
which make them excellent for their wide use at a medical and clinical level.
References
1) Ishii, K. Functional singlet oxygen generators based on phthalocyanines. Coord. Chem. Rev, 256, 1556-1568,(2012)
2) F. Vargas, C.Rivas, T. Zoltan, L. Padrón, C. Izzo, V. López, L. Gómez, F. Pujol, H. Rangel, D. Garzaro, R.
Fabbro. Comparative Antiviral (HIV) Photoactivity of Metalized mesoTetraphenylsulfonated Porphyrins.
Medicinal Chemistry4/2, 138-145 (2008).
3) Oviedo, O., Zoltan, T., Vargas, F., Inojosa, M., Vivas, J.C. Antibacterial photoactivity and photosensitizedoxidation of phenols with meso-tetra-(4-benzoate, 9-phenanthryl)-porphyrin and its metal complexes (Zn and Cu).J.
Coord. Chem. 67, 1715(2014).
4)Wadzanai, C., Ogunsipe, O.,Tebello, N.Syntheses and photophysics of new phthalocyanine derivatives of zinc,cadmium and mercury. New J. Chem,31, 377-384 (2006).
5) Barut, B,OzgürYalcin C., Demirbas Ü., Ackay H.T., Kantekin, H., Özel A. The novel Zn (II) phthalocyanines:
Synthesis, characterization, photochemical, DNA interaction and cytotoxic/phototoxic properties. J. Mol. Structure,
1218, 128502, (2020).

Acknowledgements. This work was supported by grant from Centro de Estudios Avanzados (CEA-IVIC)Book of Abstracts ������������������������������������������������������������������������������������������������ 177

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IL-4.2.1

UV-B radiation in a changing climate; can UV protect plants from drought stress?
Gaia Crestani1, Aoife Coffey1, Alexander Ač2, Karel Klem2, Otmar Urban2, Marcel A.K. Jansen1,2
1) School of Biological, Earth and Environmental Sciences, Environmental Research Institute, University College
Cork, Ireland; 2) Global Change Research Institute, Czech Academy of Sciences, Bělidla 986/4a, CZ-603 00 Brno,
Czech Republic
Cross-resistance, i.e. the concept whereby plants exposed to one particular stressor develop protection against
multiple stressors is well known in plant biology. Traditionally, Reactive Oxygen Species (ROS) were perceived as
key players in cross-resistance, with elevated antioxidant defences protecting against ROS generated as a result of
exposure to a range of different stressors. However, new evidence shows more subtle mechanisms of cross-talkbetween stress-protection pathways in plants, including a variety of chemical and/or electrical signals that transmit
information between receptor systems and signalling cascades resulting in a degree of cross-resistance. Conversely,
cross-sensitivity may arise where plant defences responses are overwhelmed by a combination of two stressors.
Indeed, in theory, impacts of combinations of two stressors can be additive, synergistic or antagonistic. A
particularly important combination of environmental factors is UV-B radiation and drought. The 2016 report by theUnited Nations Environment Programme -Environmental Effects Assessment Panel report flagged that interactions
between climate change and UV penetration in the biosphere are resulting in the exposure of plants to new
combinations of UV radiation and drought. However, lack of understanding of impacts of such combined
treatments creates uncertainties that hamper predictions of future ecological change. A meta-analysis of publishedliterature shows complex interactive effects between UV-B radiation and drought on plants, with individual studies
showing increased cross-resistance, or the opposite, increased sensitivity. Response variations are likely to reflect
exposure doses and/or kinetics. Yet, overall, in plants exposed to both UV and drought, increases in plant defence
responses are less-than-additive, and so are the damage and growth retardation. Induction of a degree of crosstolerance seems the most likely interpretation of the less-than-additive responses that are observed.
The data imply that UV sensing may play an important physiological function in fine-tuning responses to drought.
Given that realistic doses of UV-B radiation rarely cause stress in well acclimated plants, the potential to use UV-B
radiation for pre-acclimation against other stressors is attractive.
Acknowledgements. This work was supported by Science Foundation Ireland (S16/IA/4418)

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IL-4.2.2

Does ultraviolet radiation modulate plant responses to elevated CO2 concentration?
Otmar Urban1, Karel Klem1, Marcel A.K. Jansen1,2
1) Global Change Research Institute, Czech Academy of Sciences, Bělidla 986/4a, CZ-603 00 Brno, Czech
Republic; 2) School of Biological Earth and Environmental Sciences, Environmental Research Institute, University
College Cork, Cork, Ireland
The recent UNEP-EEAP (United Nations Environment Programme -Environmental Effects Assessment Panel) report
has emphasized the need to investigate interactions between climate change and ultraviolet (UV) radiation.
Depending on emission scenario, the atmospheric CO2 concentration is predicted to reach 538–936 µmol CO2 mol–1
at the end of the century. Together with increased temperature and altered precipitation patterns, elevated CO2
concentration (eCO2) is considered to be the most crucial factor affecting terrestrial ecosystems in future. However,plant responses to these factors may be further modulated by other environmental drivers, including ultraviolet (UV)radiation. Though eCO2 and UV radiation (particularly UV-B) have both direct and indirect effects on photosynthesis
and growth, their interactive effects may range from simply additive to synergistic and antagonistic.
We summarize our previous findings in woody and non-woody plants at different hierarchical and functional levels,
including changes in plant stoichiometry, physiology and morphology. Among others, we found support for the
hypothesis that the impact of eCO2 on photosynthesis is substantially modulated by UV radiation. Moreover, theeCO2×UV interaction is changing along the vegetation season: enhanced UV radiation stimulates a positive effect ofeCO2 on plant photosynthesis at the beginning of the vegetation season (short-term effect), whilst long-term
cultivation diminishes the stimulatory effect of eCO2 (a clear down-regulation of photosynthesis). Down-regulationis, however, not found in plants grown under the conditions of excluded UV radiation1,2. Such findings indicate theneed to explore interactive effects of eCO2 with other environmental drivers, including the intensity of UV radiation.
Exclusion of UV radiation in experiments simulating eCO2 may thus substantially overestimate the positive effect of
CO2 on photosynthesis and growth of future plants.
References
1) Urban O., Hrstka M. et al., Interactive effects of ultraviolet radiation and elevated CO2 concentration on
photosynthetic characteristics of European beech saplings during the vegetation season. Plant Physiology andBiochemistry 134: 20-30, 2019.
2) Uchytilová T., Krejza J. et al., Ultraviolet radiation modulates C:N stoichiometry and biomass allocation in Fagus
sylvatica saplings cultivated under elevated CO2 concentration. Plant Physiology and Biochemistry 134: 103-112,
2019.
Acknowledgements. The work was supported by the Ministry of Education, Youth and Sports of the Czech Republic
within the SustES project – Adaptation strategies for sustainable ecosystem services and food security under adverse
environmental conditions (CZ.02.1.01/0.0/0.0/16_019/0000797).

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OC-4.2.3

Short-term UV pretreatment supports cold tolerance of bell pepper seedlings
Gyula Czégény, Arnold Rácz, Kristóf Csepregi, Éva Hideg
University of Pécs, Department of Plant Biology, Pécs, Hungary
Ultraviolet radiation (UV, 280-400 nm) is an important regulator of plant growth and development1. Under UV
exposure, plants upregulate several metabolic pathways, such as the synthesis of phenolics or the activation ofantioxidant enzymes2,3. In addition, moderate UV doses stimulate energy dissipating pathways in photosystem II (PSII) by triggering zeaxanthin formation4, supporting the protection of the photosynthetic apparatus. These strategies
promote plant acclimation to other environmental challenges as well, such as high light conditions5 or chilling
temperatures6. During the following decades, extreme weather conditions are expected to appear more frequently,
thus the importance of effective techniques in plant conditioning will rise. Controlled exposure to low UV-B doses
documentedly increases tolerance to subsequent stress7, and the aim of the present study was to explore whether UVB is capable of priming the cold tolerance of bell pepper seedlings. The study was destined to model a generalagricultural practice: the springtime transfer of bell pepper seedlings from plant nurseries to open fields, which
includes a sudden change in weather conditions.
In this work, bell pepper (Capsicum annuum cv. grossum) seedlings were grown in growth chambers under 90 µmol
s-1 m-2 photosynthetically active radiation (400-700 nm), 16h/8h, 25°C/20°C day/night conditions for 3 weeks. Prior
to a 5-day long exposure to cold temperatures (15°C/10°C day/night conditions), half of the plants were illuminatedby acclimative doses of UV (6.9 kJ m-2 d-1 BED) provided by a broadband 311 nm centred UV tube (Q-Panel UVB313EL), for 5 days.
The applied UV pretreatment affected leaf metabolites, making the plants more resistant to lower temperature stress.
The latter was confirmed by measuring PS II photochemical yields and non-photochemical quenching parameters.
Results suggest a cold-induced limitation in energy transfer between the antenna and the reaction center, which waslessened by the UV-pretreatment. Priming with UV also helped to maintain higher electron transfer rates under higher
light intensities and lessen the negative effect of cold stress on these. Our results support beneficial effects of UV
priming in plant protection.
References
1) Jansen, M.A.K., Bornman, J.F., UV-B radiation: from generic stressor to specific regulator. Physiologia Plantarum
145: 501-504, 2012.
2) Schreiner, M., Mewis, I., Huyskens-Keil, S., Jansen, M.A.K., Zrenner, R., Winkler, J.B., O’Brien, N., Krumbein,A., UV-B-induced secondary plant metabolites - potential benefits for plant and human health. Critical Reviews in
Plant Sciences 31: 229-240, 2012.
3) Czégény, Gy., Kőrösi, L., Strid, Å., Hideg, É., Multiple roles for vitamin B 6 in plant acclimation to UV-B.
Scientific Reports 9: 1259, 2019.
4) Šprtová, M., Špunda, V., Kalina, J. and Marek, M.V., Photosynthetic UV-B response of beech (Fagus sylvatica
L.) saplings. Photosynthetica 41: 533–543, 2003.
5) Klem, K., Holub, P., Štroch, M., Nezval, J., Špunda, V., Tříska, J., Jansen, M.A.K., Robson, T.M., Urban, O.,
Ultraviolet and photosynthetically active radiation can both induce photoprotective capacity allowing barley toovercome high radiation stress. Plant Physiology and Biochemistry 93: 74–83, 2015.
6) Parkin, K.L., Marangoni, A., Jackman, R.L., Yada, R.Y., Stanley, D.W., Chilling injury. A review of possible
mechanisms. Food Biochemistry 13: 127-153. 1989.
7) Wargent, J.J., Elfadly, E.M., Moore, J.P., Paul, N.D., Increased exposure to UV-B radiation during early
development leads to enhanced photoprotection and improved long-term performance in Lactuca sativa. Plant Cell
& Environment 34: 1401-1413, 2011.
Acknowledgements. This work was supported by the National Research, Development and Innovation Office (grant
PD-131625), the János Bolyai Research Scholarship (BO/00362/20/4) of the Hungarian Academy of Sciences andthe New National Excellence Program of the Ministry for Innovation and Technology from the source of the National
Research, Development and Innovation Fund (grants ÚNKP-20-4-II-PTE-603 and ÚNKP-20-5-PTE-634).
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OC-4.2.4

Short daily UV exposure of Micro-Tom tomato plants favours a better stomatal
control suggesting a delayed leaf senescence
Mannucci Alessia1; Scartazza Andrea2; Santaniello Antonietta3; Mariotti Lorenzo1; Castagna Antonella1;
Santin Marco1; Quartacci Mike Frank1; Ranieri Annamaria1
1) University of Pisa, Agriculture, Food and Environment, Pisa, Italy; 2) CNR, Institute of Research on
Terrestrial Ecosystems, Pisa, Italy; 3) Valagro S.p.A., Pisa, Italy
UV radiation, unless present at high doses, is recognised to act as a regulator of plant growth and somespecific processes (Frohnmeyer and Staiger, 2003; Jenkins, 2017; Robson et al., 2019). The present study
investigated the influence of UV irradiation (0.132 and 0.015 W m-2 biologically effective UV-B and UVA irradiance, respectively, 15 minutes/day, 11 days) on leaf gas exchange and some biochemical andmolecular markers of leaf senescence in Micro-Tom tomato plants. UV-induced reduction of gs (stomatal
conductance) was associated with the modified expression of some genes involved in the control of stomatal
movement and with changes in salicylic and abscisic acid contents, which suggest a two-step regulation of
stomatal closure involving the two hormones. The effects of UV on stomatal closure were only transient,as suggested by the full recovery of gs three days after the end of the treatment. The temporal changes of gs
and Anet (steady state photosynthetic CO2 assimilation rate) along with the hormone and chlorophyll
behaviour, suggest a possible delay of leaf senescence in treated plants compared to control ones, confirmedby the expression levels of some genes related to senescence. The UV potential to induce a persistent partial
inhibition of gs without severely affecting Anet led to an increased iWUE (intrinsic water use efficiency)
during the 11-day treatment, suggesting a priming effect of low UV radiation towards drought conditions,potentially useful to reduce the excess water use in agriculture.

References
1)Frohnmeyer H., Staiger D., Ultraviolet-B radiation-mediated responses in plants. Balancing damage andprotection. Plant Physiology, 2003, 33:1420–1428
2)Jenkins G., Photomorphogenic responses to ultraviolet‐B light. Plant Cell & Environment, 2017, 40 (11):2544–2557
3)Robson TM, Aphalo PJ, Banaś AK, Barnes PW, Brelsford CC, Jenkins GI, Kotilainen TK, Łabuz J, MartínezAbaigar J, Morales LO, Neugart S, Pieristè M, Rai N, Vandenbussche F, Jansen MA, A perspective on
ecologically relevant plant-UV research and its practical application. Photochemical & Photobiological Sciences,
2019, 18(5), 970-988
Acknowledgements. This work was supported by the PhD program of Agriculture, Food and Environment
fellowship, University of Pisa.

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IL-4.2.5

Benefits of solar UV-B radiation on field crops defenses against insect pests.
Jorge A. Zavala.
University of Buenos Aires, School of Agronomy, Biochemistry unit and INBA-CONICET, Argentina.
Solar UV-B radiation has been reported to enhance plant defenses against herbivore insects in many species. However,
the mechanism and traits involved in the UV-B mediated increment of plant resistance are largely unknown in crops
species, such as soybean. Two soybean cultivars (cv.) were grown under attenuated or full solar UV-B radiation andleaves were damaged by Anticarsia gemmatalis larvae and pods attacked by the stink bug bugs Nezara viridula. We
determined in undamaged and damaged leaves and pods changes in jasmonates, ethylene, salicylic acid, trypsin
protease inhibitor (TPI) activity, flavonoids and mRNA expression of genes related with defenses. Ethylene emission
induced by herbivory was synergistically increased in plants grown under solar UV-B radiation and was positively
correlated with malonylgenistin concentration, TPI activity and expression of IFS2 and the defensive protein PR2,
while was negatively correlated with leaf consumption and stink bug damage. The precursor of ethylene ACC applied
exogenously to soybean was enough to strongly induce leaf isoflavonoids. Our results showed that in field-grownsoybean isoflavonoids were regulated by both herbivory and solar UV-B inducible ET, while flavonols were regulated
by solar UV-B radiation and not by herbivory or ET. Our study suggests that although ET can modulate UV-Bmediated priming of inducible plant defenses, some plant defenses, such as isoflavonoids are regulated by ET alone.

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IL-4.2.6
UV-B LED priming for reduced biotrophic disease susceptibility in Lettuce seedlings
Jason J. Wargent1,2, Ana Clara Pontaroli2, Emily R. McLay1,2
1) School of Agriculture and Environment, College of Sciences, Massey University, Palmerston North, New
Zealand; 2) BioLumic Limited, Palmerston North, New Zealand
Biotrophic disease is one of the largest causes of decreased yield in agriculture. While exposure to ultraviolet B (UVB) light (280–320 nm) has been previously observed to reduce plant susceptibility to disease, there is still a paucity
of information regarding underlying biological mechanisms. In addition, recent advances in UV-LED technology
raise the prospect of UV light treatments in agriculture which are practical and efficient. Here, we characterized thecapability of UV-B LED pre-treatments to reduce susceptibility of a range of lettuce (Lactuca sativa) cultivars todowny mildew disease caused by the obligate biotroph Bremia lactucae. Innate cultivar susceptibility level did notseem to influence the benefit of a UV-B induced disease reduction with similar reductions as a percentage of thecontrol observed (54–62% decrease in conidia count) across all susceptible cultivars. UV-B-induced reductions toconidia counts were sufficient to significantly reduce the infectivity of the diseased plant. Secondary infections
caused by UV-B pre-treated plants exhibited yet further (67%) reduced disease severity. UV-B-induced flavonoids
may in part mediate this reduced disease severity phenotype, as B. lactucae conidia counts of lettuce plants negatively
correlated with flavonoid levels in a UV-B-dependent manner (r = -0.81). Liquid chromatography–mass spectrometry(LC-MS) was used to identify metabolic features which contribute to this correlation and, of these, quercetin 3-O(6”-O-malonyl)-b-D-glucoside had the strongest negative correlation with B. lactucae conidia count (r = -0.68).
When quercetin 3-O-(6”-Omalonyl)- b-D-glucoside was directly infiltrated into lettuce leaves, with those leaves
subsequently infected, the B. lactucae conidia count was reduced (25–39%) in two susceptible lettuce cultivars. We
conclude that UV-B induced phenolics, in particular quercetin flavonoids, may act as phytoanticipins to limit theestablishment of biotrophic pathogens thus delaying or reducing their sporulation as measured by conidia count.
These findings highlight the opportunity for UV-B morphogenesis to be exploited through the application of UVLED technology, as part of the development of next-generation, sustainable disease control tools.
References
1) McLay ER, Pontaroli AC and Wargent JJ, UV-B induced flavonoids contribute to reduced biotrophic diseasesusceptibility in lettuce seedlings. Front. Plant Sci. 11:594681. doi: 10.3389/fpls.2020.594681, 2020
Acknowledgements. This work was supported by a Ph.D. scholarship award from Massey University and BioLumic
Limited to EM. The scholarship was funded at 50% by each of the two organizations.

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OC-4.2.7

The role of ultraviolet radiation in colonization of plants by endophytes
Aleksandra Giza1, Paweł Hermanowicz1, Rafał Ważny1, Piotr Rozpądek1, Justyna Łabuz1
1) Jagiellonian University in Kraków, Malopolska Centre of Biotechnology, Kraków, Poland
Light is one of the most important environmental factors for growth and development of plants. Recent studies havedemonstrated the crucial role of light in the plant defense system and in the control of the trade-off between growthand defense. Natural and artificial plant communities must deal with crowding, which results in canopy shade andaffect the resistance to pathogens1.
Ultraviolet light (in particular UV-B) has a positive effect on plant defense mechanisms in leaves against biotic
stressors2. UV-B enhances host resistance to fungal pathogens and infections by necrotrophs3. Secondary metabolites
such as flavonoids serve as screening compounds protecting photosynthetic apparatus from damage, but also may
improve antifungal resistance4. Plant immune responses are mediated by photoreceptors: red/far-red sensitive
phytochromes, UV-B sensitive UVR8 and blue and UV-A proteins: cryptochromes and phototropins5. Plant
hormones such as jasmonic acid (JA), salicylic acid (SA) and ethylene activate immune responses4.
In this study, we will analyze the impact of the UV-A light on the host resistance to fungi and their colonization
capacity. Symbiotic (and parasitic) interactions will be assessed on Arabidopsis thaliana plants inoculated withendophytes collected from shoots of Arabidopsis arenosa. The experiment will focus on yeast: Sporobolomyces
ruberrimus, and fungi species: Mucor sp., Paraphoma chrysanthemicola, Phomopsis columnaris, Diaporthe eres.
We have employed WGA-TexasRed staining followed by a Confocal Microscopy analysis to confirm thecolonization status of roots and shoots by fungi. Now we will test the overall plant fitness in terms of morphology(weight, shoot and root length), phenolic content (anthocyanins) and expression of genes of phenolic pathway andplant hormones.
References
1) Moreno J. E., Ballaré C.L., Phytochrome Regulation of Plant Immunity in Vegetation Canopies. Journal ofChemical Ecology, 40:848–857, 2014.
2) Ballaré C.L., Mazza C.A., Austin A.T., Pierik R., Canopy Light and Plant Health. Plant Physiology, Vol.
160, Issue 1, pp. 145–155, 2012.
3) Demkura P.V., Ballare C.L., UVR8 Mediates UV-B-Induced Arabidopsis Defense Responses against
Botrytis cinerea by Controlling Sinapate Accumulation. Molecular Plant, Vol. 5, no 3, pp. 642-652, 2012.
4) Ballare C.L., Light Regulation of Plant Defense. Annual Review of Plant Biology, 65:15.1–15.29, 2014.
5) Griebel T., Zeier J., Light Regulation and Daytime Dependency of Inducible Plant Defenses in Arabidopsis:
Phytochrome Signaling Controls Systemic Acquired Resistance Rather Than Local Defense. Plant
Physiology, Vol. 147, pp. 790–801, 2008.

Acknowledgements. This work was supported by grant POBBios for Jagiellonian University ‘The role of ultraviolet
radiation in colonization of plants by endophytes’. Endophytes were isolated as a part of the grant ‘The role ofendophytic fungi in confering Arabidopsis arenosa toxic metal tolerance’ UMO-2017/27/B/NZ8/01199.

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OC-4.2.8
UVB-dose and Salt stress response on the accumulation of secondary metabolites on
Bell pepper leaves
Shyam Pariyar1, Simone Röhlen-Schmittgen2
1) Bonn University, Department of Plant Nutrition, Bonn, Germany; 2) Hochschule Geisenheim University,
Department of Vegetable Production, Geisenheim, Germany
The accumulation of secondary metabolites (SMs) on crop leaves could add value to the crop green biomass byincreasing the nutritional quality of produce or increasing high value molecules content for industrial uses. These
SMs can be extracted from crop green biomass and sold. Crop plants accumulate higher amounts of these SMs when
they are exposed to abiotic stresses such as salt and UV. These SMs can be anti-oxidative, antifungal or antimicrobial,
thus enhancing plant defense itself or they enrich the nutritional value and market potential of specific metabolites
that have high economic demand. Thus, we evaluated, i) how UVB and Salt stress affect Bell pepper plants at
different doses of UVB application and ii) how these UVB and Salt stress treatments affect SMs accumulation suchas flavones cynaroside and graveobioside A in leaves on Bell pepper leaves when exposed to salt stress (100 mM
NaCl) and 3 different UVB doses (1.7 W m-2 for 1, 2 and 3 h per day) for a week. SMs accumulations were comparedusing non-destructive Fluorescence sensors (Dualex, Multiplex), Field Spectrometer and analysed using HPLC. We
hypothesized that the enhanced accumulation of SMs under stress conditions. Since, the SMs analysis is currently
being analysed, we expected to find out the optimal thresh hold exposure to the single (UVB) or combined stresses(UVB and Salt) to have maximum SMs accumulation in Bell pepper leaves. Thus, in this paper, some of the major
results of this research will be presented and discussed. The outcome of the study might be useful for implication of
abiotic stresses such as UV, salt stress or combined on enhancing crop quality and plant health management.
Acknowledgements. This research is funded by Federal Ministry of Education and Research (BMBF), Germany
under the framework of the „National Research Strategy Bioeconomy 2030“

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P-4.2.9

Postharvest UV-treatment induces changes in grapevine berry skin photosynthesis,
phenolic profiles and antioxidant capacities
Kristóf Csepregi1, Péter Teszlák2, Arnold Rácz1, Gyula Czégény1, László Kőrösi2, Éva Hideg1
1) Department of Plant Biology, Faculty of Sciences, University of Pécs, Hungary; 2) Research Institute for
Viticulture and Oenology, University of Pécs, Hungary
Biosynthesis of phenolic compounds is stimulated by ultraviolet (UV) light, and this regulatory role has been utilized
for modifying metabolic profiles of fruits and vegetables1. In this study, we irradiated harvested berry clusters of one
red (cv. Emperor) and two white (cv. Queen of Vineyard and White Sultana) table grape varieties with broad band
UV (either 312 nm centred UV-B or 355 nm centred UV-A) for 30 min. UV-induced changes in berry skin antioxidant
capacities and phenolic profiles were measured regularly, between 2 and 48h after irradiation. Caftaric acid was thedominant phenolic acid in all cultivars, but its content was unaffected by the UV-treatment. UV-induced changes inphenolic profiles included a decrease in the amounts of the two major quercetin-glycosides 2h after irradiation in all
cultivars. This decrease was temporal in two of the studied varieties (Emperor and Queen of Vineyard), and it was
followed by a recovery to pre-treatment-levels during storage at 20°C under low intensity photosynthetically active
light. Berry skin UV-A and UV-B absorbing capacities showed the same, transitional decrease regardless of the
wavelength of the irradiation. Antioxidant capacities and quercetin-glycoside contents showed strong and positive
correlations. Experiments with the red variety suggested that the observed decrease was due to a transient, UVinduced activation of peroxidases resulting in increased flavonoid oxidation2. Experiments with the two whitevarieties suggested that recovery of oxidized compounds was supported by berry skin photosynthesis. Recovery ofquercetin-glycoside contents after the temporal UV-induced decrease was only observed in one of the two whitegrape varieties, in Queen of Vineyard, the one with higher base (pre-treatment) photochemical activity. In addition,
this variety showed a temporal increase in berry skin photochemical activity 2 and 8h after the UV-treatment, while
the same parameter decreased permanently in the White Sultana cultivar. These differences were the consequence ofdistinct UV-responses of regulated and nonregulated nonphotochemical quenching in the two cultivars. Results
showed that mature grapevine berries have photosynthetically active tissues capable of dynamic changes even several
hours postharvest, and suggest that changes in photochemistry may contribute to postharvest metabolic responses ofberry skins3.
References
1) Schreiner M, Mewis I, Huyskens-Keil S, Jansen MAK, Zrenner R, Winkler JB, O’Brien N, Krumbein A, UV-Binduced secondary plant metabolites - Potential benefits for plant and human health, Critic. Rev. Plant Sci. 31, 229240. 2012.
2) Csepregi K, Kőrösi L, Teszlák P, Hideg É., Postharvest UV-A and UV-B treatments may cause a transient decreasein grape berry skin flavonol-glycoside contents and total antioxidant capacities. Phytochem. Lett. 31: 63-68, 2019.
3) Csepregi K, Teszlák P, Rácz A, Czégény Gy, Kőrösi L, Hideg É., Changes in grapevine berry skin photochemistry
may support metabolic responses to postharvest treatment by ultraviolet light. Photosynth. 59, 286-293, 2021.
Acknowledgements. This work was supported by the National Research, Development and Innovation Office (grantKH-129615 to ÉH and KCs).

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IL-4.3.2
Regulatory T cell induction prolongs the efficacy for the treatment of psoriasis
Akimichi Morita1
1) Department of Geriatric and Environmental Dermatology, Nagoya City University Graduate School of
Medical Sciences, Nagoya 467-8601, Japan
Phototherapy utilizes the beneficial effects of ultraviolet (UV) wavelengths to affect immunoregulatory functions.
UV light phototherapy using narrowband (NB) UVB and bath-psoralen UVA (bath-PUVA) therapy are wellestablished treatments. Dual-action operative mechanisms of UV phototherapy have been identified: apoptosis andimmune suppression. NB-UVB depletes pathogenic T cells by inducing apoptosis and regulatory T cells. Other
wavelengths are also utilized for phototherapy, i.e., 308-nm excimer light and 312-nm flat-typed NB-UVB. Excimer
light (308 nm) therapy effectively targets the affected skin without unduly exposing other areas and increases thelevels of T regulatory cells. Phototherapy improves impaired resting regulatory T cells1 and increases activatedregulatory T cells in patients with psoriasis2. In mouse, UVB increases regulatory T cells by clustering with dermal
CD11b-type Langerin- DCs to induce immunological self-tolerance3. Furthermore, UVB-expanded skin regulatory
T cells are tissue regulatory T cells and express proenkephalin (PENK), an endogenous opioid precursor, andamphiregulin (AREG), the epidermal growth factor receptor ligand4. UVB-expanded skin regulatory T cells play a
key role in promoting wound healing in vivo. However, UVB-expanded skin regulatory T cells have not been proven
yet in human.
More rational designs of phototherapy devices and irradiation protocols are under development. Biologics are highly
effective for the treatment of psoriasis. The long-term safety of biologic therapies, however, is not well established,
and they are expensive. Phototherapy takes advantage of wavelengths found in natural sunlight, which, despite its
well-known deleterious effects, such as premature skin aging and increased risk of cancer, also has beneficial effects
on diseased skin. Sunlight exposure has been historically recommended to maintain health and to treat disorders, andnatural sunlight comprises beneficial wavelengths, such as 311-nm NB-UVB. Several ongoing studies areinvestigating various wavelength-dependent effects on both skin disease and the underlying immunomechanisms.
The TL01 lamp is widely used as the light source of 311-nm NB-UVB. This lamp contains mercury, however, theuse of which will soon be limited. Therefore, a light-emitting diode that emits UV will be a more desirable andfeasible light source, particularly if it has a high enough intensity level for the treatment of skin diseases.
Phototherapy is demonstrated to be a highly effective treatment option for patients with skin disease such as psoriasis,and developments to individualize treatments and improve the efficacy and safety of light therapies are ongoing.
References
1) Takuya Furuhashi, Chiyo Saito, Kan Torii, Emi Nishida, Sayuri Yamazaki, Akimichi Morita: Bath-psoralen UVA
and narrowband UVB reduce circulating Th17 cells and restore circulating regulatory T cells in psoriasis, PLoS
ONE, 8 (1), art. no. e54895, 2013
2) Ryoji Kubo, Shinnosuke Muramatsu, Yoko Sagawa, Chiyo Sait1, Saori Kasuya, Akiko Nishioka, Emi Nishida,
Sayuri Yamazaki, Akimichi Morita: Bath-PUVA therapy improves impaired resting regulatory T cells andincreases activated regulatory T cells in psoriasis, J Dermatol Sci, 86: 46-53, 2017.
3) Sayuri Yamazaki, Mizuyu Odanaka, Akiko Nishioka, Saori Kasuya, Shime Hiroaki, Hiroaki Hemmi, Masaki Imai,
Dieter Riethmacher, Tsuneyasu Kaisho, Naganari Ohkura, Shimon Sakaguchi, Akimichi Morita: Ultraviolet Binduced maturation of CD11b-type Langerin− dendritic cells control the expansion of Foxp3+ regulatory T cells
in the skin, J Immunol 200:119-129, 2018.
4) Hiroaki Shime, Mizuyu Odanaka, Makoto Tsuiji, Takuma Matoba , Masaki Imai, Yoshiaki Yasumizu, Ryuta Uraki,
Kiyoshi Minohara, Maiko Watanabe, Anthony John Bonito, Hidehiro Fukuyama, Naganari Ohkura, Shimon
Sakaguchi, Akimichi Morita, and Sayuri Yamazaki: Proenkephalin+ regulatory T cells are expanded clonally byultraviolet-B exposure and maintain skin homeostasis with a healing function, Proc Natl Acad Sci 117: 2069620705, 2020
Acknowledgements. This work was funded by a Grant-in-Aid for Scientific Research B (AM) from the Japan
Society for the Promotion of Science.

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IL-4.3.3

UVA-1 in fibrotic diseases and beyond
Calzavara-Pinton Piergiacomo1, Gelmetti Alessandra1, Arisi Mariachiara1, Venturini Marina1
1) University of Brescia, Department of Dermatology, Brescia, Italy
Ultraviolet A1 (UVA1) is the electromagnetic waveband confined in the 340-400 nm range. Unlike other UV
wavebands, UVA1 works almost exclusively via oxidative mechanisms and penetrates down to the reticular dermis
and beyond. UVA1 phototherapy was introduced as a treatment option in the 1980s when powerful metal halide
lamps became available. Afterwards, less expensive and more practical excimer lamps and LED lamps contributed
to the widespread diffusion of this treatment modality. The treatment modality is very simple, because, unlike otherphototherapies, the risk of excessive phototoxic reactions is very low and the treatment protocol is usually withoutor with very few dose increments. Since the early beginning, UVA1 phototherapy was investigated for the treatment
of atopic dermatitis, particularly acute flares of atopic dermatitis. Results were very good but if and when it is superior
to the more practical NB-UVB is still not fully clarified. Afterwards, it was studied in inflammatory skin disorders
like skin lesions of lupus erythematosus, graft versus host disease, and psoriasis in HIV+ patients that cannot betreated with NB-UVB and PUVA. UVA1 is increasingly being used also in the treatment of inflammatory andfibrosclerotic dermal diseases like morphea, hypertrophic scars, granuloma annulare and necrobiosis lipoidica.
We present an updated review of the photobiological and photoimmunological effects of UVA1 on human skin and
In addition, we will present a retrospective review of our experience with about 500 patients in the 2000-2020 timeinterval with this modality and we will discuss our results on light of a review of the literature.

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OC-4.3.4
Simulated Penetration Depth of Ultraviolet Radiation in Skin with Varying Stratum
Corneum Thicknesses as an Aid for Phototherapy
Louise Finlayson1, Isla Barnard 1 , Lewis McMillan1 , Ewan Eadie 3 , Sally H. Ibbotson2 , Christian Thomas
Alcuin Brown 1 , Kenneth Wood1
1) University of St Andrews, SUPA School of Physics and Astronomy, St Andrews, United Kingdom; 2) Universityof Dundee, Photobiology Unit, Ninewells Hospital, Dundee, United Kingdom; 3) NHS Tayside, Photobiology Unit,
Ninewells Hospital, Dundee, United Kingdom
Phototherapy has been used for many years as a convenient and non-invasive method of treatment. Ultraviolet (UV)radiation, either alone or in combination with a photosensitiser, is currently used to effectively treat skin conditions
such as psoriasis, eczema, vitiligo, scleroderma and cutaneous T-cell lymphoma.1,2 Several different types of UV
based phototherapy exist including, broadband UVB (BBUVB), narrowband UVB (NBUVB) and the combinationof the photosensitiser Psoralen and UVA (PUVA). Studies have demonstrated the effectiveness of interchanging the
choice of light treatment for the same condition based on its severity and stage in individual circumstances bychoosing the phototherapeutic source able to penetrate to the most effective depth3,4. It is clear then that accessible
data that allows images, showing the penetration depth of light into skin as a function of wavelength, to be easily
produced would be of great benefit for either selecting the best treatment option for an individual or developing new
phototherapies.
We used Monte Carlo radiative transfer (MCRT) to simulate the path of light with wavelengths 200 – 400 nm througha 6-layer skin model. A simulation was run for each wavelength, outputting a fluence rate grid which was used todetermine penetration depth. The model was adapted to investigate the effect of different thicknesses of stratum
corneum, the outermost protective layer of the skin. A web application was then developed to allow the produced
data to be easily accessed.

Fig. 1 – Penetration depth of light into skin as a function of wavelength for a stratum corneum thickness of 20µm. Data is
produced by plotting the depth at which the light fluence rate reaches 90%, 50%, 10%, 1% and 0.1% of its incident value. Datais plotted over a scaled image of the skin model where each colour represents a different layer.

The results show the penetration depth of light varying in accordance with the selected optical properties. (Fig. 1)
Predictably, the results also show that a thinner stratum corneum results in a larger penetration depth. (Fig. 2) The
results are shown for a direct light source applied to skin of Fitzpatrick scale type I. However, the simulation is fully
adaptable, allowing other light sources and skin types to be investigated given the availability of the appropriate
optical properties.

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Fig. 2 – Penetration depth of light into skin as a function of wavelength for four different stratum corneum thicknesses. Data is
produced by plotting the depth at which the light fluence rate reaches 90%, 10% and 1% of its incident value for stratum corneum
thicknesses of 20µm, 14.8µm, 10µm and 0µm. Chosen thicknesses correspond closely with common thicknesses found at different
body sites5 and skin with no stratum corneum is also investigated. Data is plotted over a scaled image of the skin model where
each colour represents a different layer.

References
1.
Vangipuram, R. & Feldman, S. R. Ultraviolet phototherapy for cutaneous diseases: A concise review. Oral
Dis. 22, 253–259 (2016).
2.
Krenitsky, A., Ghamrawi, R. I. & Feldman, S. R. Phototherapy: a Review and Update of Treatment Optionsin Dermatology. Current Dermatology Reports vol. 9 10–21 (2020).
3.
Ling, T. C. et al. British Association of Dermatologists and British Photodermatology Group guidelines forthe safe and effective use of psoralen-ultraviolet A therapy 2015. Br. J. Dermatol. 174, 24–55 (2016).
4.
Kreuter, A. et al. A randomized controlled study of low-dose UVA1, medium-dose UVA1, and narrowband
UVB phototherapy in the treatment of localized scleroderma. J. Am. Acad. Dermatol. 54, 440–447 (2006).
5.
Sandby-Møller, J., Poulsen, T. & Wulf, H. C. Epidermal Thickness at Different Body Sites: Relationship toAge, Gender, Pigmentation, Blood Content, Skin Type and Smoking Habits. Acta Derm. Venereol. 83, 410–
413 (2003).
Acknowledgements. Louise Finlayson acknowledges financial support from EPSRC Industrial Doctorate Centre
Scheme (2262922) and the Laser Research and Therapy Fund (registered charity SC030850).

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OC-4.3.8

RATIONALIZING PHOTOTHERAPY SERVICES DURING THE COVID-19
PANDEMIC: STRATEGIES AND IMPACTS ON PATIENT ACCESS ANDOUTCOMES
Tashmeeta Ahad1, 2, Sunil Kalia1, 2, 3, 4, Harvey Lui1, 2.
1) Department of Dermatology and Skin Science, University of British Columbia, Vancouver, Canada;
2) Photomedicine Institute, Vancouver Coastal Health Research Institute, Vancouver, Canada;3) Department of Cancer Control Research, BC Cancer, Vancouver, Canada;
4) BC Children’s Hospital Research Institute, Vancouver, Canada
Background: Public health directives throughout the COVID-19 pandemic significantly constrained the provisionof phototherapy services to patients with light-responsive dermatoses. At our centre, phototherapy was unavailable
from March - June 2020, and then resumed thereafter at 50% capacity.
Objectives: We will assess the impacts of COVID-19 on local phototherapy services including patient outcomes,
and the efficiency of prioritization strategies to maximize service delivery while adhering to strict infection control
protocols.
Methods: To objectively prioritize and monitor patients before and during phototherapy, nursing staff were trained
to serially assess disease severity using a validated Investigator Global Assessment times Body surface area
(IGAxBSA) cross-product scoring tool. Dermatology Life Quality Index (DLQI) questionnaires were completed
periodically by patients to monitor disease impacts. Patients were treated using either of two phototherapy regimensthat incorporated curtailed treatment frequencies (once or twice a week only) over 10 week exposure cycles; higher
incremental dosing (up to 20%) at successive sessions was used.
Results: A total of 657 patients were undergoing phototherapy prior to clinic closure. The proportion of contacted
patients declining phototherapy following re-opening was 31% (n=142). Baseline assessment of patients withpsoriasis (n=192) and eczema (n=71), revealed median IGA x BSA scores of 20 (psoriasis) and 24 (eczema). Capacity
modelling revealed a 75% cumulative percentile of IGA x BSA scoring distribution as the most optimal thresholdfor choosing treatment frequency. Based on this, patients had twice a week treatment if IGA x BSA scores were ≥30(psoriasis) and ≥40 (eczema) for ten week treatment courses. DLQI ≥21 (extremely large effect on quality of life)
qualified for twice weekly treatment regardless of disease severity. Preliminary analysis on efficacy after ten weeks
showed median IGA x BSA scores of 9 (psoriasis; n=129) (55% median improvement) and 16 (eczema; n=40) (33%
median improvement).
Preliminary Conclusions: Our ongoing review will provide unique insights into the impacts of reduced access tocore dermatological services on patient morbidity and the efficacy of objective service rationalization strategies.

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P-4.3.9

Global verification of a model for determining daylight photodynamic therapy dose
Paul O’Mahoney1,2,3, Marina Khazova4, Ethan LaRochelle5, Brian Pogue5, Sally Ibbotson1,2,3, Ewan Eadie2,31) University of Dundee, School of Medicine, Dundee, UK; 2) NHS Tayside, Photobiology Unit, Dundee, UK; 3)
The Scottish Photodynamic Therapy Centre, Dundee, UK; 4) Public Health England, Didcot, UK; 5) Thayer
School of Engineering at Dartmouth, Hanover, USA
Daylight photodynamic therapy (DPDT) is a widely used, effective treatment for field change actinic keratoses (AK).
Successful treatment does, however, rely upon a minimum daylight exposure during treatment. This daylight
exposure is quantified as the protoporphyrin-IX (PpIX)-effective exposure dose, with current guidelines suggestinga minimum of 8 J cm-2 to be delivered over a 2-hour treatment period [1]. Measurement of daylight exposure during
DPDT is an important parameter, which can help assist practitioners in delivering optimal therapy.
We recently published a model for DPDT dosimetry (the O’Mahoney model) [2], allowing relatively simple
measurements of illuminance from a widely available lux-meter to be accurately converted to PpIX-effective
exposure dose (omahoney.shinyapps.io/daylightpdt). Originally validated for use within the UK, in order for themodel to be used confidently in other locations, we verified its use globally.
We obtained daylight spectral irradiance measurements from 4 locations: Lebanon NH, USA (43.7 °N, 72.40 °W);
Paris, France (48.86 °N, 2.35 °E); Nakhon Pathom, Thailand (13.82 °N, 100.05 °E); and Ribeirão Preto, Brazil (21.18°S, 47.81 °W). Using these spectral irradiance data, we derived both the illuminance and the PpIX-effective irradiancefor each data point (n=33,602). The conversion model was applied to the illuminance data and compared to the PpIXeffective irradiance under all weather conditions.
The median percentage deviation of the model from the true PpIX-effective irradiance was within ±10%, and 95%
confidence intervals within ±20%. These results are within expectations with respect to a similar analysis carried out
on UK spectral irradiance from our previous publication. The results of this study verify that this conversion modelfor DPDT dosimetry is suitable for use across the globe and can very simply enhance the confidence of DPDT
practitioners with respect to daylight dosimetry.
References
1) Morton et al., Journal of the European Academy of Dermatology and Venerology, 2019; vol 33, 2225-2238
2) O’Mahoney et al., British Journal of Dermatology, 2016; vol 176, 1607-1616
Acknowledgements. Dr Paul O‘Mahoney is funded by Medi-lase (registered charity SC 037390) and the Alfred
Stewart Trust. The authors would like to thank the following for their contributions to the provision of data used inthis work: Prof Luciano Bachmann and Prof Gilberto Braga (University of São Paulo, Brazil), Dr Sumaman Buntoung(Silpakorn University, Thailand) and Dr Einapak Boontaveeyuwat (Chulalongkorn University, Thailand)Book of Abstracts ������������������������������������������������������������������������������������������������ 193

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P-4.3.10

Redox processes in pathogenesis and phototherapy of vitiligo
T. Telegina1,2, Yu. Obukhov1,2, Yu. Vechtomova1, A. Buglak2,
1) Bach Institute of Biochemistry, Research Center of Biotechnology of the Russian Academy of Sciences. Moscow,
Russia; 2) Saint Petersburg State University, Saint Petersburg, Russia
Disturbances of homeostasis in skin cells lead to various diseases, including vitiligo, in which melanogenesis in
epidermal melanocytes is impaired. Melanocytes like all living cells maintain a balance between oxidation andreduction, and perturbations of this balance are known to contribute to vitiligo pathogenesis. Vitiligo is an
autoimmune disease and oxidative stress plays a significant role in melanocyte disappearance. With vitiligo, an
excess of H2O2 accumulates in melanocytes. Then H2O2 induces the synthesis of a 3-5-fold excess oftetrahydrobiopterin (H4Bip) through γ-interferon, since cyclohydrolase is γ-interferon-induced enzyme. In turn, theinevitably proceeding process of H4Bip autooxidation leads to the formation of H2O2. In this way, the autocatalytic
cycle increases oxidative stress. Research objectives for the treatment of vitiligo are as follows: 1) to stop the
autocatalytic cycle and 2) to restore balanced redox system. The autocatalytic cycle can be stopped by exposure to
UVB radiation. We have shown that UVB radiation converts H4Bip into dihydropterin dimers (quantum yield
equals 2.4%) and thus it is possible to remove the excessive H4Bip, which arises during the autocatalytic cycle. The
mechanism of the process was studied and the maximum of the UV action spectrum was found. Good therapeutic
effects were achieved with the combined action of UVB and an antioxidant enzyme that decomposes H2O2, forexample, pceudocatalase. Restoration of a balanced redox system in melanocytes appears to be resolved by
activation of the Nrf2-ARE signaling pathway, which is impaired in vitiligo. The Nrf2-ARE pathway protects
melanocytes from H2O2 damage through the induction of the antioxidant enzyme system. A number of Nrf2
activators have been used in treating vitiligo with certain therapeutic effects.
Acknowledgements. The research was supported by the Russian Science Foundation grant 20-73-10029.

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P-4.3.11

UVA photoprotective effect of adenine derivate kinetin
Denisa Škařupová1*, Alena Ryšavá1, Jitka Vostálová1, Alena Rajnochová Svobodová1, Jitka Ulrichová1, Lucie
Plíhalová2, Bohumil Zálešák3
1) Department of Medical Chemistry and Biochemistry, Faculty of Medicine, Palacký University Olomouc,Hněvotínská 3, 775 15 Olomouc, Czech Republic; 2) Laboratory of Growth Regulators, Faculty of Science,
Palacký University Olomouc & Institute of Experimental Botany AS CR, Šlechtitelů 27, 783 71 Olomouc, Czech
Republic; 3) Department of Plastic and Aesthetic Surgery, University Hospital Olomouc, I. P. Pavlova 6, 775 20
Olomouc, Czech Republic.
*denisa.skarupova@upol.cz
Overexposure of human skin to solar UV radiation (295-400 nm) is associated with many detrimental effects thatcan result in premature skin aging (photoaging), various skin disorders and photocarcinogenesis. UVA radiation(320-400 nm), the major part of UV light, reaches both epidermal and dermal layer of the skin and triggers massive
generation of reactive oxygen species (ROS). Overflow of ROS results in oxidative modification of biomolecules(proteins, nucleic acids, lipids and their structural units), further production of reactive compounds and finally inoxidative stress (OS).
Several antioxidants and cytoprotective proteins are under control of NF-E2-related factor 2 (Nrf2). Thistranscription factor plays a key role in the defence against xenobiotics, heavy metals and oxidative stress due to its
ability to induce the expression of detoxifying enzymes, antioxidant proteins and the suppression of proinflammatory signalling pathways. Under normal condition, Nrf2 retains in the cytoplasm via interaction with the
Kelch-like ECH associated protein (Keap1). In response to oxidative damage, Nrf2 translocates to the nucleus
where it forms a complex with other transcription factors and binds to the promoters of its target genes, antioxidant
response element (ARE). This leads to activation of phase 2 detoxifying enzymes and stress-response related
proteins such as heme oxygenase 1 (HO-1), NAD(P)H:quinone oxidoreductase 1 (NQO1), catalase (CAT),glutamate-cysteine ligase catalytic subunit (GCLC)1.
One possibility to strengthen skin photoprotection is topical application of dermatological preparations containing
active ingredients that neutralize UV photons, improve skin regeneration and restore tissue homeostasis. For thispurpose, several naturally occurring compounds derived from plants have been investigated/used.
In this study, we examined adenine derivate kinetin (KIN, N6-furfuryladenin), a plant hormone that controls cell
growth and differentiation. KIN has found application as the active ingredient in several anti-ageing products incosmetics but minimal knowledge about its mechanism of action is available2.
The aim of this study was to determine whether KIN is able to modulate OS and the signalling pathway controlled
by Nrf2 in normal human dermal fibroblasts after UVA radiation. KIN has decreased UVA induced ROS
production and glutathione depletion and activated the Nrf2 signalling pathway by increasing the nuclear level of
Nrf2, which was detected by immunohistochemical analysis. This effect was further evaluated by measuring thelevel of HO-1, NQO1, GLCL and CAT by western blot analysis. Kinetin UVA photoprotective properties will be
discussed in our poster contribution.
References
1) Boo Y. Ch. (2020): Natural Nrf2 modulators for skin protection. Antioxidants 9 (9), 812.
2) Jablońska-Trypuć A., Matejczyk M., Czerpak R (2016): N6-benzyladenine and kinetin influence antioxidative
stress parameters in human skin fibroblasts. Molecular and Cellular Biochemistry 413, 97-107.
Acknowledgements. This work was financially supported by the grant IGA_LF_2021_013 and by the Institutional
Support of Palacký University in Olomouc.

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IL-4.4.1
Understanding the Sequence of Events During the Water Oxidation Reaction in
Photosystem II using Crystallography/X-ray Spectroscopy
Junko Yano1, Jan Kern1, Nicholas K. Sauter1, Holger Dobbek2, Uwe Bergmann3, Johannes Messinger4, Athina
Zouni2, Vittal K. Yachandra1
1) MBIB, Lawrence Berkeley National Lab., Berkeley, USA; 2) Institut für Biologie, Humboldt-Universität zu
Berlin, Berlin, Germany; 3) Dept. Physics, Univ. Wisconsin, Madison, USA; 4) Dept. Chemistry, Uppsala Univ.
Uppsala, Sweden.
Light-driven oxidation of water to molecular oxygen is catalyzed by the oxygen-evolving complex (OEC) in
Photosystem II (PS II). This multi-electron, multi-proton catalysis requires the transport of two water molecules toand four protons from the OEC. Recent advances in X-ray free-electron laser (XFEL)-based room temperature (RT)crystallography enabled us to study the dynamics of the structure of PS II under functional conditions (1, 2). The
ability to take snapshots of the structure at the various time points at RT during the reaction allows for theinvestigation of the structural changes at the OEC, amino acid sidechain movements, and changes in hydrogenbonding networks in the enzyme (Figure 1). These studies can provide new insights into the reaction mechanism in
PS II by potentially identifying water and proton pathways.
The results show that the catalytic reaction at
the OEC in PS II is well coordinated with the
motion of both close and distant residues fromthe OEC, playing roles in shuttling substratewater and protons essential for the wateroxidation reaction.
Fig. 1 – Schematic of the S2 ® S3 transition. The
sequence of events in time leading to the insertionof OX between Mn1 and Ca. Mn1 oxidation from
(III) to (IV) is shown as a color change from orange
to purple. The other Mn atoms are all in oxidation
state (IV) and are shown in purple. Ca is shown ingreen. The ligands of Mn and Ca, YZ and otherneighboring residues and the water ligands of Mn4
and Ca are shown. Possible pathways for protontransfer are depicted as well.

References
1) Ibrahim, M. et al. Untangling the Sequenceof Events During the S2 ® S3 Transition in
Photosystem II: Implications for the Water
Oxidation Mechanism, PNAS, 117, 12624,
2020.
2) Kern, J. et al. Structures of the intermediatesof Kok’s photosynthetic water oxidation clock, Nature, 563, 421, 2018.
Acknowledgements. This work was supported by the Director, Office of Science, Office of Basic Energy Sciences(OBES), Division of Chemical Sciences, Geosciences, and Biosciences (CSGB) of the Department of Energy (DOE),
U.S.A., for X-ray methodology and instrumentation, and spectroscopy and crystallography data collection andanalysis.

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IL-4.4.2
Identity of EPR-detected intermediates in biological water oxidation
Dimitrios A. Pantazis
Max-Planck-Institut für Kohlenforschung, Department of Molecular Theory and Spectroscopy,
Kaiser-Wilhelm-Platz 1, 45470 Mülheim an der Ruhr, Germany.
The Mn4CaOx cluster of the oxygen-evolving complex (OEC) of photosystem II adopts different electronic andgeometric forms as it progresses physiologically through the catalytic water-oxidizing cycle, but also as a result ofvarious physicochemical treatments used to probe specific aspects of its structure and function. Crystallographic
studies have achieved reliable atomistic representations of the cluster, and X-ray free electron laser (XFEL)
techniques promise even greater achievements. Nevertheless, the information content of such data is still insufficient
to produce geometric models that are uniquely defined and chemically accurate, or that satisfy in obvious ways theelectronic structure constraints provided by spectroscopy. Among the spectroscopies employed in the study of theOEC, magnetic resonance techniques offer the most detailed and selective insight into the electronic structure of thecatalyst, as they provide highly local information unaffected by heterogeneity. The structural interpretation ofmagnetic resonance data for any system as complex as the OEC necessitates the use of quantum chemical modelling.1
Here I present selected experimental and theoretical studies from my group that focus on the identity of different
EPR-detected forms of the OEC in the S1, S2, and S3 states of the catalytic cycle, highlighting the different types ofisomerism and heterogeneity (e.g. Jahn–Teller isomerism, valence isomerism, hydration equilibria) that are present
in each one of these states,2,3,4 and discussing possible implications for the mechanism of biological water oxidation.

Fig. 1 – A) Orientational Jahn–Teller isomerism in the S1 state explains two EPR signals in dark-adapted samples and signals
attributed to S1YZ• metalloradical intermediates.2 B) The Jahn–Teller isomers naturally evolve into distinct valence isomericforms in the S2 state.2,3 C) EPR studies of the S3 state indicate the presence of a water-unbound high-spin and high-anisotropy
species as a major structural form, alongside multiple intermediate-spin low-anisotropy water-bound populations.4

References
1) M. Orio, D. A. Pantazis, “Successes, challenges, and opportunities for quantum chemistry in understanding
metalloenzymes for solar fuels research”, Chem. Commun. 2021, 57, 3952-3974.
2) M. Drosou, G. Zahariou, D. A. Pantazis, “Orientational Jahn–Teller Isomerism in the Dark-Stable State of Nature’s
Water Oxidase”, Angew. Chem., Int. Ed. 2021, DOI: 10.1002/anie.202103425.
3) D. A. Pantazis, W. Ames, N. Cox, W. Lubitz, F. Neese, “Two Interconvertible Structures that Explain theSpectroscopic Properties of the Oxygen-Evolving Complex of Photosystem II in the S2 State”, Angew. Chem., Int.
Ed. 2012, 51, 9935-9940.
4) G. Zahariou, N. Ioannidis, Y. Sanakis, D. A. Pantazis, “Arrested substrate binding resolves catalytic intermediates
in higher-plant water oxidation”, Angew. Chem., Int. Ed. 2021, 60, 3156-3162.

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IL-4.4.3

Electron and Proton Releasing Sites in the Oxygen-Evolving Complex of
Photosystem II
Hiroshi Ishikita1
1) Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Meguro-ku,
Tokyo 153-8904, Japan
In photosystem II (PSII), O2 evolution proceeds at the Mn4CaO5 cluster, releasing four protons. Formation of the
proton-conducting wire via H-bonds is prerequisite for proton transfer. In particular, when the pKa values of the Hbond donor and acceptor moieties are nearly equal, a short low-barrier H-bond forms and the release of the proton
from the H-bond donor moiety occurs most efficiently along the barrier-less potential (Figure 1). The O…O distancesin typical low-barrier H-bonds are < ~2.5 Å, which are significantly shorter than the O…O distances of ~2.8 Å fortypical standard H-bonds. According to Warshel (2013 Nobel Prize in Chemistry), a low-barrier H-bond cannot be
defined by only the distance or strength of an H-bond: identification of a low-barrier H-bond can be valid only if the
shape of the potential-energy profile of the H-bond is symmetric (i.e., the pKa values for the two moieties are nearly
equal) (1). Among short H-bonds identified in the PSII crystal structures, the following H-bonds, TyrZ…D1-His190(2.47 Å) (2), QB…D1-His215 (2.47 Å) (3), and O4…W539 (2.45 Å) (4) are recognized as low-barrier H-bonds.
LBHB

pKa

energyenergystandard

DpKa

Oacceptor
long

Odonor

DpKa ≈ 0

O

O
short

Fig. 1 –Typical potential-energy profiles of H-bonds. (left) Standard H-bond. (right) Low-barrier H-bond (LBHB).

The X-ray free electron laser (XFEL) structures also show significantly short O…O distances, in particular, at the
Mn4CaO5/6 moiety, e.g., O5…O6 (1.90 Å) (5) and O4…H2O (2.25 Å) (5). In this talk, the chemical origins of theshort O...O distances will be disucssed (6, 7).

References
1.
C. N. Schutz, A. Warshel, The low barrier hydrogen bond (LBHB) proposal revisited: the case of the Asp...
His pair in serine proteases. Proteins 55, 711-723 (2004).
2.
K. Saito, J.-R. Shen, T. Ishida, H. Ishikita, Short hydrogen-bond between redox-active tyrosine YZ and D1His190 in the photosystem II crystal structure. Biochemistry 50, 9836-9844 (2011).
3.
K. Saito, A. W. Rutherford, H. Ishikita, Mechanism of proton-coupled quinone reduction in Photosystem II.
Proc Natl Acad Sci U S A 110, 954-959 (2013).
4.
K. Saito, A. W. Rutherford, H. Ishikita, Energetics of proton release on the first oxidation step in the wateroxidizing enzyme. Nat. Commun. 6, 8488 (2015).
5.
M. Suga et al., An oxyl/oxo mechanism for oxygen-oxygen coupling in PSII revealed by an X-ray freeelectron laser. Science 366, 334-338 (2019).
6.
M. Mandal, K. Saito, H. Ishikita, The nature of the short oxygen-oxygen distance in the Mn4CaO6 complexof the photosystem II crystals. J. Phys. Chem. Lett. 11, 10262-10268 (2020).
7.
M. Mandal, K. Saito, H. Ishikita, Two Distinct Oxygen-Radical Conformations in the X-ray Free Electron
Laser Structures of Photosystem II. J. Phys. Chem. Lett. 10.1021/acs.jpclett.1c00814, 4032-4037 (2021).

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OC-4.4.4
The rise and fall of the photoinhibition-related energy dissipation qI
Wojciech J. Nawrocki1,2,*, Xin Liu1,2, Bailey Raber3, Chen Hu1,2, Sandrine Bujaldon4, Catherine de Vitry4, Doran I.
G. Bennett3, Roberta Croce1,21) Biophysics of Photosynthesis, Department of Physics and Astronomy, Faculty of Science, Vrije Universiteit
Amsterdam, 1081 HV Amsterdam, The Netherlands; 2) LaserLaB Amsterdam, Vrije Universiteit Amsterdam, 1081
HV Amsterdam, The Netherlands; 3) MesoScience Lab, Department of Chemistry, Southern Methodist University,
Dallas TX, USA; 4) Institut de Biologie Physico-Chimique, UMR 7141 CNRS-Sorbonne Université, 75005 Paris,
France; *w.j.nawrocki@vu.nl
Plants use light energy to oxidise water and reduce carbon dioxide in the process of photosynthesis. However,while light is necessary for carbon fixation, it can also damage the photosynthetic machinery - in particular thewater-splitting complex - causing photoinhibition of Photosystem II.
While the inactive PSII in the closed state (QA-) exhibits high fluorescence yield due to the lack of ongoing
photochemical quenching, upon photoinhibition the level of fluorescence decreases. This slowly-relaxing
quenching is termed qI. Here, we investigate the fate of the excitation energy after the establishment of qI in thepresence of photodamaged PSII reaction centres, in a context of a wild-type or altered proteostasis. We use timeresolved and steady-state fluorescence together with oxygen evolution measurements and biochemistry, which we
combine with genetic approaches using the green alga Chlamydomonas reinhardtii. Finally, we integrate the results
using membrane-scale structure-function modelling of excitation energy transfer to reveal the site and possible
mechanism of the qI quenching.

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IL-4.4.5
Fast substrate water exchange in the S2 state of photosystem II is controlled byproteins surrounding the Mn4CaO5 cluster – implications for the water oxidation
mechanism
Casper de Lichtenberg1,2, Christopher J. Kim3, Petko Chernev2, Richard J. Debus3 & Johannes Messinger1,21) Umeå University, Department of Chemistry, Umeå, Sweden; 2) Uppsala University, Department of Chemistry –Ångström Laboratory, Uppsala, Sweden; 3) University of California – Riverside, Department of Biochemistry,
Riverside, California, USA
The molecular oxygen we breathe is produced from two water-derived oxygen species bound to the Mn4CaO5 clusterin photosystem II (PSII). Present research points to the central oxo-bridge O5 as one of these ‘substrate waters’,
while, in the S2 state, the terminal water ligands W2 and W3 both remain candidates for the second substrate.1-3
Intense efforts are also ongoing to identify the main access pathway for water to the catalytic site, and to determine
whether specific amino acids limit the exchange rate of substrate with bulk water. In this study, we measured therates of H216O/H218O substrate water exchange in the S2 and S3 states of wild-type (WT), D1-E189Q and D1-D61A
PSII core complexes from Synechocystis sp. PCC 6803. We found that the exchange rates were unaffected by the
E189Q mutation (O1 channel), but strongly perturbed by the D61A mutation (Cl1/O4 channel). It is concluded thatD61 forms a rate-limiting barrier for the isotopic equilibration of the inner water pool near the Mn4CaO5 cluster thatis alliviated in the D1-D61A mutant (Figure 1). This finding removes the main argument against the Ca-bound W3
as fast substrate water in the S2 state, namely the indifference of its exchange rate towards Ca/Sr substitution. Ourdata are thus in full agreement with the proposal that W3 forms the new Ox/O6 bridge in the S3 state, and that the OO bond is formed via oxo-oxyl radical coupling between O5 and Ox in the S4 state. The role of multiple conformations
in each S state as well as possible alternative assignments of the substrate waters will be discussed.
Fig. 1 – Schematic of
the Cl1-channel inphotosystem II withestimated energy
barriers for waterexchange. The
truncation of the D1D61 to D1-A61 is
proposed to open up
the major bottleneck
for the equilibrationof the inner waterpool that includes W3.

References
1) N. Cox, J. Messinger, Reflections on substrate water and dioxygen formation, Biochim. Biophys. Acta 1827,
1020-1030, 2013.
2) C. de Lichtenberg, J. Messinger, Substrate water exchange in the S2 state of photosystem II is dependent on theconformation of the Mn4Ca cluster, Phys. Chem. Chem. Phys. 22, 12894-12908, 2020.
3) C. de Lichtenberg, A. P. Avramov, M. Zhang, F. Mamedov, R. L. Burnap and J. Messinger, The D1-V185N
mutation alters substrate water exchange by stabilizing alternative structures of the Mn4Ca-cluster in photosystem
II, Biochim. Biophys. Acta 1862, 148319, 2021.
Acknowledgements. This work was supported by ÅForsk (to CDL), Vetenskapsrådet (2016-05183 and 2020-03809;
to JM) and the US Department of Energy (DE-SC0005291; to RJD).
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IL-4.4.6

Infrared study on the mechanisms of water oxidation at the Mn cluster and its
photoassembly in photosystem II
Takumi Noguchi
Graduate School of Science, Nagoya University, Nagoya, Japan
Photosynthetic water oxidation is carried out in photosystem II (PSII) embedded in the thylakoid membranes ofplants and cyanobacteria. The catalytic center is the Mn4CaO5 cluster, which is assembled on electron donor side of
PSII by light-driven process called photoactivation. Recent crystallographic studies using an X-ray free electron
laser have shown the structures of the Mn4CaO5 cluster and the surrounding protein moiety in the dark stable state(S1 state) and flash-induced intermediates (S2, S3, and S0 states).1 However, the details of the reactions in theintermediate transitions and the process of the photoassembly of the Mn4CaO5 cluster remain unresolved. In thisstudy, we have investigated the molecular mechanisms of the photoassembly of the Mn4CaO5 cluster and the wateroxidation reaction using light-induced Fourier transform infrared (FTIR) difference and time-resolved infrared
spectroscopies.
(1) Photoassembly process of the Mn4CaO5 cluster. We applied rapid-scan time-resolved FTIR spectroscopy
combined with the attenuated total reflection (ATR) technique to monitor the photoassembly process of the
Mn4CaO5 cluster.2 Rapid-scan ATR-FTIR spectra of apo-PSII with Mn2+ upon flash illumination showed spectralfeatures typical of the carboxylate stretching vibrations. These features were attributed to the two carboxylategroups from D1-D170 and D1-E189 coordinating to the high affinity Mn2+ ion by quantum chemical calculations.
The FTIR signal decayed with a time constant of ~0.7 s, showing that the following “dark rearrangement” step
occurred with a low quantum yield and the oxidized Mn3+ ion was mostly released during this decay. Simulation of
the kinetic process provided a slow intrinsic rate (τ = ~20 s) of the dark rearrangement, which was attributed to thelarge protein conformational change of the C-terminus of the D1 protein and relocation of the Mn3+ ion. The
structural fluctuations of the lumenal domain of the CP43 protein visualized by high-speed atomic force
microscopy3 is suggested to facilitate this dark process. The model of the early process of the photoassembly of the
Mn4CaO5 cluster was proposed (Figure 1).

Fig. 1 – Schematic model of theearly steps of the photoassembly of
the Mn4CaO5 cluster.

(2) Water and proton pathways during the S2→S3 transition. The S2→S3 transition, which involves concerted waterand proton transfer, is a key process in the water-oxidation mechanism. To identify the water and proton pathways
during the S2→S3 transition, we examined the effects of D1-N298A mutation and NO3− substitution for Cl−, which
perturbed the O1 and Cl channels, respectively, on the S2→S3 kinetics using time-resolved infrared spectroscopy.
The S2→S3 transition was retarded both upon NO3− substitution and upon D1-N298A mutation, whereas it was
unaffected by further NO3− substitution in the N298A PSII. The H/D kinetic isotope effect in the N298A PSII was
small, revealing that water transfer is a rate-limiting step in this mutant. From these results it was suggested thatduring the S2→S3 transition, water delivery and proton release occur through the O1 and Cl channels, respectively
References
1) M. Suga, et al., Nature, 543, 131−135 (2017); J. Kern, et al., Nature, 563, 421−425 (2018).
2) A. Sato, Y. Nakano, S. Nakamura, and T. Noguchi, J. Phys. Chem. B, 125, 4031–4045 (2021).
3) T. Tokano, Y. Kato, S. Sugiyama, T. Uchihashi, and T. Noguchi, J. Phys. Chem. B, 124, 5847–5857 (2020).

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IL-4.4.7
Light-driven formation of high-valent manganese oxide by photosystem II supports
evolutionary role in early bioenergetics
Petko Chernev,1,2 Sophie Fischer,1 Jutta Hoffmann,1 Nicholas Oliver,1 Ricardo Assunção,1 Boram Yu,1 Robert L.
Burnap,3 Ivelina Zaharieva,1 Dennis J. Nürnberg,1 Michael Haumann,1 Holger Dau1
1) Freie Universität Berlin, Physics Department, Berlin, Germany; 2) present address: Uppsala University,
Department of Chemistry, Uppsala, Sweden; 3) Oklahoma State University, Dept. of Microbiology and Molecular
Genetics, Stillwater, United States
Water oxidation and concomitant O2-formation by the Mn4Ca cluster of oxygenic photosynthesis has shaped thebiosphere, atmosphere, and geosphere. It has been hypothesized that at an early stage of evolution, before
photosynthetic water oxidation became prominent, photosynthetic formation of Mn oxides from dissolved Mn2+ ions
may have played a key role in bioenergetics and possibly facilitated early geological manganese deposits. Here theexperimental evidence for the ability of photosystems to form extended Mn oxide particles, lacking until now, isreported. We tracked the light-driven redox processes in spinach photosystem II (PSII) particles devoid of the Mn4Ca
cluster by UV-vis and X-ray spectroscopy. We find that oxidation of aqueous Mn2+ ions results in PSII-bound
Mn(III,IV)-oxide nanoparticles of the birnessite typecomprising 50-100 Mn ions per PSII.
Having shown that even today's photosystem-II canform birnessite-type oxide particles efficiently, we
propose an evolutionary scenario, which involves Mnoxide production by ancestral photosystems, later
followed by down-sizing of protein-bound Mn-oxide
nanoparticles to finally yield today's Mn4CaO5 clusterof photosynthetic water oxidation. Moreover, we
discuss the hypothesis of an early quasi-respiratory
cycle that involves formation of Mn(III/IV) oxide
particles followed by utilization of the oxidizing
equivalents stored in the Mn oxide for an efficientquasi-respiratory activity in the Archean or early
Paleoproterozoic, when the Earth’s atmosphere had
been essentially O2-free, as detailed in ref1.

Fig. 1 – Photosystem II depleted of the PsbO protein (yellowvoid) and proposed formation site of Mn oxide nanoparticles(schematically shown as beeing spherical).

The above results, reported in ref2, are complemented
by very recent findings addressing inter alia thequestion why Nature did choose manganese at thecatalytic site of photosynthetic water oxidation(versus Fe, Co, and Ni; Oliver et al, unpublished).

References
1) Dau, H., Nürnberg, D.J., and Burnap, R.L. (2019). Local cycle of photosynthesis and quasi-aerobic respiration
facilitated by manganese oxides – a hypothesis on the evolution of phototrophy. In Bioenergetics: Oxygen Production
and Reduction, J. Barber, P. Nixon, and A.V. Ruban, eds. (World Scientific Publishers), pp. 367-396 (2019)
2) Chernev, P., Fischer, S., Hoffmann, J., Oliver, N., Assuncao, R., Yu, B., Burnap, R.L., Zaharieva, I., Nurnberg,
D.J., Haumann, M., and Dau, H. (2020). Light-driven formation of manganese oxide by today's photosystem II
supports evolutionarily ancient manganese-oxidizing photosynthesis. Nature Communications 11, ARTN 19378
10.1038/s41598-020-75397-8 (2020)

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OC-4.4.8

Harvesting the far-red with plant antenna complexes incorporating chlorophyll d & f
Eduard Elias1, Nicoletta Liguori1, Judith Schäfers1, Yoshitaka Saga2, Roberta Croce1
1) Vrije Universiteit Amsterdam, Faculteit der Exacte Wetenschappen, AmsterdamThe Netherlands; 2) Kindai
University, Faculty of Science and Engineering, Osaka, Japan
Increasing the absorption cross-section of plants by introducing far-red absorbing chlorophylls (Chls) has been
proposed as a strategy to boost crop yields. To make this strategy effective, these Chls should bind to thephotosynthetic complexes without altering their functional architecture. To investigate if plant-specific antennacomplexes can provide the protein scaffold to accommodate these Chls, we have reconstituted the main lightharvesting complex of plants LHCII, with the red-shifted Chls d & f. The results demonstrate that LHCII can naturallybind these exogenous Chls, shifting the maximum absorption ~30 nm towards the red with respect to the wildtype
complex (LHCII with Chl a and b), while maintaining the native LHC architecture. A variety of spectroscopic
measurements show that the complexes binding the red-shifted Chls are functional in light harvesting and excitationenergy transfer. Overall, we here demonstrate that it is possible to obtain plant LHCs with enhanced far-red
absorption and intact functional properties.
Acknowledgements. This work is supported by The Netherlands Organization for Scientific Research (NWO), via
a TOP grant (to R.C). N.L. was supported by a NWO Veni grant. Y.S by a JSPS KAKENHI grant (JP17KK0106).

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PL-8
Photoremediation
Mathias O. Senge1
1) Trinity College Dublin, The University of Dublin, School of Chemistry, Trinity Biomedical Sciences InstituteDublin, Ireland
Light can kill and light can heal, as already exemplified in Herodotus’ heliotherapy. Light also has global relevancethrough photosynthesis and the importance of atmospheric photochemical reactions for life on earth. Critical global
problems are weapons of mass destruction, pandemics, food production, climate change, environmental protection,clean energy production, green chemistry, and developing a circular economy. Throughout their development thephotosciences have contributed advances to many of these areas. Critical contributions of photobiology were madeto agriculture, health, photosynthesis, biorefineries or of photochemistry to photovoltaics, photonics materials andphotocatalysis, and more.
In photomedicine light is used to promote and restore human health and similarly light plays a central role in restoring
the health of our planet. Light can mitigate, repair, and remediate many of the global problems outlined above. In theclassic photobiological field of photodynamic therapy we have seen a recent resurgence of studies on antimicrobial
photodynamic therapy driven by increased antibiotic resistance and new threats like the Covid-19 pandemic. The
pressing need to tackle environmental issues has prompted research on protection and remediation using lightactivated systems for the identification, degradation or transformation of pollutants. Classic approaches onphotodecomposition or disinfection using photosensitizers are now combined with detection systems and one canenvisage complex devices that can detect, identify, and combat pollutants, CBRN materials, and microbes. The goal
is not only to degrade or sequester pollutants but to transform them into benign and value-added materials in a mannersuitable for a circular economy.
It is proposed that photoremediation can serve as a broader discipline to unify the currently often uncoordinatedefforts to ‘detect, identify, mitigate, transform/remediate’ harmful entities. Combined with the established field ofbioremediation, which also requires light for photosynthesis, and perhaps aligned with the biorefinery concept it
allows a more strategic approach – akin to the One Health initiative – towards tackling pollution, photochemical
synthesis, energy conversion and more.

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PL-9
Plant based biohybrid systems
Eleni Stavrinidou1
1) Laboratory of Organic Electronics, Department of Science and Technology, Linköping University, 601 74,
Norrköping, Sweden
Plants convert solar energy to chemical energy, sequester carbon, sample the environment, and synthesize a variety
of materials. Augmenting functionalities to plants with smart materials and devices can result in biohybrid systems
for energy harvesting, environmental monitoring and in-vivo biofabrication. Previously we demonstrated a
conjugated oligomer that was uptaken by the vascular tissue of the plant and in vivo polymerized forming conductors.
The thiophene-based molecule polymerized within the living tissue, without external chemical or physical stimuli
only due to the physiochemical environment of the plant. We used the biohybrid system for energy storage. Recently,
we unravelled the mechanism that drives the polymerization reaction and found that the polymerization is driven
from the defence mechanism of the plant through enzymes that are involved in modulating cell wall density. Within-vitro and in-vivo studies we identified the key components, limiting factors and kinetics of the polymerization
reaction. Currently we are functionalizing rooted plants forming conductors in parallel with the growth of the plant,with the conjugated polymers integrating into the plant cell wall. In this talk I will present our latest progress on
augmenting plants with electronic functionality.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-5.1.1

Quenched-phosphorescence oxygen sensing platforms for biomedical research andphotobiology
Dmitri B.Papkovsky
School of Biochemistry and Cell Biology, University College Cork, Cork, Ireland
Molecular oxygen (O2) plays major roles in biology and photobiology as key environmental parameter, metabolic
substrate, signalling molecule and source of ROS, therefore monitoring and precise control of O 2 concentration inexperimental settings with biological samples is of paramount importance. Quenched-phosphorescence O2 sensing,particularly in lifetime based mode, provides the basis for a variety of simple, robust and accurate detection platforms
that can perform different analytical tasks1. In recent years, we have designed a number of such sensor systems basedon the longwave emitting Pt-porphyrin and Pt-benzoporphyrin dyes, demonstrated them in physiological studies withvarious in vitro and ex-vivo cell and tissue models, and commercialised.
Thus, the solid-state O2 sensor stickers in conjunction with a handheld optical reader facilitate macroscopic controlof O2 by means of non-invasive contactless measurements, e.g. in environmental chambers or sealed vessels. The
low-cost calibration-free sensors can be used in convenient and flexible manner, on continuous or disposable basis.
The cell-impermeable water-soluble O2 probes and sensor coatings provide high-throughput analysis of respirationof mammalian and bacterial cells, which can be measured on standard multi-label readers and in standard microwellplates and customised substrates for cell analysis. Multiplexed with the corresponding pH sensors, they enabledetailed multi-parametric assessment of cell metabolism and metabolic rewiring in disease states2.
The cell-permeable small molecule and nanoparticle based O2 probes, which provide efficient phosphorescent
staining of various mammalian cells, can inform on in situ oxygenation in samples containing respiring mammaliancells and 3D tissue models. They enable high-resolution mapping of O2 concentration in cells and 3D micro-tissue
samples and monitoring of their responses to metabolic stimulation by PLIM (Phosphorescence Lifetime Imaging)microscopy and macroscopy2. A separate stream of applications includes in vivo O2 imaging using dedicatedintravascular probes and 2-photon PLIM microscopy3.

This talk will describe the operation principles of these O2 sensing probes and measurement methodologies, andprovide representative examples of their applications.
References
1) Papkovsky DB, Dmitriev RI (Eds.), Quenched-phosphorescence Detection of Molecular Oxygen:
Applications in Life Sciences, RSC Detection Sciences, 2018, ISBN 978-1-78801-175-4.
2) Papkovsky DB, Zhdanov AV, Phosphorescence based O2 sensors – essential tools for hypoxia research,cell and tissue physiology (Review), Free Rad Biol Med, 2016, 101: 202–210.
3) Esipova et al., Oxyphor 2P: a high-performance probe for deep-tissue longitudinal oxygen imaging, Cell
Metabolism, 2019, 29: 736–744.
Acknowledgements. This work was supported by the Science Foundation Ireland, grant 12/RC/2276_P2.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-5.1.2

Mapping microscopic viscosity and temperature using molecular rotors
Marina K Kuimova1
1) Imperial College London, Chemistry Department, London, UK;
Viscosity is one of the main factors which influence diffusion in condensed media. In a cell viscosity can play a rolein several diffusion mediated processes, such as drug delivery, signalling and mass transport. Previously, alterations
in viscosity in cells and organs have been linked to malfunction; however, mapping viscosity on a single-cell scaleremains a challenge.
We have imaged viscosity inside lipid mono- and bi-layers, in cells and in atmospheric aerosol particles using
fluorescent probes, called molecular rotors.1, 2 In molecular rotors the speed of rotation about a sterically hinderedbond is viscosity-dependent, which strongly affects fluorescence lifetime or spectra of rotors, allowing fluorescenceimaging. This approach enabled us to measure both the microscopic viscosity and temperature2, 3 and monitor theirtemporal changes in real time. The talk will cover our recent developements of this technique, such as genetic andpassive targeting of rotors4, 5 and applications to monitoring neurodegeneration.6
References
1) M. K. Kuimova, Phys Chem Chem Phys, 2012, 14, 12671
2) A. Vyšniauskas et al, Int. Rev. Phys. Chem. 2018, 37:2, 259
3) A. Vyšniauskas, et al, Chem. Sci., 2015, 6, 5773
4) J. E. Chambers et al, ACS Nano, 2018, 12, 4398
5) M. Kubankova et al, Soft Matter, 2018, 14, 9466
6) M. Kubankova et al, ACS Appl. Mater Interfaces, 2019, 11, 36307

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-5.1.3
Fluorescent nanosensors for the measurement of biological systems
Jonathan W. Aylott1, Reem Abohujel1, Veeren M. Chauhan1
1) University of Nottingham, School of Pharmacy, Nottingham, UK.
Optical nanosensors are the primary focus of this research and utilise the sensitivity of fluorescence to makequantitative real-time measurements of biological systems. Nanosensor devices can be made in the size range 50–
500 nm diameter, dependent on the matrix they are synthesised from and the application requirements. The small
size of the nanosensors allows them to be delivered directly into the biological system of choice, with minimal
physical perturbation, to measure changes in small molecule concentrations. Nanosensors exhibit advantages overwidely used fluorescence dye-based methods as they have been designed to be ratiometric so effectively contain aninternal standard. The nanosensor matrix also imparts two key benefits:
1. Protection of the sensing component from interfering species within the intracellular environment
2. Protection of the intracellular environment from any toxic effects of the sensing component.
An additional attractive feature of the nanosensors is that they can be imaged and quantified using standard
technologies, e.g., confocal microscopy.
Nanosensors capable of measuring a range of analytes including glucose, oxygen, calcium, zinc, pH and proteases
have been demonstrated and work is ongoing to expand the range of analytes that can be detected using thistechnology. In this presentation I will focus on pH and oxygen nanosensors used in our laboratory and discuss a range
of applications for nanosensors. These include the delivery of nanosensors to mammalian cell systems, wholeorganisms and bacterial biofilms.
References
1) M. Elsutohy, V.M. Chauhan, R. Markus, M.A. Kyyaly, S.J.B. Tendler, J.W. Aylott ‘Real-time measurement of
the intracellular pH of yeast cells during glucose metabolism using ratiometric fluorescent nanosensors’ Nanoscale,2017, 9(18), 5904-5911
2) V.M Chauhan, F. Giuntini, J.W. Aylott ‘Quadruple labelled dual oxygen and pH-sensitive ratiometricnanosensors’ Sensing & Bio-sensing Research, 2016, 8, 36-42
Acknowledgements. This work was supported by grant EP/N009126/1 from the Engineering and Physical Sciences
Research Council (EPSRC), UK

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-5.1.4

A new method of assessing chloroplast movements using hyperspectral imaging
Justyna Łabuz1, Paweł Hermanowicz1
Malopolska Centre of Biotechnology, Jagiellonian University, Krakow, Poland
Chloroplast movements are an important adaptation optimising photosynthetic efficiency of plants. At low lightintensities, chloroplasts gather at cell walls perpendicular to the direction of incident light. This response, called
chloroplast accumulation, increases light capture. At high light intensities, chloroplast avoidance is observed, as they
move to cell walls parallel to the direction of incident light, to protect the photosynthetic apparatus. Seed plants movechloroplasts in response to blue light, however, also red light is active in some ferns, mosses and green algae. Several
methods for tracking chloroplast positioning have been developed. Microscopic observations play a pivotal role forassessing chloroplast positions within leaf cells, but they are laborious1. Measuring transmittance changes withphotometric devices is a convenient quantitative approach, however it is still limited in its throughput2. Apart fromabsorption and transmission, a fraction of light impinging on a leaf undergoes reflection3. Red light leaf reflectance
was shown to be useful for examining of chloroplast positions4.
We propose a new method of assessing chloroplast movements based on leaf reflectance, in which spectral and spatialreflectance characteristics is used for improved detection accuracy. In this study, we employed a halogen lamp and a
hyperspectral camera to investigate chloroplast positions after low and high intensity blue light irradiation, which
induced the chloroplast accumulation and avoidance response, respectively. We acquired spectra of leaf parts thatwere either non-irradiated or irradiated with blue light, for Arabidopsis thaliana wild type and photoreceptor mutants.
The mean reflectance of dark adapted leaves is 11% in visible light (400-700 nm). The average difference ofreflectance in this range between wild type leaves with chloroplasts in avoidance and accumulation positions is ca.
2.3%. This difference is clearly visible in the hyperspectral images of partly irradiated leaves. To examine whether
the changes in reflectance spectra induced by chloroplast movements are species-specific, we performed further
experiments on Nicotiana benthamiana leaves. We employed machine learning methods, using classifiers trained on
spectra from Arabidopsis, to distinguish between spectra recorded on low and high blue light irradiated Nicotiana
leaves. We obtained a low rate of misclassification in this cross-species setting, which suggests that it may be feasible
to automatically assess chloroplast positioning using classification algorithms on hyperspectral images ofheterogenous vegetation patches.
References

1.
2.
3.
4.

Kagawa, T. & Wada, M. Blue Light-Induced Chloroplast Relocation in Arabidopsis thaliana as
Analyzed by Microbeam Irradiation. Plant and Cell Physiology 41, 84–93 (2000).
Walczak, T. & Gabrys, H. New type of photometer for measurements of transmission changes
corresponding to chloroplast movements in leaves. Photosynthetica 14, 65–72 (1980).
Baránková, B., Lazár, D. & Naus, J. Analysis of the effect of chloroplast arrangement on opticalproperties of green tobacco leaves. Remote Sensing of Environment journal 174, 181–196 (2016).
Dutta, S. et al. Non-invasive, whole-plant imaging of chloroplast movement and chlorophyll
fluorescence reveals photosynthetic phenotypes independent of chloroplast photorelocation defects inchloroplast division mutants. Plant Journal 84, 428–442 (2015).

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-5.1.5

It’s Getting Hot in Here: Intracellular Temperature Sensing Through Light
Emission
L. D. Carlos
Phantom-g, CICECO-Aveiro Institute of Materials, Physics Department, University of Aveiro, 3810-193 Aveiro,
Portugal
The emergence of luminescent nanothermometry during the last decade opened up the possibility of measure thermal
flows at spatial scales below 10 μm, unreachable by conventional electrical methods1. Diverse phosphors capable ofproviding a contactless thermal reading through their light emission properties have been examined, e.g., polymers,
DNA or protein conjugated systems, organic dyes, quantum dots, and trivalent lanthanide (Ln3+) ions incorporated
in organic-inorganic hybrids, multifunctional heater-thermometer nanoplatforms, upconverting, downconverting anddownshifting nanoparticles. The implementation of these Ln3+-based phosphors (with an emphasis in upconverting
nanoparticles) as ratiometric thermometers was extensively reviewed in the past five years1.
In the last couple of years, the focus of luminescence thermometry has gradually shifted from the fabrication of moresensitive nanoarchitectures towards the use of the technique as a tool for thermal bioimaging and the unveiling ofproperties of the thermometers themselves and their local surroundings, as, for instance, the instantaneous ballistic
velocity of Brownian nanocrystals suspended in both aqueous and organic solvents2.
After a general perspective of the work done on luminescence nanothermometry since the explosion of the field at
one decade ago, the lecture will be focused on a recent example3 illustrating the potential of the technology to measure
the intracellular temperature.
References
1) C. D. S. Brites, S. Balabhadra, L. D. Carlos, Lanthanide-Based Thermometers: At the Cutting-Edge of
Luminescence Thermometry. Adv. Opt. Mater. 7, 1801239, 2019.
2) C. D. S. Brites, X. Xie, M. L. Debasu, X. Qin, R. Chen, W. Huang, J. Rocha, X. Liu, L. D. Carlos, Instantaneousballistic velocity of suspended Brownian nanocrystals measured by upconversion nanothermometry. Nature
Nanotech. 11, 851, 20163) R. Piñol, J. Zeler, C. D. S. Brites, Y. Gu, P. Téllez, A. N. Carneiro Neto, T. E. da Silva, R. Moreno-Loshuertos,
P. Fernandez-Silva, A. Isabel Gallego, L. Martinez-Lostao, A. Martínez, L. D. Carlos, A. Millán, Real-Time
Intracellular Temperature Imaging Using Lanthanide-Bearing Polymeric Micelles. Nano Lett. 20, 9, 6466, 2020.

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for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-5.1.6

Colorimetric and luminescent nanomaterials as new nanoplatforms for sensing anddrug delivery in biological environments.
Elisabete Oliveira1,2, Carlos Lodeiro1,2, Maria P. Duarte3, and Jose Luis Capelo1,21) BIOSCOPE Group, LAQV-REQUIMTE, Chemistry Department, NOVA School of Science and Technology, FCTNOVA, Universidade NOVA de Lisboa, 2829-516 Caparica, Portugal; 2) PROTEOMASS Scientific Society, Rua
dos Inventores, Madam Parque, Caparica Campus, Portugal; 3) MEtRICs/DCTB, NOVA School of Science andTechnology, NOVA University of Lisbon, 2829-516, Caparica, Portugal.
Inorganic silica mesoporous nanoparticles (MNs) emerged as a multifunctional new generation of nanocarriers as adrug delivery device due to their low toxicity, high specific surface area, large pore volume, tunable pore structures,and sizes [1-5].
From all nanoparticles systems that have been used as drug delivery agents, luminescent inorganic mesoporous silica
nanoparticles and silicon nanocrystals emerged as a multifunctional new generation of nanocarriers acting as all-inone diagnostic and therapeutic tools [1, 2, 3]. Despite the wide utility of MNs for drug delivery/theragnostic, they
can also be explored as sorbents for pollutants removal and sensing, while enabling new strategies for separation andpurification. The multiple applications of MNs will be addressed such as: (i) cytotoxic evaluation and drug delivery
in cancer cells [1, 2]; (ii) antibacterial properties and synergetic effect (antibiotic vs nanoparticles) against bacteria[3] (iii) use as solidstate
supported
devices for metal
remediationvia
colorimetricandfluorimetric responses
in aqueous solutions[4].

Fig.1. Luminescent MNs
and their applications.

References
1) G. Marcelo, E. Oliveira, et. al. Materials 11 (2018) 1310.
2) E. Oliveira, H.M. Santos, et al., Dyes and Pigments 161 (2019) 360.
3) S. Nuti, J. Fernández-Lodeiro, et al. Front. Chem. 6 (2018) 554.
4) G. Marcelo, S. M. G. Pires,et al. Dyes and Pigments 161 (2019) 427-437.
5) G. Marcelo, C. Lodeiro et al., Materials Science and Engineering C 106 (2020) 110104.
Acknowledgements. This work was supported by the Associate Laboratory for Green Chemistry - LAQV which isfinanced by national funds from FCT/MCTES (UIDB/50006/2020 and UIDP/50006/2020) as well as the Scientific
Society PROTEOMASS (Portugal) for funding support (General Funding Grant). G.M thanks to FCT/MEC(Portugal) for his doctoral grant PD/BD/142865/2018. E.O thanks FCT/MEC (Portugal) for the individual contract,
CEECIND/00648/2017.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-5.1.7
Innovative strategies for luminescent chemosensing coupled to exquisite molecular
recognition: illustrative examples in food mycotoxins detection
Guillermo Orellana1, María C. Moreno-Bondi2, Ana B. Descalzo1, Elena Benito-Peña2, Javier L. Urraca2
1) Complutense University of Madrid, Department of Organic Chemistry, Madrid, Spain; 2) Complutense
University of Madrid, Department of Analytical Chemistry, Madrid, Spain.
The combination of nanosized materials with luminescence (fluorescence, phosphorescence) interrogation is a
powerful tool to design analytical methods and devices with the sensitivity required to determine environmental andfood contaminants. In the lab practice, the associated indispensable selectivity is typically provided by sophisticated,
expensive chromatographic techniques coupled to mass spectrometric detection. In contrast, straightforward,
affordable chemical sensors rely on biological elements (antibodies, aptamers, …) or bioinspired materials (e. g.
molecularly imprinted polymers or MIPs) to achieve the analyte recognition. The challenge to the sensor scientist is
to effectively couple recognition of the target to luminescence-based reporting.
The first example to illustrate our solutions to this challenge for the analysis of food mycotoxins such as tenuazonicacid (TeA) is the optical sensing of the contaminant with a luminescent MIP2. Both the emission intensity and, forthe first time, the emission lifetime can be used to determine the TeA levels thanks to the synthesis of a novel
trifunctional Ru(II) complex containing 2,2’-biimidazole as TeA-binding moiety, and two polymerizable 2,2'bipyridin-4,4’-diyldimethyl diacrylate ligands. A 9-nm thick MIP shell was grown onto 200-nm SiO2 beads. The
sensor shows fast response (< 5 s) to TeA with no cross-sensitivity to other common β-hydroxycarbonyl foodmycotoxins (alternariol, β-zeranol, cyclopiazonic acid). Detection limits as low as 63.8 and 75.2 μg L⁻1 under steadystate and time-resolved luminescence detection, respectively, have been measured without optimization.
O
O

O

TeA

OH

O
H
N
H

O

CO2H
CO2H

OH OH

FB1
HO2C
HO2C

O

OH

NH2

O

Fig. 1 – Chemical structure of the food mycotoxins analyzed by the novel luminescent sensors illustrated in this presentation.

The second example to illustrate the talk topic is a homogeneous fluorescence quenching immunoassay based ongold nanoparticles (AuNPs) and a recombinant epitope-mimicking fusion protein (“mimotope”) to detect andquantify the wheat mycotoxin fumonisin B1 (FB1)3. The fumonisin mimotope was cloned with a yellow fluorescentprotein allowing its direct use as the tracer for FB1 detection without requiring a labelled analyte or secondary
antibody. Furthermore, due to the fluorescence quenching ability of the AuNPs, the homogeneous immunoassay canbe carried out in a single step without further washing to separate the unbound tracer. The quenching immunoassay
displays no matrix effect in the wheat extract and high sensitivity for FB1 detection (7.3 to 22.6 ng mL−1), with adetection limit of 1.1 ng mL−1. Only the closely similar FB2 among other mycotoxins tested was also recognized.
References
1) A. Rico-Yuste, R. Abouhany, J. L. Urraca, A.B. Descalzo, G. Orellana, M.C. Moreno-Bondi, Eu(III)-Templated
molecularly imprinted polymer used as a luminescent sensor for the determination of tenuazonic acid mycotoxin infood samples. Sensors Actuat. B: Chem. 329, 129256, 2021.
2) J. Quílez-Alburquerque, A.B. Descalzo, M.C. Moreno-Bondi, G. Orellana, Luminescent molecularly imprinted polymer
nanocomposites for emission intensity and lifetime rapid sensing of tenuazonic acid mycotoxin, 2021 (submitted).
3) R. Peltomaa, F. Amaro-Torres, S. Carrasco, G. Orellana, E. Benito-Peña, M.C. Moreno-Bondi, Homogeneous
Quenching Immunoassay for Fumonisin B1 Based on Gold Nanoparticles and an Epitope-Mimicking Yellow
Fluorescent Protein. ACS Nano 12, 11333-11342, 2018.
Acknowledgements. This work was supported by grants RTI2018-096410-B-C22/21 from the Spanish Ministry of
Science and Innovation.
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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-5.1.8

Fluorescent Metal Nanoclusters for MicroRNA Detection
Marina A. Kapitonova1, Polina S. Malova1, Alexei I. Kononov1
1) Department of Molecular Biophysics and Polymer Physics, Saint Petersburg State University, Saint Petersburg,
RussiaMicroRNA are small (18-25 nucleotides) non-coding single stranded RNA sequences. The main function of
microRNA is transcriptional and post-transcriptional regulation of gene expression by RNA-interference.
MicroRNA is considered to be a promising biomarker of different diseases such as heart diseases, different types ofcancer (“oncomirs”), and diseases of neural system. Particularly miR-210, which was presented in this work as amodel target, is mostly associated with hypoxia and cancer progression including breast, lung, head, pancreaticcancer, or glioblastoma1. Measuring levels of circulating miR-210 in fluids could be a non-invasive method of
identifying cancer on early stages. Unfortunately, microRNA detection remains a challenge, because of its length.
Traditional methods of microRNA detection have some disadvantages such as poor sensitivity and time consumingof northern blotting or false positive results in real-time qPCR2. Last decade new variations and improvements ofreal-time PCR have been developed: it includes additional steps with reverse transcription or rolling-circle
amplification. Also, nanomaterials such as gold nanoparticles3, magnetic nanoparticles, or quantum dots wereapplied in microRNA detection.
In this work, we suggest a new method of microRNA detection based on fluorescent silver nanoclusters (AgNCs).
Metal nanoclusters are groups of small number (2-100) of metal atoms or ions. Nanosized silver demonstrates
unique electrooptical properties, which are different from bulk metals. Stabilized by DNA oligonucleotides, AgNCs
with a size about 1-2 nm can develop fluorescent species with a high quantum yield4, photostability, in a wide
range of fluorescence from near UV to IR, and stokes shift up to 200 nm5.
In order to make AgNCs sensitive to microRNA we used and improved special split model for DNA matrix, thatmakes fluorescence to turn-on only by an interaction with the target. This technique reduces background signal andfalse positive probability. The DNA matrix can be divided into three logical parts: the hybridization sequence,
which binds complementary to the target, the clusters formation sequence, which is separated into two different
DNA-strands (5’-CCCGTTTT-3’and 5’-CCCCACCCCT-3’6), and the spacer, which links these two parts together.
The optimization of the spacer’s length and sequence shows that the best spacer consists of single adenine on eachmatrix strands. It was found that the most optimal protocol for clusters formation is: 50 mM sodium phosphatebuffer at the room temperature and concentration ratio [DNA]:[AgNO3]:[NaBH4] = 1:8:4. The hsa-miR-210-3p (5’rCrUrGrUrGrCrUrGrUrGrArCrArGrCrGrGrCrUrGrA-3’)
and
it’s
DNA
analog(5’CTGTGCGTGTGACAGCGGCTGA-3’) served as a model target of microRNA. The sensor was successfully
tested with DNA- and RNA-targets: the bright green fluorescence, with an excitation at 490 nm and emission at
560 nm, appeared only in the presence of targets. The limit of detection was 3 nM. The linear dependence offluorescence intensity on target concentration was observed up to 500 nM.
References
1) Bavelloni, A. et al. MiRNA-210: A Current Overview. Anticancer Res. 37, 6511–6521, 2017.
2) Ye, J., Xu, M., Tian, X., Cai, S. & Zeng, S. Research advances in the detection of miRNA. J. Pharm. Anal. 9,
217–226, 2019.
3) Miao, X. et al. Label-Free Platform for MicroRNA Detection Based on the Fluorescence Quenching of
Positively Charged Gold Nanoparticles to Silver Nanoclusters. Anal. Chem. 90, 1098–1103, 2018.
4) Gwinn, E., Schultz, D., Copp, S. M. & Swasey, S. DNA-Protected silver clusters for nanophotonics.
Nanomaterials 5, 180–207, 2014.
5) Bogh, S. A. et al. Unusually large Stokes shift for a near-infrared emitting DNA-stabilized silver nanocluster.
Methods Appl. Fluoresc. 6, 2018.
6) He, C., Goodwin, P. M., Yunus, A. I., Dickson, R. M. & Petty, J. T. A Split DNA Scaffold for a Green
Fluorescent Silver Cluster. J. Phys. Chem. C 123, 17588–17597, 2019.
Acknowledgements. This work was supported by RFBR grant № 19-53-51005.
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for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-5.1.9

Optogenetic high-throughput screening
Kathrin Brenker12, Annik Jakob12, Luis Köbele121) Opto Biolabs, Freiburg, Germany; 2) Albert-Ludwigs-Universität Freiburg, Freiburg, Germany
Optogenetic tools allow the remote control of cellular pathways using light. Originating from the field of
neurobiology using light-gated ion channels to study neuronal behavior, more recently developed non-neuronal
optogenetic tools enable the isolated, functional investigation of almost any intracellular signaling molecule withincomplex signaling pathways. This remote control of cellular pathways has major implications for therapeutic tools,
including targeted immunotherapy.
A major obstacle for optogenetic researchers is the controlled delivery of light to the cell sample. Hence, the mostpopular tools for optogenetic studies are microscopy-based experiments or using self-made LED devices. A widely
neglected, yet very powerful analytical tool is flow cytometry. The flow cytometer has major advantages over a
microscope, including the ability to rapidly measure thousands of cells at single cell resolution. However, it is not
yet widely used in optogenetics due to a lack of suitable illumination.
Opto Biolabs, a spinoff from the University of Freiburg, develops specialized illumination devices enabling the
combination of optogenetics and flow cytometry. Optogenetic flow cytometry enables the easy analysis of e.g.
calcium kinetics and the developed illumination device is compatible with most existing flow cytometers. Our future
work will focus on the expansion of our technology to optogenetic cell sorting. This will enable large scale single
cell analysis and greatly simplify the discovery and development of novel, cell-based optogenetic tools and therapies.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-5.1.10

A novel NIR excitable fluorescent probe for intracellular nitric oxide detection: frommouse macrophages to human leukemic cells
Carla Arnau del Valle1, Paul Thomas2, M. Paz Muñoz-Herranz1, Francisco Galindo3, María J. Marín1
1) University of East Anglia, School of Chemistry, Norwich, UK; 2) University of East Anglia, School of Biological
Science, Norwich, UK; 3) Universitat Jaume I, Departamento de Química Inorgánica y Orgánica, Castellón, Spain
Understanding intracellular dynamics and concentrations of diverse intracellular analytes is key to develope novel
diagnostic tools. Fluorescence-based molecular probes have proben to be key for intracellular imaging and sensing
analytes of biological interest such as pH, Ca2+, nitric oxide (NO), etc. Most of the currently available fluorescentprobes are excited using ultraviolet or visible light, which results in high photodamage to living cells. Alternatively,
the use of near-infrared (NIR) light provides high photostability, low autofluorescence, high biological tissue
penetration and minimal photodamage. In the development of diagnostic tools for cancer, one of the most relevant
reactive nitrogen species is NO since its effect is strongly related to its concentration. For example, elevated NO
levels have been found in tumour cells including lung, breast and leukaemic.1 Fluorescent NIR excitable molecular
probes seem to be ideal candidates for the intracellular detection of NO and could ultimately yield to valuable insights
on the biological role played by NO.2
The aim of this work is to develop a NIR excitable molecular probe for the intracellular detection of NO via a
photoinduced electron transfer (PET) mechanism. The probe showed good sensitivity (LOD = 133 nM) andselectivity towards NO, with a fast detection response (2 min). Additionally, the fluorescence emission intensity ofthe probe was stable in a range of pHs from 4 to 9; and the detection of NO in acidic environments was succesfully
evidenced. The NO probe was able to detect NO in a variety of macrophages. In RAW264.7 cells, the fluorescenceemission intensity of the probe increased in the presence of NO as confirmed by confocal microscopy images andwith the intracellular fluorescence emission spectrum (Fig. 1). The probe was also able to detect NO in primary bonemarrow mouse cells and in THP-1 human leukaemia monocytic cells, used as human model. These intracellularstudies confirm the versatility of the NO probe to monitor the presence of NO in a variety of cellular environments.
The NO probe will next be used to detect intracellular NO in other cancer and healthy cell lines aiming to investigate
the differences in NO concentrations between these two environments.

NO probe + NO

NO probe

a)

i)

ii)

iii)

b)

NO
iv)

v)

vi)

Fig. 1 – a) Confocal microscopy images of RAW264.7 cells treated with NO probe: i - iii) before and iv - vi) after stimulation of
the cells to produce NO; and b) fluorescence emission spectra of RAW264.7 cells treated with the NO probe before (black) andafter (green) stimulation and the control cells without the NO probe before (red) and after (blue) stimulation.

References
1) S. K. Choudhari, M. Chaudhary, S. Bagde, A. R Gadbail, V. Joshi, Nitric oxide and cancer: a review. World J.
Surg. Oncol., 11, 118, 2013
2) H. Yu, Y. Xiao, L. Jin, A Lysosome-targetable and two-photon fluorescent probe for monitoring endogenous andexogenous nitric oxide in living cells. J. Am. Chem. Soc., 134, 42, 2012
Acknowledgements. The authors would like to thank the School of Chemistry and Faculty of Science at the
University of East Anglia for financial support. CAdV would like to specially thank Mr and Mrs Whittaker for
the FJW Oncology Scholarship.
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P-5.1.11

Detection of nitric oxide in live cells using a gold-based near-infrared excitable
fluorescent nanoprobe
Carla Arnau del Valle1, Paul Thomas2, M. Paz Muñoz-Herranz1, Francisco Galindo3, María J. Marín1
1) University of East Anglia, School of Chemistry, Norwich, UK; 2) University of East Anglia, School of Biological
Science, Norwich, UK; 3)Universitat Jaume I, Departamento de Química Inorgánica y Orgánica, Castellón, Spain
Nitric oxide (NO) is involved in the regulation of diverse physiological and pathological mechanisms of the
cardiovascular, nervous and immune systems.1 NO regulates many physiological processes such as antiinflammatory effects when present in the cell at low concentrations; however, at high levels, it is related to many
pathologies including Alzheimer’s disease and cancer.2 Intracellular NO concentrations can be monitored using
fluorescent molecular probes in combination with fluorescence imaging techniques. Most of the currently available
NO molecular fluorescent probes are excited using ultraviolet or visible light, which results in poor penetration andhigh photodamage to living cells. Alternatively, the use of near-infrared (NIR) light provides high photostability, lowlevel of autofluorescence, high biological tissue penetration and minimal photodamage upon long-term irradiation.
Although fluorescent probes to image intracellular NO have been reported,3 further research is needed in thedevelopment of novel NIR excitable NO probes for biological applications; and the incorporation of nanoparticles(NPs) to the game seems an exciting approach due to their great versatility.
This work describes the development of a NIR excitable nanoprobe, consisting of a gold nanoparticle functionalisedwith a NIR excitable NO probe (NOAuNPs), for the detection of NO in live cells. The NO probe contains an ophenylenediamine moiety that reacts with NO forming the corresponding benzotriazole product and resulting in an
increase of the fluorescence emission intensity of the probe. The sensitive NOAuNPs showed good selectivity
towards NO over other potential cellular interferences and an excellent stability in aqueous medium over time. The
intracellular NO detection by the NOAuNPs has been successfully achieved in MDA-MB-231 breast cancer cells(Fig. 1a) and it is currently under investigation using THP-1 human leukaemia monocytic cells. The intracellular
fluorescence emission spectrum of the NOAuNPs confirmed a significant enhancement in the fluorescence emission
intensity in the presence of NO (Fig. 1b). The NOAuNPs will be next used to report intracellular NO in other cancer
and healthy cell lines to investigate the differences in NO concentrations between these two environments.
i)

ii)

iii)

iv)

v)

vi)

NOAuNPs
NOAuNPs + NO

a)

b)

NO

Fig. 1 – a) Confocal microscopy images of MDA-MB-231 cells treated with NOAuNPs, i – iii) before and iv – vi) after additionof NO; b) fluorescence emission spectra recorded using confocal microscope of MDA-MB-231 cells treated with NOAuNPs
before (grey) and after (green) addition of NO and the control cells without NOAuNPs (black).

References
1) A. R. Butler, D. L. H. Williams, The physiological role of nitric oxide. Chem. Soc. Rev., 22, 233, 1993
2) S. K. Choudhari, M. Chaudhary, S. Bagde, A. R Gadbail, V. Joshi, Nitric oxide and cancer: a review. World J.
Surg. Oncol., 11, 118, 20133) M. J. Marín, P. Thomas, V. Fabregat, S. V. Luis, D. A. Russell, F. Galindo, Fluorescence of 1,2‐Diaminoanthraquinone and its nitric oxide reaction product within macrophage cells. ChemBioChem, 12, 2471-2477, 2011
Acknowledgements. The authors would like to thank the School of Chemistry and Faculty of Science at the
University of East Anglia for financial support. CAdV would like to thank Mr and Mrs Whittaker for
the FJW Oncology Scholarship.
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P-5.1.12

DNA G-quadruplexes recognition by cationic zinc(II) phthalocyanine and graphene
oxide hybrid materials
Ana R. Monteiro1,2, Catarina I. V. Ramos2, Leandro M. O. Lourenço2, Sara Fateixa1, Joana Rodrigues3, Maria G. P.
M. S. Neves,2 Tito Trindade1
1) CICECO, Department of Chemistry of University of Aveiro, Aveiro, Portugal; 2) LAQV-REQUIMTE, Departmentof Chemistry of University of Aveiro, Aveiro, Portugal; 3) i3N, Department of Physics of University of Aveiro, Aveiro,
Portugal.
The stabilization of deoxyribonucleic acid (DNA) G-quadruplex structures promotes the inactivation of telomerase
in cancer cells, being considered a valuable anticancer therapy1. Several molecules and materials have been used as
potential G-quadruplex ligands, such as porphyrins, phthalocyanines and graphene-based materials2,3. The current
work evaluates the performance of non-covalent hybrids consisting of graphene oxide (GO) and a tetracationiczinc(II) phthalocyanine (ZnPc) as G-quadruplex stabilizing agents (Fig. 1). The Q-bands in the absorption spectrumof the ZnPc@GO hybrid are red-shifted when compared to the counterparts in the free ZnPc. In addition, GO acts asa fluorescence quencher, significantly decreasing the fluorescence emission of ZnPc. Such distinctive opticalproperties allowed to carry out spectrofluorimetric titrations of the non-immobilized ZnPc and hybridized ZnPc@GOwith both the DNA G-Quadruplex (T2G5T) and the DNA duplex (ds26) oligonucleotide sequences. Noteworthy,
fluorescence recovery was observed in the titration of ZnPc@GO with T2G5T, suggesting progressive release of ZnPc
from the ZnPc@GO hybrids to stabilize the DNA G-quadruplex structures. This behaviour was not evident in thetitration of ZnPc@GO with the DNA duplex. We suggest that GO might act as a nanoplatform for the selective
delivery of ZnPc towards different DNA structures, which can be controlled by a “switch off-on” fluorescence model,
as already reported for a porphyrin@GO system3.

Fig. 1 – Development of ZnPc@GO hybrids for potential recognition of DNA G-quadruplexes.

References
1) Kosiol, N.; Juranek, S.; Brossart P.; Heine, A.; Paeschke K.; G-Quadruplexes: a promising target for cancer
therapy. Molecular Cancer, 2021, 20, 40.
2) Ramos, C. I. V.; Almeida, S.; Lourenço, L. M. O.; Pereira, P. M. R.; Fernandes, R.; Faustino, M. A. F.; Tomé, J.
P. C.; Carvalho, J.; Cruz, C.; Neves, M. G. P. M. S., Multicharged Phthalocyanines as Selective Ligands for GQuadruplex DNA Structures. Molecules, 2019, 24, 733.
3) Monteiro, A. R.; Ramos, C. I. V.; Fateixa, S.; Moura, N. M. M.; Neves, M. G. P. M. S.; Trindade, T., Hybrids
Based on Graphene Oxide and Porphyrins as Tools for Detection and Stabilization of DNA G-Quadruplexes. ACS
Omega, 2018, 3, 1184.

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Acknowledgements. The authors are grateful to the University of Aveiro and to the Fundação para a Ciência e a
Tecnologia (FCT) for the financial support to LAQV-REQUIMTE (Ref.UIDB/50006/2020), CICECO-Aveiro
Institute of Materials (Ref.UID/CTM/50011/2020), FCT project P2020-PTDC/QUI-QOR/31770/2017 throughnational funds and, where applicable, co-financed by the FEDER, within the PT2020 Partnership Agreement. Ana
R. Monteiro thanks the FCT for the PhD grant SFRH/BD/137356/2018. Sara Fateixa and Catarina I. V. Ramos
acknowledge the costs of their research contracts (REF.-069-88-ARH/2018 and REF.-047-88-ARH/2018,
respectively) resulting from the FCT hiring funded by National funds (OE), through FCT, I.P., in the scope of theframework contract foreseen in the numbers 4, 5 and 6 of the article 23, of the Decree-Law 57/2016, of August 29,changed by the law 57/2017, of July 19. J. Rodrigues acknowledges i3N project UIDB/50025/2020 &UIDP/50025/2020 and project PTDC/NAN-MAT/28755/2017 (POCI-01-0145-FEDER-028755), financed bynational funds through the FCT/MEC and supported by FEDER funds through the COMPETE 2020 Programme.

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P-5.1.13

Synthesis of luminescent gold nanoclusters on aromatic amino acids
Varvara Kubenko1, Zakhar Reveguk1, Alexei Kononov1, Andrey Buglak1
1) St. Petersburg State University, Department of Molecular Biophysics and Polymer Physics, Saint Petersburg,
RussiaTyr is involved in the synthesis of neurotransmitters, catecholamines, thyroid hormones, etc. There is anumber of pathologies associated with impaired Tyr metabolism: phenylketonuria, hypothyroidism, tyrosinemia,
alkaptonuria, and vitiligo. Therefore, it is important to be able to detect tyrosine in biological fluids, as well as todetermine its concentration. Our aim was to develop a new detection method using luminescent metal nanoclusters.
Tyrosine (Tyr), dioxyphenylalanine (DOPA), and tryptophan (Trp) were chosen as compounds with similar
chemical structures to assess the selectivity of the method.
A typical protocol of nanocluster synthesis was as follows: amino acids were dissolved in water (C = 2.25mM) at pH 1, controlled with HNO3, then AgNO3 (1.25 мМ) or HClAu4 (1.25 мМ) was added; the samples wereplaced in a thermostat at 37° C. The spectra were recorded in 24 h1.
It was found that luminescent gold clusters are formed at pH 1 on tyrosine, tryptophan, and DOPA. Fig. 1
shows absorption spectra (left) and luminescence spectra (right) of tyrosine complexes with silver and gold. One
luminescent fraction of tyrosine complexes with gold nanoclusters with excitation/emission maxima of 360/470 nm
was found (at right). No luminescent complexes with silver nanostructures were formed (at left).

Fig. 1. Absorption spectra (left) and luminescence spectra (right) of tyrosine complexes with silver and gold.

Fig. 2 shows 2D luminescence spectra of DOPA (right) and its complex with gold (left). The complex
possesses excitation/emission maximum at 330/450 nm. Also, tryptophan stabilizes the luminescent fraction ofgold clusters with excitation/emission maxima at 320/410 nm, 330/500 nm and 370/500 nm (Fig. 3). No
luminescent silver structures were observed.

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Fig. 2. 2D luminescence spectra of DOPA (right) and its complex with gold (left).

Fig. 3. 2D luminescence spectra of tryptophan (right) and its complex with gold (left).

The luminescence of gold nanoclusters synthesized on tyrosine, DOPA, and tryptophan differs in
excitation/emission parameters, which suggests the possibility of the amino acid selective detection.
References
1) Xiaoming Yang, Yawen Luo, Yan Zhuo, Yuanjiao Feng, Shanshan Zhu, Novel synthesis of gold nanoclusters
templated with l-tyrosine for selective analyzing tyrosinase, Analytica Chimica Acta, 840, 87-92, 2014.
Acknowledgements. This work was supported by the Russian Science Foundation (project 20-73-10029).

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P-5.1.14

Colorimetric detection of L-DOPA
Yana Chuiko1, Zakhar Reveguk1, Alexei Kononov1, Andrey Buglak1
1) St Petersburg State University, Department of molecular biophysics and polymer physics, Saint Petersburg,
RussiaDopamine is known as an important neurotransmitter of the human central nervous system and affects
many brain functions and behavioral responses. Lack of dopamine in the brain may lead to neurological disorders,such as schizophrenia and Parkinson’s disease. Considering the biochemical significance of dopamine precursor LDOPA and its significance for medicine, it is highly desirable to develop a new efficient method for quantitative
detection of L-DOPA with improved selectivity and sensitivity.
The colorimetric method is quite effective for detection of various analytes in solution. It is well known
that under certain conditions amino acids can reduce ions of noble metals, forming nanoclusters (NCs)1 andnanoparticles (NPs)2 with characteristic absorption bands. To check the selectivity of the method, Tyrosine (Tyr),
dioxyphenylalanine (DOPA), phenylalanine (Phe), and tryptophan (Trp) were studied as compounds with similar
chemical structures.
Typical synthesis protocol proceeded as follows: silver nitrate was added to the amino acid dissolved in
distilled MiliQ water (pH 6.5), NaOH was added after 10 minutes. We observed the reduction process a couple ofminutes after the addition of the alkali. The samples were placed in a thermostat (20 0C) for 2 hours. Thenabsorption spectra were recorded.
At pH 7-10, complexes of silver nanoparticles with DOPA exhibited a notable absorption band with amaximum at 450 nm. For other amino acids, this band was not observed (Fig. 1), which indicates the selectivity ofthis method in relation to DOPA. It is noteworthy that at pH 11.5, all selected molecules showed the ability to
reduce silver and form nanoparticles.

Fig. 1. Absorption spectra of Tyr, DOPA, Phe and Trp complexes with silver at different pH.

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Fig. 2(on the left) shows the absorption spectra of silver NP complexes with DOPA, recorded 2 h after thesynthesis with various DOPA concentrations in solution (pH 7.5). The optical density recorded at 410 nm showed alinear dependence in the range of DOPA concentrations from zero to a few micromoles per liter (Fig. 2, right). The
limit of detection (LOD) was about a few nM.

Fig. 2. Absorption spectra of silver NPs with different concentrations of DOPA (left); dependence of the optical density ofsolutions of silver NPs recorded in a 4 mm cuvette at 410 nm on the concentration of DOPA (right).

Further optimization is currently in progress. In this way it is also possible to detect Tyr using the oxidationof Tyr into DOPA by the enzyme tyrosinase.
References
1) Tomash S. Sych, Andrey A. Buglak, Which Amino Acids are Capable of Nucleating Fluorescent Silver Clusters
in Proteins? J. Phys. Chem. C, 122 (45), 26275-26280, 2018.
2) Sharma R. Synthesis of Amino Acid Capped Silver Nanoparticles, Characterization, and Biological Application.
Orient J Chem 2020;36(2).
Acknowledgements. This work was supported by the Russian Science Foundation (project 20-73-10029).

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P-5.1.15

The light-mediated effects on photoluminescence of hydrophilic CdSe/ZnS-COOH
quantum dots: dependence on biological mediumAgnė Kalnaitytė1,2, Emilija Januškaitė1, Osvaldas Vilutis1 and Saulius Bagdonas1,2
1) Vilnius University, Biophotonics Group of Laser Research Centre, Vilnius, Lithuania; 2) Nature of ResearchCentre, Institute of Ecology, Vilnius, Lithuania
Semiconductor nanoparticles (NPs) or quantum dots (QDs) are used in optical instruments or in monitoring
techniques, because QDs photoluminescence (PL) can be easily detected and tracked. Due to sensitivity of thespectral properties to environmental factors (temperature, pH, presence of ions or proteins, other parameters ofliving systems), hydrophilic quantum dots can also be used as indicators of environmental conditions. However, as
the physicochemical properties of NPs can be affected by illumination conditions [1] and surrounding medium, thephotoinduced changes in the PL also become important for monitoring their environmental fate and estimation of
the threat of toxicity in various biological environments [2].
In this study, the spectroscopic measurements were performed on aqueous suspensions of hydrophilic CdSe/ZnSCOOH quantum dots (625 nm, Invitrogen, USA). The QDs samples (4 nM) were prepared for experiments bydiluting QDs powder with distilled (DW), deep well (DWW) and lake water, MWC medium and also with PBS and
TES buffer solutions (pH 6, 7, 8). A part of experiments was done on the samples keeping them under controlled
illumination conditions, while the control samples were kept in the dark at +4℃ and at room temperature. The
samples were either constantly illuminated during 24 hours using a white fluorescent lamp (11W/827) at room
temperature, or a 8/16 h “light/dark” cycle was applied for those being kept in an aquarium (Aquael, Poland) (at+22℃) using a combined white LED lamp (6 W) intended for the cultivation of aquatic plants.
In general, initial irradiation doses accelerate the decrease of PL intensity of QDs, the bathochromic spectral shiftof the PL peak in comparison with unexposed samples, and also increase the scattering in the optical density (OD)
spectra due to the formation of unstable QDs aggregates. However, when higher doses were reached, depending on
the surrounding medium and its pH, the decrease slows down, the PL intensity stabilizes, or even partially recovers,
which is also followed by the decreasing relative intensity of the excitonic absorption band in the OD spectra andthe appearing hypsochromic shift of the PL spectral peak. Thus, higher irradiation doses break the formed unstable
QDs aggregates and disrupt the structure of the NPs. Although the destruction of light-induced QDs aggregates andpartial recovery of spectroscopic properties were more pronounced in a more alkaline environment, the recovery of
the PL intensity was not detected in DWW (where the pH was about 8) demonstrating that photoinduced indicativechanges in the spectral properties of CdSe/ZnS QDs strongly depend on the composition of surrounding medium.
In addition, the violet LED (404 ± 9 nm, 30 mW/cm2 at a distance of 8 cm) was used to expose the QDs samples in
MWC and DWW to 108 J/cm2 irradiation dose. Afterwards, some irradiated samples of both types were mixed
with a small amount of concentrated suspension of green freshwater unicellular microalgae cells and kept inaquarium under “light/dark” illumination cycle. During the first week, the following effects were observed in thecase of irradiated QDs in MWC solution: the decrease of scattering in the OD spectra, a slight increase of the PL
intensity and narrowing of the PL spectrum, as compared to the spectroscopic parameters of QDs measured
immediately after irradiation, which could be related to the degradation of the light-mediated unstable QDs
aggregates. Although the PL intensity of the irradiated QDs in the sample with algae cells was lower, thesubsequent recovery of intensity was more pronounced and the narrowing of the spectrum in the red side wasobserved, presumably due to interaction of algae cells and/or their extracellular materials with QDs surface.
References
1) A. Kalnaitytė, S. Bagdonas, R. Rotomskis. The dose-dependent photobleaching of CdTe quantum dots inaqueous media. J Lumin, 201, 434–44, (2018).
2) A. Kalnaitytė, S. Bagdonas, Light-mediated effects of CdTe-MSA quantum dots on the autofluorescence offreshwater green microalgae: Spectroscopic studies, J Photoch Photobio B, 199, 111629, (2019).
Acknowledgements. This work was funded by the Research Council of Lithuania, Project No. S-MIP-20-22.
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P-5.1.16

Organic dyes as luminescent probes for protein aggregates
Samuel Guieu1,2, Raquel Nunes da Silva1,3, Francisco Cardona1, Catarina C. Costa1, Maria J. G. Santos1,
Mariana Q. Alves3, Susana S. Braga1, Sandra I. Vieira3, João Rocha2, Artur M. S. Silva1
1) LAQV-REQUIMTE, Chemistry Department, University of Aveiro, Portugal; 2) CICECO-Aveiro Institute ofMaterials, Chemistry Department, University of Aveiro, Portugal; 3) IBiMED-Institute for biomedicine, Universityof Aveiro, Portugal
All-organic luminescent dyes have attracted much attention for biological imaging, due to their ease of use, andbecause they are easily modified to fine-tune their properties. Unfortunately, they usually present a high sensitivity
to their environment, and can loss their emission properties upon aggregation or interaction with their surroundings.
Interestingly, some fluorophores present the opposite effect, and exhibit an enhanced emission intensity in confinedenvironments.1 These modifications of emission properties can be advantageous to detect a change in theenvironment of the dye.
A series of organic dyes based on a double hemi-salen core has been prepared and characterized (Figure 1).2 They
are almost non emissive in dilute solution, but become highly fluorescent when they are in the crystalline state, in aphenomenon known as Aggregation-Induced Emission Enhancement.1 This observation has been rationalized
through the study of their crystal structures. The emission wavelength of the dyes has been fine-tuned by modifying
the substituents.
To broaden the scope of application of these fluorophores, water soluble derivatives have been prepared, presenting
the same photophysical properties. Their interaction with cyclodextrins and DNA has been studied: upon formationof inclusion complex or interaction with DNA, the emission intensity is enhanced, in a way similar to when thedyes crystallize. The dyes have then been applied to the staining of protein aggregates in live cells, demonstrating a
selectivity similar to probes commercially available.3

Fig. 1 – Probe with tunable colour, presenting an emission enhancement upon interaction with cyclodextrins or protein
aggregates.

References
1) Y. Hong, J. W. Y. Lam, B. Z. Tang, Chem. Soc. Rev. 2011, 40, 5361-5388. B.-K. An, J. Gierschner, S. Y. Park, Acc. Chem.

Res. 2012, 45, 544-554.

2) S. Guieu, F. Cardona, J. Rocha, A. M. S. Silva, Chem. Eur. J. 2018, 24, 17262-17267.
3) R. Nunes da Silva, C. C. Costa, M. J. G. Santos, M. Q. Alves, S. S. Braga, S. I. Vieira, J. Rocha, A. M. S. Silva,
S. Guieu, Chem. Asian J. 2019, 14, 859-863.

Acknowledgements. This work was supported by Portuguese Fundação para a Ciência e a Tecnologia (FCT) via
the CICECO-Aveiro Institute of Materials (UID/CTM/50011/2013, UIDB/50011/2020 & UIDP/50011/2020),
QOPNA (UID/QUI/00062/2019), LAQV-REQUIMTE (UIDB/50006/2020), Institute for Biomedicine iBiMED(UID/BIM/04501/2013, UID/BIM/04501/2019 - POCI-01-0145-FEDER-007628), by Centro 2020 and Portugal
2020, and by the European Union (FEDER program) via R. Nunes da Silva post-Doctoral fellow grant(BPD/UI98/6327/2018) from the pAGE project (CENTRO-01-0145-FEDER-000003). S. Guieu is supported bynational funds (OE), through FCT, I.P., in the scope of the framework contract foreseen in the numbers 4, 5, and 6of the article 23, of the Decree-Law 57/2016, of August 29, changed by Law 57/2017, of July 19.
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P-5.1.17
Liguori, Nicoletta

Poster withdrawn

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P-5.1.18
Luminescent complexes of gold clusters with amino acids
Mizintsev Artem1, Sych Tomash1, Kononov Alexei1, Buglak Andrey1
1) St Petersburg University, Department of molecular biophysics and polymer physics, Saint Petersburg, Russia
Amino acids (AAs) are relatively small organic molecules which play an important role in various biochemical
processes. Their concentration in natural biological fluids can indicate some diseases of human organism in earlystages. In this work, we develop simple and selective method of AAs detection using the amino acids as stabilizing
matrices for luminescent gold clusters. In comparison with luminescent silver clusters stabilized by AA matrices1,
here we used the approach of gold atom chemical reduction without any external reducing agent. In the case of goldclusters, the reduction of metal atoms occurred due to the stabilizing matrix.
In this work, we tested the possibility of synthesizing luminescent gold clusters on almost twenty different aminoacids under conditions of strongly alkaline pH values (12.8). Here we used a “double synthesis” strategy.
Chloroauric acid was added twice to the alkaline AA solutions with an interval of 24 hours. The studied complexes
were incubated at +37℃ in the dark. Among the whole variety of considered amino acids, only two demonstrated
the possibility of stabilization of luminescent gold clusters – histidine (His) and phenylalanine (Phe) (Fig. 1).

Fig. 1 – 2D luminescence spectra of gold clusters stabilized by phenylalanine (A-D) and histidine (E-H). A,E: Au/AA = 1:2;
B,F: Au/AA = 1:1; C,G: Au/AA = 3:2; D,H: pure amino acid.

Analyzing the obtained luminescence spectra, one can notice that the spectral properties of clusters stabilized bydifferent amino acids differ markedly. Thus, in the case of phenylalanine, a strong luminescence band is observed
at 380 nm. In the case of histidine, a noticeably weaker luminescence band is observed at 450 nm.
Significant spectral differences between two obtained gold clusters make it possible to unambiguously determine
the presence of His and/or Phe solution.
References
1) Tomash S. Sych, Andrey A. Buglak, Which Amino Acids are Capable of Nucleating Fluorescent Silver Clusters
in Proteins? J. Phys. Chem. C, 122 (45), 26275-26280, 2018.
Acknowledgements. This work was supported by the Russian Foundation for Basic Research (Project No. 19-5351005 NIF_а RFFI-Korea).

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IL-5.2.1
Chances and limitations of light-based antimicrobial approaches in dentistry – anupdateFabian Cieplik1
1) University Hospital Regensburg, Department of Conservative Dentistry and Periodontology, Regensburg,
Germany 1
The 2016 Review on Antimicrobial Resistance has predicted that the number of annual deaths attributable toantimicrobial resistance will increase globally from the current 700,000 to 10 million in the year 2050 if noappropriate action is taken immediately. In light of this increasing threat of resistance towards conventional
antibiotics and antiseptics, alternative antimicrobial approaches are desperately needed. In particular in the field ofdentistry, where usually no life-threatening diseases need to be treated, it seems reasonable to promote research forsuchlike alternative antimicrobial approaches. In this context, especially light-based approaches including
antimicrobial photodynamic therapy (aPDT), photothermal therapy (PTT) or low-level light (or laser) therapy(LLLT), have increasingly been proposed in the last two decades.
This talk aims to give an overview about these approaches by summarizing evidence from in vitro studies as well
as recent clinical trials and to discuss the chances and limitations for application of light-based antimicrobial
approaches in dental practice.
Acknowledgements. This work was supported in part by grants from the Deutsche Forschungsgemeinschaft (DFG
CI 263/1-1, CI 263/1-3, CI 263/3-1).

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IL-5.2.2

Light-Responsive Coatings for Biofilm Disruption
Devatha P. Nair1,2,3, Gannon M. Kehe1, Christopher J. Carpenter4, Michael J. Schurr51) University of Colorado-School of Dental Medicine, Craniofacial Biology, Aurora, U.S.A 1; 2) University ofColorado-Materials Science and Engineering program, Boulder, U.S.A; 3) University of Colorado-Skaggs Schoolof Pharmacy and Pharmacological Sciences, Aurora, U.S.A; 4) Metropolitan State University, Biology, Denver
U.S.A; 5) University of Colorado-Immunology and Microbiology, Aurora, U.S.A.
Bacterial biofilms are ubiquitous- be it in the oral cavity, within pipes, or on the surface of a ship’s hull and marine
structures.1 The ability of bacteria to form biofilms is a potent evolutionary mechanism that allows them to thrive
and proliferate in a variety of environments and enable them to evade antibiotics. The rise of pathogenic biofilms andtheir role in growing antibiotic resistance observed globally has urgent implications in the current and future
treatments against infectious disease. Biofilms offer protection to a consortium of bacteria and can be 10−1000 times
more resistant to antibacterial agents. To address the emerging crisis, our group studies alternative strategies to disrupt
biofilms on resin surfaces by engineering functionalized coatings that can: 1) prevent and/or delay biofilm formationand 2) repeatedly detach biofilms via transient changes in surface topography (photofluidization effect, Fig.1).2

Fig. 1 – Biofilm disruption via light-responsive azobenzene coatings.

Functionalized azobenzene molecules were synthesized and characterized to undergo 1) Light-induced molecular
movement via (400-500 nm, 700 mW/cm2, 10s pulsed exposures) and 2) as coatings to ascertain biofilm growth over
24-48 hrs. Biocompatibility of the molecules was ascertained (ISO-10993) and biofilm disruption of Pseudomonas
aeruginosa (PA01) and Streptococcus mutans (SM) biofilms (presence and absence of 1wt% sucrose in BHI) on
glassy surfaces were evaluated. Biofilms were imaged and quantified via CFU counts. While molecular movement
was able to achieve a 3-log reduction in SM biofilms grown in the absence of sucrose, no reduction in biofilms
formation or removal was seen in the presence of sucrose. The light-activated coatings completely removed mature
PA01 biofilms from surfaces, indicating that light-induced biofilm removal can be a viable tool for biofilm disruption.
References
1) Algburi A, Comito N, Kashtanov D, Dicks LMT, Chikindas ML. Control of Biofilm Formation: Antibiotics
and Beyond. Appl. Environ. Microbiol. 20172) Kehe GM, Mori DI, Schurr MJ, Nair DP. Optically Responsive, Smart Anti-Bacterial Coatings via the
Photofluidization of Azobenzenes ACS Appl. Mater. Interfaces. 2019.
Acknowledgements. This work was supported by grant K25DE027418 from the National Institutes of Health National Institute of Dental and Craniofacial Research (NIH-NIDCR).

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IL-5.2.3

Photocrosslinking of collagen and antimicrobial applications
Krister Gjestvang Grønlien1, Mona Elisabeth Pedersen2, Håkon Valen3, Ellen Bruzell3, Hanne Hjorth Tønnesen1
1) Section for Pharmaceutics and Social Pharmacy, Department of Pharmacy, Oslo, Norway; 2) Department of
Raw Material and Process Optimization, Nofima AS, Ås, Norway; 3) Nordic Institute of Dental Materials (NIOM)
AS, Oslo, Norway
Collagen is extensively used in fabrication of scaffolds for pharmaceutical and biomedical applications. Scaffolds
can be used in drug delivery (e.g., in ophthalmology, topical and dental applications), as wound dressings or as cell
templates in tissue engineering1. However, low viscosity, slow gelation and low mechanical strength of the prepared
constructs are the major limitations for such use. Crosslinking by chemical or photochemical methods could enhancethese properties. However, chemical crosslinkers are often toxic and the crosslinking reaction is difficult to control.
Photoactivated crosslinkers have been considered as good biocompatible alternatives. Photocrosslinking of collagenhas been achieved by modification of the protein with methacryloyl-substitution or by addition of photocrosslinkers
to collagen solutions combined with radiation prior to or during thermogelation2,3. The presentation will give an
introduction to photocrosslinking of collagen and results from a present work on a new natural photocrosslinker ofcollagen.
Examples of photosensitizers commonly used in photocrosslinking are Igracure 2959 (mainly used forphotocrosslinking of methacrylates), Eosin Y, ruthenium compounds, riboflavin (vitamin B2) and rose bengal4. The
mechanisms behind photocrosslinking of collagen have been thoroughly studied. The crosslinking effect is postulated
to involve the formation of reactive oxygen species (ROS) and singlet oxygen (1O2) by the exited photosensitizer,
resulting in photo-oxidation and formation of covalent bonds between the amino acids in the collagen protein chains3.
In the present work, collagen was crosslinked with two naturally occurring photosensitizers; riboflavin (RF) and a
newly discovered photocrosslinker (NXL). Photocrosslinking of collagen by RF is commonly applied in corneal
crosslinking in the treatment of the eye disease keratoconus and has been investigated as a potential technique for
production of collagen-based biomaterials3,5. However, riboflavin photocrosslinking requires a long irradiation time,
limiting its use in applications such as bioprinting. The newly discovered natural photocrosslinker NXL reduced theirradiation time from minutes to only 10 seconds and did not require incubation and self-assembly of collagen prior
to crosslinking. The resulting hydrogels were compared to RF photocrosslinked hydrogels and showed an
approximately 20-fold higher water holding capacity (44.97 and 2.57%, respectively). The NXL hydrogels werehighly elastic with a Young’s modulus over twice the value of the RF photocrosslinked hydrogels (62.30 and 30.88
kPa, respectively). Fibroblasts seeded on the hydrogels had a migratory phenotype. These are properties favorable inpharmaceutical and biomedical applications. Further, the hydrogels photocrosslinked with the new photosensitizer
NXL were tested for their antimicrobial properties, and the results were promising for further development into abiomedical product.
References
1) Lee, C. H., Singla, A., and Lee, Y. Biomedical applications of collagen. Int. J. Pharm., vol. 221, no. 1, 2001.
2) Drzewiecki, K. E. et al. A thermoreversible, photocrosslinkable collagen bio-ink for free-form fabrication ofscaffolds for regenerative medicine. Technology, vol. 5, no. 4, 20173) Heo J. et al. Riboflavin-induced photo-crosslinking of collagen hydrogel and its application in meniscus
tissue engineering. Drug Deliv. Transl. Res., vol. 5, no. 2, 2016.
4) Mu, X., Sahoo, J. K., Cebe, P., and Kaplan, D. L. Photo-Crosslinked Silk Fibroin for 3D Printing. Polymers,
vol. 12, no. 12, 2020.
5) Wollensak, G., Spoerl, E., and Seiler, T. Riboflavin/ultraviolet-A–induced collagen crosslinking for thetreatment of keratoconus. Am. J. Opthalmol., vol. 135, no. 5, 2003.
Acknowledgements. Turkey tendon by-products and financial support for isolation of collagen from turkey were
contributed by Norilia AS (Oslo, Norway).
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OC-5.2.4
Blue light risk evaluation of dental operating lights
Ellen Bruzell1, Bjørn Johnsen2, Jeff Zawada3
1) Nordic Institute of Dental Materials, Materials Science Group, Oslo, Norway; 2) Norwegian Radiation and
Nuclear Safety Authority, Dept. of Emergency Preparedness and Response, Østerås, Norway; 3)A-dec, Inc., Dept.
of Technical Research, Newberg, OR, USA.
Dentistry is an occupation that places great demands on the lighting conditions. Thus, the operating light must deliver
sufficient intensity in addition to ensuring specific lighting parameter values, such as contrast and colour fidelity.
The illuminance of an operating light is typically 20-30 klx.
Following the substitution of incandescent light sources to light emitting diodes (LED) in nearly all light products
including medical devices, the public, researchers and organisations raised concern about the blue light portion of
the LED spectra. This concern was aimed at a potential risk of photochemical hazard to the eyes and disturbance of
circadian rhythms1,2. No limit values exist for the blue light level needed to affect or disrupt circadian rhythms;
however, melatonin depression by blue light can be considered an advantage for the dental team during examinationand treatment procedures.
The aim of this study was to determine the blue light hazard (retinal injury) risk group class of operating lights as
described in the standards for photobiologcial safety3 and dental operating lights4. The spectral power of threeoperating lights (one manufactured in USA, two in Japan) were measured using a double monochromator
spectroradiometer5. Appropriate weighting functions were applied to obtain blue light-weighted radiance. Prior tothe measurement the operating lights had undergone similar measurements provided by the respective manufacturers(pre-measurements), and the values obtained from the two measurements for each device were compared.
The blue light-weighted irradiance values of the operating lights were lower than the limit of 10000 W/m2/sr
separating Risk Group 1 from Risk Group 2 with the exception of one value obtained for one device in the premeasurement. This operating light exceeded Risk Group 1 limit by <1% in the pre-measurement. Comparable results
were obtained irrespective of optical laboratory. Due to the conservative requirement of the exposure conditions,occupational exposure of the investigated operating lights did not pose a blue light risk for retinal injury.
References
1) SCHEER (Scientific Committee on Health, Environmental and Emerging Risks), Opinion on Potential risks tohuman health of Light Emitting Diodes (LEDs), 6 June 2018.
2) ANSES (the French Agency for Food, Environmental and Occupational Health & Safety), Opinion on the “effects
on human health and the environment (fauna and flora) of systems using light-emitting diodes (LEDs)”, 5 April 20193) CIE S009:2002. Photobiological Safety of Lamps and Lamp Systems. Vienna, 2002.
4) ISO/DIS 9680. Dentistry – Operating lights. Geneva, 2021.
5) Terje Christensen, Bjørn Johnsen, Ellen Bruzell, Violet-blue light exposure of the skin: is there need for protection?
Photochemical & Photobiological Sciences, 20(5), 615-625, 2021

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IL-5.2.5
Lasers in DentistryFirst author: Peter Fahlstedt, DDS, MSc Lasers in Dentistry, PhD candidate.
Faculty of Medicine, Department of Clinical Dentistry, University of Bergen, Bergen, Norway; Institute for Laser
Supported Dentistry, Akersberga, Sweden
Lasers of different wavelengths have been used as support in dentistry for decades. Diode laser 445 nm up to 980
nm, free-running pulsed lasers 2780/2940 nm and CO2 lasers are the most common in daily practices. Constituting awide range of different interactions in soft- and hard tissues, we find lasers having biomodulating and antibacterial
effects and the ability to remove tissue and foreign materials without adverse effects on the following process ofhealing and regeneration. This lecture focuses on the daily practice of laser support in dental clinics, the benefits, thelimitations, and challenges in convincing not the patients but the colleagues and universities why the integration oflasers in dentistry is essential, even in a broader context.

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IL-5.2.6

Abstract for ESP2021: Laser in Public dental healthcare, experience and reflections.
Are Kristofer Hjeltnes, DDS, MSc Laser Dent11) Ålesund kompetanseklinikk, Public dental healthcare, Ålesund, Norway;
Introduction:    
ͳǤͳΨ ǡ
  Ǥ     
Ǥ    
  ʹͲͳ͵Ǥ
Settings:   %  ȋʹͺǡ͹ͲǤͲͲͲȌǡ %Ǥ
Methods and proceduresǣThe dental lasers used are ǣ ȋʹͻͶͲȌǡǣ ȋͳͲ͸ͶȌǡ
ȋͶͶͷǡ͸͸ͲǡͺͳͲǡͻ͹ͲȌǡ Ǥ
 Ǥ
Results:   
ǤǦǡ Ǧǡ ǡ ǡ  ǡǡ ǡ
ǡǡ ʹǡ͵ǡͶǡͷǡ͸Ǥ
been graded as “good”, “fair” and “bad” according to my own experience and patient satisfacǡ
 Ǥ  
    ǡ Ǥ
  ͳǦʹͳͲ Ǥ  ǡ Ǥ
Discussion and conclusion: 
 ǣ ǡ 
treatment when used on the mentioned diagnoses and indications, given that it’s executed by a laser Ǥ        ͹Ǥ
References

1) Leknes KN, Alstad ME, Angelsen S, Eriksen ES, Espervik A, Bruzell E. “Laser treatment of periodontal andperi-implant infections”, Nor Tannlegeforen Tid 2009; 119: 700-62) Liu JF, Lai YL, Shu WY, Lee SY. Acceptance and efficiency of Er:YAG laser for cavity preparation inchildren. Photomed Laser Surg. 2006;24(4):489-493.
3) Daneswari V, Nandlal B. Restorative dentistry for children using a hard tissue laser - a review. J Int Oran
Health. 2011;3(3):1-8.
4) Eren F, Altinok B, Ertugral F, Tanboga I. The effect of Er,Cr:YSGG laser therapy on pain during cavity
preparation in paediatric dental patients: a pilot study. Oral Health Dent Manag. 2013;12(2):80-84.
5) Boj J, Galofre N, Espana A, Espasa E. Pain perception in pediatric patients undergoing laser treatments. J Oral
Laser Applications. 2005;5(2):6) 85-89 Polonsky M, Gutknecht N, Franzen R. Review of possible predictors for pain perception with class 1-5
cavity preparations using Er,Cr:YSGG laser: a retrospective clinical in vivo study. Laser Dent Sci. 2017;1(1):921.
7) Polinsky M, “Laser-assisted restorative dentistry: Can we afford not to use it?”, Dental Economics, apr 2018,https://www.dentaleconomics.com/science-tech/article/16385053/laserassisted-restorative-dentistry-can-weafford-not-to-use-it

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IL-5.2.7

Evaluation of upper labial frenectomy - a randomized, controlled comparative studyof conventional scalpel technique and Er:YAG laser technique
Sarmadi Roxana1, 3, Gabre Pia2, 3, Thor Andreas4

Department of Paediatric Dentistry, Public Dental Health, Uppsala County Council, Sweden.

1

Department of Preventive Dentistry, Public Dental Health, Uppsala County Council,
Sweden.

2

Department of Cariology, Institute of Odontology, The Sahlgrenska Academy of Gothenburg, Gothenburg, Sweden.

3

Department of Plastic and Oral and Maxillofacial Surgery, Department of Surgical Sciences, Uppsala University

4

Objectives: Abnormalities in the maxillary frenum may lead to aesthetic or functional limitations and need to becorrected with a surgical intervention called frenectomy. The aim of the study was to compare frenectomies
performed using Er: YAG laser technology with those using a conventional scalpel technique. Comparisons wereof patients´ experiences, treatment times, bleeding during treatment and wound healing.
Material and methods: The trial was performed as a prospective, randomized and controlled, single-blindinvestigation. A total of 40 patients requiring frenectomy were randomly assigned to groups which underwent
either conventional or Er:YAG laser treatment. Patients´ experiences, treatment time, bleeding and wound healing
were evaluated immediately after surgery and five days, twelve days and three months after surgery.
Results: Significant increase in time spent in surgery and bleeding was seen with conventional scalpel surgery.
Directly after surgery the wound area was significantly larger in the laser group but at the five-day evaluation nodifference could be observed between the groups. Finally, patients were satisfied with both methods, giving themthe same assessments.
Conclusions: In the frenectomy procedure, laser surgery is faster and causes less bleeding and may be
advantageous in frenectomies.
Acknowledgement
This project was supported by grants from the Public Dental Service in Uppsala County Council. We thank the study
coordinator Anna Maria Carlberg for careful planning of the practical parts of the study We also thank Niclas
Eriksson for help with statistical analyses and Gwyneth Olofsson for revising the English text.

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OC-5.2.8

With blue light against bacteria and viruses
Magdalena Metzger1,2, Lisa Karner1,2, Kristjan Plaetzer3, Barbara Holzer4, Kurt Schicho5, Johannes Grillari1,2,6,
Peter Dungel1,21) Ludwig Boltzmann Institute for experimental and clinical Traumatology, Vienna, Austria; 2) Austrian Cluster
for Tissue Regeneration, Vienna, Austria; 3) University of Salzburg, Department of Biosciences, Laboratory of
Photodynamic Inactivation of Microorganisms, Salzburg, Austria; 4) Austrian Agency for Health and Food Safety,
Mödling, Austria; 5) Department of Oral and Maxillofacial Surgery, Medical University of Vienna, Vienna,
Austria; 6) Department of Biotechnology, Institute of Molecular Biotechnology, BOKU-University of Natural
Resources and Life Sciences, Vienna, Austria.
While the germicidal properties of UV radiation have been known for a long time, its negative side effects, e.g.
DNA damage, photoaging and carcinogenicity, limit the possibilities and potential fields of medical application.
Alternatively, blue light can be a promising option for the inactivation of a variety of microorganisms such asbacteria and viruses with the additive potential to improve wound healing (1). While the underlying mechanisms ofblue light therapy have not been completely resolved yet, it has been proposed, that intrinsic chromophores e.g.
porphyrins or flavins, generate ROS upon irradiation which destroy lipids, proteins and nucleic acids. The aim of
this study was to investigate the effects of a LED device emitting “soft” blue light with wavelengths >430 nm(kindly provided by Repuls Lichtmedizintechnik, Vienna, Austria) on the inactivation of Gram-negative
Escherichia coli, Gram-positive Staphylococcus capitis and the corona virus type SARS-CoV-2.
Treating bacteria of different strains with blue light, the present study revealed that strains lacking the gene recA
were more susceptible towards illumination than those with an intact copy of the gene. In most bacterial strains, theprotein RecA is responsible for homologous recombination, DNA repair and induction of the SOS response.
Therefore, we hypothesized that the inactivation of the protein would increase the susceptibility of resistant strains
and enhance the antimicrobial effects of the treatment. Although the potentiation of the therapy via RecA inhibitors
was only partially successful, the influence of recA was confirmed by rescuing susceptible bacterial strains with theinsertion of a plasmid carrying the gene.
Regarding the antiviral properties of blue light, it was shown that the number of infectious SARS-CoV-2 viruses
drops by around half after a short exposure of 20 +/- 5 mW/cm² (5 minutes) and by one order of magnitude (by90%) after 30 minutes. An examination of the viral load as a function of the exposure time to blue light shows analmost linear decrease between 3 and 12 minutes.
In conclusion, “soft” blue light was able to significantly reduce bacterial growth which was associated with the lackof recA gene. However, further experiments are needed to uncover the mechanisms behind the toxicity of blue lighttherapy and the importance of the SOS response. Blue light from the Repuls illumination device had also virucidal
effects on SARS-CoV-2, i.e. the viral load was already reduced by light alone. Further studies should investigate
the probable enhancement of these effects in combination with suitable photosensitizers.
References
1) Dungel P, Hartinger J, Chaudary S, Slezak P, Hofmann A, Hausner T, Strassl M, Wintner E, Redl H, Mittermayr
R. Low level light therapy by LED of different wavelength induces angiogenesis and improves ischemic woundhealing. Lasers Surg Med. 2014
Acknowledgements. This work was supported by a grant funded by the Vienna Business Agency. A fund of theCity of Vienna, Austria.

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IL-5.3.1
Dynlt3 controls melanosome movement, distribution, acidity and transfer tokeratinocytes
Zackie Aktary1, Lionel Larue1
1) Normal and Pathological Development of Melanocytes, Institut Curie, CNRS UMR3347, Inserm U1021, Orsay,
FrancePigmentation of the appendage including skin and hair depends on fundamental processes such as biogenesis,
transport, and maturation of melanosomes and their transfer to keratinocytes. The control of these different precise
processes in space and time remains poorly understood. Here, we show that Dynlt3, a subunit of the cytoplasmic
dynein complex, is required to efficiently transport, acidify and transfer melanosomes. When the level of Dynlt3 isdecreased in melanocytes, the movement of melanosomes is more directional, and they are consequently mainly
located at the periphery of these cells. The acidity of stage IV melanosomes is higher when Dynlt3 is reduced andresults in less efficient melanosome transfer, yet they remain highly pigmented. Finally, the level of mRNA
corresponding to Dynlt3 is dependent on β-catenin activity, Finally, the level of mRNA corresponding to Dynlt3 isdependent on β-catenin activity, revealing a function of the Wnt/β-catenin signalling pathway during melanocyte
and skin pigmentation, coupling transport, positioning in the cell and acidity of melanosomes, a process that isessential for their efficient transfer.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-5.3.2

Sun-induced pigmentation: relative contribution of UVA1 and Visible Light color
domains
Piffaut Virginie1, Marionnet Claire1, Sasai Jun2, Nouveau Stéphanie1, Jouni Hussein1, Ortiz Maria1,
Huguet Etienne1, Planel Emilie1, Duteil Luc3, Bernerd Francoise1
1) L'Oréal, Research and Innovation, Aulnay Sous-Bois, France; 2) L'Oréal, Research and Innovation, Kawasaki,
Japan ; 3) Centre de Recherche Clinique Appliquée à la Dermatologie, Hôpital de l'Archet 2, Nice, France
Hyperpigmentation and pigmentary disorders are known to be induced or aggravated by sun exposures. While therole of UVB and UVA2 rays is indisputable, data emphasize a contribution of UVA1 wavelengths and also Visible
Light (VL) for which High Energy Visible Light (HEV) seems to play a major role through activation ofmelanogenesis. To better understand the contribution of UVA1, VL and the different VL color wavelengths bands(HEV, “Blue”, “Green” and “Red”) in induction of pigmentation, two clinical studies were performed using
physiological sources (solar simulators) and specific cut off filters. Results could be illustrated using computerbased simulations on full-face models. In the first study, the back of volunteers (n=27) was exposed to various
wavelengths ranges: UVA1+HEV (350-450nm), UVA1 (350-400nm) or HEV (400-450 nm). In the second study,
volunteers (n=25) were exposed to VL (400-700nm), HEV (400-450nm), “Blue” color domain (400-500nm),“Green” color domain (500-600nm) and “Green + Red” color domains (500-700nm). Pigmentation was assessed byvisual scoring and chromametric measurements (L*, a*, b*, ΔITA) at several time points (from day 1 to day 43).
In the first study, pigmentation was induced in all the exposure conditions as attested by ΔL* and ΔITA values andclinical soring. The higher intensity was observed at 2h with a progressive decline over time. At 2h post exposure
the pigmentation induced by UVA1+HEV, UVA1 and HEV corresponded to a ΔL*= -6,70; -4,48 and -2,45
respectively, indicating an additive effect of UVA1 and HEV.
In the second study, VL exposure induced a significant and persistent pigmentation (ΔL*=5.54 at 2h), as well as
exposure to HEV (400-450nm) (ΔL*=-1,90 at 2h) or “Blue” light (400-500 nm) (ΔL*=-2,84 at 2h). The study
revealed that “Green” light was also responsible for induction of pigmentation although its contribution was lower(ΔL*=1,16 at 2h) than the “Blue” light’s one. By comparing pigmentation induced by “Green” and “Green + Red”
color domains, we confirmed that “Red” light doesn’t significantly contribute to pigmentation. Clinical scoring
confirmed these results. Based on the colorimetric measurements, a contribution percentage could be calculated foreach color domain in VLinduced pigmentation 24 hours post exposure with: 400-700nm full VL= 100%, “Blue”
400-500nm= 71%; HEV 400-450nm= 47%; “Green” 500-600nm=37 %; “Red” 600-700nm <1%.
Altogether, the two studies reinforced the basis for the need of UVA1 photoprotection up to 400 nm andhighlighted the importance to also protect the skin from solar visible wavelengths, especially the shortest
wavelengths HEV/ “Blue” but also “Green” color band, to limit skin hyperpigmentation.

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IL-5.3.3
UV-induced growth and migration of melanoma cells in a spheroid model.
Linda Vainikka, Petra Wäster, Linda Rößler and Karin Öllinger1
1) Linköping University, Department of biomedicine and Clinical Sciences, Linköping, Sweden

Cutaneous melanoma is a highly aggressive cancer with a propensity for metastasis to various organs.
The most important risk factor to develop the disease is exposure to UV radiation. UVB wavelengths has
ability to cause tumor-initiating mutations while UVA exposure generates free radicals and may act
immunosuppressive. How the microenvironmental effects of UV radiation influence melanoma
pathogenesis and its ability to metastasize remains to be elucidated. In a previous report we have shownthat UVA exposure triggers increased secretion of extracellular vesicles. UVA cause plasma membrane
damage which is promptly repaired by transportation of lysosomes to the damaged membrane to form a
resealing patch. Subsequently, extracellular vesicles are released outside the cell (see Figure 1). The
vesicles promote signaling event when taken up by non-irradiated cells and increased growth andmigration is noted1.
Figure 1. UVA-radiation induces plasma
membrane damage resulting in calcium influx,
which trigger transport of lysosomes to thedamaged site. The lysosomal membranes forms
a resealing patch and lysosomal content isreleased outside the cell. Subsequently, excess
membrane is shredded as extracellular vesicles.

In this study results from UV exposure of malignant melanoma cells will be presented and the possible
signaling pathways induce by UV will be discussed. We generated melanoma tumor spheroids from four
different cell lines and exposed to sub-lethal doses of UVA- and UVB-radiation. We found spontaneous
secretion of extracellular vesicles by all melanoma cell lines, but after UVA, increased number of large
size vesicles were found. Following radiation, the spheroids were embedded in Matrigel and observed
microscopically to follow cell migration and invasion. Cell migration out from the spheroid was higher in
spheroids exposed to UVA compared to control and UVB exposed samples. A transcriptome analysis was
performed 24 h after irradiation of the spheroids, and revealed significantly increased mRNA levels of
matrix metalloprotease-1 (MMP1; +3.3-fold, p=0.0009) after UVA exposure. MMP-1 is a collagenase
involved in extracellular matrix degradation and the protease is secreted and activated outside the cell.
We found elevated protein level of MMP1 and secretion to the medium in three out of four tested
melanoma spheroids after UVA exposure and in one of four after UVB. We conclude that melanoma
migration and invasion is elevated more efficiently after UVA than UVB radiation and MMP1 is acandidate protein to promote this effect.
References
1. Wäster P, Eriksson I, Öllinger K. Intercellular cross-talk mediated by extracellular vesicles promotes survival andproliferation via miR-21-signaling. Pigment Cell and Melanoma Research 2020; 33(4):542
Acknowledgements. This work was supported by grant from Swedish Cancer Society and Welander Finsten Skin
Foundation.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-5.3.4

Photoprotective and Antigenotoxic Properties of Actinobacteria Indigenous Strainsfrom Deep Subsurface Environments of Colombia
Carlos Adolfo Pedraza Barrera1, Elena E. Stashenko1 and Jorge Luis Fuentes Lorenzo1
1) Universidad Industrial de Santander, Santander, Bucaramanga, Colombia.
Introduction: Actinobacteria are known to produce a variety of secondary metabolites with clinical properties.
Purpose: The study was aimed at investigating photoprotective and antigenotoxic properties against UVB of ten
extracts from Cutibacterium acnes and Microbacterium paraoxydans strains collected from subsurface habitats ofColombia. Materials and Methods: The bacterial growth was measured spectrophotometrically in different culture
media. In vitro photoprotection efficacy was evaluated using in vitro indices such as sun protection factor (SPFin vitro)and critical wavelength (λc). The percentage of UVB– antigenotoxicity estimates (%GI) in the SOS Chromotest wasalso evaluated. Correlation analysis was used to examine the relation between SPFin vitro and %GI estimates. Results:
The Reinforced Clostridium Medium showed the highest bacterial growth under anaerobic conditions with yields(weight/volume) of lyophilized extracts between 200 and 500 mg/mL. Among the strains under study, one (1) showedlow (6.0 ≤ SPFin vitro ≤ 14.9) and nine (9) medium (15.0 ≤ SPFin vitro ≤ 29.9) UVB photoprotection efficacy, but all ofthem resulted in broad-spectrum (UVA–UVB) photoprotection (λc > 370 nm). SPFin vitro was correlated (R = 0.76, p≤ 0.05) with %GI estimates. Conclusions: We demonstrated that extracts of Actinobacteria strains have properties
as sunscreen. UVB photoprotection efficacy depended on concentration and %GI estimates in strain extracts.
Acknowledgments. The authors thank funding from the Ministry of Science, Technology and Innovation,
the Ministry of Education, the Ministry of Industry, Commerce and Tourism, and
ICETEX, Program Ecosistema Científico-Colombia Científica, from the Francisco José de Caldas Fund,
Grant RC-FP44842-212-2018. The Ministry of Environment and Sustainable Development of Colombia
supported the Universidad Industrial de Santander through access permits to genetic resources andderivatives for bioprospecting (Contract N o. 270).

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-5.3.5

Role of the AhR transcription factor in reprogramming melanoma cells
Marie-Dominique Galibert
University of Rennes, Faculty of Medicine, CNRS UMR6290, Institute of Genetics and Development of Rennes,
FranceCutaneous melanoma originates from melanocytes, the pigment-producing cells. They are the most aggressive anddeadliest form of skin cancer. The establishment of the genomic landscape of cutaneous melanoma revealed theimportance of the NRAS-BRAF driver mutations to fuel tumor development through the downstream activation of
the MEK/ERK pathway. Selective inhibitors of the BRAF V600 driver mutation in combination with MEK
inhibitors constitute now front-line treatment of metastatic BRAF V600 melanoma. This double blockade leads to a
massive and initial tumor shrinkage, unfortunately patient ultimately relapse. It is now admitted that the highplasticity of melanoma cells mediates resistance to MAPK pathway inhibitors. Phenotypic drug resistance isdynamically linked to cell cycle heterogeneity, cell differentiation and metabolic reprogramming. Understanding
how melanoma cells undergo cell reprogramming is thus critical to overcome resistance. We recently, showed thatsustained activation of AhR, a ligand binding transcription factor, mediates melanoma resistance through specificcell reprogramming. Together, these results revealed new drug vulnerabilities to combat resistance.

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IL-5.3.6

MITF and the chromatin modifiers PRDM7 and SETDB2 in melanoma
Ramile Dilshat1, Hong Nhung Vu1, Eiríkur Steingrímsson1
1) University of Iceland, Faculty of Medicine, BioMedical Center, Department of Biochemistry and Molecular
Biology, Reykjavik, Iceland;
The microphthalmia associated transcription factor (MITF) is a critical regulator of melanocyte development anddifferentiation. It also plays an important role in melanoma where it has been described as a molecular rheostat that,
depending on activity levels, allows reversible switching between different cellular states. We have shown that
MITF directly represses the expression of many genes involved in establishing the extracellular matrix and focal
adhesions in human melanoma cells1, thus affecting cell morphology and cell-matrix interactions. However, MITF
also indirectly affects the expression of many other genes raising the question of how it mediates these effects. We
have identified the chromatin modifiers PRDM7 and SETDB2 as targets of MITF, suggesting that MITF may inpart mediate its effects on gene expression through these modifiers. The role of both modifiers in melanoma cells
and their link to MITF function will be presented.
References
1) Dilshat R, Fock V, Kenny C, Gerritsen I, Lasseur RMJ, Travnickova J, Eichhoff OM, Cerny P, Möller K,
Sigurbjörnsdóttir S, Kirty K, Einarsdottir BÓ, Cheng PF, Levesque M, Cornell RA, Patton EE, Larue L, de Tayrac
M, Magnúsdóttir E, Helga Ögmundsdóttir M, Steingrimsson E. 2021. MITF reprograms the extracellular matrix
and focal adhesion in melanoma. Elife. 10:e63093. PMID: 33438577
Acknowledgements. This work was supported by grants 207067 and 217768 from the Icelandic Research Fund.

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IL-5.3.7
B cells in human melanoma: spatiotemporal changes in phenotypes andantibody-independent functions
Stephan N Wagner
Medical University of Vienna, Department of Dermatology, Vienna, Austria
My research group is interested in the spatiotemporal dynamics of B cell responses in human cancer,
particularly human melanoma. We and others have recently shown that the presence of tumor associated B
cells (TAB) either scattered within the TME or localized within mature tertiary lymphoid structures (TLS) is
associated not only with increased survival but also with enhanced treatment response to immune checkpoint
blockade in melanoma patients.
B cells typically undergo a sequence of developmental changes in phenotype and function and do occur inhuman primary melanoma scattered within the TME preferentially at the invasive tumor-stroma front andalso in the intratumoral stroma in metastatic melanomas. In addition, B cells can also aggregate to form withother cells tertiary lymphoid structures (TLS) in which, similar to secondary lymphoid organs, B cells arehypothesized to undergo local maturation by antigen-dependent activation, affinity maturation and classswitch recombination before leaving the TLS to exert immunological functions at local tumor sites.
However, compared with canonical secondary lymphoid tissues, human melanomas exhibit significant
differences in B cell and lymphoid follicle maturation and distribution. These observations suggest a further
evaluation of the role of local TLS versus locoregional lymph nodes in generating and enhancing local
adaptive anti-tumor B cell responses.
Furthermore, the functional roles of B cells in human cancer remain poorly understood. These include thepresence of B cell subpopulations with differences in phenotype and function at different disease stages andthe mechanisms by which B cells can support T cell mediated anti-tumor immunity in an antibodyindependent manner, including immune checkpoint blockade-mediated activation.
Here, we will discuss our recent work identifying the spatiotemporal dynamics of different phenotypic andfunctional B cell subpopulations during human melanoma disease progression, address the putative link
between the presence of TLS and mature B cell subtypes in human melanoma samples, and describe
potential mechanisms of B cell support for anti-tumor T cell responses. These mechanisms include T cell
recruitment to tumor sites, secretion of immuno-stimulatory cytokine, cross-presentation of tumor antigens,and direct T cell activation.
Acknowledgments. This work was supported by the Austrian Science Fund (FWF) Project P30325-B28 and
IPPTO Project No. DOC 59-B33 to SNW.
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OC-5.3.8
Photoreactive properties of melanin from different skin phototypes and thecontribution of melanin subunits
Krystian Mokrzyński1, Shosuke Ito2, Kazumasa Wakamatsu2, Theodore G. Camenisch3, Tadeusz Sarna1, Michał
Sarna1
1) Faculty of Biochemistry, Biophysics and Biotechnology, Department of Biophysics, Jagiellonian University,
Cracow, Poland; 2) Institute for Melanin Chemistry, Fujita Health University, Toyoake, Aichi, Japan; 3)
Department of Biophysics, Medical College of Wisconsin, Milwaukee, WI, USA
Photoreactivity of melanin has become a major focus of research due to the postulated involvement of the pigment
in UVA-induced melanoma. However, most of the previous studies were carried out using synthetic melaninpigments. In this research we examined the photoreactive properties of natural melanin pigments isolated from hairsamples obtained from donors of different skin phototypes (I, II, III, and V). Dynamic Light Scattering (DLS) alongwith X-band and W-band electron paramagnetic resonance (EPR) spectroscopy were used to determine the physicalproperties of examined pigments. Alkaline hydrogen peroxide degradation and hydroiodic acid hydrolysis were
employed to identify melanin subunits. EPR oximetry, EPR spin trapping and time-resolved near infrared
phosphorescence was employed to determine the singlet oxygen photogeneration by the melanins. The efficiency ofboth superoxide and singlet oxygen photogeneration was found to be related to the chemical composition of thestudied melanins. Melanins from blond and chestnut hair (phototypes II and III) exhibited the highest photoreactive
potential among all examined pigments. Moreover, the highest values of quantum efficiency of singlet oxygen
photogeneration for natural melanins were demonstrated at 332 nm and 365 nm supporting the postulated pigment
contribution in the UVA-induced melanoma.
References
1) Mokrzyński K et al. Photoreactivity of Hair Melanin from Different Skin Phototypes - Contribution of Melanin
Subunits to the Pigments Photoreactive Properties. International Journal of Molecular Sciences, 22, 2021.
2) Szewczyk G et al. Aerobic photoreactivity of synthetic eumelanins and pheomelanins: Generation of singlet
oxygen and superoxide anion, 29, 2016.
3) Zadlo A et al. Photobleaching of pheomelanin increases its phototoxic potential: Physicochemical studies ofsynthetic pheomelanin subjected to aerobic photolysis. Pigment Cell and Melanoma Research, 32, 2019.
Acknowledgements. This research was funded by National Science Center of Poland, grant number Sonata2015/19/D/ST4/01964, Opus-2017/27/B/ST5/02631 and Preludium-2020/37/N/NZ1/01054.

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IL-5.4.1
Flow-induced shear stress and chemoresistance in ovarian cancer: A role for targeted
PDT and combinations
Imran Rizvi1,2, Utkan Demirci3, Tayyaba Hasan4, Marcela G. del Carmen5, Huang-Chiao Huang6,7, Jonathan P.
Celli8, Shubhankar Nath4, Mustafa K. Ruhi1, Christina Conrad6.
1) University of North Carolina Chapel Hill and North Carolina State University, UNC-NC State Joint Departmentof Biomedical Engineering, Chapel Hill, NC, USA
2) University of North Carolina Chapel Hill School of Medicine, Lineberger Comprehensive Cancer Center,
Chapel Hill, NC, USA
3) Stanford University School of Medicine, Bio-Acoustic MEMS in Medicine (BAMM) Laboratory, Canary Centerat Stanford for Cancer Early Detection, Department of Radiology, Palo Alto, CA, USA
4) Massachusetts General Hospital, Harvard Medical School, Wellman Center for Photomedicine, Department of
Dermatology, Boston,MA, USA
5) Massachusetts General Hospital, Harvard Medical School, Division of Gynecologic Oncology, Department of
Obstetrics and Gynecology, Boston, MA, USA
6) University of Maryland, Fischell Department of Bioengineering, College Park, MD, USA
7) University of Maryland School of Medicine, Marlene and Stewart Greenebaum Cancer Center, Baltimore, MD,
USA8) University of Massachusetts at Boston, Department of Physics, College of Science and Mathematics, Boston,
MA, USA
Ascites results from the abnormal accumulation of fluid due to disease or abnormal pathology, and is most frequently
associated with advanced-stage epithelial ovarian cancer. The presence of ascites portends the poorest outcomes andis associated with significant co-morbidities. The movement of fluid in the peritoneum is also known to promotedisease progression and to facilitate the spread of tumor cells. An understudied area remains elucidating the role ofphysical stress (due to fluid flow) on treatment failure. Recent findings on flow-induced shear stress and resistanceto platinum-based chemotherapy in 3D models for ovarian cancer will be presented. The role of targeted
photodynamic therapy (PDT) and PDT-based combinations will be discussed.
References
1) Nath S, Pigula M, et al. Flow-induced Shear Stress Confers Resistance to Carboplatin in an Adherent ThreeDimensional Model for Ovarian Cancer: A Role for EGFR-Targeted Photoimmunotherapy Informed by PhysicalStress. J Clin Med, 2020.
2) Rizvi I, Gurkan UA, et al. Flow induces epithelial-mesenchymal transition, cellular heterogeneity and biomarker
modulation in 3D ovarian cancer nodules. Proc Natl Acad Sci USA, 2013.
3) Sorrin AJ, Kemal Ruhi M, et al. Photodynamic Therapy and the Biophysics of the Tumor Microenvironment.
Photochem Photobiol, 2020.
4) Conrad C, Gray KM, et al. Mechanical Characterization of 3D Ovarian Cancer Nodules Using Brillouin
Confocal Microscopy. Cell Mol Bioeng, 2019.
Acknowledgements. This work is supported by the following funding sources: Dr. Imran Rizvi - NIH R00
CA175292, NC Translational and Clinical Sciences Institute (NCTraCS), supported by the National Center for
Advancing Translational Sciences (NCATS), National Institutes of Health (NIH), through Grant Award Number
UL1TR002489, and UNC-NC State Joint BME Department Start-up Fund. Dr. Utkan Demirci - Stanford University
Canary Center Seed Grant, NCI-NIH grant to the Center for Cancer Nanotechnology Excellence for Translational
Diagnostics (CCNE-TD) at Stanford University (grant number: NIH U54 CA199075). Dr. Tayyaba Hasan – NIH
R01 CA158415, R01 CA160998, R01 CA119388 Dr. Huang-Chiao Huang - NIH R00 CA194269, UMD Start-up
Fund, UMD-UMB Research and Innovation Seed Grant, the Caring Together, NY, Ovarian Cancer Research grant,
and the UMD-NCI Partnership for Integrative Cancer Research Grant. Dr. Jonathan Celli – NIH U54CA156734
UMass Boston-Dana Farber/Harvard Cancer Center PartnershipBook of Abstracts ����������������������������������������������������������������������������������������������� 243

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IL-5.4.2
Role of photooxidative damage in the acquired resistance of cancer cells
Valentina Rapozzi, Mariachiara Gani, Francesca D’Este, Luigi E. Xodo
University of Udine, Department of Medicine, Udine, Italy.
In recent years, we have investigated acquired tumor resistance after photodynamic therapy (PDT) in PC3 prostatecancer cells. We followed step by step what happened after chronic photooxidative stimulation until the developmentof a more aggressive and resistant tumor population (1). We examined some of the molecular signaling pathways
involved in the oxidative stress response. We discovered that the modulation of these signaling pathways depend on
the intensity of the oxidative insult.
Since the study of these different signaling pathways and their interactions could provide clues for understanding theevents leading to an optimal PDT tumor response, our aim was to find a potential therapeutic target to prevent
acquired resistance and resensitize prostate cancer cells to PDT.
We focused on Raf Kinase Inhibitor Protein (RKIP), which plays a pivotal role in tumor survival, invasion, and drug
resistance (2). We showed that RKIP is involved in several PDT tumor response pathways, such as Mitogen ActivatedProtein Kinase (MAPK), Nuclear Factor Kappa-Light Chain-Enhancer of Activated B Cell (NF-kB), Nucleic Factor
Erythroid 2-related factor 2(Nrf2), BTB and CNC homology (BACH -1) and Glycogen Synthase Kinase 3 Beta (GSK3ß).
We found that during the development of acquired PDT resistance, the expression of pro-apoptotic RKIP wasinhibited, while the expression of pro-survival KRAS, NF-kB, BACH -1 and Nrf2 genes was upregulated.
With increasing photooxidative stress, RKIP was overexpressed and MAPK, NF-kB, BACH -1, pGSK 3B were
inhibited. Moreover, the subsequent increase in GSK 3ß expression led to a decrease in Nrf2 expression, which resensitized prostate cancer cells to PDT.
Our results highlight the role of RKIP as a potential convergence point of redox signaling pathways and suggest its
use as a potential diagnostic marker to determine the efficacy of PDT treatment by preventing cell resistance.
References
1) Della Pietra E, Simonella F, Bonavida B, Xodo LE, Rapozzi V. Repeated optimal photodynamic treatments withpheophorbide a induce an epithelial mesenchymal transition in prostate cancer cells via nitric oxide. Nitric Oxide
2015.
2). Zaravinos A, Bonavida B, Chatzaki E, Baritaki S. RKIP: a key regulator in tumor metastasisinitiation andresistance to apoptosis: therapeutic targeting and impact. Cancers 2018.
Acknowledgements. This work was supported by AIRC “Associazione Italiana per la Ricerca sul Cancro”, IG 2017,
Project Code 19898.

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OC-5.4.3
Effect of oxidative states on the efficiency of porphyrinoids in Photodynamic
Therapy of Prostate Cancer
Mariana Q. Mesquita1,2, Ana Rita Ferreira2, Maria G. P. M. S. Neves1, Margarida Fardilha2, Maria A. F. Faustino1
1) University of Aveiro, Department of Chemistry & LAQV-REQUIMTE, Aveiro, Portugal; 2) University of Aveiro,
Institute of Biomedicine – iBiMED & Department of Biomedical Sciences Institution, Aveiro, Portugal
Prostate cancer (PCa) is the second most common cancer in men worldwide, counting for more than 1.25 million
new diagnoses in 2018.1 Current treatment options for men with localized PCa include active surveillance, surgery(radical prostatectomy) and radiotherapy. Unfortunately, despite the clinical advances made in the last years
regarding the treatment modalities for PCa, there is a high rate of functional complications associated with theaforementioned therapeutic approaches, which affect patient’s quality of life.2,3 The need for reducing thesecomplications led to the development of PCa focal therapy. This type of therapy aims to minimize the side effects
observed with radical therapy to the structures that surround the prostate, which are required for normal erectile
function and urinary continence (specifically the bladder, rectum, neurovascular bundles and urinary sphincter)without compromising the oncological outcomes.2,4 Among several focal therapies, photodynamic therapy (PDT) has
been considered a promising local therapy to treat PCa.4
In the last decades, porphyrins have received increasing attention from the scientific community as favorable
photosensitizers (PS) for PDT due to their good absorption in the visible range of the electromagnetic spectrum,
effective production of reactive oxygen species (ROS), low dark toxicity, and stability/biocompatibility.5 Reduced
porphyrinoids, such as chlorins and isobacteriochlorins are also being explored as potential PSs, mainly due to theirhigher absorption bands in the red, when compared with porphyrin macrocycles.5 To address the oxidative state effecton the photosensitising properties, we synthesized three fluorinated porphyrinoid derivatives (porphyrin, chlorin andisobacteriochlorin) and incorporated them into polyvinylpyrrolidone (PVP). The obtained PS@PVP formulations
were fully characterized and their effectiveness as PS agents was evaluated in vitro against PCa cells (PC-3 cells). In
addition, the molecular mechanisms underlying the photocytotoxicity of the formulations towards PC-3 cells werealso investigated. In summary, in this communication, the synthesis, the characterization and the biologicalevaluation obtained with the three derivatives will be presented and discussed.
References
1) P. Rawla, Epidemiology of Prostate Cancer. World J. Oncol. 10, 63–89, 2019.
2) A. Kovács, M. Pinkawa, Interventional therapy in malignant conditions of the prostate. Radiologe. 59, 28–39,
2019.
3) O. Lodeizen, M. de Bruin, S. Eggener, S. Crouzet, S. Ghai, I. Varkarakis, A. Katz, J. L. Dominguez-Escrig, S.
Pahernik, T. de Reijke, J. de la Rosette, Ablation energies for focal treatment of prostate cancer. World J. Urol. 37,
409–418, 2019.
4) T. Gheewala, T. Skwor, G. Munirathinam, Photosensitizers in prostate cancer therapy. Oncotarget. 8, 30524–
30538, 2017.
5) S. Singh, A. Aggarwal, N. V. S. D. K. Bhupathiraju, G. Arianna, K. Tiwari, C. M. Drain. Glycosylated Porphyrins,
Phthalocyanines, and Other Porphyrinoids for Diagnostics and Therapeutics. Chem. Rev. 115, 10261–10306, 2015.
Acknowledgements. Thanks are due to University of Aveiro and FCT/MCTES (Fundação para a Ciência eTecnologia/Ministério da Ciência e Tecnologia, Portugal) for the financial support to LAQV-REQUIMTE(UIDB/50006/2020) and Institute for Biomedicine – iBiMED (UID/BIM/04501/2013 and POCI-01-0145-FEDER007628) through national funds and, when applicable, co-financed by European Union, FEDER, QREN, and
COMPETE within the PT2020 Partnership Agreement. Thanks are also due to the Portuguese NMR and Mass
Networks. This work was also financially supported by an individual fellowship grant from FCT to M.M(SFRH/BD/112517/2015).

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OC-5.4.4, P-5.4.14

Photodynamic efficacy of micelles containing a porphyrinic photosensitizer andpotassium iodide against the resistant melanoma cell line B16F10
Letícia D. Costa1, Kelly A. D. F. Castro1,2, Juliana A. Prandini2, Juliana C. Biazzotto2, Augusto C. Tomé1, Michael
R. Hamblin3, Maria da Graça P. M. S. Neves1, Maria A. F. Faustino1 and Roberto S. da Silva2
1) LAQV-REQUIMTE, Department of Chemistry University of Aveiro, Aveiro, Portugal; 2) Department of Physics
and Chemistry, Faculty of Pharmaceutical Sciences of Ribeirão Preto University of São Paulo, Ribeirão Preto, SãoPaulo, Brazil; 3) Laser Research Centre, Faculty of Health Science University of Johannesburg, Doornfontein 2028,
South Africa.
Melanoma is an aggressive and invasive form of skin cancer, that is characterized by being resistant to conventional
therapies1. Photodynamic therapy (PDT) is a promising approach to treat oncological and non-oncological skinpathologies, and an interesting alternative to increase the response of melanotic cancers to the treatment1. In the lastyears, several strategies have been studied to overcome this resistance to treatments, and almost all of theseapproaches aim at the melanosome disruption to hinder the melanin production1. Notwithstanding its crucial role in
the photoprotection of skin, melanin acts as powerful antioxidant, and the consequent decrease in the level of ROS
allows the cancer cells to survive and proliferate1. So, enhance the treatment response it is mandatory to overcome
the tolerance of these cells to the oxidative stress1.
Porphyrins have been widely utilized as photosensitizing agents (PS) against cancer cells or microorganisms mainly
due its ability to produce reactive oxygen species (ROS)2. In an attempt to increase the production of reactive species
and increase the death of cancer cells, the 1O2 produced after the PS photoactivation was combined with potassium
iodide (KI). To the best of our knowledge, this is the first approach about the combined action of these two species
against cancer cells, and in particular against the B16F10 melanoma cell line3. The porphyrinic PS and KI wereencapsulated into micelles, not only to avoid solubility constrains associated to the poor solubility of porphyrins in
the biological media, but also to increase the photodynamic effect due the proximity of all reactive species involved(Fig. 1)3. The biological results obtained with this approach will be presented and discussed in this communication.

Fig. 1 – Photodynamic efficacy of micelles containing a porphyrin PS and KI against B16F10 melanoma cells.

References
1) Y. Huang, D. Vecchio, P. Avci, R. Yin, M. Garcia-Diaz and M. R. Hamblin, Melanoma Resistance to
Photodynamic Therapy: New Insights, Biol. Chem. 2013, 394, 239-250.
2) M. Ethirajan, Y. Chen, P. Joshi and R. K. Pandey, The role of Porphyrin Chemistry in Tumor Imaging and
Photodynamic Therapy, Chem. Soc. Rev. 2011, 40, 340-362.
3) K. A. D. F. Castro, L. D. Costa, J. A. Prandini, J. C. Biazzotto, A. C. Tomé, M. R. Hamblin, M. G. P. M. S. Neves,
M. A. F. Faustino and R. S. da Silva, The Photosensitizing Efficacy of Micelles Containing a Porphyrinic
Photosensitizer and KI against Resistant Melanoma Cells, Chem. Eur. J. 2021, 27, 1990.
Acknowledgements: Thanks are due to Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq),
Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), Fundação de Amparo à Pesquisa do Estadode São Paulo (FAPESP- 2016/12707-0). We also thank the Fundação para a Ciência e a Tecnologia (FCT) for funding
the project PTDC/QEQ-QOR/6160/2014, the LAQV-REQUIMTE (UIDB/50006/2020) through national funds and,
when applicable, co-financed by FEDER within the PT2020 Partnership Agreement. K.A.D.F. Castro thanks CAPES
for the post-doctoral scholarship granted (PNPD/CAPES). L.D. Costa thanks FCT for the doctoral grant
SFRH/PD/BD/114578/2016. MRH was supported by US NIH Grants R01AI050875 and R21AI121700.
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SP-5.4.5, P-5.4.5
Discovery of a novel mitochondrial-targeted photosensitiser for photodynamic cancer
therapy
Radovan Krejcir1, Tomas Briza2,3, Martin Sterba1, Oliver Simoncik1, Petr Muller1, Philip J. Coates1, Pavel
Martasek4,5, Borivoj Vojtesek1, Pavlina Zatloukalova1
1) Masaryk Memorial Cancer Institute, RECAMO, Brno, Czech Republic; 2) Charles University, First Faculty of
Medicine, BIOCEV, Vestec, Czech Republic; 3) University of Chemistry and Technology Prague, Department ofAnalytical Chemistry, Prague, Czech Republic; 4) Charles University, First Faculty of Medicine, Department ofPediatrics and Adolescent Medicine, Prague, Czech Republic; 5) General University Hospital, Prague, Czech
Republic
Photodynamic therapy (PDT) uses light to activate an inactive drug (so-called photosensitiser) in patients´ tissues.
Its mechanism of action is based on the photodynamic generation of toxic molecular radicals, particularly reactive
oxygen species (ROS). In cancer, PDT enables easy targeting of the tumour mass in organs that are accessible forillumination. An ideal photosensitiser is not toxic without illumination but generates a high yield of ROS upon
photoactivation. Here, we present a newly identified photosensitiser from the polymethine class, salt 1, which shows
an inherent anticancer effect and mediates a substantial phototoxic reaction with the potential to strengthen its
therapeutic efficacy (1). Salt 1 selectively accumulates in the mitochondria of cancer cells and disrupts their functions,causing metabolic stress and inhibiting pro-growth signalling. As a result, autophagy is induced and, after depletionof cellular energy reserves, cells die in a non-apoptotic manner (2). Salt 1 was successfully tested in vivo in a mouse
xenograft model, where it showed a strong antitumour effect with no apparent toxicity for animals in the study (3).
As a photosensitiser, salt 1 interacts with 630nm light in the red region of the spectrum that penetrates well into
tissues. Combined with light, salt 1 generates ROS in the mitochondria of treated cells, which leads to disruption of
mitochondrial structure and a decrease in its membrane potential. Eventually, apoptosis is triggered in illuminatedcells. The difference in the viability of illuminated and non-illuminated cells in vitro reaches as much as 14-fold.
Moreover, the effective phototoxic reaction is induced with sub-micromolar concentrations of salt 1 (for A375 cells,
IC50=121nM) and low doses of activating light (5J/cm2), which is convenient for therapeutic applications (1).
Therefore, salt 1 is a candidate therapeutic molecule for exploiting the aberrant mitochondrial metabolism of cancer
cells with two modes of action – antimetabolic and photodynamic.
References
1) Krejcir R., Briza T., Sterba M. et al. Anticancer pentamethinium salt is a potent photosensitizer inducing
mitochondrial disintegration and apoptosis upon red light illumination. J. Photochem. Photobiol. B, Biol. 2020.
2) Krejcir R., Krcova L., Zatloukalova P. et al. A cyclic pentamethinium salt induces cancer cell cytotoxicity
through mitochondrial disintegration and metabolic collapse. Int. J. Mol. Sci. 2019.
3) Briza T., Kralova J., Dolensky B. et al. Striking antitumor activity of a methinium system with incorporated
quinoxaline unit obtained by spontaneous cyclization. ChemBioChem. 2015.
Acknowledgements
This work was supported by the Grant Agency of the Czech Republic (Grant No. 17-07822S), the European Regional
Development Fund (Project ENOCH, No. CZ.02.1.01/0.0/0.0/16_019/0000868) and by the MH CZ - DRO (MMCI,00209805).

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SP-5.4.6, P-5.4.6

Quantitative Imaging of Pancreatic Microtumors on Alginate Hydrogels for
Photodynamic Therapy Optimization
Nazareth Milagros Carigga Gutierrez, Tristan Le Clainche, Jean-Luc Coll, Lucie Sancey, Mans Broekgaarden
Institute for Advanced Bioscience, Univ. Grenoble Alpes, INSERM U1209, CNRS UMR5309, 38000 Grenoble,
FrancePancreatic adenocarcinoma is a highly resistant form of cancer for which new treatments are needed. Still, thedevelopment of new cancer therapeutics faces a major challenge when promising results obtained from 2D in vitro
systems fail to be confirmed in vivo. Three-dimensional culture models in which microscale tumors (microtumors)
are grown on hydrogel scaffolds may solve this issue, as they bridge the gap between conventional 2D cell culturesand in vivo experiments. The 3D architecture of these microtumors mimics biochemical- and biomechanical cues, aswell as cell-cell interactions more faithfully than 2D cultures. However, for culturing such microtumors, there is astrong need for inexpensive hydrogels that are easy to prepare and have low batch-to-batch heterogeneity. The aimof this study is to explore the use of sodium alginate-based hydrogels for culturing pancreatic microtumors in amanner that is compatible with state-of-the-art imaging assays1.
We evaluated the impact of new hydrogel formulations composed of different percentages of sodium alginate andgelatin. These hydrogels were assessed for generating pancreatic microtumos from the PANC-1 cell line. We
demonstrate that 2% alginate complemented with 0.5% gelatin formed hydrogels with similar performance as
Matrigel: PANC-1 cells formed a large population of microtumors after 17 days of culture. Furthermore, thesehydrogels were compatible with high-throughput image analysis1, 2, enabling us to apply these models to studying
the nano-bio interface of novel lipid nanocarriers for photodynamic therapy.
In conclusion, alginate-based hydrogels are inexpensive and effective scaffolds for 3D culture models of cancer, withversatile applications in research on novel cancer therapeutics. However, hydrogels need to be further evolved tomatch the complexity of native tissues3. For that purpose, new applications should be assessed for modeling thepancreatic cancer stroma using co-cultures. Ongoing work will focus on investigating cancer-stroma interactions andthe impact of cancer therapies.
References:
1. Carigga Gutierrez NM, Le Clainche T, Coll J-L, Sancey L, Broekgaarden
M. Microtumor cultures on Alginate-Gelatin hydrogels: Versatile 3D
models to quantify the uptake, dark toxicity, and efficacy ofphotosensitizing agents. Methods Mol Biol.:Accepted.
2. Bulin A-L, Broekgaarden M, Hasan T. Comprehensive high-throughput
image analysis for therapeutic efficacy of architecturally complex
heterotypic organoids. Sci Rep 2017; 7: 16645.
3. Caliari SR, Burdick JA. A practical guide to hydrogels for cell culture. Nat
Methods. 2016 May;13(5):405–14.
Figure 1. PANC-1 microtumors
growth after 14 days on alginategelatin hydrogel.

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IL-5.4.7

PCI-based strategies for overcoming resistance to cancer therapy
Pål Kristian Selbo1, Judith Jing Wen Wong1, Anette Weyergang1, Cathrine Elisabeth Olsen1, Anders Høgset2,
Kristian Berg1
1) Oslo University Hospital – The Norwegian Radium Hospital, Department of Radiation Biology, Oslo, Norway;
2) PCI Biotech, Oslo, Norway
Cancers develop in a Darwinian manner. Genetic and epigenetic changes cause origin of a malignant heterogeneous
cell population, of which the fittest cells survive selective pressure, e.g., cancer stem cells (CSCs) resisting cancer
therapy. Indeed, it has been demonstrated by independent labs that resistant progenies emerge from photodynamic
therapy (PDT)-treated cancer cells. Resistance to PDT arises over time after multiple treatments of the same cell
population and seems to be independent of photosensitizer type. However, the mechanism of resistance isdependent on the structure and intracellular localisation of the photosensitizer. We have shown that the MA11
breast cancer cells, overexpressing the efflux transporter and cancer stem cell (CSC) marker ABCG2, areintrinsically resistant to Pheophorbide A (PhA) PDT and that the cancer cells acquire stronger resistance withrepeated sequences of PhA-PDT. Enhanced resistance to PhA-PDT was associated with enrichment of cells
expressing ABCG2 on the cell surface1. The MA11 cells also acquired resistance to PDT using di-sulphonatedmeso-tetraphenylchlorin (TPCS2a) as photosensitizer, which is not a substrate of ABCG22. However, here theresistance was linked to dysregulated p38MAPK signalling (loss of p38-induced death signal) and enhanced
EGFR-expression compared to MA11 wild-type cells1. The PDT-resistant cells did neither develop cross-resistanceto chemotherapy (doxorubicin), radiotherapy nor the drug delivery method Photochemical Internalisation (PCI).
PCI-based targeting of EGFR with EGF-saporin was demonstrated to bypass the PDT resistance in the MA11cells1. In another study that will be presented, we show that 5-FU-resistant EMT-like pancreatic cancer cells arehighly sensitive to TPCS2a-PDT and hypersensitive to PCI of anti-CD105-saporin3. And finally, we recently
demonstrated that high activity of the CSC marker aldehyde dehydrogenase (ALDH) does not protect colon cancer
cells against TPCS2a-PDT as opposed to radiotherapy4. Altogether, our accumulating data on using PCI toovercome therapy resistance further strengthen the use of TPCS2a-PCI for targeting and elimination of CSCs.
References
1) C. E. Olsen, A. Weyergang, V. T. Edwards, K. Berg, A. Brech, S. Weisheit, A. Høgset and P. K. Selbo,
Development of resistance to photodynamic therapy (PDT) in human breast cancer cells is photosensitizerdependent: Possible mechanisms and approaches for overcoming PDT-resistance. Biochemical Pharmacology,
2017.
2) P. K. Selbo, A. Weyergang, M. S. Eng, M. Bostad, G. M. Mælandsmo, A. Høgset and K. Berg, Strongly
amphiphilic photosensitizers are not substrates of the cancer stem cell marker ABCG2 and provides specific andefficient light-triggered drug delivery of an EGFR-targeted cytotoxic drug. J.Control.Release, 2012.
3) K. Lund, C. E. Olsen, J. J. W. Wong, P. A. Olsen, N. T. Solberg, A. Hogset, S. Krauss and P. K. Selbo, 5-FU
resistant EMT-like pancreatic cancer cells are hypersensitive to photochemical internalization of the novel
endoglin-targeting immunotoxin CD105-saporin. J Exp Clin Cancer Res, 2017.
4) J. J. W. Wong and P. K. Selbo, High aldehyde dehydrogenase activity does not protect colon cancer cells
against TPCS2a-sensitized photokilling, Photochem Photobiol Sci, 2020.
Acknowledgements. These works were supported by grants 2017068 (P. K. S.), 2016023 (J. J. W. W.) and2014030 (C.E.O) from the South-Eastern Norway Health Authority and FU0803 (P. K. S.) from The Norwegian
Radium Hospital Research Foundation.

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IL-5.4.8

Resistance to Photodynamic Therapy of non melanoma skin cancer. Strategies toovercome it.
Juarranz A1, González S2, Gilaberte Y3
1) Department of Biology, Faculty of Sciences, Autónoma University of Madrid (UAM), Instituto Ramón y Cajal deInvestigación Sanitaria (IRYCIS), Madrid, Spain; 2) Department of Medicine and Medical Specialties, Alcalá deHenares University, Madrid, Spain; 3) Miguel Servet University Hospital, IIS Aragón. Zaragoza Spain.
Photodynamic therapy (PDT) employing methyl-aminolevulinate (MAL) is being used for actinic keratosis, in situ
squamous cell carcinoma (SCC) and basal cell carcinoma (BCC), precancerous and cancerous situations of nonmelanoma skin cancer (NMSC). However, the persistence of some tumors after treatment can occur. In fact, atclinical level, it has been described that some cutaneous SCC resistant to MAL-PDT become more aggressive, lessdifferentiated and/or acquire an infiltrative pattern1. The problem is not only due to characteristics of tumoral cells,
but also to the complex microenvironment around them (tumor microenvironment, TME) and particularly tocytokines secreted by one of the TME components, cancer associated fibroblasts (CAFs). CAFs have been reported
to be crucial in the resistance to therapies. Therefore, we are studying the PDT resistance mechanisms of NMSC andlooking for strategies to avoid it.
In our research we are using different in vitro and in vivo models for the evaluation of resistant cells alone or in
combination with CAFs, isolated from patients. Resistant cells obtained by repeated MAL-PDT treatments, grownas 2D or 3D (spheroids), constitute an excellent system to evaluate factors implicated in the response to the treatment.
In the same way, mice models, such as cell transplantation, permit evaluating tumorogenicity and aggressiveness ofresistant cells. We have observed alteration in genes implicated in the resistance response of SCC and BCC to MALPDT, including: ATPase, GAPDH, PKM2, CCND1, EFGR, MAP3K1 and SMADs. We consider that complementary
therapies to modulate them constitute potential strategies for the optimization of PDT in the treatment of NMSC.
References
1) Gilaberte Y et al., Cellular Intrinsic Factors Involved in the Resistance of Squamous Cell Carcinoma to

Photodynamic Therapy. J Investigativ Dermatol, 134: 2428-2437. 2014.

Acknowledgements. This research was funded by Spanish grants from Instituto de Salud Carlos III
MINECO and Feder Funds (PI18/00708).

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IL-5.4.9
Multifunctional platforms for PDT in hypoxic environment
Ludivine Larue1, Nolwenn Breton2, Bertrand Vileno2, Valérie Jouan-Hureaux3, Tataye Moussounda Moussounda
Koumba4, Samir Acherar5, Philippe Arnoux1, Cédric Boura3, Sylvain Marque4, Gérard Audran4, Céline Frochot1
1) CNRS, University of Lorraine, LRGP UMR 7274, Nancy, France 2) CNRS, University of Strasbourg, UMR
7177, Strasbourg, France 3) CNRS, University of Lorraine, CRAN UMR 7039, Nancy, France 4) CNRS, Universityof Marseille, ICR UMR 7273, Marseille, 5) CNRS, University of Lorraine, LCPM UMR 7274, Nancy, France
PDT can suffer from two drawbacks: the lack of selectivity of the PSs as well as the need for oxygen to beeffective. To remedy the lack of selectivity, it is possible to design photosensitizers or nanoparticles coupled tovectors that will target receptors over-expressed onto tumor cells1 or neovessels that allows the destruction of thevascularization and cause asphyxia of the tumor2. We already proved that KDPPR peptide has a good affinity for
the NRP-1 receptors overexpressed on endothelial cells3. To compensate for the lack of oxygen, several strategies
are possible4 and we focused on photoactivatable alkoxyamines, molecules capable of generating toxic radicals by
light activation. We will describe the synthesis of a multifunctional platform combining three units: a PS for an
oxygen-dependent PDT, a peptide to target NRP-1 receptors and an alkoxyamine for an oxygen-independent
activity. The study of the photophysical properties showed that the PS retained its capacity to induce the formationof singlet oxygen, the affinity tests confirmed the compound affinity for NRP-1. Thanks to the Electron
Paramagnetic Resonance (EPR) spectroscopy we could detect the radicals generated after illumination of thealkoxyamine. The proof of concept was thus successfully established5.

Fig. 1 – Structure of the new multifunctional platform.

References
1) Baydoun M., Morales O., Frochot C., Ludovic L., Leroux B., Thecua E., Ziane L., Grabarz A., Kumar A., de
Schutter C., Collinet P., Azaïs H., Mordon S., Delhem N. Photodynamic Therapy on Peritoneal Ovarian Cancer
Cells Induces the Release of Extra-cellular Vesicules with Immunoactivating Properties, Journal of Clinical
Medicine, 2020.
2) Thomas E., Colombeau L., Gries M., Peterlini T., Mathieu C., Thomas N., Boura C., Frochot C., Vanderesse R.,
Lux F., Barberi-Heyob M., Tillement O. Ultrasmall AGuIX Theranostic Nanoparticles for Vascular-TargetedInterstitial Photodynamic Therapy of Glioblastoma. International Journal of Nanomedicine, 2017.
3) Youssef Z., Yesmurzayeva N., Larue L., Juan-Hureaux V., Colombeau L., Acherar S., Frochot C. New targeted gold
nanorods for the treatment of glioblastoma by photodynamic therapy, Journal of clinical medicine, 2019.
4) Larue L., Myrzakhmetov B., Ben-Mihoub A., Moussaron A. Thomas N., Arnoux P., Baros F. Vanderesse R.,
Acherar S. , Frochot C. Fighting Hypoxia to Improve PDT, Pharmaceuticals, 2019.
5) Larue L., Moussounda Moussounda Koumba T., Le Breton N., Vileno B., Arnoux P., Jouan-Hureaux V., BouraC., Audran G., Bikanga R., Marque S.R.A., Acherar S., Frochot, C. The design of a targeting and Oxygenindependent platform to improve Photodynamic Therapy: A proof of concept, ACS applied Biomaterials, 2021.
Acknowledgements. This work was supported by the french ministry of research.

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OC-5.4.10

Metabolic reprogramming involved in resistance to photodynamic therapy in skinsquamous cell carcinoma. Metformin as adjuvant.
Mascaraque-Checa M1, Santiago JL2, Terrén S1, Nájera L3, González S4, Gilaberte Y5, Juarranz Á1.
1) Universidad Autónoma de Madrid, Dept. Biology, Madrid, Spain; 2) Hospital General Universitario de Ciudad
Real, Dept. Dermatology, Ciudad Real, Spain; 3) Hospital Universitarios Puerta de Hierro-Majadahonda, Dept.
Pathology, Madrid, Sapin; 4) Universidad de Alcalá, Dept. Medicine and Medical Specialities, Madrid, Spain; 5)
Hospital Universitario Miguel Servet, Dept. Dermatology, Zaragoza, Spain.
Squamous Cell Carcinoma (SCC) is the second most frequent type of skin cancer among the population1.
Within the clinically approved treatments, photodynamic therapy (PDT) with the methyl aminolevulinate acid (MAL)
is an extended non-invasive modality. Nevertheless, PDT is not always effective and resistant cells may appear after
treatment2. Normal differentiated cells depend primarily on mitochondrial oxidative phosphorylation to generateenergy, but cancer cells change this metabolism to an aerobic glycolysis (Warburg effect), which could influence inthe response to antitumor drugs3. A potential option to avoid recurrences is to combine PDT with other treatments.
In this work we have evaluated the metabolic changes that occur in PDT-resistant squamous cell carcinoma(SCC) cells. We have also analyzed metformin treatment, an antidiabetic type II compound, to sensitize SCC
PDT-resistant to PDT. For this we have used the human SCC cell lines: A-431 and SCC-13, which we have calledparental (P). From these cell lines, PDT resistant cells were generated by exposure to 10 cycles of PDT (1 mM MAL
followed by irradiation with red light). Resistant cells were inoculated in immunosuppressed mice, the induced

tumours were sub-cultured by explants and a cell population called 10GT was obtained. PDT resistant cells
showed a metabolic reprogramming (aerobic glycolysis increasing and OXPHOS reduction), which could
influence the response to PDT. This result was also confirmed in the induced tumors by SCC13 cells (Pand PDT-resistant) inoculation in mice. The metformin treatment caused a reduction in aerobic glycolysis
and an increases in oxidative phosphorylation in SCC resistant cells. Finally, the metformin addition to
PDT (combined treatment) improved the cytotoxic effect on P and resistant cells. The mechanism of actionof combined treatment is caused for increased protoporphyrin IX production, reactive oxygen species
generation and inhibition of AKT/mTOR pathway, through AMPK activation. In conclusion, we canindicate that PDT resistance implicate a metabolic reprogramming towards aerobic glycolysis andmetformin treatment reversed it. Therefore, metformin constitutes an excellent adjuvant for PDT in SCC.

References
1) Barton V, Armeson K, Hampras S… Alberg AJ. Nonmelanoma skin cancer and risk of all-cause and cancerrelated mortality: a systematic review. Archives of dermatological research, 2017.
2) Gracia-Cazaña T, Salazar N, Zamarron A… Juarranz A. Resistance of Nonmelanoma Skin Cancer to NonsurgicalTreatments. Part II:Photodynamic Therapy, Vismodegib, Cetuximab, Intralesional Methotrexate, andRadiotherapy. Actas Dermo-Sifiliograficas, 2016.
3) Bhattacharya B, Mohd Omar MF, Soong R. The warburg effect and drug resistance. British Journal of
Pharmacology, 2016.
Acknowledgements. This work was supported by Spanish grants from Instituto de Salud Carlos III MINECO andFeder Funds (PI18/00708).

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P-5.4.11

Could the length of the alkyl chain affect the photodynamic activity of tetraalkylpyridylporphyrins?
Matteo Rugiero, Enrico Caruso, Miryam Chiara Malacarne, Marzia Bruna Gariboldi
Department of Biotechnology and Life Sciences (DBSV), University of Insubria, Via J.H. Dunant 3, 21100 Varese(VA), Italy.
Photodynamic therapy (PDT) is a minimally invasive treatment that uses the combination of a photosensitizing
agent (PS) and light to selectively target solid tumors, as well as several nonneoplastic proliferating cell diseases.
Following systemic administration, PSs are activated by localized irradiation with visible light; in the presence ofadequate concentrations of molecular oxygen, this results in reactive oxygen species (ROS) formation andsubsequent tissue damage [1].
The family of porphyrins and their derivatives encompass the majority of photosensitizers and studies concerning
the synthesis and activity of new porphyrin derivatives, aiming the improvement of chemical-physical or activity
characteristics, are abundant in the scientific literature [2,3].
In this study, two series of tetra cationic alkyl pyridyl porphyrins, including four derivatives, were synthesized,
differing for the presence or absence of the zinc atom in the tetrapyrrole core; the four derivatives are characterized
by a different length of the alkyl chain (from one to twelve carbon atoms). The compounds were fully chemicallycharacterized, their light stability and singlet oxygen production determined. To determine a possible relationshipbetween the photodynamic activity and the length of the alkyl chain, the effects on cell viability were also
evaluated on a panel of tumor cell lines, following PSs activation.
References[1] Macdonald, I.J.; Dougharty, T.J. Basic principles of photodynamic therapy. J. Porphyrins Phthalocyanines
2001, 5(2), 105-129.
[2] Banfi, S.; Caruso E.; Buccafurni, L.; Murano, R.; Monti, E.; Gariboldi, M.B.; Papa, E.; Gramatica, P.
Comparison between 5,10,15,20-tetraaryl- and 5,15-diarylporphyrins ad photosensitizers: Synthesis, photodynamic
activity, and quantitative structure-activity relationship modeling. J. Med. Chem. 2006, 49(11), 3293–3304.
[3] Kou, J.; Dou, D.; Yang, L. Porphyrin photosensitizers in photodynamic therapy and its applications. Oncotarget
2017, 8(46), 81591-81603

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P-5.4.12

Zn(II) and free base N-methylated tripyridylporphyrins: impact of solubility andlight excitation wavelength on PDT effectMartina Mušković1, Iva Džeba2, Nikola Basarić3, Martin Lončarić4, Branka Mihaljević2, Ivana Ratkaj1, Nela
Malatesti1
1) University of Rijeka, Department of biotechnology, Rijeka, Croatia; (e-mail: nela.malatesti@biotech.uniri.hr)
2) Ruđer Bošković Institute, Division of Material Chemistry, Radiation chemistry and Dosimetry Laboratory,
Zagreb, Croatia
3) Ruđer Bošković Institute, Division of Organic Chemistry and Biochemistry, Laboratory of Synthetic Organic
Chemistry, Zagreb, Croatia
4) Ruđer Bošković Institute, Centre of excellence for advanced material and sensors, Photonics and quantum optics
unit, Zagreb, Croatia
Amphiphilic cationic porphyrins have shown high potential in use as photosensitizers (PSs) for photodynamic
therapy (PDT). Our research group has shown that compounds conjugated with long chain (C18) have an exceptional
PDT effect compared to hydrophilic analogues on different tested cell lines. Unfortunately, highly effective
amphiphilic compounds showed reduced selectivity to tumour cells, relatively high dark toxicity, and theiraggregation was observed in high concentration1. It must be kept in mind that the optimal ratio of hydrophilic andhydrophobic moieties is necessary to facilitate passage through the membrane bilayer, while maintaining goodsolubility in water and avoiding formation of aggregates that reduce the PDT effect2.
Chelation with Zn(II) alters the properties of free base porphyrins; it changes lipophilicity of the compounds and may
increase the lifetime of the excited triplet state (3PS*). The change also occurs in the absorbance spectra, as expected
the number of Q bands reduces from four to two and there is a negligible absorption above 610 nm for thesecompounds to be efficiently activated with red. Therefore, we suggest photoactivation using LED based source oforange light, which better corresponds to the optical properties of Zn(II) porphyrins while maintaining good
penetration through the tissue4.In this work, we will present the synthesis of two groups of N-methylated
tripyridylporphyrins, free base and chelated with Zn(II), both conjugated to alkyl chains of different length. Their
photophysical and photochemical properties studied in different solvents, by laser flash photolysis (LFP) and timeresolved fluorescence spectroscopy (TRF) will be presented. The impact of small changes in chain length will also
be evaluated in vitro, using MTT assay to detect cytotoxicity and measuring cellular uptake by using fluorescencemicroscopy on melanoma cells (MeWo) and human foreskin fibroblasts (HFF). To study the effect of the irradiation
wavelength, the cytotoxicity of the compounds will be evaluated using red light (645 nm; fluence rate 2.0 mW / cm2)and orange light (605 nm; fluence rate 1.5 mW / cm2).
References
1) M. Jelovica, P. Grbčić, M. Mušković, M. Sedić, S. Kraljević Pavelić, M. Lončarić, In Vitro Photodynamic
Activity of N-Methylated and N-Oxidised Tripyridylporphyrins with Long Alkyl Chain and Their Inhibitory
Activity in Sphingolipid Metabolism. ChemMedChem, January 2018.
2) N. Malatesti,I. Munitić, I. Jurak, Porphyrin-based cationic amphiphilic photosensitizer as potential anticancer
antimicrobial and immunosuppressive agents. Biophysics Reviews, April 2017.
3) L. Benov, J.Craik, I. Batinić-Haberle, The Potential of Zn(II) N-Alkylpyrdylporphyrins for Anticancer
Therapy. Anti-Cancer Agents in Medicinal Chemistry, February 2012.
4) M. Kim, A. Darafsheh, Light Sources and Dosimetry Techniques for Photodynamic Therapy.
Photochemistry and Photobiology, March 2020.
Acknowledgements. This work was supported by the University of Rijeka grants uniri-prirod-18-173. The
equipment used in this work is part of the project “Research Infrastructure for Campus-based Laboratories at the
University of Rijeka”, financed by European Regional Development Fund (ERDF) and the Ministry of Science,
Education and Sport of the Republic of Croatia (RC.2.2.06-0001).

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P-5.4.13
Polyvinylpyrrolidone formulations of porphyrinoids as photosensitizers for
Photodynamic Therapy of Prostate Cancer
Mariana Q. Mesquita1,2, Ana Rita Ferreira2, Maria G. P. M. S. Neves1, Margarida Fardilha2, Maria A. F. Faustino1
1) University of Aveiro, Department of Chemistry & LAQV-REQUIMTE, Aveiro, Portugal; 2) University of Aveiro,
Institute of Biomedicine – iBiMED & Department of Biomedical Sciences Institution, Aveiro, Portugal
Prostate cancer (PCa) is the second most frequent cancer diagnosed in men worldwide.1 Conventional treatment
options to treat this cancer include active surveillance, radical prostatectomy, radiotherapy, and/or androgendeprivation therapy. Unfortunately, all these treatments may cause several side effects that may seriously impair thequality of life of each patient.2,3 The necessity of reducing these complications led to the development of PCa focal
therapy, which aims to reduce the side effects seen with radical therapy, while maintaining oncological control.2
Among the common treatment options, photodynamic therapy (PDT) is being considered a promising local therapy
to treat PCa.4 Although PDT is an established treatment modality approved for several types of cancer, the low
solubility, the reduced tumor selectivity, the absorption in the therapeutic window and the poor clearance from thebody of the currently approved photosensitizers (PS) hampers its wide clinical application.5 In this regard, herein we
synthesized three fluorinated porphyrinoid derivatives (porphyrin, chlorin and isobacteriochlorin derivatives) andentrapped them into polyvinylpyrrolidone (PVP) to prevent the aggregation and preserve the desirable photophysicalproperties of the compounds under physiological environment. The photodynamic efficacy of the synthesized
formulations as PS was subsequenytly assessed in vitro against a PCa cell line (PC-3 cells). Furthermore, themolecular mechanisms underlying the photocytotoxicity of the formulations towards PC-3 cells were also elucidated.
References
1) P. Rawla, Epidemiology of Prostate Cancer. World J. Oncol. 10, 63–89, 2019.
2) A. Kovács, M. Pinkawa, Interventional therapy in malignant conditions of the prostate. Radiologe. 59, 28–39,
2019.
3) O. Lodeizen, M. de Bruin, S. Eggener, S. Crouzet, S. Ghai, I. Varkarakis, A. Katz, J. L. Dominguez-Escrig, S.
Pahernik, T. de Reijke, J. de la Rosette, Ablation energies for focal treatment of prostate cancer. World J. Urol. 37,
409–418, 2019.
4) T. Gheewala, T. Skwor, G. Munirathinam, Photosensitizers in prostate cancer therapy. Oncotarget. 8, 30524–
30538, 2017.
5) X. Zhao, J. Liu, J. Fan, H. Chao, X. Peng, Recent progress in photosensitizers for overcoming the challenges ofphotodynamic therapy: from molecular design to application. Chem. Soc. Rev. 50, 4185–4219, 2021.
Acknowledgements. Thanks are due to University of Aveiro and FCT/MCTES (Fundação para a Ciência eTecnologia/Ministério da Ciência e Tecnologia, Portugal) for the financial support to LAQV-REQUIMTE(UIDB/50006/2020) and Institute for Biomedicine – iBiMED (UID/BIM/04501/2013 and POCI-01-0145-FEDER007628) through national funds and, when applicable, co-financed by European Union, FEDER, QREN, and
COMPETE within the PT2020 Partnership Agreement. Thanks are also due to the Portuguese NMR and Mass
Networks. Image acquisition was performed in the LiM facility of iBiMED, a node of PPBI (Portuguese Platform of
BioImaging): POCI-01-0145-FEDER-022122. This work was also financially supported by an individual fellowshipgrant from FCT to M.M (SFRH/BD/112517/2015).

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IL-6.1.2

The Origin of the Phytochrome A Functions.
Akira Nagatani, Misato Kikuchi, Yukiko Yoshikawa, Kanako Shinohara, Tomomi Suzuki, Nobuyoshi Mochizuki
Kyoto University, Graduate School of Science, Kyoto, Japan
PHYA and PHYB are major members of the phytochrome family. They have diverged very early in the history of
seed plants to have distinctive functions (Mathews, 2010). PHYA responds to continuous far-red (FR-HIR) and very
low fluence red light (VLFR), whereas PHYB mediates responses to low fluence red light (LFR). In addition, PHYA
but not PHYB is rapidly degraded upon photo-activation. Nevertheless, their overall structures are very similar. All
phytochromes consist of the photo-sensory N-terminal and the dimerizing C-terminal moieties. The N-terminal half
is further divided into 4 domains, namely the N-terminal extension, PAS, GAF and PHY. The high structuralsimilarity between different phytochromes allow us to study functions of the chimeric phytochromes. We havesystematically expressed 16 chimeric phytochromes between PHYA and PHYB in Arabidopsis and examined how
they behave in vivo. The results indicate that the PAS domain is important for nuclear localization under continuous
FR, whereas PHY domain tunes the light sensitivity of phytochrome (Oka et al., 2012). In addition, GAF is important
for rapid degradation upon exposure to light.
The PHY domain is spatially separated from the PAS-GAF domain to which the chromophore is attached.
Interestingly, a peculiar “tongue” structure is protruded from the PHY domain towards the chromophore (Essen etal., 2008: Yang et al., 2008). Furthermore, a few amino acid substitutions in this region are known to affect the activityof phytochrome. Hence, we constructed chimeric phytochromes with PHYA or PHYB “tongue” structure andcompared their activities. Consequently, the PHYA tongue sequence was shown to confer higher light sensitivity tothe phytochrome. Next, we replaced several amino acid residues, one by one, in the PHYA tongue structure withrespective PHYB amino acid residues in a PHYB-based chimeric phytochrome. The activity of the original chimeric
phytochrome to respond to weak R and continuous FR were simultaneously diminished in three of the mutants,
thereby 3 amino acid residues were indicated to be involved in the sensitization of phytochrome within the “tongue”
structure.
Intriguingly, two of those residues are predicted to face each other in the basal region of the tongue structurein the 3 D model of phytochrome molecule, whereas one of them is situated in the vicinity of the chromophore.
Hence, insights into how the light sensitivity is controlled by the tongue structure are provided. We then examined
the phylogenetic tree of phytochromes (Li et al., 2015) to infer when those amino acid substitutions took place during
the evolution of seed plants. To our surprise, the amino acid residues in these three positions are highly conserved
among phytochromes not only in ferns, mosses and liverworts but also in gymnosperms, indicating that the lightsensitivity adjustment through the mutations of these amino acid residues happened only recently after the split ofangiosperms. Secondly, the first position is mutated only in the PHYA linage whereas the second position is mutatedonly in PHYB/E. The third position is mutated in both the linages but to different amino acids. Taken together, lightsensitivity is inferred to be modified both in PHYA and PHYB but to the opposite directions early in the history ofangiosperms. As discussed by Mathews (2010), the ability to respond to a wider light intensity under the sun andcanopy might be beneficial for ancestral angiosperms to spread in the forest dominated by non-vascular plants.
References
Sarah Mathews, Evolutionary Studies Illuminate the Structural-Functional Model of Plant Phytochromes, Plant
Cell, 22:4-16, 2010
Yoshito Oka et al., Arabidopsis Phytochrome A Is Modularly Structured to Integrate the Multiple Features That Are
Required for a Highly Sensitized Phytochrome, Plant Cell, 24:2949-62, 2012
Lars-Oliver Essen et al., The structure of a complete phytochrome sensory module in the Pr ground state, Proc.
Natl. Acad Sci. USA, 105:14709-14, 2008
Xiaojing Yang et al., Crystal structure of Pseudomonas aeruginosa bacteriophytochrome: photoconversion andsignal transduction, Proc. Natl. Acad Sci. USA, 105:14715-20, 2008
Fay-Wei Li et al., Phytochrome diversity in green plants and the origin of canonical plant phytochromes, Nat
Commun.,6:7852, 2015

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IL-6.1.3
Structural Basis for Light Control of Cell Development: New Insights from Bacterial
Phytochrome Proteins
Emina A. Stojković1, Sebastian Westenhoff2, Marius Schmidt3
1) Northeastern Illinois University, Department of Biology, Chicago, IL, USA; 2) University of Gothenburg,
Department of Chemistry and Molecular Biology, Gothenburg, Sweden; 3) University of Wisconsin-Milwaukee,
Department of Physics, Milwaukee, WI, USA
Phytochromes (PHYs) are photoreceptor proteins first discovered in plants, where they control a variety ofphotomorphogenesis events. PHYs as photochromic enzymes that usually contain C-terminal kinase domain canreversibly switch between a red light absorbing (Pr) and a far-red light absorbing (Pfr) state. Light signals aretransduced as structural change through the entire protein to modulate its activity. The unexpected discovery ofbacteriophytochromes (BphPs) in non-photosynthetic bacteria has opened new frontiers in our understanding of themechanisms by which these natural photoswitches can control single cell development. BphPs from the nonphotosynthetic myxobacterium Stigmatella aurantiaca are of special interest. Myxobacteria are distinguished amongprokaryotes by a multicellular stage in their life cycle known as fruiting bodies, which in S. aurantiaca is controlled
by light. We determined the crystal structures of the two BphPs from S. aurantiaca, denoted SaBphP1 and SaBphP2.
They have distinct photochemistry, although they bind the same bilin chromophore and share a large sequence
identity. Unlike classical BphPs, wild-type SaBphP1 lacks a conserved histidine (His289) that stabilizes thecovalently-bound bilin chromophore and undergoes limited Pr/Pfr photoconversion that can be restored by a single
Thr289His mutation. Furthermore, the crystal structures of SaBphP1 wild-type and Thr289His mutant in comparison
to classical SaBphP2 wild-type protein differ in the conformation of the essential bilin chromophore as well as
important protein dimer interface in the dark-adapted Pr state. Recently, we determined short-lived crystal structuresof the photosensory core modules of SaBphP2 captured by an X-ray free electron laser 5 ns and 33 ms after lightillumination of the Pr state1. We observe large structural displacements of the covalently bound bilin chromophore,
which trigger a bifurcated signaling pathway that extends through the entire protein. The snapshots show with atomic
precision how the signal progresses from the chromophore, explaining how plants, bacteria, and fungi sense red light.
References
1) M. Carrillo, S. Pandey et al, High-resolution Crystal Structures of Transient Intermediates in the Phytochrome
Photocycle. Structure 2021 Mar 20: S0969-2126(21)00077-0. doi: 10.1016/j.str.2021.03.004. Online ahead of print.
Acknowledgements. This work was supported by National Science Foundation (NSF) Science and Technology
Centers (STC) grant NSF 1231306 (Biology with X-ray Lasers).

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OC-6.1.4
QM/MM Simulations of Spectral Tuning in Phytochrome-like Photoreceptors
Christian Wiebeler1
1) Institute for Analytical Chemistry, Faculty of Chemistry and Mineralogy, Leipzig University, Germany
Phytochromes constitute a diverse family of photoreceptors and one of its subfamilies are Cyanobacteriochromes(CBCRs), which are composed of a single GAF domain. CBCRs are promising candidates for biotechnologicalapplications, owing to their photochromism, compactness and spectral diversity. In case of the CBCR Slr1393, one
isomer absorbs red light (Pr) and the other one green (Pg).1 These two forms can be interconverted into each otherby light illumination. Slr1393 binds phycocyanobilin (PCB) as chromophore and the crystal structures of both forms
have been obtained recently.2 In this contribution, we will focus on results from hybrid quantum mechanics/molecularmechanics (QM/MM) simulations for the Pr and Pg forms of Slr1393.3
Our QM/MM studies started from the crystal structures. First, the structures were optimized in several stages,
followed by classical molecular dynamics (MD) for thermalization and backbone relaxation. During these steps, it
was checked that the non-covalent interactions of PCB with the protein remained intact. The snapshots for excitedstate calculations were then generated via QM/MM MD. The final spectrum is an average of the spectra from thedifferent geometries of each form and the results are complemented with wave function analysis.

Fig. 1 – The difference in absorption between the red light-absorbing dark state and the green light-absorbing photoproduct of
Slr1393 could be traced back to changes in the effective conjugation lengths of the chromophore in the two conformations.3

In addition, also results from a benchmark study for this protein will be presented.4 Its focus is on the choice of an
appropriate semiempirical method for QM/MM MD, on methods for excited state calculations and on the importanceof sampling. More recently, we have carried out similar simulations for further proteins, e.g. for All2699,5 andpublished a review article on computational studies of the photochemistry in phytochromes.6
References
1) X. Xu et al., Combined Mutagenesis and Kinetics Characterization of the Bilin-Binding GAF Domain of theProtein Slr1393 from the Cyanobacterium Synechocystis PCC6803. ChemBioChem 15, 1190–1199, 2014.
2) X. Xu, A. Höppner, C. Wiebeler, K.-H. Zhao, I. Schapiro, and W. Gärtner, Structural Elements Regulating the
Photochromicity in a Cyanobacteriochrome. Proc. Natl. Acad. Sci. USA 117, 2432–2440, 2020.
3) C. Wiebeler, A. G. Rao, W. Gärtner, and I. Schapiro, The Effective Conjugation Length Is Responsible for the
Red/Green Spectral Tuning in the Cyanobacteriochrome Slr1393g3. Angew. Chem. Int. Ed. 58, 1934–1938, 2019.
4) C. Wiebeler and I. Schapiro, QM/MM Benchmarking of Cyanobacteriochrome Slr1393g3 Absorption Spectra.
Molecules 24, 1720, 2019.
5) C. Slavov et al., The Interplay between Chromophore and Protein Determines the Extended Excited State
Dynamics in a Single-Domain Phytochrome. Proc. Natl. Acad. Sci. USA 117, 16356–16362, 2020.
6) J. R. Church, A. G. Rao, A. Barnoy, C. Wiebeler and I. Schapiro, Computational Studies of Photochemistry inPhytochrome Proteins. Challenges and Advances in Computational Chemistry and Physics 31, 197–226, 2021.
Acknowledgements. We acknowledge funding by the German Research Foundation via grants: WI 4853/1-1 andWI 4853/2-1.
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IL-6.1.5

Bridging the language barrier between sensor and effector domains inbacteriophytochromes
Cornelia Böhm1, Nikolina Todorovic1, Marco Balasso1, Geoffrey Gourinchas1 and Andreas Winkler1
1) Graz University of Technology, Institute of Biochemistry, Graz,
Bacteriophytochrome sensors are versatile entities that during evolution have frequently been repurposed to control
a variety of biological functions. Among other modes of operation, the direct allosteric regulation of enzymaticfunctionalities exists for histidine kinases, diguanylate cyclases, cyclic-dimeric-GMP phosphodiesterases and metal
dependent protein phosphatases1. Therefore, repurposing bacteriophytochrome sensors to control enzymaticfunctionalities that have not been created during evolution is one interesting approach for the design of novel
optogenetic tools. To this end, however, the molecular mechanisms of how different sensors are functionally coupled
to diverse effectors need to be understood in more detail.
In an order to tackle this challenging question, we have recently characterized chimeric proteins between two related
phytochrome-activated diguanylate cyclases, with pronounced functional differences of the parent proteins, in detail.
Thereby we have obtained insights into the functional roles of different structural elements and their coupling witheach other. Among other important findings, the central regulatory role of the PHY domain dimer interface has
manifested itself in being one of the key determinants for efficient light signal integration and transduction to theeffector domain. Understanding how a particular structural element is modulated by upstream and/or downstream
elements provides important functional insight and the major strength of our chimeras is to highlight elements that,when targeted for modifications, influence specific phytochrome properties. In the end, any novel optogenetic tool
is a chimeric system generated from a photosensory module and an effector of interest. Being able to build uponinsights from chimeras within closely related systems will therefore also be beneficial for more ambitious new
optogenetic tool developments. Appreciating the fact that more than just linker length and/or linker composition isneeded for the creation of a functional sensor-effector system is one of the key-findings that will be discussed in thispresentation.
References
1) Geoffrey Gourinchas, Stefan Etzl, Andreas Winkler, Bacteriophytochromes – from informative model systems ofphytochrome function to powerful tools in cell biology. Curr. Opin. Struct. Biol. 57, 72–83. 2019.
Acknowledgements. This work was supported by the Austrian Science Fund (FWF): P32022 to AW.

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IL-6.1.6

Biochemical activity of a model bacteriophytochrome from Deinococcus radiodurans
Elina Multamäki1, Rahul Nanekar2, Dmitry Morozov3, Topias Lievonen2, David Golonka4, Weixiao Yuan
Wahlgren5, Brigitte Stucki-Buchli2, Jari Rossi1, Vesa P. Hytönen6, Sebastian Westenhoff5, Janne A. Ihalainen2,
Andreas Möglich4, Heikki Takala1,2
1) University of Helsinki, Department of Anatomy, Helsinki, Finland; 2) University of Jyvaskyla, Department of
Biological and Environmental Science and Nanoscience Center, Jyvaskyla, Finland; 3) University of Jyvaskyla,
Department of Chemistry and Nanoscience Center, Jyvaskyla, Finland; 4) Universität Bayreuth, Lehrstuhl für
Biochemie, Bayreuth, Germany; 5) University of Gothenburg, Department of Chemistry and Molecular Biology,
Gothenburg, Sweden;6) University of Tampere, Faculty of Medicine and Health Technology, Tampere, Finland
Bacteriophytochrome photoreceptors usually belong to two-component signaling systems, which transmit
environmental stimuli to a response regulator through a histidine kinase domain. Phytochromes switch between red
light-absorbing (Pr) and far-red light-absorbing (Pfr) states. Despite exhibiting extensive structural responses during
this transition,1,2 no kinase activity of the model bacteriophytochrome from Deinococcus radiodurans (DrBphP) has
been reported. Here, we resolve this long-standing conundrum by comparatively analyzing the interactions and output
activities of DrBphP and a bacteriophytochrome from Agrobacterium fabrum (Agp1).3 Whereas Agp1 acts as aconventional histidine kinase, we identify DrBphP as a light-sensitive phosphatase (Fig. 1). While Agp1 binds its
cognate response regulator only transiently, DrBphP does so more strongly, which is rationalized at the structurallevel. Our data pinpoint two key residues affecting the balance between kinase and phosphatase activities in twocomponent signaling. The opposing output activities in two highly similar bacteriophytochromes suggest the use oflight-controllable histidine kinases and phosphatases for optogenetics.

Fig. 1 – Proposed signalling mechanism of two bacteriophytochromes. Agp1 acts as an active histidine kinase byphosphorylating its cognate response regulator AtRR1 in Pr state. DrBphP de-phosphorylates its cognate response regulator
DrRR in Pfr state, therefore acting as a phyohatase in two-component signalling.

References
1) Takala H, et al., Signal amplification and transduction in phytochrome photosensors. Nature, 509: 245-248, 2014
2) Björling A, et al., Structural photoactivation of a full-length bacterial phytochrome. Science Advances, 2(8):e1600920, 20163) Multamäki E, et al., Illuminating a Phytochrome Paradigm – a Light-Activated Phosphatase in Two-Component
Signaling Uncovered. bioRxiv, 2020
Acknowledgements. This work is supported by Academy of Finland grants 285461 and 330678, Three-year grant2018–2020 from the University of Helsinki, and Bayreuth Humboldt Centre Senior Fellowship 2020.

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OC-6.1.8

Characterising Phytochrome-Activated Diguanylyl Cyclases
Böhm Cornelia1,2, Todorovic Nikolina1, Balasso Marco1, Hujdur Elvira1, Gourinchas Geoffrey1, Sensen Christoph
Wilhelm1, Winkler Andreas1,2
1) Graz University of Technology, Department of Biochemistry, Graz, Austria; 2) BioTechMed-Graz, Graz, Austria
Protein dynamics play a major role for the catalytic function of enzymes, the interaction of protein complexes orsignal integration in regulatory proteins. In the context of multi-domain proteins involved in light-regulation of
enzymatic effectors, the central role of conformational dynamics is well established. Light activation of sensory
modules is followed by long-range signal transduction to different effectors; rather than domino-style structural
rearrangements, a complex interplay of functional elements is required to maintain functionality.
These sensor-effector systems are frequently linked to second messenger production influencing lifestyle decisions
in bacteria or higher organisms. One family of such photoreceptor systems are red-light-regulated phytochromes that
control diguanylate cyclases involved in cyclic-dimeric-GMP formation. Based on structural and functional studies
of one prototypic family member, the central role of a coiled-coil sensor-effector linker element was established.
Interestingly, subfamilies of the phytochrome-activated diguanylate cyclases feature strongly varying biochemical
characteristics, especially with respect to the linker length. The evolutionary reasons for these differences and thefunctional implications as well as the dynamic interplay of the domains involved, however, are presently not fully
understood. We have found that the PHY domain dimer interface plays an essential role in signal integration, andthat a functional coupling with the coiled-coil linker element is crucial; the phytochrome-spanning helical spine is an
essential structural element involved in light-dependent upregulation of enzymatic turnover. Moreover, a structurally
asymmetric light state1, even though potentially advantageous for substrate binding and product formation2, does notappear to be an absolute requirement for red light induced increase in GTP turnover in bacteriophytochromes.
However, isolated structural elements can frequently not be assigned to specific properties of thebacteriophytochrome sensors, which further emphasises the importance of global conformational dynamics indefining photocycle properties as well as structural heterogeneity.
We provide insights into the intricate processes at play during light signal integration and transduction in thesephotosensory systems. Since deep tissue penetration properties of red light render phytochrome-regulated systems
particularly attractive candidates for the rational design of optogenetic tools, our results provide additional guidelines
for a more directed design of novel sensor-effector combinations by detailing the complex mechanisms of signal
integration as well as inter-domain conformational dynamics in phytochrome systems.

References
1) Gourinchas, G., Heintz, U., Winkler, A, Asymmetric activation mechanism of a homodimeric red light-regulated
photoreceptor. eLife, 2018.
2) Gourinchas, G., Vide, U., Winkler, A, Influence of the N-terminal segment and the PHY-tongue element on lightregulation in bacteriophytochromes. JBC, 2019.
Acknowledgements. This work was supported by the Austrian Science Fund (FWF): P32022 (AW).
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IL-6.2.1

How the Montreal Protocol Saved Earth’s Skin (and our own)
Richard McKenzie1
1) NIWA Lauder, PB 50061 Omakau, Central Otago, New Zealand
I’m proud to have played a small part in the success of the Montreal Protocol on Protection of the Ozone Layer, boththrough my research at Lauder and through participation in the Ozone Science Assessments and Environmental
Effects Panel Assessments over the last four decades. Our high quality measurements of spectral UV irradiances havebeen made continuously at Lauder, New Zealand, since 1989. An analysis of these data, and data from other NDACCcompliant sites throughout the world, which was published in 20191, was an important end-point to my formal science
career, culminating with the publication last year of my book, “Saving our Skins” 2.
The analysis showed that because of compliance to the Montreal Protocol, UV levels at Lauder and throughout mostof the world were no longer increasing and were in fact beginning to show signs of recovery at mid to high southernlatitudes (the picture was less clear-cut in the northern hemisphere, probably due to its larger changes in clouds andaerosols). Without the Montreal Protocol, damage to our atmosphere would have already been severe, with ozone
levels diminished by around 20 percent, leading to UV increases of a similar magnitude. That damage to Earth’s skinwould have flowed on to damage to our own skins. New Zealand and Australia have the world’s highest rates of skincancer, and for every 1 percent reduction in ozone, that rate is expected to increase by 2 or 3 percent. The health costs
would have been considerable, and the flow on economic costs of treatment. And, as everybody in this audience will
be aware, human health is just one of many environmental implications. In the two years that have passed since that
paper was published, the recovery in ozone has continued at Lauder, as shown in the figure below.

Fig. 1 – Relative changes in summer UV at Lauder from the late 1970s when satellite ozone data became available to the present.
Measured changes are compared with model calculations with and without compliance to the Montreal Protocol on protectionof the Ozone Layer (preliminary data update from ref 1).

The situation is more complex at polar latitudes, where severe ozone loss has been seen both in the Arctic and theAntarctic springs in recent years. This is no cause for despair. Two criteria must be met for rapid ozone depletion
through the heterogeneous chemical processes that take place there on the surfaces of Polar stratospheric clouds(PSCs). Firstly, chlorine levels must be elevated above about a threshold around 1 ppb, and secondly temperatures
must be lower than the threshold needed for the formation of PSCs. Although chlorine levels have already passed
their peak, it will be decades before they return to the levels present before the Antarctic ozone hole first formed inBook of Abstracts ����������������������������������������������������������������������������������������������� 262

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the 1980s. In the meantime, as climate change progresses due to the continued release of Greenhouse Gases, energy
is trapped in the lower atmosphere leading to reduced temperatures in the stratosphere. With those lower
temperatures, the volume of air cold enough for PSCs to form has increased. Consequently, years with anomalously
large ozone losses at high latitudes are expected for the next few years. Meanwhile ozone levels should continue torecover outside polar regions, and when chlorine levels return below 1 ppb, the problem in polar regions will also beover. Projected changes for the future UV irradiances will depend on changes in clouds and aerosols as much aschanges in ozone.
We’re still on track. A summary of Lauder’s wider contribution to ozone research was published last year 3, but if
you’re interested in the UV story, please get a copy of my book.3

References
1) McKenzie R, Bernhard G, Liley B et al. Success of Montreal Protocol demonstrated by comparing high-quality
UV measurements with “World Avoided” calculations from two chemistry-climate models. Scientific Reports 2019;9: 12332.
2) Richard McKenzie, Saving our Skins, available in three formats from Amazon, kindle, black & white paperback
ISBN 979863473987, and colour paperback ISBN 9798654069511, 2020
3) McKenzie R. Lauder and its part in the ozone success story. SPARC Newsletter 2021: 16-21.
Acknowledgements. I’m grateful to Germar Bernhard for his assistance producing the final version of Figure 1. I’dalso like to take this opportunity express my gratitude to all the colleagues I’ve worked with over the last 40 years.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-6.2.2
Plant and animal response to UV radiation and climate change in the Antarctic
Sharon A. Robinson1, Georgia Watson1
1) University of Wollongong, Securing Antarctica’s Environmental Future & Global Challenges Program,
Wollongong, Australia

Ozone depletion above Antarctica has resulted in increased surface ultraviolet (UV)-B radiation across theregion. Ozone depletion coupled with the heating associated with globally increasing atmosphericgreenhouse gases, has also resulted in pronounced changes in Southern Hemisphere climate1-3,5. Antarctica
is also experiencing extreme events such as the continent wide heatwaves over the summer of 2019/20
summer4. We review how increasing UV radiation over the past four decades, along with changes inSouthern Hemisphere climate have impacted the biodiversity of Antarctica and the Southern Ocean.

Fig. 1 – A satellite image showing melting on the ice cap of Eagle Island, Antarctica, on February 13. NASA EARTH

References
1) Barnes, P. W., Williamson, C. E., Lucas, R. M., Robinson, S. A., Madronich, S., Paul, N. D., Bornman, J.
F., Bais, A. F., Sulzberger, B., Wilson, S. R., Andrady, A. L., McKenzie, R. L., Neale, P. J., Austin, A. T.,
Bernhard, G. H., Solomon, K. R., Neale, R. E., Young, P. J., Norval, M., Rhodes, L. E., Hylander, S., Rose,
K. C., Longstreth, J., Aucamp, P. J., Ballare, C. L., Cory, R. M., Flint, S. D., de Gruijl, F. R., Hader, D. P.,
Heikkila, A. M., Jansen, M. A. K., Pandey, K. K., Robson, T. M., Sinclair, C. A., Wangberg, S. A., Worrest,
R. C., Yazar, S., Young, A. R., & Zepp, R. G. Ozone depletion, ultraviolet radiation, climate change andprospects for a sustainable future. Nature Sustainability 2: 569-579, 20192) Bornman, J. F., Barnes, P. W., Robson, T. M., Robinson, S. A., Jansen, M. A. K., Ballare, C. L., & Flint, S.
D. Linkages between stratospheric ozone, UV radiation and climate change and their implications forterrestrial ecosystems. Photochemical & Photobiological Sciences 18: 681-716. 20193) Robinson, S. A., Erickson, D. J., III. Not just about sunburn - the ozone hole's profound effect on climate hassignificant implications for Southern Hemisphere ecosystems. Global Change Biology 21: 515-527. 2015
4) Robinson, S.A., Klekociuk, A.R., King, D.H., Rojas, M.P., Zúñiga, G.E., Bergstrom D.M. The 2019/2020
summer of Antarctic heatwaves. Global Change Biology 26 3178-3180. 2020
5) Williamson, C. E., Neale, P. J., Hylander, S., Rose, K. C., Figueroa, F. L., Robinson, S. A., Hader, D. P.,
Wangberg, S. A., & Worrest, R. C. The interactive effects of stratospheric ozone depletion, UV radiation,and climate change on aquatic ecosystems. Photochemical & Photobiological Sciences 18: 717-746. 2019
Acknowledgements. This work was supported by grant DP200100223 from the Australian Research Council.

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IL-6.2.3
Biodiversity, climate change and UV radiation
Janet F. Bornman
Food Futures Institute, Murdoch University, Perth, Australia
Climate change, stratospheric ozone depletion, changes in UV radiation, and ecosystem and biodiversity loss are allinterlinking factors affecting life on Earth. Biodiversity of flora and fauna is being adversely affected by changes inclimate, which in turn pose many threats to sustainable livelihoods, including food security and human health.
However, other critical interacting drivers of biodiversity loss include for example, unsustainable agricultural foodsystems, invasive species, pollution, land clearing, and urbanisation. Globally, biodiversity is at serious risk of rapid
decline from the large-scale conversion of land for agriculture and infrastructure. The increase in agricultural land inturn adds to the emissions of greenhouse gases along the food chain, accounting for ca 30% of the total anthropogenic
emissions1 and thereby accelerating climate change. These determinants are already causing shifts in species habitator species extinctions, changes in species composition, and diminishing suitability of habitat, although colonisationof new habitats can also afford advantages2. Furthermore, biodiversity is vulnerable to increased exposure to UV
radiation, although there are currently few studies on the UV radiation-climate effects. Increased exposure to UV
radiation can be due to climate-related reduced cloud cover in some regions and species migrations to higher
elevations as temperatures rise3. Loss of biodiversity through increasing deforestation also exposes dead plantmaterial to solar UV radiation. This results in photodegradation of the plant litter, again contributing to emissions ofcarbon dioxide and other greenhouse gases4,5. Proposed actions for reversing or reducing the negative impacts of the
intricate, indivisible web of interacting drivers on the fate of species biodiversity range from our lifestyle changes totechnological interventions, such as geoengineering for reducing solar radiation to the Earth’s surface. However,geoengineering may have serious consequences for biodiversity, natural ecosystem functioning, human health, andagriculture, due to the expected abrupt changes in environmental conditions that would occur6. UV radiation can beboth reduced and enhanced by aerosol injections into the stratosphere, with the enhancement resulting from multiple
scattering7. Depletion of stratospheric ozone may also occur from solar geoengineering, depending on the aerosol
used8,9, with the increased UV radiation likely resulting in many undesirable consequences for biodiversity andecosystems through changes in environmental photobiology. The role of UV radiation and climate change onterrestrial biodiversity will be discussed in the context of the challenges of maintaining a sustainable Earth.
References
1) Benton, T.G. et al. Food system impacts on biodiversity loss. Three levers for food system transformation in
support of nature. Chatham House, London, pp. 1-71, 2021.
2) Neale, R.E. et al. Environmental effects of stratospheric ozone depletion, UV radiation, and interactions withclimate: UNEP Environmental Effects Assessment Panel, Update 2020. Photochemical & Photobiological
Sciences 20(1), 1-67, 2021.
3) Bornman, J.F. et al. Linkages between stratospheric ozone, UV radiation, and climate change and theirimplications for terrestrial ecosystems, Photochemical & Photobiological Sciences 18(3), 681-716, 2019.
4) Hansen, M.C. et al. High-resolution global maps of 21st-century forest cover change. Science, 342(6160), 850853. 2013.
5) Berenstecher, P. et al. Sunlight doubles above-ground carbon loss in a seasonally dry woodland in Patagonia.
Current Biology 30(16), 3243-3251, 2020.
6) Trisos, C.H. et al. Potentially dangerous consequences for biodiversity of solar geoengineering implementationand termination. Nature Ecology & Evolution 2(3), 475-482, 2018.
7) Madronich, S. et al. Response of surface ultraviolet and visible radiation to stratospheric SO2 injections.
Atmosphere, 9(11), 432, 2018.
8) Fang, X. et al. Challenges for the recovery of the ozone layer. Nature Geoscience, 12(8), 592-596, 2019.
9) Zarnetske, P.L. et al. Potential ecological impacts of climate intervention by reflecting sunlight to cool
Earth. Proceedings of the National Academy of Sciences, 118(15), 2021.

1
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OC-6.2.4

Re-discovering the history for new tools for information about skin cancer
prevention. The shadow projected by an object related to UV index is an universal
photoprotection tool.
José Aguilera Arjona1, María Victoria de Gálvez Aranda1, Enrique Herrera Ceballos1,21) Dermatological Photobiology Laboratory. Bio Medical Research Center. Department of Dermatology.
Unversity of Málaga. Málaga. Spain.
2) Dermatological Unit of the Hospital Clínico Universitario Virgen de la Victoria. Málaga. Spain.
Introduction: Every day better information solutions are offered to the general population to educate on theprevention against overexposure to solar ultraviolet radiation and its negative and positive effects. The informationon the incidence of the UV index is based on specific photodetectors that are not easily accessible to society. Since
the daily evolution of the UV index follows a pattern dependent on the solar arc, and the length of our shadow
varies depending on the solar angle that arc forms with respect to our height, is it possible to relate the shadow
distance with the UV index? Taking a point at earth surface, the solar arc angle varies with the date of the year andwith the latitude. So, the projected shadow of an object will be longer depending on time of the day, day of the year
and the latitude and therefore, the UV index, in conditions of clear sky can be predicted with a very high level ofsensibility.
The main objective of the present work was to find a universal equation relating the UV index with the distance ofthe projected shadow of an object independently of the daytime, date or latitude and secondly to develop a system
to create a rapid and a easy system that with a simple view we can predict UV index related to de distance of the
shadow of an object.
Methods: A shadow length tracking device has been constructed consisting of a rod 100 cm long by 5 cm indiameter located in the centre and perpendicular to a surface that contains a template based on concentric circles
drawn with intervals of 20 cm distance between them. The device was placed on the roof of the Medical ResearchCenter of University of Málaga and the daily variation of the length of the shadow of the rod was followed with a
web camera connected to the computer and next to it a UV index sensor and it images with an interval of 15 min
were followed along a period of 6 months. In order to analyse the mathematical model between shadow distancewith UV index including the effect of latitude, UV index was also correlated to solar zenith angle at different
latitudes from Tenerife to Oslo.
Results: Regardless of the latitude or time of year, an equation has been obtained that relates the projection distanceof a shadow, which is given by the height of the sun throughout the daily cycle and the height of the projection
object of the same. The angle formed by the sun with the shadow is the solar height and the daily cycle formed aparabolic arc while the evolution of the UV index followed a cycle with the shape of a normal curve. It has beenpossible to relate both models of daily evolution, so the date of the year or latitude does not matter. The shadow
distance of the object is related to the UV index following a polynomial model. Therefore, with this equation it has
been possible to construct a universal solar target to give us the UV index value at a given moment only byobserving the shadow distance by using concentric circles around the gnomon with the same colours as the OMS
recommendations for different risk levels of UV index (See Fig.1).

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Conclusion: We are faced with an informative and educational tool in prevention against the harmful effects of thesun, versatile for any location and time of year, at a minimum cost and a potential for universal dissemination for
the entire population.
Acknowledgements. This work was supported by a grant of the II Plan Propio Smart Campus of the University of
Málaga.

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IL-6.2.5

Interactive effects of ocean acidification and UV radiation on aquatic primary
production
Cristina Sobrino1; Patrick J. Neale2; Julia Puig-Fábregas1; Paulo Alcaraz- Rocha1
1) University of Vigo, Department of Ecology and Animal Biology, Vigo, Spain; 2) Smithsonian Environmental
Research Center, Edgewater (MD), USA
Enhanced CO2 in the aquatic environment affects phytoplankton physiology and metabolism. As the source of mostaquatic primary production, phytoplankton make an important contribution to global climate and biogeochemical
cycles. However, many aspects of their physiology and ecology in future global change scenarios are poorly
understood. Here we show that the increase in atmospheric CO2 regulates carbon incorporation mechanisms andpromotes a cascading response in the whole cell that triggers a decrease of cellular energy-consuming pathways.
This capability of marine phytoplankton to adjust their resources and energy to cellular requirements translates into
the optimization of growth rate under high CO2 conditions. Nonetheless, the physiological regulation of cell
metabolism in ocean acidification scenarios affects pigment concentration and cell photoprotection, increasing its
sensitivity to stressing conditions such as UVR. The response of primary productivity to UVR under different CO 2
levels has been measured as Biological Weighting Functions, which allow predicting responses under realistic
conditions. As a general response the results using phytoplankton cultures under controlled conditions in the labshow that phytoplankton acclimated to high CO2 under stable steady-state conditions are more susceptible to stress,
but there is little study of this response in natural populations. More of these types of studies are needed in the
future and we will briefly show how natural environments, such as the Rhode River subestuary (Maryland, USA)or the Ría de Vigo (Northwest Spain), can be used as advanced experimental areas for testing the effects of ocean
acidification on phytoplankton.

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IL-6.2.6
Coral reefs, UV radiation and climate change
Anastazia T. Banaszak1, Sergio D. Guendulain García2
1) Universidad Nacional Autónoma de México, Unidad Académica de Sistemas Arrecifales, Puerto Morelos,
Mexico; 2) Universidad Autónoma Metropolitana, Ciencias Biológicas y de la Salud, Mexico City, Mexico
The key to the success of reef-building corals is attributed to the symbiosis they form with photosynthetic
dinoflagellates and the tight recycling of nutrients and compounds that occurs in an otherwise oligotrophic
environment. The dependence of these organisms on solar radiation signifies exposure to high UVR levels, due tothe low solar zenith angles, natural thinness of the ozone layer and high transparency of the water column
characteristic of tropical, coral reef environments. As such, corals have evolved photoprotective mechanisms that
include the presence of mycosporine-like amino acids to reduce potential UVR damage. However, over recent
decades, coral reefs have been subjected to an ever-increasing number of above average seawater temperature events
attributed to climate change. Thermal stress has provoked bleaching, the breakdown of the symbiotic relationships,
in corals and other symbiotic coral reef dwelling species, and, if the event is prolonged, can lead to widespreadmortality of corals. The photosynthetic dinoflagellates play a crucial role in the photoprotection of corals. DNA
photo-damage, as measured by cyclo-butane pyrimidine dimer (CPD) formation is higher in bleached relative tounbleached specimens. We propose that the breakdown of the symbiosis results in multiple scattering of UVR
wavelengths by the underlying skeleton, which increases the probability of DNA damage within the transparent
tissue. To lower the likelihood that coral reefs undergo repeated bleaching events, stronger steps must be taken to
curb CO2 emissions and reduce the global temperature increase.
Acknowledgements. This work was supported by grant IN2041053 from the Dirección General de Asuntos delPersonal Académico, Universidad Nacional Autónoma de México to ATB.

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OC-6.2.7

Quantifying the Change in Degradation of Plastic Materials Due to Environmental
Exposure and Climate Change
Christopher C White1
1) Exponent, Bowie, MD, USA
Finding plastic waste is not difficult. In the oceans, atmosphere, Arctic, and even in the food we eat, plastic waste isever-present. Recent studies calculate that, on average, 8M tons of plastic waste enter the oceans every year.1
Understanding the dynamic accumulation of plastic waste in the environment is a complex scientific undertaking,perhaps only superseded by the challenge of developing strategies for its removal. This presentation will focus onunderstanding the reduction of plastic waste using tools and techniques from polymer chemistry and the economics
of quantifying the disposal of this waste.
Recent studies indicate that ~250MT to ~300MT of plastic waste is generated every year.2 This plastic waste is
primarily discarded with only small quantities of it either recycled (<10%) or incinerated (<10%). The persistenceor lifetime of these plastics significantly contributes to the overall build-up of plastic waste. Knowledge of thedegradation rate is critical in managing plastic waste, yet these predictions remain imperfect science. Understanding
the limitations of estimating the degradation of plastics is the core of this contribution.
Plastic waste is often considered a singular entity, but plastics and plastic waste are a complex assortment of widely
varying chemistries, formulations, assemblies, and size distributions. Each component contributes to the overall
plastic waste problem and requires a unique determination of its specific degradation rate. The majority of plastic
waste is composed of purely synthetic polymers.
There are two primary mechanisms of degradation of plastic waste: biological or chemical degradation. Biologicaldegradation of purely synthetic polymers is a prolonged process.3 The rate of chemical degradation is a function ofboth the composition of the plastic formulation and the environmental exposure, thus increasing the difficulty inquantifying the decomposition rate.
Determining the degradation rate ofplastics and polymers is primarily
driven by estimating the expectations
for the design life and associatedwarranty guarantees in the intended
serviceenvironment.
The
corresponding standards and test
methods align with two strategies:
timed outdoor exposure in a
representative environment or alaboratory-derived protocol to assess
relative performance.
The timed outdoor exposure ofplastic materials is performed in a
representative environment that isjudged to produce faster degradation
Figure 1 Plastic Waste Litters the Landscape.
than typical in-service environments.
Arizona, Florida, Australia, or
another similar global location is selected for the extreme environmental conditions such as some combination ofhot, humid, or bright Ultra Violet (UV) radiation. Devices employing mirrors increase the UV radiation andBook of Abstracts ����������������������������������������������������������������������������������������������� 270

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temperature to accelerate the degradation process in these locations. Typically, the samples are exposed on racks
tilted to maximize the solar exposure and monitored over at least several years. These exposures benchmark
durability for the formulation tested but did not estimate the degradation in other environments.
The second type of durability test simplifies and accelerates specific aspects of the outdoor environment with
exposure instruments that produce UV, temperature, and moisture exposures. These tests can effectively compare
formulations in the performance in these tests, but correlation to actual outdoor performance has remained elusive.
These tests establish baseline performance expectations for the formulations and allow for monitoring of the relative
durability.
The impetus to develop test methods that can predict degradation historically has been to reduce the barrier tointroducing new innovative materials. The attributes of a new formulation can all be determined immediately, except
for durability. Establishing the durability with existing test methods yields a relative performance using laboratory
methods in a matter of months that has a limited correlation to actual design life or a threshold performance in an
exemplar environment after years of exposure. Either approach significantly increases expense and time related to
new product introduction while not substantially reducing the liability risk.
Increasing interest in sustainability may spur innovation in these test methods. Scope 3 4 reporting requirements
require assessing the environmental fate of sold plastic products, including the costs associated with greenhouse gas
emissions from the end-of-life treatment of sold products. These assessments become an expense or liability on thecompany's regular financial disclosures. Knowledge of the degradation rate and then taking steps to increase thatrate will reduce the expense or liability for the company. This financial pressure will increase the innovation in thetest methods used to determine the degradation rate of plastic waste.

References

1.

2.
3.
4.

Law, K. L.; Starr, N.; Siegler, T. R.; Jambeck, J. R.; Mallos, N. J.; Leonard, G. H., The UnitedStates' contribution of plastic waste to land and ocean. Science Advances 2020, 6 (44).
Geyer, R.; Jambeck, J. R.; Law, K. L., Production, use, and fate of all plastics ever made. Science
Advances 2017, 3 (7), e1700782.
Shah, A. A.; FarihaHasan; AbdulHameed; SafiaAhmed, Biological degradation of plastics: A
comprehensive review. Biotechnology Advances 2008, 26 (3), 246-265.
https://www.epa.gov/climateleadership/scope-3-inventory-guidance.

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OC-6.2.8

PHOTOPHYSIOLOGICAL RESPONSE OF MARINE PHYTOPLANKTONUNDER OCEAN ACIDIFICATION CONDITIONS
Alcaraz-Rocha P.1, Puig-Fàbregas J.1, Garrido J.L.2 & Sobrino C.1
1) Grupo de Oceanografía Biolóxica, Centro de Investigación Mariña (CIM), Universidade de Vigo, Vigo, Spain;
2) Grupo de Fotobiología y Toxinología de Fitoplancton, Instituto de Investigaciones Marinas (IIM), Consejo
Superior de Investigaciones Científicas (CSIC), Vigo, Spain
ABSTRACT
Ocean acidification, due to the atmospheric CO2 increase, produces significant changes on phytoplankton
physiology that can affect the photosynthetic metabolism. Among them, a down-regulation of the enzymatic
activity and the production of several metabolites, including Chl a and RuBisCO, have been observed in high CO2
cultures under acclimated conditions. However, the extent of how phytoplankton photophysiology regulates under
ocean acidification conditions is poorly understood. This study shows the effect of the atmospheric CO2 increase on
pigment concentration of 3 important marine primary producers: Thalassiosira pseudonana, Skeletonema costatum
and Emiliania huxleyi. Cultures grown under saturating PAR irradiance were aerated during at least 3 weeks withcurrent concentrations of atmospheric CO2 (~400 ppmv CO2) and with CO2 concentrations expected for futurescenarios of climate change (~1000 ppmv CO2) to assess the effect of CO2 under stable conditions and acclimatedmetabolism. Moreover, cultures were also subjected to a CO2 perturbation (4 h without aeration) to assess
responses under non-stable conditions. The results showed that both, light harvesting and photoprotective pigment
concentrations (i.e. Chl a, Chl c2, β-carotene, Diadinoxanthin, Diatoxanthin, Fucoxanthin and its derivatives, among
others) decreased significantly under high CO2 and acclimated conditions, but the response reversed when themetabolism was perturbed. The results also demonstrate that the CO2 concentration can modify the photoprotective capability of the phytoplankton through changes in the photoprotective pigment content and in the
deepoxidation state of the xanthophylls. They also identify Fucoxanthin and Chl c2 as potential biomarkers ofphytoplankton under ocean acidification conditions.
Key words: Ocean acidification, CO2 acclimation, phytoplankton, pigments, photoprotection

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SP-6.2.9, P-6.2.9

EFFECTS OF OCEAN ACIDIFICATION IN PHOTOPHYSIOLOGY OF MARINEBACTERIA FROM TWO DIFFERENT MARINE HABITATS
Alcaraz-Rocha P.1, Puig-Fàbregas J.1, Garrido J.L.2 & Sobrino C.1
1) Grupo de Oceanografía Biolóxica, Centro de Investigación Mariña (CIM), Universidade de Vigo, Vigo, Spain;
2) Grupo de Fotobiología y Toxinología de Fitoplancton, Instituto de Investigaciones Marinas (IIM), Consejo
Superior de Investigaciones Científicas (CSIC), Vigo, Spain
ABSTRACTPrevious studies have shown that high CO2 levels expected for future scenarios of global change might result in
phytoplankton with higher sensitivity to photoinhibitory conditions than those exposed to present CO2 levels. This
response, mainly observed in diatoms, has been attributed to a decrease in the repair rates caused by the lower
levels of enzymatic activity and cellular pools observed, but could also be attributed to greater damage related todecreased protection. In this study, we will focus on the effects of elevated CO2 on pigment content on marine
bacteria. The CO2 effect will be tested on a globally distributed cyanobacteria species, Synechococcus spp., isolated
from two different marine habitats: the strain WH7803, typical from coastal areas, is observed during transition
periods between mixing and stratification, while the strain WH8102 is adapted to oligotrophic environments andappears in open-ocean waters. The cultures were acclimated during at least 3 weeks with current concentrations ofatmospheric CO2 (~400 ppmv CO2) and with CO2 concentrations expected for future scenarios of climate change(~1000 ppmv CO2) to assess the effect of CO2 under stable conditions and acclimated metabolism. Moreover,cultures were also subjected to a CO2 perturbation (4 h without aeration) to assess responses under non-stableconditions. HPLC results showed that in Synechococcus spp. the Zeaxanthin is the predominant pigment, which
relates to cell protection. Chlorophyll a and β-carotene concentrations were lower than Zeaxanthin concentration inboth strains. However, Zeaxanthin concentration increased significantly under high CO2 conditions in the coastal
strain after a perturbation but did not change in the open-ocean strain under similar conditions. The results from thecoastal strain were more similar to those observed in coastal diatoms, which might imply that differences infunctional traits between phytoplankton assemblages from open-ocean vs. coastal waters are more significant thantheir taxonomic features regarding responses to ocean acidification.
Key words: Ocean acidification, CO2 acclimation, marine bacteria, pigments, photoprotection

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IL-6.3.3

New Development in Sunscreens
Henry W. Lim
Photobiology and Photomedicine Unit, Department of Dermatology, Henry Ford Health System, Detroit,
Michigan, USA
Topics that are actively discussed and developing in sunscreens will be presented:
1. Percutaneous penetration of active ingredients of sunscreens. This has caused significant confusion and concerns
among the general public. It should be noted that the authors of the two studies clearly indicated that the clinicalsignificance is unclear, and that further studies need to be done (1,2).
2. Environmental impact of UV filters. The concentrations reported to be toxic to coral reefs in laboratory settings
were 1000 to one million-folds higher that the concentrations measured in sea water, and there are many otherfactors that contribute to bleaching of coral reefs, including warming and acidification of the ocean water (3).
3. Protection against the photobiologic effects of visible light (VL). VL is known to induce persistent and intense
pigmentation in dark-skinned subjects, contributing to the development of melasma and post-inflammatory
pigmentation. Except for pigmentary-grade mineral filters (which result in unacceptably whitish discoloration on
the skin), the rest of currently available UV filters do not protect against VL Tinted sunscreens containing ironoxides and pigmentary titanium dioxide, and novel filters currently under various stages of development, offer VL
protection (4).
References
1) Matta MK, Zusterzeel R, et al. Effect of Sunscreen Application Under Maximal Use Conditions on Plasma
Concentration of Sunscreen Active Ingredients: A Randomized Clinical Trial. JAMA. 2019 Jun 4;321(21):20822091..PMID: 31058986
2) Matta MK, Florian J, et al. Effect of Sunscreen Application on Plasma Concentration of Sunscreen Active
Ingredients: A Randomized Clinical Trial. JAMA. 2020 Jan 21;323(3):256-267. PMID: 31961417
3) Mitchelmore CL, He K, et al. Occurrence and distribution of UV-filters and other anthropogenic contaminants incoastal surface water, sediment, and coral tissue from Hawaii. Sci Total Environ. 2019 Jun 20;670:398-410. PMID:
30904653
4) Narla S, Kohli I, et al. Visible light in photodermatology. Photochem Photobiol Sci. 2020 Jan 22;19(1):99-104.
PMID: 31922171

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria, D. Douki

19th Congress of the European Society
for Photobiology
OC-6.3.4

A new sun care product containing Phenylene Bis-Diphenyltriazine (TriAsorB)
provides full-spectrum photoprotection against sunlight radiation induced skindamageD. Bacqueville1, C. Jacques-Jamin1, N. Roullet1, J. Sereno1, S. Delassan1, P. Lapalud1, T. Douki2, D. Redoulès3, S.
Bessou-Touya1 and H. Duplan1
1) Laboratoire Pierre Fabre, Centre R&D, France, Toulouse; 2) INAC/SyMMES/CIBEST, CEA Grenoble, France;3) Laboratoires Dermatologiques Avène, Lavaur, France
Photoprotection is a major issue in public health to prevent the harmful effects of sunlight radiations (actinic
keratosis, skin cancers and photoaging). Recently, we have developed a new generation of non-soluble organic
sunfilter called Phenylene Bis-Diphenyltriazine or TriAsorB (CAS N°55514-22-2). It absorbs/reflects the solar
radiations mainly in the ultraviolet UVB and UVA spectral range but also in the visible/infrared light (VIS/IR),
especially in the high energy visible (HEV)/Blue light. TriAsorB protects the skin against the genotoxicity of solarsimulated radiation (SSR), suggesting that it could be used in combination with specific sunfilters in sun care products
to provide photoprotection in the entire spectrum of sunlight (ESP congress Barcelona 2019).
Thus, the aim of this work was to formulate the TriAsorB in a new photoprotective system in association with 3
sunfilters (Bis-Ethylhexyloxyphenol Methoxyphenyl Triazine, Diethylamino Hydroxybenzoyl Hexyl Benzoate,
Ethylhexyl Triazone) and to explore the photoprotective efficacy of this innovative SPF50 + sun care products toprevent skin photodamage from UV to VIS/IR light including HEV/Blue light.
Genoprotection was first assessed in vitro by using a reconstructed human epidermis (RHE) model exposed to asingle acute dose of SSR (UV+ VIS light, UV total 16.5 J/cm2). Liquid chromatography/mass spectrometry analysis
revealed that SSR generates cyclobutane pyrimidine dimers (CPD) and that the topical application of the TriAsorB
sunscreen at a standard dose of 2 mg/cm2 before SSR exposure prevented 95.2% DNA lesions. The efficacy of thesunscreen was maintained in extreme UV condition up to 55 J/cm2 with 89.9% DNA photoprotection. DNA damage
was also correlated to tissue toxicity and apoptosis induction 24h after SSR irradiation as assayed by sunburn cell
histology and CPD/active caspase-3 confocal microscopy in an ex vivo human skin model. TriAsorB sun care productprovided an almost complete photoprotection in response to SSR. Interestingly, it was also effective to limit themalondialdehyde (MDA, lipid peroxidation) production and superoxide dismutase activity (SOD, anti-oxidative
defence) in the RHE model (39.8% and 30.7% protection respectively). Immunostaining of 8-hydroxy-2’deoxyguanosine (8OHdG), a biomarker of DNA oxidation, confirmed genome photoprotection in human skin after
SSR exposure. Therefore, sunscreen prevented SSR-induced oxidative stress in skin models.
TriAsorB sun care product efficacy was then studied beyond UV in both the VIS and IR spectral range of the sunlight.
Since TriAsorB has unique absorption/reflection properties in the HEV/Blue light, a RHE model was exposed to a
LED 427 nm at 80 J/cm2 and 8OHdG staining was performed. HEV light induced tissue oxidation 24h after exposure
and TriAsorB sunscreen protected the skin against the 8OHdG oxidative lesions induced by HEV light with 95.3%
efficacy. Thus, the sun care product provided photoprotection in the VIS light.
Finally, a clinical study was done to analyze IR-induced oxidative stress and photoprotection. The forearm of 18
healthy volunteers was exposed to an IR lamp (750 to 3000 nm) at 900 J/cm2 and the sunscreen was applied at 2
mg/cm2 before irradiation. Skin were harvested by using swabs and MDA was quantified 1h30 after IR exposure.
MDA was detected in non-irradiated forearm and IR induced an increase of MDA production up to 82 ng/mg total
protein. The TriAsorB sunscreen completely protected the forearm of the volunteers with 120% efficacy against IR
radiation-induced lipid peroxidation.
Altogether, an innovative photoprotective system combining TriAsorB with 3 sunfilters provided full-spectrum
photoprotection from UV to VIS/IR spectral range of the solar radiation. The new sun care product protected the skinagainst sunlight radiation-induced skin damage including genotoxicity and oxidative stress.

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IL-6.3.5
Photoprotection for persons with albinims
Yolanda Gilaberte11) Miguel Servet University Hospital, Department of Dermatology, IIS Aragón, Zaragoza, Spain.

Introduction: Skin cancer is the main cause of death in persons with albinism (PWA) in Africa.
Education would minimize sun damage and the use of sunscreens specifically designed for theirnecessities would help to reduce the incidence of skin cancer in the future.
Objective: to evaluate the efficacy and acceptance of a photoprotection educative and sunscreen (Umozi
Max) package specifically designed for PWA in reducing sunburns and skin cancer.
Patients and Methods: A multicenter, non-controlled, before-and-after, interventional was conducted in
PWA > 12 years of age, from February to May 2019, in Malawi. During the baseline and the follow-up
visits (at 8 and 15 weeks) subjects received an educational program designed to PWA and use of Umozi
Max. This sunscreen is an O/W emulsion that turn into W/O during application, with and SPF 50.6,
UVA-PF 19.6 and critical wavelength 380.3 nm. The filters used do not penetrate through the skin andare safe for the environment. At every visit, photoprotection behaviour and knowledge were checked andcutaneous lesions were recorded. Univariate and bivariate analysis were performed.
Results: 210 PWA were analysed, 50% males, with a mean age of 24.5 (SD 11.29) years. The percentageof people using sun protective clothing increased from 80% to 100% and sunscreen from 81.9% to99.5%. People avoiding the midday sun increased by 38.9% (p<0.05). Participants that erroneously
applied the sunscreen at night diminished from 40% to 4% (p<0.001). Absent erythema on the face
increased from 40% to 90% (p<0.05). The percentage of patients with actinic keratoses (AK) on all
locations significantly decreased during the study. All the participants preferred Umozi Max to previously
used sunscreens. The satisfaction with the program was unanimous.
Conclusion: the educational program enhanced the use of all photoprotection measures, improved
behaviours and decreased the incidence of solar erythema and contributed to decreasing the incidence of
new AKs. Umozi Max was effective in terms of prevention of sunburn and also very well accepted by theparticipants.

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IL-6.3.6

Visible Light Photoprotection
Indermeet Kohli,1,2 Iltefat H. Hamzavi,1 Henry W. Lim1
1) Henry Ford Health System, Photomedicine and Photobiology Unit, Department of Dermatology, Detroit, USA;
2) Wayne State University, Department of Physics and Astronomy, Detroit, USA
The impact of ultraviolet (UV) radiation on skin and associated photodamage have been studied extensively overthe past several years. Recent studies have demonstrated the ability of other parts of sunlight, particularly visible
light (VL), in inducing biologic effects. VL has been shown to induce erythema and pigmentation amongindividuals with dark skin.1, 2 The responses were shown to be more exuberant and persistent when combined withlong-wavelength UVA1 (VL+UVA1) demonstrating synergistic effects.3 The combination, VL+UVA1, has also
been shown to induce erythema in light-skinned population.4 The presentation will review the impact of VL on skinhealth, VL phototesting methodologies and currently available means for VL photoprotection.
References
1. Mahmoud BH, Ruvolo E, Hexsel CL, Liu Y, Owen MR, Kollias N et al. Impact of long-wavelength UVA andvisible light on melanocompetent skin. The Journal of investigative dermatology 2010;130:2092-7.
2. Duteil L, Cardot-Leccia N, Queille-Roussel C, Maubert Y, Harmelin Y, Boukari F et al. Differences in visible
light-induced pigmentation according to wavelengths: a clinical and histological study in comparison with UVB
exposure. Pigment Cell Melanoma Res 2014;27:822-6.
3. Kohli I, Chaowattanapanit S, Mohammad TF, Nicholson CL, Fatima S, Jacobsen G et al. Synergistic effects oflong-wavelength ultraviolet A1 and visible light on pigmentation and erythema. The British journal of dermatology
2018;178:1173-80.
4. Kohli I, Zubair R, Lyons AB, Nahhas AF, Braunberger TL, Mokhtari M et al. Impact of Long-Wavelength
Ultraviolet A1 and Visible Light on Light-Skinned Individuals. Photochemistry and photobiology 2019;95:1285-7.
Acknowledgements. The fundamental VL work was internally funded by the Photomedicine and Photobiology Unit
at Henry Ford Health System, Department of Dermatology. Results from studies supported by Ferndale Laboratories,
La Roche-Posay Laboratoire Dermatologique, and Beiersdorf (previously known as Bayer) will also be presented.

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IL-6.3.7

UVA1 harmful effects. Benefit of an enlarged photoprotection efficiently covering thewhole UV spectrum.
Claire Marionnet 1, Romain de Dormael 1, Xavier Marat 1, Angélina Roudot 2, Emilie Planel 1, Carine Tornier 3,
Philippe Bastien 1, Didier Candau 2 and Françoise Bernerd 1.
1) L’Oréal Research and Innovation, Aulnay sous Bois, France; 2) L’Oréal Research and Innovation, Chevilly
Larue, France; 3) Episkin, Lyon, France.
UVA1 rays (340–400 nm) account for at least 75% of ultraviolet wavelengths reaching the Earth. They are able toinduce epidermal and dermal damage, and alteration of gene and protein expression involved in diverse biologicalpathways such as immunity, development, oxidative stress response, inflammation or dermal matrix organization1.
Clinically, they contribute to immunosuppression, carcinogenesis, and photoaging 2,3,4.
Today, state of the art sunscreen formulas can efficiently filter UV wavelengths up to 370/380 nm, but lack sufficientabsorption in the 370/380–400 nm wavelengths range. Recently, a new cyclic merocyanine UVA1 absorber,
Methoxypropylamino Cyclohexenylidene Ethoxyethylcyanoacetate (MCE), exhibiting a maximal peak of absorption
at 385 nm was approved by the SCSS for use in sunscreen products.
The studies presented here aimed at evaluating, in vitro and in vivo, the protection afforded by state of the art
sunscreen formulations enriched with MCE.
In vitro, UV absorption spectra of formulations were determined and showed that MCE addition in a state of the art
reference formula, enlarged the profile of absorption up to 400 nm, resulting in a full coverage of the whole UV
spectrum. Then, formulations efficacy was evaluated through morphological, biochemical and gene expression
analysis in 3D skin model exposed to UVA1. Compared to the reference sunscreen, formulations containing MCE
enabled significantly higher protection of the epidermis and the dermis, with a better protection against dermal
fibroblasts alterations, and a significantly higher reduction of gene and protein expression modulation. In an in vivo
randomized controlled trial, pigmentation was used as a clinical surrogate for impact of UVA1 exposure using
colorimetric measurements (L*, a*, b*, ΔE and ΔITA) and visual scoring5, in 19 volunteers with skin types III/IV.
UVA1-induced pigmentation was significantly reduced by the formulations containing MCE compared to the stateof art formulation. This superiority of protection with formulations containing MCE compared to that of references
sunscreen was also proven in a second clinical trial on 25 volunteers (skin types III/IV), following repeated exposures
to realistic daily UV radiation (UVB+UVA).
In conclusion, thanks to MCE filtering properties, a full coverage of the whole UV spectrum up to 400 nm wasreached, leading to a higher photoprotection against UV-induced biological and clinical impacts. The data alsostrongly supports benefits regarding long-term sun-induced harmful effects.
References
1)Marionnet C, Pierrard C, Golebiewski C, Bernerd F. Diversity of biological effects induced by longwave UVA
rays (UVA1) in reconstructed skin. PLoS One 9(8):e105263, 2014.
2)Damian DL, Matthews YJ, Phan TA, Halliday GM. An action spectrum for ultraviolet radiation-induced
immunosuppression in humans. Br J Dermatol 164(3):657-9, 2011.
3)Tewari A, Grage MM, Harrison GI, Sarkany R, Young AR. UVA1 is skin deep: molecular and clinicalimplications. Photochemical & Photobiological Sciences;12(1):95-103, 2013.
4)Wang F, Smith NR, Tran BA, Kang S, Voorhees JJ, Fisher GJ. Dermal damage promoted by repeated low-level
UV-A1 exposure despite tanning response in human skin. JAMA Dermatology;150(4):401-6, 2014.
5)Marionnet C, Nouveau S, Hourblin V, Pillai K, Manco M, Bastien P, et al. UVA1-induced skin darkening is
associated with molecular changes even in highly pigmented skin individuals. J Invest Dermatol;137(5):1184-7,
2017.
Acknowledgements. We warmly thank Julie Gizard and Christelle Golebiewski for their expert technical assistance.

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OC-6.3.8

Evaluation of the biological effect of a high broad spectrum sunscreen with
nicotinamide and panthenol repairing photodamaged skin.
Gemma Tell1, Neus Calbet1, Judit Mateu1, Javiera Pérez2, Bea Alejo2, Pablo Iglesias2, Josep Malvehy1,2, Ann’Laure
Demessant-Flavigny3, Caroline Le Floc’h3, Leonor Prieto4, Susana Puig1,2
1. Centro de Investigación Biomédica en Red de Enfermedades Raras, CIBERER, Instituto de Salud Carlos III,
Barcelona, Spain, 2. Dermatology Department, Melanoma Unit, Hospital Clínic de Barcelona, IDIBAPS, Universitat
de Barcelona, Barcelona, Spain, 3. International Scientific Direction, Laboratoire Dermatologique La Roche-Posay,
France, 4. Scientific Direction, Laboratoire Dermatologique La Roche-Posay, L’ORÉAL Cosmética Activa, Madrid,
Spain
Nicotinamide is a precursor of nicotinamide adenine dinucleotide (NAD), co-enzyme essential in the production of
adenosine triphosphate (ATP), main source of cellular energy. Previous studies in mice showed that the oral or topicalapplication of nicotinamide prevents immunosuppression and reduce the number of UV-induced tumors. In humans,
topical application of nicotinamide prevents UV induced immunosuppression, but not sunburn. Nicotinamide
increases the production of ATP, increases DNA repair and prevents/avoids apoptosis.
The main objective of the present study was to determine the biological effect of a high broad spectrum UVB-UVA
sunscreen containing Nicotinamide and Panthenol, in photodamaged skin analyzing, immunohistochemistry (p53,
PCNA, p21 and pyrimidine dimers) and RNA profiles using high throughput technology.
Fifteen healthy patients (> 40 years) were included. Two areas in photodamaged skin in the dorsum of the forearm(area 1 and 2) and one area in the inner part of the forearm less photodamaged (area 3) were identified, imaged withlineal confocal OCT and biopsies performed in area 1 and 3. During four weeks, a high broad UVB-UVA sunscreenwith nicotinamide and panthenol was applied daily. After that, a new biopsy was performed in area 2 after imaging
with lineal confocal OCT. Each biopsy was split into two halves. One was fixed in formalin for ulterior be embedded
in paraffin and immunostaining with p21, p53, PCNA and CPD, and the other one was included in RNAlater andfrozen to extract RNA subsequently and carried out the sequencing.
From the 14 patients included that ended the study only 13 were assessable for the immunohistochemistry comparison(in one patient the post treatment biopsy was not evaluable because of the absence of epidermis in the sample).
100% of cases showed an increased expression of p21, PCNA, p53 and CPD in photodamaged skin (L01) whencompared with less damaged skin (L03). After 4 weeks of product application, we identified a slightly decrease of
p21 expression in treated photodamaged skin compared to non-treated photodamaged skin (L02 vs L01), but thistendency did not reach statistical significance. Direct comparison pre-post treatment showed improvement of p21
expression in 6 cases (1, 6, 7, 8, 13, 14). Differential gene expression analysis (DGEA) from the RNA-Seq data
showed an overexpression of Collagen Type I Alpha 1 Chain (COL1A1) gene (adjusted p-value < 0.001) in treatedphotodamaged skin in comparison with non-treated photodamaged skin (L02 vs L01 biopsies). Furthermore, thegene set enrichment analyses (GSEA), which took into account the fold-change value for each gene in order to rank
them, identified 40 biological pathways significantly dysregulated after treatment (adjusted p-value < 0.001). Amongthe 20 upregulated pathways, there are some related with the SUMOylation process, extracellular matrixorganization, cell adhesion molecule activity and keratinization. Otherwise, among the 20 downregulated pathways,
several are involved in the reactive oxygen species generation and mitochondrial function.
Four weeks of daily application of high broad spectrum UVB-UVA sunscreen with nicotinamide and panthenol inphotodamaged skin induces overexpression of COL1A1 and influences the regulation of 40 biological pathways
involved in the skin homeostasis.

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P-6.3.9
Plants growing in Colombia as sources of natural actives sunscreen ingredients
Diego Armando Villamizar Mantilla1, Silvia Juliana Flores González1, Luis Alberto Núñez1, Elena E. Stashenko1,
and Jorge Luis Fuentes Lorenzo1.
1) Universidad Industrial de Santander, Santander, Bucaramanga, Colombia.
Introduction: Plants can be sources of photoprotective/antigenotoxic compounds that prevent cellular mutations
involved in skin cancer and aging by regulating UV-induced mutability. Purpose: The study was aimed atinvestigating the sunscreen properties of plants growing in Colombia. Materials and Methods: Ultraviolet radiation(UVR)-absorption capability of different plant extracts was examined. In vitro photoprotection efficacies were
evaluated using in vitro indices such as sun protection factor (SPF in vitro) and critical wavelength (c). Pearson
correlation analysis was used to examine the relationship between SPF in vitro and complementary UVBantigenotoxicity estimates (%GI) based on SOS Chromotest database. The cytotoxicity in human fibroblasts wasstudied using the trypan blue exclusion assay. Major compounds of promising plant extracts were determined by gas
chromatography coupled to mass spectrometry (GC/MS). Results: We showed that plant extracts have sunscreen
properties against UVB, whereas broad-spectrum radiation protection efficacy was poor. SPFin vitro and %GI werecorrelated (R = 0.71, p  0.0001) for the plant extracts under study. Three extracts obtained from Achyrocline
satureioides, Chromolaena pellia, and Lippia origanoides species resulted to possess high protection efficacy andrelatively low cytotoxicity in human fibroblasts. These plant extracts had major compounds such as -pinene, transβ-caryophyllene, -muurolene, -cadinene and caryophyllene oxide in A. Satureioides extract, squalene and -amyrin
in C. pellia extract, and p-cymene, carvacrol, trans-β-caryophyllene and pinocembrin in L. origanoides extract.
Conclusions: Plants growing in Colombia contain compounds which ca be useful as a sunscreen. SPFin vitro and %GI
estimates were correlated, but %GI estimates were more sensitive to detecting activity at lower plan extract
concentrations. Our results supported the need to use DNA damage detection assays as a complement tophotoprotection efficacy measurement.
Acknowledgements. The authors thank funding from the Ministry of Science, Technology and Innovation,
the Ministry of Education, the Ministry of Industry, Commerce and Tourism, and
ICETEX, Programme Ecosistema Científico-Colombia Científica, from the Francisco José de Caldas Fund,
Grant RC-FP44842-212-2018. The Ministry of Environment and Sustainable Development of Colombia
supported the Universidad Industrial de Santander through access permits to genetic resources andderivatives for bioprospecting (Contract N o. 270).

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P-6.3.10

Scoring Levels To Corroborate The Booster Effect Of A Natural Extract Of
Polypodium Leucotomos (Fernblock®) In Topical Sunscreens In Improving The UV
Barrier Activity And Immune Protection Capability
Azahara Rodríguez-Luna1, María Villalba2, María Victoria de Gálvez3, Enrique Herrera-Ceballos3, Salvador
González4, José Aguilera3.
1) Cantabria Labs, Innovation and Development Dept, Madrid, Spain.
2) Cantabria Labs, Medical Affairs Dept, Madrid, Spain.
3) University of Malaga, Photobiological Dermatology Laboratory, Medical Research Centre, Málaga, Spain.
4) Alcalá de Henares University, Dept of Medicine and Medical Specialties, Madrid, Spain.
Different in vitro and in vivo techniques have been used to determine the photoimmunoprotection potential ofsunscreens. Nevertheless, the diversity of experimental designs, data management and different forms of assessment
make the identification of a suitable reference method difficult. The aim of the study is to complete the existingscientific evidence of Fernblock® about its photoimmunoprotection potential factor evaluating its physicochemical
activity. Thus, to validate the physiochemical level, we evaluated by means of UV solar simulated transmission
measurements the immunoprotection factor (IPF) of different sunscreens based on Fernblock®, an aqueous extractfrom Polypodium leucotomos. Four different sunscreen formulations containing Fernblock® were selected and theirtransmittance of solar simulated UV was calculated by spectroradiometric measurement. In vitro SPF, UVAPF and
IPF based on the action spectrum for systemic contact hypersensitivity (CHS-IPF) were measured.
Fernblock® added to different combinations of organic and mineral sunscreens has a booster effect with a mean
increase of SPF, CHS and HIF factor over 10 arbitrary units and more than 10 % of average boost of all factors. In
addition to these physiochemical results, Fernblock has demonstrated extensive biological immunoprotection
capabilities in vitro and in vivo (1-4). Thus, we elucidate a scoring system to evaluate the photoimmunoprotection
potential consisting in three study levels of evidence for any sunscreen to validate the immunoprotection factor (IPF)a) the physicochemical level protection, b) the biological in vitro level protection and c) the validation by clinicalevidence. In summary, new generations of topical sunscreens need to promote additional effects, not only with filters,
but with compounds that promote both regeneration and/or immunoprotection activities, among others. Taken
together with the evidence displayed, new scoring systems should be implemented beyond SPF. Different studies
should be considered to evaluate the IPF including not only barrier/photon blocking function but also
immunoprotective biological activities.
References:
1. Capote R, Alonso-Lebrero JL, et al. Polypodium leucotomos extract inhibits trans-urocanic acidphotoisomerization and photodecomposition. J Photochem Photobiol B. 2006.
2. Middelkamp-Hup MA, Pathak MA et al. Orally administered Polypodium leucotomos extract decreases
psoralen-UVA-induced phototoxicity, pigmentation, and damage of human skin. J Am Acad Dermatol. 2004.
3. Middelkamp-Hup MA, Pathak MA, Parrado C, Goukassian D, Rius-Díaz F, Mihm MC, Fitzpatrick TB,
González S. Oral Polypodium leucotomos extract decreases ultraviolet-induced damage of human skin. J
Am Acad Dermatol. 2004.
4. Schalka S, Donato LC. Evaluation of effectiveness of a sunscreen containing Polypodium leucotomos extract
in reducing the sun damage to the skin. Surg Cosmet Dermatol. 2019
Acknowledgements. This work was supported by Cantabria LabsBook of Abstracts ������������������������������������������������������������������������������������������������ 281

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P-6.3.11

Photoprotective and Antigenotoxic Properties of Serratia marcescens Indigenous
Strains from Eastern Cordillera of Colombia
José Duban Daniel Cediel Becerra1, Jhon Alexander Suescún Sepúlveda1, Jorge Luis Fuentes Lorenzo1
1) Universidad Industrial de Santander, Santander, Bucaramanga, Colombia
Introduction: Serratia marcescens is a bacterial species that produces antibacterial pigment (prodigiosin) that showsa wide adaptive response to environmental stresses. This pigment has a strong UV– absorption capability, suggestingthat it can be useful as a sunscreen. Purpose: The study was aimed at investigating photoprotective and antigenotoxic
properties against UVB of forty extracts from S. marcescens strains collected across an altitude gradient in the Eastern
Cordillera of Colombia. Materials and Methods: Prodigiosin production was spectrophotometrically assayed in
extracts of bacterial strains grown in different culture media and temperatures. A calibration curve of pure prodigiosin
was used to estimate the compound concentration in each extract. In vitro photoprotection efficacy was evaluatedusing in vitro indices such as sun protection factor (SPFin vitro) and critical wavelength (λc). The percentage of UVB–
antigenotoxicity estimates (%GI) in the SOS Chromotest was also evaluated. Correlation analysis was used to
examine the relationship of the SPFin vitro with prodigiosin concentration, %GI, and environmental markers (altitude,
temperature, rainfall, and solar irradiance). Results: Salt minimum culture medium supplemented with glycerol anda growing temperature of 28 °C showed higher prodigiosin production. Among the strains under study, six showed
medium (15.0 ≤ SPFin vitro ≤ 29.9) and two high (30.0 ≤ SPFin vitro ≤ 59.9) photoprotection efficacy against UVB
radiation, while only one strain resulted in broad-spectrum (UVA–UVB) photoprotection (λc > 370 nm). SPFin vitro
was well correlated with prodigiosin concentration (R = 0.79, p ≤ 0.05) and with %GI values (R = 0.80, p ≤ 0.05) inthe bacterial strains being studied, but no to environmental markers (R = 0.0 – 0.16, p ≥ 0.05, n.s). Conclusion: We
demonstrated that S. marcescens strains producer of prodigiosin could be useful as source of sunscreen. Extracts
from S. marcescens strains with prodigiosin concentration higher than 17 g/mL showed good quality as sunscreen(SPFin vitro  15.0) that reduced UVB– induced genotoxicity in the SOS Chromotest. UVB photoprotection efficacy
depended on prodigiosin concentration and on %GI values of the S. marcescens extracts, but not on the environmental
markers.
Acknowledgements. The authors thank funding from the Ministry of Science, Technology and Innovation, theMinistry of Education, the Ministry of Industry, Commerce and Tourism, and ICETEX, Programme Ecosistema
Científico-Colombia Científica, from the Francisco José de Caldas Fund, Grant RC-FP44842-212-2018. The
Ministry of Environment and Sustainable Development of Colombia supported the Universidad Industrial deSantander through access permits to genetic resources and derivatives for bioprospecting (Contract No. 270).

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P-6.3.12

The hydroalcoholic extracts of Ipomoea horsfalliae, Posoqueria latifolia and Rosa x
centifolia contain compounds with filter and antigenotoxic effects against UV radiation
Jorge Luis Fuentes Lorenzo1, Carlos Adolfo Pedraza Barrera1, Silvia Juliana Flórez González1, Lady Johanna Sierra
Prada1, and Elena E. Stashenko1
1) Universidad Industrial de Santander, Santander, Bucaramanga, Colombia.
Introduction: Plants are sources of photoprotective/antigenotoxic compounds that prevent cellular mutations
involved in skin cancer and aging by regulating UV-induced mutability. Purpose: The study was aimed atinvestigating the sunscreen properties of plants growing in Colombia. Materials and Methods: Ultraviolet radiation(UVR)-absorption capability of nineteen flower extracts obtained from eleven plant species was examined. In vitro
photoprotection efficacies were evaluated using in vitro indices such as sun protection factor (SPFin vitro) and critical
wavelength (c). UVB- antigenotoxicity estimates (%GI) using the SOS Chromotest were also obtained forpromising extracts. Cytotoxicity of these extracts was studied in human fibroblasts using the trypan blue exclusion
assay. Major compounds of promising plant extracts were determined by UHPLC-ESI+-Orbitrap-MS. Results:
Among the studied plant extracts, thirteen showed low (6.0 ≤ SPFin vitro ≤ 14.9), two medium (15.0 ≤ SPFin vitro ≤ 29.9)
and four high (30.0 ≤ SPFin vitro ≤ 59.9) photoprotection efficacy against UVB radiation, while only two resulted inbroad-spectrum (UVA–UVB) photoprotection (λc > 370 nm). These extracts were obtained from Ipomoea
horsfalliae, Posoqueria latifolia, and Rosa x centifolia plant species, they also showed UVB- antigenotoxicity andpossessed relatively low cytotoxicity (LD50  125 µg/mL) in human fibroblasts. These plant extracts had major
compounds as follows: chlorogenic acid, dicaffeoylquinic acid, and scopoletin (I. horsfalliae), caffeic acid andchlorogenic acid (P. latifolia), and cyanidin-3,5-glucoside, quercetin-3-rhamnoside, kaempferol-3-glucoside,kaempferol-rhamnoside, and kaempferol (R. centifolia). Several of these compounds also presented highphotoprotection efficacy and antigenotoxicity against UVB radiation, indicating that they were responsible for thecorresponding extract activity. Conclusions: I. horsfalliae, P. latifolia, and Rosa x centifolia plant species containphotoprotective compounds with low cytotoxicity in human fibroblasts and could be used as natural sunscreen
ingredients.
Acknowledgments. The authors thank funding from the Ministry of Science, Technology and Innovation,
the Ministry of Education, the Ministry of Industry, Commerce and Tourism, and
ICETEX, Program Ecosistema Científico-Colombia Científica, from the Francisco José de Caldas Fund,
Grant RC-FP44842-212-2018. The Ministry of Environment and Sustainable Development of Colombia
supported the Universidad Industrial de Santander through access permits to genetic resources andderivatives for bioprospecting (Contract N o. 270).

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IL-6.4.1
Adjuvant PDT after surgical resection:
murine modeling of applications to breast cancer, mesothelioma and beyond
Theresa M. Busch
University of Pennsylvania, Perelman School of Medicine, Department of Radiation Oncology, Philadelphia, USA
In the combined modality setting, the intraoperative delivery of photodynamic therapy (PDT) provides opportunity
to treat microscopic and/or unresectable disease at surgical margins. As an adjuvant that can intraoperatively bedelivered directly to the surgical field, PDT provides local cytotoxic effect in the absence of mutagenicity and thepotential for an anti-tumor immune response1. Murine models of PDT in the context of surgery provide for the studyof new regimens and an understanding of their limitations. Two models are discussed. One evaluates light dosimetry
and delivery as variables in the treatment of thoracic malignancies with pleural spread, such as mesothelioma andpleural dissemination of non-small cell lung carcinoma. In illumination of the murine thoracic cavity, fluence rate is
spatially distributed across the cavity, not unlike that found in patients who receive intraoperative PDT of their chest
cavity following macroscopic resection of pleural disease. As shown in Figure 1, fluence rate distributions can bemeasured in patients, identifying a range of treatment-averaged moderate fluence rates to which tissues are exposed2.
In murine models, intrathoracic fluence rate has been studied as a variable, demonstrating there to be fluence ratedependencies in PDT response, as known to occur with external beam PDT.
Fluence rate (mW/cm2)

150

100

50

0

Apex ACW PCW

PM Diaph PeriAS

PS

Fig. 1 – Illumination is delivered to the thoracic cavity during
intraoperative PDT for malignant pleural mesothelioma, resulting intreatment-averaged light exposures of ~40-60 mW/cm2 for most tissuesof the chest. Fluence rate is measured by isotropic detectors in the chest
cavity at tissues sites of the apex, anterior chest wall (ACW), posterior
chest wall (PCW), posterior mediastinum (PM), diaphragm (Diaph),
pericardium (Peri), posterior sulcus (PS), and anterior sulcus (AS).
Figure reprinted from Photochem Photobiol. 2020 Mar;96(2):417-425

Tissue typeIn a second murine model of intraoperative PDT, photosensitizer delivery is studied as a variable, in this casecomparing systemic dosing versus topical application of 5-aminolevulinic acid (ALA) in PDT of surgical margins
after the complete resection of murine tumor (TUBO) from the mammary fat pad. PDT in conjunction with either
systemically delivered ALA (oral gavage) or local administration (topical application to the resection site) after
complete resection of TUBO tumor provided for similar long-term tumor control. However, ALA-PDT with oral
drug delivery produced more vascular damage, a greater proportion of tissue-resident neutrophils and stronger
inflammation than that resulting from photosensitization with topical ALA application. Notably, these effects wereachieved after only a 10 min topical ALA exposure.
Each of the above-described murine models provide opportunity to explore questions relevant to the delivery of PDT
as an adjuvant to surgery. Observations generated in studies of such models are hypothesis generating–providing
guidance on novel approaches or new applications that can be taken back to the clinic.
References
1) Mahmoudi K, Garvey KL, Bouras A, Cramer G, Stepp H, Jesu Raj JG, Bozec D, Busch TM, Hadjipanayis
CG. 5-aminolevulinic acid photodynamic therapy for the treatment of high-grade gliomas. J Neurooncol.
2019 Feb;141(3):595-607. PMID: 30659522.
2) Dupre PJ, Ong YH, Friedberg J, Singhal S, Carter S, Simone CB 2nd, Finlay JC, Zhu TC, Cengel KA, Busch
TM. Light Fluence Rate and Tissue Oxygenation (StO2) Distributions Within the Thoracic Cavity of Patients
Receiving Intraoperative Photodynamic Therapy for Malignant Pleural Mesothelioma. Photochem
Photobiol. 2020 Mar;96(2):417-425. PMID: 32048732.
Acknowledgements. This work was supported by NIH grants R01 CA236362 and P01 CA087971.
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IL-6.4.2
Molecular and Functional Photoacoustic Imaging for personalizing Photodynamic
Therapy
Srivalleesha Mallidi
Department of Biomedical Engineering, Tufts University, Medford, MA USA 02155
Photodynamic therapy (PDT) is a photochemistry based cytotoxic technique that imparts cellular damage via
excitation of a photosensitizer with drug-specific wavelength of light. The dose at the treatment site for type II
PDT is determined by three factors: photosensitizer (PS) concentration, oxygenation status and delivered lightirradiance. Most of the FDA approved photosensitizers in their triplet-excited state generate cytotoxic species byreacting with the ground state oxygen that is available in the surrounding environment. Given the inter- and intrasubject variability in the uptake of the photosensitizer and the distribution of oxygen in the tumor, understanding
the interplay between these dose parameters could aid in determining photodynamic therapy efficacy. Previously
several studies have discussed the interplay between the dose parameters using shown point measurements and 2D
imaging systems. In this presentation, we will discuss the utility of molecular and functional photoacoustic imaging
for personalizing photodynamic therapy. Using various subcutaneous and orthotopic mouse models we will
demonstrate the utility of a non-invasive non-ionizing photoacoustic imaging modality to determine efficacy andpredict treatment response in Benzoporphyrin derivative (BPD), Aminolevulinic acid (ALA) based PDT or other
PDT based combination therapies. We further compare the predictive capability of photoacoustic imaging with themore predominantly used fluorescence imaging and immunohistochemistry techniques. Finally the clinicaltranslation of photoacoustic image-guided PDT will be discussed.

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IL-6.4.3
The Role of Mechanical Stress in Targeted PDT Combinations for Ovarian Cancer
Imran Rizvi1,2, Huang-Chiao Huang3, Utkan Demirci4, Tayyaba Hasan5, Marcela G. del Carmen6, Jonathan P.
Celli7, Shubhankar Nath4, Mustafa K. Ruhi1, Christina Conrad3.
1) University of North Carolina Chapel Hill and North Carolina State University, UNC-NC State Joint Departmentof Biomedical Engineering, Chapel Hill, NC, USA
2) University of North Carolina Chapel Hill School of Medicine, Lineberger Comprehensive Cancer Center,
Chapel Hill, NC, USA
3) University of Maryland, Fischell Department of Bioengineering, College Park, MD, USA
4) Stanford University School of Medicine, Bio-Acoustic MEMS in Medicine (BAMM) Laboratory, Canary Centerat Stanford for Cancer Early Detection, Department of Radiology, Palo Alto, CA, USA
5) Massachusetts General Hospital, Harvard Medical School, Wellman Center for Photomedicine, Department of
Dermatology, Boston,MA, USA
6) Massachusetts General Hospital, Harvard Medical School, Division of Gynecologic Oncology, Department of
Obstetrics and Gynecology, Boston, MA, USA
7) University of Massachusetts at Boston, Department of Physics, College of Science and Mathematics, Boston,
MA, USA
Mechanical stress (such as flow-induced shear stress) plays an important role in tumor growth and survival, withincreasing implications for therapy design and drug delivery. The role of physical stress in the tumor
microenvironment remains understudied in the context of photodynamic therapy (PDT) and PDT-based
combinations. A focus of this presentation will be to discuss the role of mechanical stress in the context of targeted
PDT with an emphasis on ovarian cancer, the leading cause of death from gynecologic malignancies. Advancedstage epithelial ovarian cancer is most frequently associated with the production of ascites, the abnormal
accumulation of fluid. Despite decades of evidence that the accumulation of peritoneal fluid portends the poorest
outcomes for cancer patients, the role of fluid flow in promoting metastases and therapy resistance, remains poorly
understood. The effects of flow-induced shear stress on chemoresistance and modulation of molecular survival
pathways that could be exploited for targeted PDT, and combinations, will be discussed.
References
1) Nath S, Pigula M, et al. Flow-induced Shear Stress Confers Resistance to Carboplatin in an Adherent ThreeDimensional Model for Ovarian Cancer: A Role for EGFR-Targeted Photoimmunotherapy Informed by PhysicalStress. J Clin Med, 2020.
2) Rizvi I, Gurkan UA, et al. Flow induces epithelial-mesenchymal transition, cellular heterogeneity and biomarker
modulation in 3D ovarian cancer nodules. Proc Natl Acad Sci USA, 2013.
3) Sorrin AJ, Kemal Ruhi M, et al. Photodynamic Therapy and the Biophysics of the Tumor Microenvironment.
Photochem Photobiol, 2020.
4) Conrad C, Gray KM, et al. Mechanical Characterization of 3D Ovarian Cancer Nodules Using Brillouin
Confocal Microscopy. Cell Mol Bioeng, 2019.
Acknowledgements. This work is supported by the following funding sources: Dr. Imran Rizvi - NIH R00
CA175292, NC Translational and Clinical Sciences Institute (NCTraCS), supported by the National Center for
Advancing Translational Sciences (NCATS), National Institutes of Health (NIH), through Grant Award Number
UL1TR002489, and UNC-NC State Joint BME Department Start-up Fund. Dr. Utkan Demirci - Stanford University
Canary Center Seed Grant, NCI-NIH grant to the Center for Cancer Nanotechnology Excellence for Translational
Diagnostics (CCNE-TD) at Stanford University (grant number: NIH U54 CA199075). Dr. Tayyaba Hasan – NIH
R01 CA158415, R01 CA160998, R01 CA119388 Dr. Huang-Chiao Huang - NIH R00 CA194269, UMD Start-up
Fund, UMD-UMB Research and Innovation Seed Grant, the Caring Together, NY, Ovarian Cancer Research grant,
and the UMD-NCI Partnership for Integrative Cancer Research Grant. Dr. Jonathan Celli – NIH U54CA156734
UMass Boston-Dana Farber/Harvard Cancer Center PartnershipBook of Abstracts ����������������������������������������������������������������������������������������������� 286

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OC-6.4.4
Optimizing Photodynamic Therapy: A Tour of the Death Pathways
David Kessel1, Imran Rizvi2 and Girgis Obaid3
1
Wayne State University, Department of Pharmacology, Detroit MI, USA; 2 UNC-NC State Joint Department of
Biomedical Engineering, University of North Carolina at Chapel Hill and North Carolina State University;
3
Department of Bioengineering, University of Texas at Dallas.
Photodynamic therapy (PDT) for the treatment of cancer involves the light-catalyzed formation of reactive molecularspecies (RMS) at sites of neoplasia. The precise death mode is a function of sub-cellular localization of thephotosensitizer since RMS do not migrate far from the site of formation. Apoptosis and necrosis are often cited as
death pathways but investigators tend not to examine cellular morphology and may thereby miss another common
death pathway: paraptosis. This occurs when the ER is among the targets for photodamage and is independent of
apoptosis. Paraptosis is characterized by the formation of a series of vacuoles derived from the ER that gradually fill
the entire cytoplasm, which eventually results in only bare nuclei appearing in microscopic fields.1 When PDT
targeting involves mitochondria or lysosomes, this leads to apoptosis. When both are targeted, release of lysosomal
calcium ions that can activate calpain, ultimately leading to a considerable promotion of the apoptotic consequencesof mitochondrial photodamage. The ideal targeting profile therefore appears to be a combination of mitochondria,
lysosomes and ER. This is not easy to achieve with a single photosensitizer but can be done via nanotechnology.
Because of solubility considerations, benzoporphyrin derivative (BPD, Visudyne) is formulated in a liposomal
environment and targets mitochondria and the ER. Adding a second nano-formulation provides the ability to target
lysosomes.2 Fig. 1 indicates how combining these two formulations can lead to the optimizing of PDT targeting so
as to promote maximum efficacy.

Fig. 1 – Ability of different BPD formulations to alter the sub-cellular targeting pattern

Details providing a comparison of apoptosis and paraptosis are shown in Fig. 2A. Apoptosis is characterized bycellular blebbing and fragmentation along with a corresponding fragmentation of nuclei. This can readily be revealed
by both phase-contrast imaging and observing fluorescence of the nuclear probe Ho33342. Fig. 2B provides acomparison of the photodynamic efficacy, individually and together, using the two BPD formulations described inFig. 1, reveals the synergistic effect of the combination of mitochondrial + lysosomal photodamage. Studies with the
OVCAR-5 cell line in 3D culture show that PDT doses of the free vs. lipid-anchored formulations individually led
to an undetectable level of photokilling but that the combination resulted in a very substantial decrease in viability.

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Fig. 2. A) Characteristics of apoptosis vs. paraptosis as indicted by phase-contrast and Ho33342 fluorescent labelling patterns.
B) PDT efficacy of free vs. anchored BPD preparations as defined in Fig. 1. Effects on viability of OVCAR-5 cells in 3D culture
are indicated.

References
1) Kessel D, Death pathways associated with photodynamic therapy. Photochem. Photobiol. In Press 2021
2) Rizvi R, Shubhanker N, Obaid G, Ruhl MK, Moore K, Bano S, Kessel D and Hasan, T. A combinationof Visudyne and a lipid-anchored liposomal formulation of benzoporphyrin derivative enhances photodynamic therapy efficacy in a 3D model for ovarian cancer. Photochem. Photobiol. 95, 419-429, 2019.

Acknowledgements. This work was partially supported by grants R01CA23378, K99CA215301 and R00CA175292
from the NIH.

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IL-6.4.5

Anti-angiogenics overcome tumor endothelial cell anergy and improve
immunotherapy outcomes
Zowi R. Huinen1, Elisabeth J.M. Huijbers1, Judy R. van Beijnum1, Patrycja Nowak-Sliwinska2,3,
Arjan W. Griffioen1*
1) Angiogenesis Laboratory, Department of Medical Oncology, Amsterdam University Medical Center, Vrije
Universiteit Amsterdam, Cancer Center Amsterdam, Amsterdam, The Netherlands; 2) Molecular Pharmacology
Group, School of Pharmaceutical Sciences, University of Geneva, Geneva, Switzerland; 3) Institute ofPharmaceutical Sciences of Western Switzerland, University of Geneva, Geneva, Switzerland.
Immunotherapy, in particular immune checkpoint inhibition (ICI), is becoming a pillar in cancer treatment enabling
unprecedented clinical benefits in a variety of malignancies, including durable responses in aggressive,
metastasized, or previously untreatable malignancies. Currently, however, only a subset of treated patients
responds. A compelling body of evidence shows that anti-angiogenic therapy has the capacity to ameliorate antitumor immunity, owing to the inhibition of various immunosuppressive features of angiogenesis. Hence, antiangiogenic- and immunotherapy combinations are currently tested in >80 clinical trials and 5 FDA approvals wererecently achieved. We describe the angiogenesis-induced endothelial immune cell barrier, which limits immune
cell infiltration and hampers anti-tumor immune responses and immunotherapy efficacy. We thereby highlight themechanism of endothelial cell anergy, which we describe as the vascular counterpart of immune checkpoints. We
review the anti-tumor immunity promoting effects of anti-angiogenic therapy, and give an update on the current
clinical successes achieved by the combination therapy.
The combination of photodynamic therapy (PDT) with anti-angiogenic agents has also been shown to be beneficial.
This has been described to be the result of the PDT-induced onset of angiogenesis. It remains to be demonstrated
that the PDT induced anti-tumor immune response also benefits from overcoming endothelial cell anergy by
combination with anti-angiogenic agents. We hypothesize that endothelial anergy is a vascular immune checkpointthat can be overcome by angiogenesis inhibition.

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OC-6.4.6
Protondynamic Therapy: A radical hybridization of PDT and proton radiotherapy
Theodossis A. Theodossiou1, Mantas Grigalavicius1, Maria Mastrangelopoulou1, Delmon Arous2, Jonas Asperud1,
Tine T. Henriksen Raabe1, Mathilde Ménard1, Efim Brondz2, Sunniva Siem2, Andreas Görgen2, Nina F. J. Edin2,
Eirik Malinen1,2, Kristian Berg1.
1

Department of Radiation Biology, Institute for Cancer Research, Oslo University Hospital, Oslo 0379, Norway.

2

Department of Physics, University of Oslo, Blindern, 0315, Oslo, Norway

Deep seated tumours as for example the central nervous system (CNS) cancer glioblastoma multiforme (GBM) arepractically incurable by the current standards-of-care. Even though photosensitive drugs (PSs) are used to guide theresection of GBM by their characteristic fluorescence, photomedical treatments like photodynamic therapy (PDT)or photochemical internalisation (PCI) are severely restricted by the shallow penetration of light into tissue (< 5mm), even at red/far-red wavelengths. Proton radiotherapy, can deposit a therapeutic amount of energy, in the socalled "Bragg peak", in varying depths within the tissue (even in excess of 20 cm), depending on the initial energyof the accelerated protons. Here we present proof of principle of a novel technology based on the radical
hybridisation of the principles behind PDT and proton therapy: Protondynamic Therapy. The underlying hypothesisof this hybridization was that accelerated protons can excite the electrons of PSs by attractive Coulomb forces, andthat the PSs can generate singlet oxygen and eliminate difficult to cure cancers like GBM by a dual action. Indeed,
upon irradiation of PS solutions and gels with accelerated protons at a research cyclotron, we verified the PS
proton-activation, by fluorescence, i.e. radiative de-excitation of the photosensitive molecules.
Furthermore, we verified the population of PS triplet states in dry gels where there was no oxygen quenching.
Finally we registered the production of singlet oxygen, either directly through its characteristic luminescence at1270 nm or indirectly through the formation of the singlet-oxygen-associated PpIX photoproduct:
PhotoProtoporphyrin IX. We subsequently tested our hypothesis in GBM cell cultures: M059K, T98G and U87, bycomparing the survival in cell groups irradiated by various proton doses in the abscence/presence of a PS. The
results revealed an enhanced cell death in the presence of a PS in M059K and T98G cells. In parallel to theexperimental work we are conducting theoretical calculations on the mechanism of electronic excitations byaccelerated protons.

Fig. 1. Principles of Protondynamic Therapy

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OC-6.4.7

Spatiotemporal controlled drug release with light to overcome chemotherapy
resistance in pancreatic cancer.
T. Le Clainche, N. Carigga Gutierrez, J-L. Coll and M. Broekgaarden
Institute for Advanced Biosciences, Université Grenoble-Alpes, INSERM U 1209, CNRS UMR 5309, Grenoble,
FranceDespite its low incidence, pancreatic ductal adenocarcinoma (PDAC) is the fourth-leading cause of cancer-related
mortality [1]. Cancer desmoplasia, which forms a protective barrier of stromal cells and extracellular matrix aroundcancer tissues, is widely known for its implication in the high resistance of PDAC against chemotherapeutic
treatments [2,3]. To improve the efficacy and safety of such treatments, we aim to design innovative liposomal vectors
to control the release of the drug in a spatiotemporal-controlled manner. In the field of photodynamic therapy,
photosensitizers such as benzoporphyrin derivative (BPD) are used to produce reactive oxygen species (ROS) uponlight excitation [4]. By including photosensitizers in liposomes composed of oxidation-susceptible unsaturated
phospholipids, we provide compelling proof for the spatiotemporal-controlled release of drugs with light (Figure 1)[5,6]. We have systematically optimized the liposome composition and discovered that liposomes surface properties
were of major impact on drug release efficiency. In parallel, we are developing new 3D culture models of PDAC on
alginate hydrogels that are compatible with state-of-the art, high-content imaging assays [7,8]. These models are used
to investigate the uptake and toxicity of the liposomes, and to determine the penetration depth of the released
therapeutics. These exciting findings open new possibilities for the controlled release of cancer therapeutics and pave
the way to in vivo experiments.

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Fig. 1-light-sensitive liposomes for spatiotemporal-controlled drug release.
References
1) J. Ferlay, M. Colombet, I. Soerjomataram et al. Cancer incidence and mortality patterns in Europe: Estimates for
40 countries and 25 major cancers in 2018. Eur. J. Cancer Oxf. Engl. 1990, 2018.
2) D. Fukumura, R. K. Jain, Tumor microenvironment abnormalities: Causes, consequences, and strategies tonormalize. J. Cell. Biochem, 2007.
3) P. P. Adiseshaiah, R. M. Crist, S. S. Hook et al. Nanomedicine strategies to overcome the pathophysiologicalbarriers of pancreatic cancer. Nat. Rev. Clin. Oncol, 2016.
4) P. Agostinis, K. Berg, K. A. Cengel et al. Photodynamic therapy of cancer: An update. CA. Cancer J. Clin, 2011.
5) M. Broekgaarden, R. Weijer, T. M. van Gulik et al. Tumor cell survival pathways activated by photodynamic
therapy: a molecular basis for pharmacological inhibition strategies. Cancer Metastasis Rev, 2015.
6) M. Broekgaarden, A. I. P. M. de Kroon, T. M. van Gulik et al. Development and in vitro proof-of-concept ofinterstitially targeted zinc-phthalocyanine liposomes for photodynamic therapy. Curr. Med. Chem, 2014.
7) A.-L. Bulin, M. Broekgaarden, T. Hasan, Comprehensive high-throughput image analysis for therapeutic
efficacy of architecturally complex heterotypic organoids. Sci. Rep. 2017.
8) M. Broekgaarden, A.-L. Bulin, J. Fredericket al. Tracking Photodynamic- and Chemotherapy-Induced RedoxState Perturbations in 3D Culture Models of Pancreatic Cancer: A Tool for Identifying Therapy-Induced
Metabolic Changes. J. Clin. Med, 2019.

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OC-6.4.8
Probing the mechanism of TPPS2a as a photosensitizing agent for PDT in 2D
monolayer and a 3D compressed collagen model of ovarian cancer
Balukova, A., MacRobert, A.J., Yaghini, E.
University College London, Division of Surgery and Interventional Science, London, United Kingdom
Introduction: In this study, the efficacy of the porphyrin-based photosensitiser TPPS2a (tetraphenylporphine
disulfonate) was evaluated in a 2D monolayer and 3D compressed collagen cancer model of a human ovarian cancer
cell line (HEY). This porphyrin which bears adjacently substituted sulfonate groups is known to be an effective
photosensitiser for both PDT and Photochemical Internalisation (PCI).
Aims: This study aimed to investigate the uptake and subcellular localisation of TPPS2a using fluorescence
spectroscopy and confocal microscopy with organelle markers, and to evaluate the photooxidative mechanism bywhich TPPS2a induces cell death by analysing 2D and 3D samples using ROS and lipid peroxidation probes.
Materials and Methods: The HEY cells were used for both the 2D and 3D culture models. The 3D cancer constructs
were manufactured using the RAFT 3D culture systems (Lonza, UK) protocol in 96-well plates, to create acompressed type 1 collagen hydrogel. Confocal microscopy (Leica SP8) using 405 nm excitation was used to assessthe uptake and localisation of TPPS2a (Frontier Scientific) in both in 2D and 3D cancer model. In 2D cultures, theuptake of 0.5 μM - 2 μM TPPS2a was assessed in fluorodishesTM. Furthermore, 5 μM 2’,7’-Dichlorofluorescin
diacetate (Sigma-Aldrich) and 5 μM BODIPYTM 581/591 C11 (ThermoFisher) were tested as ROS and lipid
peroxidation sensors. Cells were incubated for 24 h with drugs prior to imaging, with the exception of the additionof the ROS/lipid peroxidation probes which were applied 2 h prior to light exposure. For 3D constructs, 1 μM or 2
μM TPPS2a was applied on day 7 post-seeding and the constructs were left to incubate for a further 22 h.
Results: TPPS2a fluorescence was observed inside the cells and localized within the cellular membranes at the 4 htime point and inside endolysosomes at the 24 h time point as confirmed with the Lysotracker green. The mechanismof action of TPPS2a was investigated by illuminating porphyrin treated 2D and 3D samples for a longer time andimaging the PS fluorescence with increasing exposure time using a time-lapse mode of imaging. In both 2D and 3D
samples, real time imaging confirmed the fast-acting photodynamical properties of TPPS2a as exposure to blue lightincreased. To detect the formation of ROS and lipid peroxidation, 2D and 3D samples were co-treated with C11 andTPPS2a. Upon oxidation, a shift was detected in the fluorescence of C11, where the emission peak shifts from 590
nm (orange/red) to 510 nm (green). Using a sequential mode of imaging with two lasers and two detectors, a green
shift was clearly observed in the 2D and 3D cultures and was later confirmed by quantitative assessment of thefluorescence intensity of the images. Overall, this shift confirmed the presence of ROS which are a result of thephotoactivation of the TPPS2a. Dichlorofluorescein diacetate (DCFH-DA) is a cell permeable probe that becomes
fluorescent upon oxidation. In this study, 2D cultures were co-treated with DCFH-DA and TPPS2a. Sequential
imaging using two lasers and two detectors resulted in the observation of a c. 50% increase in mean fluorescenceintensity for regions of interests, which confirmed that the probe had been oxidised by the PDT-generated ROS.
Conclusion: Time-lapse confocal imaging permitted the visual observation of the PS fluorescence dynamics withtime as more ROS are generated. The PDT-induced generation of ROS was confirmed with C11 and DCFH-DA
using confocal microscopy in both 2D and 3D cancer models. The response observed using the lipid peroxidation
probe C11 is consistent with photosensitisation of endolysosomal membranes as required for the PCI mechanism.
Reference: Hadi L , Yaghini E, Stamati k , Loizidou M, MacRobert A.J. Therapeutic enhancement of a cytotoxic
agent using photochemical internalisation in 3D compressed collagen constructs of ovarian cancer. Acta
Biomaterialia, 2018.

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OC-6.4.9
Peptide-Targeted Systems for Photodynamic Therapy
Dong Wang1,2, Sandy MacRobert3, Charareh Pourzand1,2, Ian Eggleston1,2
1) University of Bath, Department of Pharmacy and Pharmacology, Bath BA2 7AY, UK.
2) Centre for Therapeutic Innovation, University of Bath, Bath BA2 7AY, UK
3) University College London, Division of Surgical and Interventional Sciences, London NW3 2PE, UK
Photodynamic therapy (PDT) is a minimally invasive approach for the treatment of cancer and various otherhuman disorders, based on the selective activation of photosensitizers (PSs) with light. At present, one of themost promising strategies for PDT and also fluorescence photodiagnosis (PDD) is to use 5-aminolevulinicacid (ALA) as a prodrug to increase intracellular levels of the endogenous PS, protoporphyrin IX (PpIX). Although ALA-PDT has been shown to be a very promising clinical approach, the physicochemical properties
and chemical reactivity of ALA present some challenges. These may be addressed by incorporation of ALA
units into a variety of prodrug systems,1 and we have previously shown that peptide-based prodrugs are an
attractive way to improve the delivery of ALA, leading to enhanced PpIX accumulation and PDT effects.2 In
this study, we present a novel and easy to assemble prodrug system to enhance the delivery of ALA to specificcell types using targeting with tumour-homing peptides. More specifically, we will describe the synthesis of
prodrug systems consisting of a molecular core to which multiple ALA units may be attached along with atargeting peptide of choice. This combines the concept of ALA dendrimers and ALA-peptide prodrugs.3 Asproof of concept of this particular approach, we have prepared systems containing a bombesin-derived peptidethat allows selective targeting of the GRP receptor (GRPR) which is overexpressed in a variety of tumours.
Targeted ALA delivery and PpIX production with these prodrugs in GRPR-expressing PC3 cells will be described.
References
1: Tewari K M, Eggleston I M. Chemical approaches for the enhancement of 5-aminolevulinic acid-based
photodynamic therapy and photodiagnosis. Photochemical & Photobiological Sciences, 2018.
2: Giuntini F, Bourré L, MacRobert A J, et al. Improved peptide prodrugs of 5-ALA for PDT: rationalizationof cellular accumulation and protoporphyrin IX production by direct determination of cellular prodrug uptake
and prodrug metabolization. Journal of Medicinal Chemistry, 2009.
3: Tewari K M, Dondi R, Yaghini E, et al. Peptide-targeted dendrimeric prodrugs of 5-aminolevulinic acid: A
novel approach towards enhanced accumulation of protoporphyrin IX for photodynamic therapy. Bioorganic
Chemistry, 2021.

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SP-6.4.10, P-6.4.10
How to use 5-ALA induced PpIX without light
Sara Sansaloni Pastor, Emmanuel Varesio, Norbert Lange
School of Pharmaceutical Sciences, Institute of Pharmaceutical Sciences of Western Switzerland, University ofGeneva, Switzerland
The selective generation of PpIX following exogenous administration of 5-ALA is one of the most selective processes
known in cancer or pre-cancer therapy and diagnosis. This feature of 5-ALA is now widely used in photodynamic
therapy (PDT) and photodiagnosis (PD), notably for the detection of bladder cancer and glioblastoma or the treatmentof basal cell carcinoma and actinic keratosis. These methodologies are restricted to body areas that are accessible tolight which tissue penetration is only sallow. Therefore, we were searching for possibilities to use this phenomenon
differently, i.e. without the use of visible light. To this end we have analyzed to proteonome of different cell lines ofcancerous origin following exposure to 5-ALA. In this presentation, our new methodology will be presented.
Furthermore, potential future applications and indications will be discussed.
Acknowledgements. This work was supported by a grant of the Swiss National Science Foundation.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-6.4.11

Conjugates of octreotide and exendin-4 with zinc-phthalocyanine TT1 forphotodynamic therapy of neuroendocrine tumors
Sanne A.M. van Lith1, Alba Fonseca3, Desiree Bos1, Annemarie Kip1, Fritz Andreae2, Thorsten Radolf2, Tomas
Torres3, Martin Gotthardt1
1) Radboud university medical center, Medical Imaging, Nijmegen, The Netherlands; 2) piCHEM, RaabaGrambach, Austria; 3) Autonoma University of Madrid, Department of Organic Chemistry, Madrid, Spain
Neuroendocrine tumors (NETs) comprise a very heterogeneous group of malignancies, as they can arise fromneuroendocrine cells in virtually every internal organ. The management of localized disease involves surgicalresection with curative intent. To ensure complete removal of the tumor, currently often unnecessary radical
resections are performed, and this is associated with substantial morbidity. Furthermore, residual disease often leads
to recurrence of these lesions. Therefore, techniques that lead to more specific and less invasive ablation of tumor
cells are warranted, and targeted photodynamic therapy is an interesting treatment option. Two G protein-coupled
receptors that are regularly overexpressed on neuroendocrine tumors are the somatostatin receptor (SSTR), and theglucagon-like peptide 1 receptor (GLP1R). Here, we developed and characterized peptide-photosensitizer conjugatesfor SSTR- and GLP1R-targeted photodynamic therapy.
Zinc-phthalocyanine based photosensitizer TT1-maleimide was conjugated to a sulfhydryl group on the
primary amine of Lys40 in GLP1R-targeting peptide exendin-4, and of dPhe1 in SSTR-targeting peptide octreotide.
properties such as absorbance and fluorescence of exendin-4-TT1 and octreotide-TT1 were characterized in dimethyl
sulfoxide (DMSO), phosphate buffered saline (PBS) and PBS with human serum or bovine serum albumin. To
determine half maximal inhibitory concentrations (IC50), chinese hamster lung cells stably transfected with GLP-1R(CHL-GLP1R) or SSTR-overexpressing AR42J cells were co-incubated with either exendin-4-TT1 and 111In-labeledexendin-4-DTPA as a competitor or octreotide-TT1 and 111In-labeled octreotide-DTPA as a competitor, respectively.
Then, light-induced toxicity by both compounds was determined. Cells were incubated for 4 hours with various
concentrations of either exendin-4-TT1 or octreotide-TT1, and after washing away the unbound fraction, cells were
illuminated with 690 nm light using a light emitting diode (LED 100 J/cm2 at 200 mW/cm2). Non-illuminated cells
and cells incubated with excess exendin-4-DTPA or octreotide-DTPA served as controls. Finally, BALB/c nude micecarrying subcutaneous xenografts of CHL-GLP1R tumors were injected with 20 µg exendin-4-TT1, and an ex vivo
fluorescent biodistribution to determine uptake in tumors and other tissues (liver, spleen, kidneys, blood, pancreas)
was performed at 4 and 16 hours post-injection.
Spectral properties of TT1 were retained after conjugation to both peptides, with excitation and emission
maxima of 682 nm and 696 nm in DMSO. Dissolving the conjugates in PBS led to a blue-shift and a decreased
intensity of the Q-band, and this could partly be rescued by adding human serum or bovine serum albumin. Bothexendin-4-TT1 and octreotide-TT1 bound to target-expressing cells with high affinity (IC50: 85.2 nM and 28.9 nM,
respectively). Cytotoxicity of CHL-GLP1R and AR42J cells was induced upon incubation with exendin-4-TT1 oroctreotide-TT1 and illumination with 690 nm light, respectively. The cytotoxic effect was blocked by adding an
excess of exendin-4-DTPA or octreotide-DTPA, indicating receptor specificity. Both conjugates did not induce
cytotoxicity without illumination. Upon injection in BALB/c nude mice carrying subcutaneous CHL-GLP1R
xenografts, relative tumor uptake of 1.46±0.52 %ID/g and 3.24±1.05 was found for exendin-4-TT1 at 4 and 18 hours
post injection, respectively. Non-target specific uptake of exendin-4-TT1 was mainly found in the liver (37.98±3.69%ID/g at 18 hours post injection) and spleen (56.04±13.36 %ID/g at 18 hours post injection).
In conclusion, the novel TT1-peptide conjugates show potency for SSTR,- and GLP1R-targeted PDT in vitro.
Furthermore, exendin-4-TT1 homes to GLP-1R expressing tumors in vivo. In ongoing experiments, we will
determine in vivo tumor targeting of the octreotide-TT1 conjugate, and light-induced ablation of target-expressing
tumor cells by both conjugates.
Acknowledgements. This work was supported by an investigator grant from the neuroendocrine tumor foundation.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-6.4.12,

Improving the efficiency of ALA-PDT of skin cells using indoor and outdoor UVAbased light fractionation approach
Dana Beiki1, Ian Eggleston1,2, Charareh Pourzand1,2
1) Department of Pharmacy and Pharmacology, University of Bath, Bath, United Kingdom; 2) Centre for
Therapeutic Innovation, University of Bath, Bath, United Kingdom
Topical 5-aminolevulinic acid-based photodynamic therapy (ALA-PDT) is widely used for the treatment ofdermatologic conditions including actinic keratoses (AK). ALA administration to skin cells results in intracellularaccumulation of the endogenous photosensitiser protoporphyin IX (PpIX). Upon exposure to a light source (usually
blue or red light), PpIX can catalyse the generation of reactive oxygen species, resulting in cell death. The major
drawback of topical ALA-PDT is the pain experienced by patients. This may be counteracted by improving theefficiency of the ALA-PDT by optimisation of the light source, irradiation dose and duration of treatment. For thispurpose, ultraviolet A (UVA, 320-400 nm) was used instead of conventional visible light sources as it is absorbed
efficiently by PpIX and is significantly more cytotoxic due to its ability to promote an increase in the intracellularlevel of harmful labile iron which can be a major contributor to cell death by acting as a catalyst for oxidative
damage to cell constituents.1 Our optimization strategy relied on exploiting the rapid release of labile ironfollowing application of short pulses of low UVA doses to ALA-treated cells instead of a continuous dose fromvisible light source, to further sensitise cells to subsequent UVA doses. The proof of concept of this new approach,referred to as “UVA-based light fractionation of ALA-PDT” was evaluated in immortalised HaCaT keratinocytes
as a model of skin cancer. In clinical settings, this protocol would have the potential to reduce considerably thetreatment time and the discomfort/pain associated with prolonged exposure to high intensity visible light.
The experimental procedure involved the evaluation of the impact of a series of sub-lethal dose combinations of
UVA, designed to promote labile iron release and exacerbate the intracellular oxidative damage following a second
UVA light dose in ALA-treated cells using both an indoor broad spectrum UVA lamp (Sellas, Germany, 340-405nm) and outdoor sunlight. In the indoor setting, cytotoxicity assays (MTT, AnnexinV-propidium iodide flow
cytometry and colony-forming), showed that: (i) ALA significantly sensitised HaCaT keratinocytes to very low
non-cytotoxic UVA doses (up to 20 kJ/m2) and (ii) applying two short pulses of UVA (1-2.5 kJ/m2) to ALA-treatedcells with a dark interval (60-120 min) was a fast and effective way to promote cell death. The outcome of thisproject also highlighted the importance of the first UVA dose applied and of the extent of labile iron release indetermining the efficiency of the subsequent UVA doses. The low split-dose UVA radiation protocol has therefore
the potential to improve the current modality for topical ALA-PDT, through a reduction of the irradiation time andhence the duration of pain endured upon the treatment.
In the outdoor light fractionation scheme, we investigated the biological impact of the solar components of UVA
and visible light together or visible light alone, using the organic UVB-specific filter, octocrylene, and thecommercially available UVB+UVA filters, SPF 50 +, respectively. The synergistic effect of the solar emission
bandwidth was significantly higher in the photokilling of ALA-treated cells, and proved to be beneficial inshortening irradiation periods, without compromising the treatment efficiency. The latter provided the first proof ofconcept study for a novel UVA-based daylight PDT with light fractionation involving two short pulses of daylight
exposure with a 60-120 min dark interval. The present work is the first of its kind demonstrating that a lightfractionation protocol with extremely low doses of UVA (e.g. 1 kJ/m2 translating to ca 30 s in sunlight) with a 1 hinterval can effectively inactivate over 80% of the colony forming ability of ALA-treated and daylight-irradiatedcells either alone or with octocrylene filtering out the harmful UVB component of sunlight. The results further
indicated that while exposing an ALA-treated patient’s skin lesions to sunlight with UVB+UVA filters SPF 50 +..isunlikely to be effective at such low doses, our UVA-based daylight PDT split-dose regimen can be a more simple
and more powerful system for the daylight PDT of AK patients.
References
1) Pourzand C, Watkin RD, Brown JE, Tyrrell RM. Ultraviolet A radiation induces immediate release of iron inhuman primary skin fibroblasts: the role of ferritin. Proc Natl Acad Sci U S A. 1999

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P-6.4.13
Carbapenem resistant Enterobacterales do not resist antibiotic treatment upon
exposure to antimicrobial blue light.
Agata Wozniak1, Izabela Zybala1, Mariusz Grinholc1
1) Laboratory of Molecular Diagnostics, Intercollegiate Faculty of Biotechnology University of Gdansk andMedical University of Gdansk, Gdansk, Poland.
Carbapenem resistant Enterobacterales (CRE) are responsible for infections associated with high mortality rates
among hospitalized patients, up to 50% in some studies1. Continuously growing antimicrobial resistance among CRE
strains forces into a finding a new method of eradication or re-sensitization to antibiotics such resistant strains.
Antimicrobial blue light (aBL) is suited for such applications. Presence of endogenously produced porphyrins andflavins, which can be easily excited by appropriate wavelength leads to production of reactive oxygen radicals 2.
Therefore, the oxygen radicals in appropriate amount can lead to the damage of bacterial cells probably resulting inincreased susceptibility to antibiotics or cell death.
In present study two clinical isolates of Enterobacter cloacae (no. 2640/19, 4986/12) and two isolates Klebsiellapneumoniae (no. 680, 479) were exposed to sub-lethal doses of aBL and the influence of light on the level ofantimicrobial resistance CRE strains was investigated with multiple methods (E-test, checkerboard assay,
postantibiotic effect). Moreover, the presence of endogenous porphyrins and flavins in CRE strains was confirmedwith the UV-vis spectroscopy. Production of ROS and singlet oxygen after exposure of bacterial cells to aBL was
examined with fluorescent probes (AMDA, HPF) and additional SYTOX green and Propidium iodide (PI) probes
were involved in the cell membrane integrity assays.
E. cloacae isolate no. 2640/13 after exposure to low dose of blue light was more susceptible to doxycycline,
tigecycline and imipenem, and second strain no. 4986/12 to tigecycline, ciprofloxacin and fosfomycin - these results
were confirmed with disk diffusion method. However, the checkerboard assay revealed that both E. cloacae andK. pneumoniae strains after exposure to aBL had increased susceptibility to ceftazidime and colistin. Postantibiotic
effect confirmed also increased susceptibility after blue light exposure only for E. cloacae isolate no. 4986/12 forcolistin and fosfomycin. Observed synergies in case of strains no. 680 and 2640/13 for light and antimicrobials(ceftazidime, colistin) can be explained by the increased permeabilization which occurred after combined treatment
with the implementation of SYTOX green and PI probes. Combined action of blue light and antibiotics (e.g., colistin
and ceftazidime) lead also to the increased production of ROS in case of strains no. 680 and 2640/13 and this fact
can also explain the increased susceptibility of these isolates to antimicrobials.
Antimicrobial blue light inactivation can efficiently increase susceptibility of Enterobacterales to various antibiotics
e.g. fosfomycin, ceftazidime, however this phenomenon is strain depended, thus the re-sensitization to antimicrobials
was observed between different strains and applied methods. Increased sensitivity to antibiotics after exposure toblue light can be explained by the increased permeabilization of cell membranes and ROS activity, decreased activityof carbapenemase or other not recognized within this study mechanisms of resistance.
References
1) CRE Technical Information | CRE | HAI | CDC https://www.cdc.gov/hai/organisms/cre/technical-info.html
2) Wang, Y.; Wang, Y.; Wang, Y.; Murray, C.K.; Hamblin, M.R.; Hooper, D.C.; Dai, T. Antimicrobial blue
light inactivation of pathogenic microbes: state of the art. Drug Resist. Updat. 2017
Acknowledgements. This work was supported by grant from National Centre of Science: 2015/19/B/NZ7/02487

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P-6.4.14

Photosensitizing Efficacy of Curcumin-Loaded Liposomes Following Photodynamic
Therapy on Melanoma MUG-Mel2, Squamous Cell Carcinoma SCC-25 and Normal
Keratinocyte HaCaT Cells
Marta Woźniak1, Martyna Nowak1, Izabella Jabłońska1, Krzysztof Szpadel1, Piotr Ziółkowski1
1) Wroclaw Medical University, Department of Pathology, Wroclaw, Poland
The research focused on the investigation of curcumin encapsulated in hydrogenated soya phosphatidylcholine
liposomes and its increased photoactive properties in photodynamic therapy (PDT). The goal of this study was
twofold: to emphasize the role of a natural photoactive plant-based derivative in liposomal formulation as an easily
bioavailable alternative photosensitizer (PS) for the use in PDT of skin malignances. Furthermore, to prove thedecreased cytotoxicity of phototoxic agent loaded in liposomes towards normal skin cells. Our research was
conducted on melanoma (MugMel2), squamous cell carcinoma (SCC-25), and normal human keratinocytes (HaCaT)
cell lines. The MTT study evaluated cell death after exposure to blue light irradiation after 4 hours of preincubation
with free and encapsulated curcumin. Additionally, the wound healing assay, flow cytometry, andimmunocytochemistry to detect apoptosis were performed. The malignant cells revealed increased phototoxicity after
the therapy in comparison to normal cells. Moreover, liposome curcumin-based photodynamic therapy showed anincreased ratio of apoptotic and necrotic cells. The study also demonstrated that nanocurcumin significantly
decreased malignant cell motility following PDT treatment. Our results suggest that liposomal formulation of a poor
soluble natural compound may improve photosensitizing properties of curcumin-mediated PDT treatment in skincancers and reduce toxicity in normal keratinocytes.

Fig. 1 –Scheme of encapsulated curcumin in liposomes and mechanism of photodynamic therapy.

References
1) Szlasa, W.; Supplitt, S.; Drąg-Zalesińska, M.; Przystupski, D.; Kotowski, K.; Szewczyk, A.; Kasperkiewicz,
P.; Saczko, J.; Kulbacka, J. Effects of Curcumin Based PDT on the Viability and the Organization of Actinin Melanotic (A375) and Amelanotic Melanoma (C32) – in Vitro Studies. Biomed. Pharmacotherapy, 2020.
2)
Acknowledgements. This research was funded by the Wroclaw Medical University, grant number SUB.A010.20.13.

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P-6.4.15

Photobiological studies on a novel lead BODIPY-anthracene dyad for bladdercancer therapy
Odrun A. Gederaas,1 Susan Callaghan,2 Benedikte E. Vindstad,1 Keith J. Flanagan2, Thor B. Melø,1 Mikael
Lindgren,1 Kristin G. Sæterbø,1 and Mathias O. Senge2
1.Department of Physics, Department of Physics, Norwegian University of Science and Technology(NTNU), N-7491 Trondheim (Norway)
2.Medical Chemistry, Trinity Translational Medicine Institute, Trinity Centre for Health Sciences,
Trinity College Dublin, The University of Dublin, St. James’s Hospital, Dublin 8 (Ireland)

The structural, photobiological, and photophysical properties of a lead photosensitizer PDT BODIPYanthracene dyad BAD-3 were studied to better understand the properties of this molecule and to assess it
as a possible photodynamic therapy agent for bladder cancer treatment.
The rat bladder cancer cell line (AY27) was grown in standard RPMI growth medium, and further
incubated with the photosensitizer BAD-3 before blue light illumination (LumiSource, 13mW/cm2).
Using flow cytometry (Beckman Coulter Gallios Flow Cytometer), both necrosis and apoptosis processes
were studied by using annexin V, PI, and annexin-binding buffer. The cells were imaged employing a
confocal laser scanning microscope (Leica TCS SP8 MP, Leica Microsystems, Germany) after incubationof different staining procedures.
BAD-3 gave a strong therapeutic response against the AY27 cell line and additionally cytotoxicity
was observed in the glioma F98 cell line. At a concentration of 10 µM, BAD-3 induced complete killing
after 120 s of illumination using 12.9 mW/cm2 in the AY27 cells, thus presenting as a promising andversatile photodynamic therapy agent. The flow cytometry document both necrosis and apoptosis and alarge G1 and minor S phase arrestation post PDT.
The current study confirmed that BAD-3 is a suitable BODIPY-anthracene dyad for medical
applications. The characteristics of BAD-3, with view of translating this photosensitizer to an in vivo
model, is promising.
1) Callaghan S, Vindstad BE, Flanagan KJ, Melø TB, Lindgren M, Sæterbø K G, Gederaas OA and Senge
MO. ChemPhotoChem, 2020, 5, 131.

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P-6.4.16
5-Aminolevulinic acid-Induced Porphyrin Generation in Prostate Cancer Cells with
N-Formyl and N-Acetyl Peptide Prodrugs
Nabeela Farooq1, Thomas Elliott2, Matthew Hunt2, David White2, Ian Eggleston2, Sandy MacRobert1
1) University College London, Division of Surgical and Interventional Sciences, London, W1W 7TY, United
Kingdom; 2) University of Bath, Department of Pharmacy & Pharmacology, Bath, BA2 7AY, United Kingdom
Background. Photodynamic diagnosis (PDD) and therapy (PDT) for fluorescence imaging and focal therapy of
prostate cancer (PCa) is a growing area of research. Using 5-aminolevulinic acid (ALA) as a prodrug to generate
the photosensitiser protoporphyrin (PpIX) has shown much promise. However, inadequate physicochemistry,
lipophilicity and bioavailability of ALA warrants the search for novel ALA prodrugs. Here, amino acidconjugated ALA derivatives with either acetyl or formyl N-termination have been investigated to circumventthese drawbacks. No previous studies using the conjugates investigated herein have been carried out on PCa
lines.
Methods. Human PCa lines (PC3 and LNCaP) were treated with ALA and four phenylalanine- or methionine- based
dipeptide derivatives. For each, the efficiency of PpIX synthesis as a function of concentration was investigated,assessed as intracellular fluorescence compared to an ALA standard. Next, PDT was modelled by irradiation,and cellular viability measured using metabolic activity (MTT) assays. Fluorescence and microscopy imaging
were performed to support these findings.
Results. Cell line- and time-dependent, ALA-mediated PpIX generation was exhibited in this study. Using PC3, 3of the 4 drugs tested exhibited promising PpIX synthesis at low doses, but ineffective fluorescence at molarities
0.05mM and above. In line with our previous findings with N-acetyl peptide prodrugs of ALA, Ac-Phe-ALAMe was the most effective derivative tested here.1 Methionine derivatives were less successful, with Ac-MetALA-Me marginally outperforming For-Met-ALA-Me. Experimentation with For-Phe-ALA-Me failed to elicit
measurable PpIX in either cell line (P<0.01).
Conclusion. Our studies have shown that N-formyl and N-acetyl peptide prodrugs of ALA can be readily synthesised.
ALA release from both types of prodrug can take place in PCa cells leading to PpIX formation that could be
exploited for PDT.
References
1) Giuntini F, Bourre L, MacRobert AJ, Wilson M, Eggleston IM. Improved peptide prodrugs of 5-ALA for PDT:
rationalization of cellular accumulation and protoporphyrin IX production by direct determination of cellular prodrug
uptake and prodrug metabolization. Journal of Medicinal Chemistry, July 2009.

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PL-11

Reactive Oxygen Species Go Beyond Photodynamic Therapy
Xiaoyuan Chen
1) National University of Singapore, Departments of Diagnostic Radiology, Surgery, Chemical and BiomolecularEngineering, Biomedical Engineering, Yong Loo Lin School of Medicine and Faculty of Engineering, Singapore,
Singapore
The reactive oxygen species (ROS)-mediated mechanism is the major cause underlying the efficacy ofphotodynamic therapy (PDT). The PDT procedure is based on the cascade of synergistic effects between light, aphotosensitizer (PS) and oxygen, which greatly favors the spatiotemporal control of the treatment. This procedure
has also evoked several unresolved challenges at different levels including (i) the limited penetration depth of light,
which restricts traditional PDT to superficial tumors; (ii) oxygen reliance does not allow PDT treatment of hypoxic
tumors; (iii) light can complicate the phototherapeutic outcomes because of the concurrent heat generation; (iv)
specific delivery of PSs to sub-cellular organelles for exerting effective toxicity remains an issue; and (v) side
effects from undesirable white-light activation and self-catalyzation of traditional PSs. In this talk, the current
status and the possible opportunities of nanomedicine for ROS generation for cancer therapy will be discussed indetail.
Acknowledgements. This work was supported by the start-up fund of the National University of Singapore(NUHSRO/2020/133/Startup/08) and NUS School of Medicine Nanomedicine Translational Research Programme(NUHSRO/2021/034/TRP/09/Nanomedicine)Book of Abstracts ����������������������������������������������������������������������������������������������� 302

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IL-7.1.1

Imaging and photoswitching actin dynamics by small molecule probes
Hans-Dieter Arndt**Friedrich-Schiller-Universität, Institut für Organische Chemie und Makromolekulare Chemie, Humboldtstr. 10,
D-07743 Jena, Germany; hd.arndt@uni-jena.de
Actin is the most abundant cytoskeletal protein and key to most cellular activities.1) Therefore, mastering on-target
selectivity and confining activity in its pharmacological modulation will be crucial for resolving actin functions on
the tissue and cell level. By starting from F-actin targeting jasplakinolide derivatives,2) chemical synthesis, SAR
and labeling studies led us to create cell permeable fluorescent probes such as SiR-Actin with exquisite target
staining selectivity,3) and to resolve the molecular interaction of these ligands by cryo-EM.4)
For controlling actin dynamics locally by using photopharmacology,5) jasplakinolide-derived optojasp photoswitches were developed that feature metastable azobenzene triggers.6) Late stage derivatization guided by SAR
data enabled a systematic study of photoswitch type and ligand structure. From this collection several compounds
active in either the “dark” (E) or “irradiated” (Z) form were identified that show remarkable potential for the lightdependent manipulation of actin dynamics in living cells.6) First applications of optojasps in cell biology6) andstructural properties of F-actin4) and of ligand binding in both switch states7) will be discussed. Furthermore, firstdata on photoswitch optimization8) and second generation optojasps with improved properties will be presented.9)

References
1) T. D. Pollard, G. G. Borisy, Cell 2003, 112, 453.
2) a) H. Waldmann et al., Angew. Chem. Int. Ed. Engl. 2008, 47, 6473; b) R. Tannert et al., J. Am. Chem. Soc.
2010, 132, 3063.
3) a) L. G . Milroy et al., J. Am. Chem. Soc. 2012, 134, 8480, b) G. Lukinavičius et al., Nat. Methods 2014, 11, 731.
4) F. Merino et al., Nat. Struct. Mol. Biol. 2018, 25, 528.
5) J. Broichhagen, F. A. Frank, D. Trauner, Acc. Chem. Res. 2015, 48, 1947.
6) M. Borowiak, F. Küllmer, F. Gegenfurtner, S. Peil, V. Nasufovic, S. Zahler, O. Thorn-Seshold, D. Trauner, H.D. Arndt, J. Am. Chem. Soc. 2020, 142, 9240.
7) S. Pospich, F. Küllmer, V. Nasufovic, J. Funk, A. Belvy, P. Bieling, H.-D. Arndt, S. Raunser, Angew. Chem. Int.
Ed. 2021, 60, 8678.
8) F. Küllmer, L. Gregor, H.-D. Arndt, in submission.
9) F. Küllmer, N. Veprek, M. Borowiak, V. Nasufovic, S. Barutzki, D. Trauner, H.-D. Arndt, in preparation.

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IL-7.1.2

Next-generation optogenetic tools for manipulating the membrane potential
Ramona A Kopton1, Franziska Schneider-Warme1
1) Institute for Experimental Cardiovascular Medicine, University Heart Center Freiburg – Bad Krozingen,
Freiburg, Germany;2) Medical Faculty, University of Freiburg, Freiburg, Germany
Optogenetic approaches offer unprecedented opportunities to manipulate cellular activity at high spatiotemporal
resolution. Most commonly used optogenetic tools comprise light-gated ion channels (so-called channelrhodopsins)and light-driven ion pumps that enable modulating the membrane potential in the target cells upon illumination.
More specifically, activation of cation-conducting channelrhodopsins such as ChR2 from Clamydomonas
reinhardtii leads to depolarizing photocurrents, which can be used to trigger action potentials in excitable cells,
including neurons and cardiomyocytes. Light-driven proton, chloride and sodium pumps actively transport ionsagainst the electrochemical gradient, thereby hyperpolarizing the membrane potential and reducing cellular
excitability. The hyperpolarizing effect of ion pumps, however, is limited, as maximally one ion can be transported
per absorbed photon, thus requiring constant high light intensities for sustained hyperpolarization. The more
recently found group of anion channelrhodopsins (ACR) mediates light-activated chloride currents, which can beused to “shunt” the membrane potential to the reversal potential of chloride. This has been exploited for silencing
excitable cell activity, but we and others have shown that, depending on transmembrane gradients for chloride,
which vary in different cell types and subcellular compartments, activation of chloride channels can have both
depolarizing and hyperpolarizing effects1.
The ideal tool for optogenetic silencing of excitable cells would be a light-gated potassium channel, whose
activation would draw cells towards the potassium reversal potential, close to the natural resting membranepotential. We have established the two-component optogenetic system PAC/K, combining a light-activated cyclase
and cyclic-nucleotide gated (CNG) potassium channel, which drives long-lasting (seconds to minutes) potassium
currents upon illumination with short (milliseconds) blue light pulses. Resulting photocurrents effectively inhibit
action potential initiation, as shown for cardiomyocytes and different neuronal classes, both in vitro and in vivo2.
At present, optogenetic applications of PAC/K are limited as 1) use of cAMP for activation of the CNG channel
can have detrimental side effects, and 2) the slow off-kinetics do not offer sufficient temporal control to inhibit
individual action potentials. In on-going research, we develop alternative potassium-based silencing systems, basedon rhodopsin guanylyl cyclases and custom engineered potassium channels, with the vision to be able to reversibly
silence excitable cells with millisecond precision without affecting their resting potential.
References
1. Kopton RA et al. Cardiac Electrophysiological Effects of Light-Activated Chloride Channels. Front
Physiol 2018/9:1806.
2. Bernal Sierra YA et al. Potassium channel-based optogenetic silencing. Nat Commun 2018/9(1):4611.
Acknowledgements. This work was supported by the German Research Foundation (DFG: Priority Program 1926[#315193289], and the Emmy-Noether Programme [#412853334]). FSW is a member of CRC1425 and an
associate member of the excellence cluster Centre for Integrated Biological Signalling Studies (CIBSS).

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-7.1.3
Controlling Gene Expression with Light
Lea Albert, 1 Benedikt Heinrich, 1 Olalla Vázquez 1
1) Philipps-Universität Marburg, Department of Chemistry & SYNMIKRO, Marburg, Germany
All cells in an organism contain the same DNA sequence but vary greatly in gene expression. Epigenetics deals withthese phenotype changes that retains the same DNA sequence. Importantly, misregulation of these epigenetic
processes is implicated in the pathophysiology of numerous human diseases, 1 including: cancer, autoimmune
disorders and neurodegenerative disease. Therefore, epigenetic regulation is at the core of both natural andpathological states. The current available methods do not have sufficient spatiotemporal resolution to deal with thechallenges of targeting the dynamic epigenome. We and others 2 envision that light could offer new possibilities andachieve molecular functionality. Reversible photoswitches, which have demonstrated their potential in diverse areas
such as material science, have hardly implemented as genome regulators. Modulating the epigenome to tunetranscription profiles, and cellular phenotypes in a programmable manner is of wide interest, and may ultimately leadto novel epigenetics-based therapeutics. Here, we will present an overview of our optoepigenetic tools from ourphotoswitchable peptides as histone-methyltransferase inhibitors 3 until our light-activatable nucleosome binders
with capacity to modulate DNA accessibility.4

Fig. 1 – Conceptual idea of using light to control gene expression

References
1.
Portela, A.; Esteller, M., Epigenetic modifications and human disease. Nat Biotechnol 2010, 28 (10), 105768.
2.
Hartmann, D.; Smith, J. M.; Mazzotti, G.; Chowdhry, R.; Booth, M. J., Controlling gene expression withlight: a multidisciplinary endeavour. Biochem Soc Trans 2020, 48 (4), 1645-1659.
3.
Albert, L.; Xu, J.; Wan, R.; Srinivasan, V.; Dou, Y.; Vazquez, O., Controlled inhibition of methyltransferases
using photoswitchable peptidomimetics: towards an epigenetic regulation of leukemia. Chem Sci 2017, 8 (6), 46124618.
4.
Heinrich, B.; Bouazoune, K.; Wojcik, M.; Bakowsky, U.; Vazquez, O., ortho-Fluoroazobenzene derivatives
as DNA intercalators for photocontrol of DNA and nucleosome binding by visible light. Org Biomol Chem 2019, 17(7), 1827-1833.
Acknowledgements. This work was financial supported by German Research Fundation (DFG) programs: SPP1926'Next Generation Optogenetics'; Young Investigator Program (grant # GO1011/11-1) and TRR81: “Chromatin
Changes in Differentiation and Malignancies (TRR81/3, Z04).

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for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-7.1.4

Light-Free and Self-Activating Single-Molecule Chemiluminescent Photosensitizers
for Selective Photodynamic Therapy of Cancer
Luís Pinto da Silva1, Carla M. Magalhães1, Patricia González-Berdullas1
1) Chemistry Research Unit (CIQUP), Faculty of Sciences of University of Porto, Porto, Portugal; e-mail:luis.silva@fc.up.pt
Photodynamic therapy (PDT) is a cancer treatment with significant advantages over more conventional cancer
therapies, such as its minimally-invasive nature and fewer side-effects.1 However, the low penetration of light into
biologic tissues limits PDT to tumours on/just under the skin or on the outer lining of internal organs and cavities.1
Herein, we have developed single-molecule photosensitizers able of light-free and intracellular self-activation fortumour-selective PDT, which is based on a chemiluminescent reaction involving a cancer marker (Fig. 1).2,3 Morespecifically, the photosensitizers are directly chemiexcited to a triplet excited state (T1) capable of sensitizing
singlet oxygen, without the need for a light source.2,3Fig. 1 – Proposed light-free and tumour-selective PDT mechanism based on a chemiluminescent reaction.

Cytotoxicity assays were performed toward different cancer cell lines (breast, neuroblastoma, colon and prostate),with the photosensitizers showing relevant toxicity toward these cells,2,3 with activities comparable with reference
chemotherapeutic drugs. Importantly, these novel photosensitizers did not induce toxicity toward normal cells.2,3
In conclusion, this work provides a proof-of-concept for a novel type of single-molecule photosensitizer thateliminates the current restrictions that PDT presents regarding tumour size and localization.
References
1) W. Fan, P. Huang, X. Chen, Overcoming the Achilles’ heel of photodynamic therapy. Chem. Soc. Rev. 45:
6488-6519, 2016.
2) L. Pinto da Silva, A.N. Montenegro, C.M. Magalhães, P.J.O. Ferreira, D. Duarte, P.G. Berdullas, J.E.R. Borges,
N. Vale, J.C.G. Esteves da Silva, Single-molecule chemiluminescent photosensitizer for a self-activating andtumor-selective photodynamic therapy of cancer. Eur. J. Med. Chem. 132: 111683, 2019.
3) L. Pinto da Silva, C.M. Magalhães, A.N. Montenegro, P.J.O. Ferreira, D. Duarte, J.E.R. Borges, N. Vale, J.C.G.
Esteves da Silva. Biomolecules 9: 384, 2019.
Acknowledgements. This research was funded by the Portuguese Foundation for Science and Technology (FCT)
via the projects PTDC/QUI-QFI/2870 and UIDB/00081/2020.

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SP-7.1.5, P-7.1.5

A Fluorescent β-Cyclodextrins Polymer Encapsulating Sorafenib and Releasing
Nitric Oxide Under Visible Light
Francesca Laneri1, Milo Malanga2 and Salvatore Sortino1.
1) University of Catania, Department of Drug and Health Sciences, Catania, Italy; 2) CycloLab Ltd., Budapest,
Hungary
In the last years, the combination of cytotoxic radical species with conventional drugs has encountered growinginterest in cancer treatment, with the final goal to maximize the therapeutic action and to minimize side effects. Nitricoxide (NO) has particularly been investigated since, besides its well-known involvement in physiological and physiopathological pathways, it shows anticancer properties. Nonetheless, the delivery of this gaseous transient specie tothe targeted cells has resulted particularly challenging as tumor growth inhibition is granted at micromolar
concentrations, while at picomolar concentrations of NO tumor cells proliferation is increased.1 Light-triggered NO
donors allows the fine spatiotemporal modulation of the NO dosage and hold great potential in cancer treatment.2 In
this contribution we report a macromolecular construct made of a fluorescent, water-soluble cyclodextrins branchedpolymer releasing NO under visible light3 and encapsulating Sorafenib, a drug largely administered for the treatmentof liver and kidney cancer. The photophysical and photochemical properties of this supramolecular complex are
discussed and the preliminary biological tests on different cancer cell lines are also reported.

Fig. 1 – Schematic representation of the fluorescent polymer encapsulating Sorafenib and releasing NO under visible light

References
1) A. Fraix, N. Marino, S. Sortino, Phototherapeutic Release of Nitric Oxide with Engineered Nanoconstructs.
Top Curr Chem. 2016.
2) S. Sortino, Photoactivated nanomaterials for biomedical release applications. J Mater Chem. 2011.
3) M. Malanga, M. Seggio, V. Kirejev, A. Fraix, I. Di Bari, E. Fenyvesi, M.B. Ericson, S. Sortino, A
phototherapeutic fluorescent β-cyclodextrin branched polymer delivering nitric oxide. Biomater Sci. 2019.
Acknowledgements. We thank MIUR (Italy) for the Industrial PhD grant (DOT1308125) to F.L. and AIRC – Italian
Association for Cancer Research (IG-198559) for financial support.

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IL-7.1.6
New Azobenzene-based Photoswitches for the Two-Photon light-induced control ofneuronal signalling
Gisela Cabré1, Aida Garrido-Charles2, Mercè Izquierdo-Serra2, Marta Gascón-Moya1, Ramon Alibes1, Félix
Busqué1, Jordi Hernando1, Pau Gorostiza2,3,4
1) Departament de Química, Universitat Autònoma de Barcelona (UAB), Cerdanyola del Vallès, Spain; 2) Institut
de Bioenginyeria de Catalunya (IBEC), Barcelona Institute of Science and Technology (BIST), Spain; 3) Institució
Catalana de Recerca I Estudis Avançats (ICREA), Spain ; 4) Centro de Investigación Biomédica en Red en
Bioingenieria, Biomateriales y Nanomedicina (CIBER_BBN), Spain
Presenting author: Félix Busqué
Azobenzenes have been extensively studied as photopharmacological tools for the fast and reversible optical controlof biological systems, for example for the manipulation of neuronal signaling. However, most of these current
compounds operate under near-UV light irradiation, which is not compatible with many biomedical applications
since it can cause photodamage and is absorbed and scattered by tissues, thus preventing efficient penetration depths.1
It would be desirable to shift the absorbance of these compounds to the red and near-infrared (NIR) regions. In our
research groups we have been developing new azobenzene-based light-activable drugs that could exhibit efficient
two-photon (2P) absortion under near-infrared light excitation and could act as efficient photochromic tethered
ligands (PTLs). We have been focused in a family of azobenzene-based PTLs called MAG ligands, which arecomposed of a maleimide moiety for receptor tethering, the azobenzene photoswitch and a glutamate agonist.
R

a)
R=

CH3
N

O

N

O

b)
O

NH
N

O

1
O

O

N

N

NH
O

O

NH

F
O
N
H

O
HN

2

+

H3N

N

N

NH

H3N

COOH

O
HOOC

COOH

HOOC
Control of astrocytes and hippocampal rat raneurons with NIR light+

Control of hippocampal rat neurons and, in vivo,
in the C. elegans nematode with NIR light

Fig. 1 – Synthesized MAGs PTLs operative under 2P excitation with NIR light.

In particular we have applied this strategy to the regulation of several ionotropic glutamate receptors (iGluRs), themain responsible of excitatory currents in the central nervous system. Trans-cis photoisomerisation of thesecompounds allows modulation of glutamate-receptor interaction, thus resulting in light-induced operation of the cell
membrane ionic channels governed by iGluRs. We have developed new MAG switches capable to trigger iGluRs
upon 2-photon excitation with near infrared light following two main strategies: (i) the introduction in the compoundof a photosensitizer unit, with 2P absorption capacity, that can transfer its electronic excitation energy to theazobenzene core to trigger the isomerization (Figure 1a);2 and (ii) the use of electronically asymmetric azobenzene
switches, that leads to maximize its 2-photon response (Figure 1b).3
References

1) K. Hüll, J. Morstein, D. Trauner, In vivo Photopharmacology. Chem. Rev. 2018, 118, 10710-10747.
2) M. Izquierdo-Serra, M. Gascón-Moya, J. J. Hirtz, S. Pittolo, K, E. Poskanzer, E. Ferrer, R. Alibés, F. Busqué, R. Yuste, J.
Hernando, P. Gorostiza. Two-Photon Neuronal and Astrocytic Stimulation with Azobenzene-based Photoswitches. J. Am. Chem.
Soc. 2014, 136, 8693-8701.
3) G. Cabré, A. Garrido-Charles, M. Moreno, M. Bosch, M. Porta-de-la-Riva, M. Krieg, M. Gascón-Moya, N. Camarero, R.
Gelabert, J. M. Lluch, F. Busqué, J. Hernando, P Gorostiza, R. Alibés. Rationally designed azobenzene photoswitches for
efficient two-photon neuronal excitation. Nat. Commun. 2019, 10, 297

Acknowledgements. This work was supported by projects and grants from different institutions of Spain.
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-7.1.7

The potential of G protein-coupled receptor photopharmacology for the discovery of
new biological mechanisms
Xavier Rovira
1) Institute for Advanced Chemistry of Catalonia (IQAC-CSIC), Medicinal Chemistry & Synthesis group,
Barcelona, Spain;
By definition, biological mechanisms are the dynamic producers of physiological phenomena. However, when using
drugs in research, we introduce molecules into the biological system for which the action cannot be dynamicallycontrolled, thus hampering the possibility to study their effects in specific locations and time points. G proteincoupled receptors (GPCRs) are important regulators of many physiological processes targeted by around one thirdof approved drugs. Recent reports reveal that signal transduction by GPCRs is a multidimensional process governedby molecular, spatial and temporal components. Indeed, endogenous GPCR ligands are finely regulated by biologicalprocesses that allow their temporally controlled release and reuptake in specific locations, sometimes even at thesubcellular level. In contrast, one important limitation of current drugs is that once released in the organism, theireffect cannot be selectively interrupted. To solve these limitations, GPCR photopharmacology aims at controlling
native biological systems with a high degree of temporal and spatial precision using innovative light-regulated
molecules and illumination devices. In this presentation, I will talk about recent advances in the generation ofphotopharmacological tools targeting GPCRs and their potential to uncover new biological mechanisms that cannot
be studied with conventional drugs. The possibility to dynamically control signalling pathways activated by a single
GPCR with a unique component (light-regulated drugs) may open new avenues in cell communication research field
and inspire the development of precise and personalized medicines with high efficacy and reduced side-effects.

Fig. 1 – (A) The GPCR Photopharmacology principle. (B) Dynamic control of native neuronal receptors in primary cell cultureswith Alloswitch-1, a reversible light-regulated drug targeting metabotropic glutamate receptor 51. (C) Dynamic control of native
receptors in alive animals with chronic pain using reversible photopharmacological tools targeting GPCRs2.

References
1) Pittolo S. et al., An allosteric modulator to control endogenous G protein-coupled receptors with light. Nat Chem
Biol 10(10):813-5, 2014.
2) Zussy C., et al. Dynamic modulation of inflammatory pain-related affective and sensory symptoms by opticalcontrol of amygdala metabotropic glutamate receptor 4. Mol Psychiatry 23(3):509-520, 2018.
Acknowledgements. This work was supported by the Spanish Ministry of Economy, Industry and Competitiveness(SAF2015-74132-JIN).
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-7.1.8
Opto-pharmacological Approaches for Manipulating Neurotransmitter Receptors
and Motivated Behaviors in Mice
Alexandre Mourot1
1) Brain Plasticity lab, UMR 8249 CNRS, ESPCI, Paris, France.
One fundamental goal of modern neuropharmacology is to causally link changes in receptor activity in given neuronal
pathways with alterations of circuit physiology and ultimately of behaviour. The utility of classical pharmacology invivo in achieving this goal is limited, because local drug delivery is slow, imprecise and hardly compatible withelectrophysiology. In contrast, opto-pharmacological approaches for remote-controlling neurotransmitter receptors
with light are, in principle, able to mimic the timing, amplitude and spread of naturally occurring modulatory signals(1). My lab is developing opto-pharmacological tools to study the role of the midbrain dopamine system in the controlof reward-guided behaviors and in the development of addiction. We notably developed novel caged-compounds thatare, for the first time, fully compatible with in vivo use in mice, and demonstrated optical induction of Pavlovian
conditioning upon optical stimulation of the midbrain dopamine system (2). We further have engineered lightcontrollable nicotinic acetylcholine receptors, and deployed these tools in vivo to reveal the impact of acetylcholine,
an endogenous neuromodulator, on the activity of midbrain dopamine neurons, and to manipulate nicotine-related
addictive behaviors (3). By providing acute and precise manipulation of neurotransmission, our optopharmacological approaches will lead to a better comprehension of how specific receptors contribute to themodulation of circuits and behaviors, a crucial step towards the development of novel therapies.

References
1) Paoletti P, Ellis-Davies GCR, Mourot A., Optical control of neuronal ion channels and receptors. Nat Rev
Neurosci. 20(9):514-532, 2019.
2) Durand-de Cuttoli R, Chauhan PS, Pétriz Reyes A, Faure P, Mourot A*, Ellis-Davies GCR* (co-last). Optofluidic
control of rodent learning using cloaked caged glutamate. Proc Natl Acad Sci U S A. 117(12):6831-6835. 2020
3) Durand-de Cuttoli R, Mondoloni S, Marti F, Lemoine D, Nguyen C, Naudé J, d'Izarny-Gargas T, Pons S, Maskos
U, Trauner D, Kramer RH, Faure P, Mourot A. Manipulating midbrain dopamine neurons and reward-related
behaviors with light-controllable nicotinic acetylcholine receptors. Elife. 7:e37487. doi: 10.7554/eLife.37487. 2018.
Acknowledgements. This work was supported by the Agence Nationale de la Recherche (ANR), the Brain and
Behavior Research Foundation (NARSAD) and the Human Frontier Science Program (HFSP).

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-7.1.9

Intrafollicular UVA-induced drug release from nanocapsules – a photochemical
approach for enhanced dermal drug delivery
Loris Busch1,4, Yuri Avlasevich2, Paula Zwicker3, Maxim E. Darvin1, Katharina Landfester2, Martina C. Meinke1,
Cornelia M. Keck4, Axel Kramer3, Jürgen Lademann1, Alexa Patzelt1
1) Center of Experimental and Applied Cutaneous Physiology, Department of Dermatology, Venereology andAllergology, Charité – Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, HumboldtUniversität zu Berlin, and Berlin Institute of Health, Berlin, Germany; 2) Max Planck Institute for Polymer
Research, Mainz, Germany; 3) Institute of Hygiene and Environmental Medicine, University Medicine Greifswald,
Greifswald, Germany; 4) Department of Pharmaceutics and Biopharmaceutics, Philipps University Marburg,
Marburg, Germany
The human hair follicle is known to be an important drug delivery target for the therapy of various skin diseases aswell as for skin antisepsis. Nanoparticles of approximately 600 nm in diameter are excellently suited for folliculardrug delivery as they show deep follicular penetration when applicated by massage.1 This phenomenon is based ona mechanically driven transport process, also known as the so-called ratchet effect.2 Following the penetration
process, an intrafollicular drug release can be initiated by various trigger mechanisms depending on thefunctionalization of the nanoparticulate system.3
Since UVA light can reach the connective tissue layers of the skin, and thus deeper sections of the hair follicles, it
is usable for triggering the release of drugs from photodegradable nanocapsules (NCs) within the follicularcompartment.
In the framework of our experiments, we have established a novel system, which involves the transportation of themodel drug sulforhodamine 101 (SR101) into hair follicles by NCs of 600 – 700 nm in size. These NCs werefunctionalized by a photocleavable o-nitrobenzyl linker. Thus, we were able to induce an intrafollicular UVAtriggered drug release by using a light emitting diode (LED).4 UVA radiation with a power density of 12 mW/cm2
induced a drug release of over 50% after 2 min as investigated in vitro via fluorescence spectroscopy. We further
investigated follicular penetration as well as intrafollicular drug release on ex vivo porcine skin by using confocal
laser scanning microscopy. As schematically depicted in Fig. 1, UVA-induced cleavage of the NCs along withrelease of SR101 at a mean follicular penetration depth of 509 ± 104 µm was observed after topical application andsubsequent irradiation of the skin. Furthermore, sufficient cell viability after exposition of HaCaT keratinocytes tointact and photodegraded NCs was revealed by MTT tests. The application of the presented system represents apromising approach for an optimized follicular drug delivery of various hydrophilic drugs. This might constitute a
future benefit for the therapy of miscellaneous skin diseases or could be used for optimized preoperative skinantisepsis.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

Fig. 1 – Schematic representation of the application and irradiation process on ex vivo porcine skin (top) as well as thefollicular penetration of SR101-loaded NCs (bottom left) and subsequent UVA-induced photodegradation of the NCs alongwith release of SR101 (bottom right) [4].

References
1) Patzelt A, Richter H, Knorr F, Schäfer U, Lehr CM, Dähne L, Sterry W, Lademann J, Selective folliculartargeting by modification of the particle sizes. J Control Release 150(1):45-8, 2011.
2) Radtke M, Patzelt A, Knorr F, Lademann J, Netz RR, Ratchet effect for nanoparticle transport in hair follicles.
Eur J Pharm Biopharm 116:125-130, 2017.
3) Patzelt A, Mak WC, Jung S, Knorr F, Meinke MC, Richter H, Rühl E, Cheung KY, Tran NBNN, Lademann J,
Do nanoparticles have a future in dermal drug delivery? J Control Release 246:174-182, 2017.
4) Busch L, Avlasevich Y, Zwicker P, Thiede G, Landfester K, Keck CM, Meinke MC, Darvin ME, Kramer A,
Müller G, Kerscher M, Lademann J, Patzelt A, Release of the model drug SR101 from polyurethane nanocapsulesin porcine hair follicles triggered by LED-derived low dose UVA light. Int J Pharm 597:120339, 2021.
Acknowledgments. This study was funded by the German Federal Ministry of Education and Research (BMBF)
within the “Advanced UV for Life” project “NanUVsept” [grant number 03ZZ0142A].

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P-7.1.10

Exploring Photoactive Pigments of Fungal Extracts with Feature-Based Molecular
Networking
Fabian Hammerle1, Johannes Fiala1, Luis Quirós-Guerrero2,3, Adriano Rutz2,3, Jean-Luc Wolfender2,3, Ursula
Peintner4, Bianka Siewert1
1) University of Innsbruck, Institute of Pharmacy/Pharmacognosy and Center for Molecular Biosciences Innsbruck(CMBI), Innrain 80/82, 6020 Innsbruck, Austria; 2) University of Geneva, Phytochemistry and Bioactive Natural
Products, School of Pharmaceutical Sciences, CMU - Rue Michel-Servet 1, 1211 Geneva, Switzerland; 3)
University of Geneva, Institute of Pharmaceutical Sciences of Western Switzerland, CMU, 1211 Geneva,
Switzerland; 4) University of Innsbruck, Institute of Microbiology, Technikerstraße 25, 6020 Innsbruck, Austria
Investigating the colourants of fungal fruiting bodies has led to the discovery of an activity previously unexpected
for Basidiomycetes: photoactivity [1]. Several antraquinone pigments were isolated from Cortinarius uliginosus thatwere capable of generating reactive oxygen species (i.e., singlet oxygen) after irradiation with visible light [2]. To
test whether photoactivity is a common phenomenon in the globally distributed, species-rich mushroom genus
Cortinarius [3], six colourful species belonging to the different classical subgenera Dermocybe, Leprocybe,
Myxacium, Phlegmacium, and Telamonia were selected for mycochemical analysis. Acetone extracts of fungal
basidiomes were submitted to a special workflow combining in vitro photochemical/photobiological experiments [1],ultra-high performance liquid chromatography coupled to high-resolution tandem mass spectrometry (UHPLCHRMS2), and feature-based molecular networking (FBMN) [4].
Two of the six species examined, i.e., C. rubrophyllus (Dermocybe) and C. xanthophyllus (Phlegmacium), were not
only characterised by high singlet oxygen formation values in the photochemical assay (i.e., DMA-assay), but also
exhibited light-dependent cytotoxicity in the low µg/mL-range on various cancer cell lines (i.e., A549/lung,
AGS/stomach, T24/bladder). With the help of the bioinformatics tool FBMN, the compounds responsible for theobserved photoactivity were identified as unknown polyketides. Comparative liquid chromatographic studies
indicated that they are most likely dimeric anthraquinones.
The results of this study highlight the occurrence of various photoactive pigments in Cortinarius species belonging
to different phylogenetic lineages, and thus fungi as a potential source of natural photopharmaceuticals. Furthermore,
an innovative metabolomics-based workflow is presented to screen natural extracts for photoactivity and to
subsequently identify the responsible photosensitizers.
References
1) Siewert, B., et al., A convenient workflow to spot photosensitizers revealed photo-activity in basidiomycetes. RSC
Advances, 2019.
2) Fabian, H., et al., Targeted Isolation of Photoactive Pigments from Mushrooms Yielded a Highly Potent New
Photosensitizer: 7,7’-Biphyscion. ChemRxiv (Preprint), 2021.
3) Peintner, U., J.-M. Moncalvo, and R. Vilgalys, Toward a better understanding of the infrageneric relationships in
Cortinarius (Agaricales, Basidiomycota). Mycologia, 2004.
4) Nothias, L.-F., et al., Feature-based molecular networking in the GNPS analysis environment. Nature Methods,
2020.
Acknowledgements. The Austrian Science Fund (FWF P31950, BS) and the University of Innsbruck are
acknowledged for support. L. Q-G is supported by a scholarship (N_ 214171–025) Ministerio de Ciencia, Tecnología
y Telecomunicaciones, MICITT, from Costa Rica.

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P-7.1.11

Melanopsin signalling pathway in HEK293 cells line with stable expression of humanmelanopsin
Olga Krzysztyska-Kuleta, Magdalena Olchawa, Tadeusz Sarna
Department of Biophysics, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University,
Krakow, Poland
Introduction Melanopsin, a member of the G-protein coupled receptors family, is involved in non-image-forming
functions including circadian rhythm, sleep regulation, pupil response. Although significant efforts research havebeen devoted to different cell subtypes and their behavioural responses to light activation, signalling cascade
involving melanopsin photoactivation is still poorly characterized. In this study, we analysed the effects ofphotoactivation of melanopsin on phospholipase C (PLC) and diacylglycerol in HEK293 cells with stable
expression of human melanopsin.
Methods HEK 293 cells with stable expression of human melanopsin were enriched with 11-cis-retinal and blue
light irradiated for selected time intervals. To determine the role of phospholipase C and involvement of
diacylglycerols, two approaches were employed: inhibition of the G protein and phospholipase C (using the BIM46187 and U73122 inhibitors, respectively), and gene silencing using siRNA of PLCβ1 and PLCβ4.
Results Our study showed that diacylglycerol was present in blue light irradiated HEK293 cells enriched with 11cis-retinal. Using specific inhibitors or small interfering RNA (siRNA), we found that PLCβ4 was involved inmelanopsin phototransduction pathway. Completely diminished response of diacylglycerol and calcium ions in
irradiated cells expressing human melanopsin was observed after silencing the PLCβ4 gene.
Conclusion It has been demonstrated, for the first time that diacylglycerols can be involved in melanopsin
signalling pathway. The data also indicate that PLCβ4 plays the dominant role in melanopsin transduction pathway.
These results may facilitate a better understanding of the role of phospholipase C and diacylglycerols in themelanopsin signalling pathway.

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P-7.1.12

A photoactivatable β- and γ- cyclodextrin branched co-polymer delivering nitric oxide.
Mimimorena Seggio1, Szabolcs Béni,2 Milo Malanga3, Salvatore Sortino.1
1) University of Catania, Department of Drug and Health Sciences, Catania, Italy; 2) Semmelweis University,
Department of Pharmacognosy, Budapest, Hungary 3) CycloLab Ltd., Budapest, Hungary.
Polymer based on cyclodextrins (CDs) are receiving a growing interest in the biopharmaceutical field thanks to theirwell-known complexation capability and their ability to stabilize and solubilize guest compounds. This contribution
reports the synthesis and characterization of a novel β- and -CD-based branched co-polymer covalently integrating
a nitric oxide photodonor (NOPD) and able to supramolecularly encapsulate betaxolol (BTX), a drug widely used inglaucoma treatment. (Fig 1)

Fig. 1 – Schematic representation for the functioning of the cyclodextrin-based copolymer.

The system presents remarkable advantages over the isolated photoactivatable units not linked to the CDs scaffold.
It shows excellent water solubility, good stability in the dark at room temperature and the capability to release abiologically relevant amount of NO under the control of visible light. In view of the well-known vasodilator
properties of NO,1,2 the present work may open intriguing prospects for biological studies on formulations for ocularapplication against glaucoma, addressed to explore the combinatory effect of BTX and NO rapidly released uponenvironmental light. These studies are currently underway in our laboratories.
References
1) Nitric Oxide: Biology and Pathobiology, (ed. L. J. Ignarro), Elsevier Inc, 2010.
2) S. Sortino, Light- Responsive Nanostructured Systems for Applications in Nanomedicine. Springer, 2016.
Acknowledgements. We thank the MIUR (Italy) for the Industrial PhD grant (DOT-1308125-1) to M.S and AIRC
– Italian Association for Cancer Research (IG-198559) for financial support.

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IL-7.2.2
Worldwide prevalence and incidence of the photodermatoses
Burfield L1 Rutter KJ1,2, Neale RE3, Rhodes LE1,2.
1) Centre for Dermatology Research, School of Biological Sciences, University of Manchester, Manchester, UK;
2) Photobiology Unit, Dermatology Centre, Salford Royal NHS Foundation Trust, Manchester, UK;
3) Population Health Department, QIMR Berghofer Medical Research Institute, Queensland, Australia.

The photodermatoses are a diverse group of skin diseases caused or exacerbated by solar radiation.
Despite their significant impact on quality of life, psychosocial welfare and employment1, little is known
regarding their worldwide prevalence and incidence.
This study aimed to systematically review the literature to establish what is known regarding prevalence
and incidence of the photodermatoses in adult and paediatric populations. Searches of three electronic
medical databases (Medline, CINAHL, Embase) were performed to identify original studies reporting theprevalence and/or incidence of the photodermatoses. Publications that did not include data on the
numerator and denominator populations were not eligible for inclusion.
Twenty-six studies were identifed and included: 15 reported the prevalence of photodermatoses based onstudies of samples of the general population, and 11 reported on their prevalence and/or incidence fromnational or international registry/diagnostic laboratory data. The general population studies concerned
polymorphic light eruption (PLE; 9 studies), unspecified photosensitivity (2 studies), actinic prurigo (2studies), juvenile spring eruption (1 study) and variegate porphyria (1 study), while registry studies
reported on cutaneous porphyrias (8 studies) and selected genophotodermatoses (2 studies).
While population-based studies were scarce for most individual diseases, they suggest a high prevalenceof photodermatoses at wide-ranging world locations. This includes a large paediatric study in South Sinai
reporting unspecified photosensitivity in 4% of children. The prevalence of PLE ranged from 0.65% to21.4%, with a strong positive correlation (r=0.776) between prevalence and distance from the equator.
Studies of actinic prurigo in Chimila Indians and Inuit populations showed this was prevalent in bothgroups (8% and 2% respectively), while no studies were available in other ethnic groups. Heterogeneityof study design and lack of standardised diagnostic criteria hampered comparison between studies.
Overall, study quality was assessed as high (3 studies), medium (11 studies) and low (2 studies).
Published registry data from 12 European countries demonstrated porphyria cutanea tarda is the mostprevalent porphyria in Europe. Combined data from DNA repair diagnostic centres in 5 European
countries gave incidence rates for xeroderma pigmentosum, Cockayne syndrome and trichothiodystrohyof 2.3, 2.7 and 1.2 per million births, respectively.
Crucially, we found prevalence and/or incidence studies are lacking for many of the photosensitivity
diseases. Further high-quality population-based studies, employing standardised diagnostic criteria, arerequired to better understand the disease burden of the photodermatoses.
References

1) Rutter KJ, Ashraf I, Cordingley L, Rhodes LE. Quailty of life and psychological impact in thephotodermatoses: a systematic review. Br J Dermatol. 2020; 182: 1092-102.
Acknowledgements. KR and LER acknowledge the NIHR Manchester Biomedical Research Centre.

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IL-7.2.3

Consensus in Photodiagnostic Services in the UK and Republic of Ireland
Sally Ibbotson on behalf of the British Photodermatology Group Working Party
Photobiology Unit, University of Dundee, Ninewells Hospital & Medical School, Dundee, UK
Abnormal photosensitivity can have a major adverse effect on patients’ lives and prompt, accurate diagnosis is theunderlying basis of optimal management. Whilst photodiagnostic investigations are not required in all cases, such as
in straightforward polymorphic light eruption, they are invaluable for patients with severe or atypical disease as it isoften impossible to confirm abnormal photosensitivity on clinical grounds alone. Photodiagnostic investigations
should therefore be available for patients with suspected photosensitivity diseases. However, the requirements for
specific equipment, expertise and techniques dictates that such facilities are only available through tertiary specialist
photobiology units.
The last review of Photodiagnostic Services in the UK was almost 30 years ago and therefore, through a British
Photodermatology Group/British Association of Dermatologists (BPG/BAD) Workshop we critically appraised
photodiagnostic techniques in the UK and Republic of Ireland, seeking to establish areas of consensus practice,acceptable variation and priorities for service, research and educational development. The Workshop findings basedon the 12 photodiagnostic units reviewed, reinforced the importance of availability of expertise and specialised staff,
with emphasis on the consultant-led multidisciplinary team, with dedicated photophysics input. Specialised
equipment is essential for the detailed investigation of patients with suspected photosensitivity disorders and further
clinical studies are required to assist with development of commercial optical radiation phototesting equipment torequired standards. Narrow waveband phototesting, particularly monochromator phototesting, is the Gold Standard
investigation with respect to establishing the action spectrum and degree of photosensitivity and assessing treatment
response and natural history of photosensitivity. However, this is often also supplemented by broadband phototesting
to confirm objective photosensitivity. Prioritisation of defining normal range phototesting erythemal responses(defined as the threshold minimal erythema dose; MED) in patients of skin phototype IV-VI was emphasised as thesedata are currently lacking. Most photodiagnostic units offer iterative photoprovocation testing but methodologies andassessment of reaction patterns are not standardised and this was identified as a key area for further study. Minimal
erythema dose (MED) testing to phototherapeutic sources (eg. NBUVB), patch testing and photopatch testing todefine coexistent allergens and photoallergens and other investigations to exclude less common causes ofphotosensitivity, such as lupus, cutaneous porphyrias and genophotodermatoses, should be available to allphotodiagnostic services. We identified a relative lack of defined training programmes for photophysicists, clinicalscientists and allied health care professionals in the field of photodiagnostics and identified this as a priority area foreducational and training development moving forward. The information obtained through this review assists with thedevelopment of minimum standards for photodiagnostic services and emphasises the importance of collaboration,
data sharing and multidisciplinary working to facilitate deep phenotyping and improved understanding of thephotosensitivity diseases.

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OC-7.2.4

Automated real-time monitoring of intracellular protoporphyrin IX synthesis in alive-cell plate reader
Christian Oberdanner1, Nicole Eggenhofer1, Kristjan Plaetzer2
1) Tecan Austria GmbH, Grödig, Austria; 2) Laboratory of Photodynamic Inactivation of Microorganisms,
University of Salzburg, Salzburg, Austria
Over the last couple of years the importance of 5-Aminolevulinic acid (5-ALA)-based protoporphyrin IX (PPIX)PDD and PDT has not decreased. However, the dynamics of intracellular PPIX formation after ALA application ishighly critical for a successful diagnosis and treatment (1). Therefore, time-resolved in vitro measurements of theintracellular PPIX formation represent the bedrock for identification of further target diseases or testing novel ALA
derivates.
This presentation points out the convincing advantages of automation of such analysis. A431 human epidermoid
carcinoma cells were stably transfected with green fluorescent protein (GFP) and automatically seeded into 96- or
384-well microplates using the dispensing unit of a Tecan live cell multimode reader. Automated determination of
the GFP fluorescence revealed excellent cell growth in the atmospheric chamber of the device and provided
complete data sets without night gaps. Standardized replacement of the growth medium after overnight incubation
with a microplate washer minimized the experimental error of this experimental step.
After addition of ALA the intracellular PPIX synthesis was automatically recorded by the fluorescence of thiscompound, again gaining advantage from the reader's atmospheric control features. The results demonstrate that thepeak intracellular PPIX-level, which represents the optimal point in time for tumour therapy, is a complex functionof the incubation concentration and -period. They furthermore provide clear evidence for the benefits of automation
for investigation of multi-parametric cellular processes: (i) high-resolution complete data sets, (ii) low experimental
error, (iii) high costs-effectiveness due to employment of 384 well microplates and (iv) minimized time
consumption.
1) Malik, Z. Fundamentals of 5-aminolevulinic acid photodynamic therapy and diagnosis: An overview. Transl.
Biophotonics 2020, 2, e201900022

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IL-7.2.5
Pathways of polymorphic light eruption: from potential triggers to immune responsePeter Wolf
Medical University of Graz, Department of Dermatology, Research Unit for Photodermatology, Graz, Austria
Polymorphic light eruption (PLE) is the most common and prevalent photodermatosis, particularly among young
women in temperate climates. We recently reported that the onset of disease usually occurred between young
adulthood and middle age, at a mean age of 25.9 years in females and 28.1 years in males. Moreover, we observedthat in most cases (74% of females and 65% of males), the onset of PLE symptoms occurred between the ages of
15 and 40 years.1 Our recent registry analysis indicated that though the disease improved in a substantial number ofpatients (i.e., 77% of females and 59% of males) over the years, it most often takes a long-term course.1 It took 20(95%CI, 13-26) years until 25% of patients had normalized from PLE and it took 25 (95%CI, 18-41) years until
one third of patients had normalized from PLE.
Recently, the pathophysiology of PLE has become much better understood. This includes concurrent resistanceagainst induction of UV-induced immune suppression, linked to an imbalanced micromilieu marked by low levelsof IL-4, IL-10, and TNF-alpha; failure of Langerhans cell emigration from the skin and neutrophilic infiltration into
the skin; disturbances in Treg levels and function; and potential involvement of CD11b/IL-31+ cells, and mast
cells. Also better understood now are the therapeutic mechanisms of photohardening and other preventive
measures.
Recently, as in many other conditions the skin’s microbiome came into the focus of research in PLE. For
instance, we have observed that the skin’s microbiota can protect against UV-induced immune suppression. Likeother investigators we have also seen abnormalities in the expression of antimicrobial peptides (AMP) in the skinof PLE. We recently hypothesized that a potential link between disturbances in the microbiome and UV-induced
immune suppression may play a role in the initiation and pathophysiology of the disease and are currently testing
this hypothesis in an ongoing experimental study.2 Indeed, the quality and/or quantity of the microbiota present on
human skin does vary during different periods of life and age-dependency may be linked sex hormones.
References
1. Alexandra Gruber-Wackernagel, Tanja Schug, Thomas Graier, Franz J. Legat, Hanna Rinner, Angelika Hofer,
Franz Quehenberger, Peter Wolf. Long-term course of polymorphic light eruption: a registry analysis. Frontiers in
Medicine 2021, doi: 10.3389/fmed.2021.694281
2. The significance of the skin microbiome n the pathophysiology of polymorphic light eruption(PLE_microbiome), 2021, https://clinicaltrials.gov
Acknowledgements
This work was supported by the Austrian Science Fund FWF (W1241).
Correspondence to: peter.wolf@medunigraz.at

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IL-7.2.6
The Psychological basis of photoprotective behaviour in X.P.: implications for
improving photoprotection in other patient groups
Sarkany RPE1, Norton S 3, Canfield M 3, Morgan M 2, Foster L 1, Johnstone, K2, Sainsbury K4, Araujo-Soares V. 4,
Wulf H-C 5, Weinman J 2, Walburn J 2
1. Xeroderma Pigmentosum Unit, Guys and St Thomas’ NHS Foundation Trust; 2.School of Cancer andPharmaceutical Sciences, King’s College London, UK; 3.Health Psychology Section, Institute of Psychiatry,
Psychology & Neuroscience, King’s College London, UK; 4.Institute of Health & Society, Faculty of Medical
Sciences, Newcastle University,; 5.Dermatology Department, Bispjeberg Hospital, Copenhagen
Photoprotection is central to the management of patients with photosensitive skin diseases and those at high risk ofskin cancer. It is also crucial to Public Health planning since it is the only way to reduce population skin cancer rates.
Photoprotection involves a group of behaviours all of which reduce UV exposure. These include the use of sunscreensand photoprotective clothing and hats and the use of shade, as well as adaptation of the timing and duration of goingoutdoors.
The results of previous educational and other interventions to improve photoprotection have generally been
disappointing. It has recently been shown that, even after a diagnosis of malignant melanoma, patients improve theirphotoprotection for an average of only 18-24 months before reverting to their previous behaviour.
We have recently studied 36 patients with Xeroderma Pigmentosum. In this disease, stringent photoprotection is
critical to health and life expectancy. We measured the daily dose of UV to the skin on the face and identified a huge
range of UV exposure in these patients, from very low up to levels similar to those in healthy individuals. We thensucceeded in identifying the factors creating these differences in photoprotection behaviour. We found thatpsychological and social factors are responsible for at least as much of the variation between patients as can be
attributed to demographic or clinical differences. Most of the psychological factors which we have identified ascorrelating with poor photoprotection are a group of factors including patients’ beliefs and perceptions about XP and
its treatment, which are known to contribute to poor treatment compliance in other diseases. The way in which
patients accommodate the disease into their ‘identity’ has turned out to play a crucial role in determining how well
our XP patients photoprotect. We went on to use these findings to design a Behaviour Change Intervention(‘XPAND’) to specifically address these psychosocial factors, and our controlled clinical trial has shown that theIntervention is effective at improving photoprotection behaviour.
This systematic approach to identifying and ‘treating’ the underlying psychological causes of photoprotection iseffective in this rare disease. This raises the possibility that this approach may succeed where previous attempts toimprove photoprotection in other groups have failed, and we are planning to move on to apply this approach to othergroups of patients, especially those at high risk of skin cancer.
Acknowledgements. This work was supported by a grant from the National Institute for Health Research.

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IL-7.2.7
Sun exposure and protection guidance following COVID-19 lockdowns – considerations
for the public & photosensitive patients
Yolanda Gilaberte
Miguel Servet University Hospital, Department of Dermatology,IIS Aragón, Zaragoza, Spain.
Our relationship with the environment has been affected by the COVID-19 pandemic, especially in terms of spendingtime outdoors. The lookdown has created a situation in which most people have had less exposure to sunlight thanusual. Hence, cutaneous synthesis of vitamin D in the skin, which is our greatest source of vitamin D, has beenhampered by the lockdown, during which many people have not been exposed to sunlight in several weeks. Low
vitamin D levels have been demonstrated to be associated to a more severe COVID-19 illness.
On the other hand, our group has found and association between UV index and the incidence of COVID-19 infection
in cities along the whole latitude of Spain.
Taking all of this into consideration, a healthy exposure is achieved by ensuring that the dose of UV radiation is
sufficient to obtain the positive effects of sunlight on our body minimizing the risk of damage resulting fromoverexposure. In this sense, UVI values can be used to plan the activities outdoors.
The Spanish Group of Photodermatology have made some recommendations in order to get a safe exposure to thesunlight in the context of the COVID-19 pandemic. Photoprotective measures are recommend when the incidence of
solar UV is high, since the doses of UV necessary to produce vitamin D are low and can be reached without getting
erythema. Gradual exposure to sunlight is recommended in order to facilitate adaptation of the skin and favour natural
defence mechanisms. Use of shade, clothes, hats and sunscreens are recommended as usual. Some patients withphotodermatoses have suffered less intense clinical manifestations during the pandemic due to the recommendationsof stay at home and the use of face mask which provide a very good photoprotection. However, their photoprotective
measures should be very high when they go out. Patients who had COVID-19 receiving medication should takespecial care and ask about the potential risk of photosensitivity.
References
1) Aguilera J, de Galver MV, Aguilera P, de Troya-Martín M, Gilaberte Y, del Grupo Español de Fotobiología de la
AEDV. Recommendations on sun exposure and photoprotection following easing of the COVID-19 pandemic
lockdown: Spanish Photobiology Group of the Spanish Academy of Dermatology and Venereology. Actas
Dermosifiliogr 2020;111:799-801.

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OC-7.2.8
Solar Urticaria-real life data of an international support group
Teresa Hipfl, Donja Homayoon, Angelika Hofer
Medical University of Graz, Department of Dermatology and Venerology, Graz, Austria;
Solar urticaria belongs to the idiopathic photodermatoses and is triggered by diverse wavelengths of UV-light orvisible light. Within minutes exposure to sunlight leads to itchy redness and wheals. Therefore, the disease has asignificant impact on the patient’s normal daily life. 1,2This paper aims to provide inside into the patient cohort of an international urticaria solaris support group (n >300
people), focusing on their medical care, as well as on the aspects of quality of life limitations.
A comprehensive questionnaire was provided online to all members of the urticaria solaris support group. Questions
were asked about the spectrum of therapies, their effects and side effects and the impact on quality of life due to thedisease. Of 149 participants who had completed the questionnaire 57 members (51 women, 6 men; living in 12
different countries) who reported that the diagnosis urticaria solaris had been confirmed by a doctor, were includedin this study.
The most commonly prescribed medications were antihistamines (47 out of 56 patients; 83.9%). However, only
12.8% of these patients reported a good effect with this treatment. Better improvement was seen with light therapy
or treatment with the monoclonal antibody omalizumab, good results in 37.5% versus 42.9%, respectively. However,treatment with omalizumab was only used in 14 out of 55 patients (25.5%) from 5 different countries (UK, US, IL,
CA, AT).
Most patients (77.8%) reported that they felt medically undersupplied because the symptoms could not be sufficiently
improved under therapy so far. Besides, the majority of the affected persons (68.7%) stated a severely impaired
quality of life despite therapy. Nearly 70% of the patients had the impression that their disease was not sufficiently
well known in society and that there was therefore no understanding of the severe impairment of quality of life.
However, being a member of the urticaria solaris support group, all affected feel that they are taken seriously. Whilenearly half of those affected would welcome psychological support in dealing with the disease, less than a quarter ofthese people have actually received psychological help.
In conclusion, solar urticaria patients suffer greatly from this disease. The quality of life is markedly reduced andmost of the patients are unsatisfied with their current therapy due to lack of efficacy. Better access to effective
treatment options and also to psychological care should be achieved. Especially in rare diseases such as urticaria
solaris, international networking to improve treatment strategies is urgently needed.
References
1) Harris BW, Badri T, Schlessinger J. Solar Urticaria. StatPearls. Treasure Island (FL): StatPearls Publishing,
20202) Levi A, Enk CD, Snast I. Solar Urticaria. Current Dermatology Reports, 2019

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P-7.2.9

In vitro photo(geno)toxicity assessment of gefitinib and its metabolites.
Meryem El Ouardi1,2, Lorena Tamarit1,2, Ignacio Vayá1,2, M.A Miranda1,2, Inmaculada Andreu1,2.
2) Unidad Mixta de Investigación Universitat
1) Departamento de Química/Instituto de Tecnología
Politècnica de València-Instituto de Investigación
Química UPV-CSIC, Universitat Politècnica deSanitaria (IIS) La Fe, Hospital Universitari i
València, Camino de Vera s/n, 46022 València, Spain.
Politècnic 106, 46026, Valencia, Spain.
Gefitinib (GFT) is an epidermal growth factor receptor (EGFR) inhibitor extensively used in patients with locallyadvanced or metastatic non-small cell lung cancer (NSCLC) 1. In general, GFT is a well-tolerated drug, however, aconsiderable number of patients have experienced dermatological side effects, being skin rash the most common
reaction (63% of patients)2. Most of EGFR-TKIs display similar adverse reactions although not fully studied.
Interestingly, drugs containing the quinazoline moiety are known to produce skin photosensitivity. In this context, it
has been recently reported that lapatinib, another tyrosine kinase inhibitor, can induce phototoxic disorders.3.
In addition, the pharmacokinetics of gefitinib suggests a clearance mainly through hepatic metabolism, where ODesmethyl gefitinib (DMT-GFT) is the major metabolite identified in human plasma 4. Another two compounds wereformed through Phase I metabolism in human liver microsomes, the oxidative defluorination (4-Defluoro-4-hydroxy
gefitinib, DF-GFT) and the loss of morpholine ring (O-Desmorpholinopropyl gefitinib, DMOR-GFT) 5,6. Despite theaim of metabolism is to improve the ease with which substances can be excreted by changing the molecular structure,
in some cases, this leads to the production of new compounds with higher toxicity. Based on that and the in vitro
phototoxicity results performed by AstraZeneca for Iressa® that have demonstrated a possible phototoxicity effectof this drug, it would be interesting to approach this issue by performing in vitro assays in human skin cells with GFT
and its main metabolites.
R2
Ri = OH

R3

R1

Fig. 1 – Gefitinib, GFT; 4-Defluoro-4-hydroxy gefitinib, DF-GFT (R1); O-Desmethyl gefitinib, DMT-GFT (R2) and ODesmorpholinopropyl gefitinib, DMOR-GFT (R3)

Thus, it is important to study the phototoxic potential of these compounds and to compare the behavior of themetabolites to GFT. For this purpose, a Neutral Red Uptake assay (NRU) was performed in human keratinocytes(HaCaT) for both GFT and its metabolites. The dose-response curves displayed in Fig.2 revealed a significant
phototoxic effect for both GFT and DMOR-GFT. Remarkably, this metabolite showed higher phototoxic potential
than the parent drug, whereas DMT-GFT exhibited lesser phototoxicity than GFT. In contrast, DF-GFT did notdisplay any phototoxic results. To understand the underlying photosensitizing mechanism of GFT and its metabolites
it has been evaluated the lipid peroxidation, protein photooxidation, and cellular DNA damage. As shown in Fig.2,
alkaline comet assay for both GFT and DMT-GFT displayed a considerable photogenotoxic effect, whereas DMORGFT did not trigger significant DNA damage. Contrary, DMOR-GFT showed a huge enhancement in both protein
and lipidic photooxidation. Moreover, GFT also promoted a substantial protein photooxidation, however, DMT-GFT
was observed to damage neither proteins nor lipids. The photosafety evaluation of pharmaceuticals should be
performed not only on the drug but also on its main metabolites, taking into consideration that the phototoxic
properties could be enhanced after drug metabolism as demonstrated in this study.

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P-7.2.9
Darkcell viability (%)

100

UVA Light

50

0
0

1
2
log [GFT] (M)

3Fig. 2 – From left to right: Intracellular colocalization of GFT in HaCaT cells by confocal microscopy, HaCaT Neutral Red
Uptake assay dose-response curves for GFT, Alkaline Comet assay of HaCaT cells treated with GFT.

References
1) Ghafoor Q, Baijal S, Taniere P, O'Sullivan B, Evans M, Middleton G. Epidermal Growth Factor Receptor(EGFR) Kinase Inhibitors and Non-Small Cell Lung Cancer (NSCLC) - Advances in Molecular Diagnostic
Techniques to Facilitate Targeted Therapy. Pathol Oncol Res. 24(4):723-731, 2018.
2) Annunziata MC, Ferrillo M, Cinelli E, Panariello L, Rocco D, Fabbrocini G. Retrospective Analysis of Skin
Toxicity in Patients under Anti-EGFR Tyrosine Kinase Inhibitors: Our Experience in Lung Cancer. Open Access
Maced J Med Sci. 7(6):973-977, 2019.
3) García-Lainez G, Vayá I, Marín MP, Miranda MA, Andreu I. In vitro assessment of the photo(geno)toxicity
associated with Lapatinib, a Tyrosine Kinase inhibitor. Arch Toxicol. 95(1):169-178, 2021
4) Frampton JE, Easthope SE. Gefitinib: a review of its use in the management of advanced non-small-cell lungcancer. Drugs. 64(21):2475-2492, 2004.
5) McKillop D, McCormick AD, Miles GS, et al. In vitro metabolism of gefitinib in human liver microsomes.
Xenobiotica. 34(11-12):983-1000, 2004.
6) Alfieri RR, Galetti M, Tramonti S, et al. Metabolism of the EGFR tyrosin kinase inhibitor gefitinib bycytochrome P450 1A1 enzyme in EGFR-wild type non small cell lung cancer cell lines. Mol Cancer. 10:143,
2011.
Acknowledgements. This work was financially supported by Universitat Politècnica de València (Subprograma 1,
PAID-01-19), the MICINN (FPU19/00048, BEAGAL 18/00211) and Carlos III Institute of Health (Grant
PI16/01877).

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IL-7.3.1
Porphysome Nanotechnology: From Discovery toward First-in-Patients
Michael Valic1, Juan Chen1, Gang Zheng1,21) Princess Margaret Cancer Centre, University Health Network, Toronto, Canada; 2) University of Toronto,
Department of Medical Biophysics, Toronto, Canada
Porphysomes are intrinsically multifunctional liposome-like nanoparticles self-assembled from a single porphyrinlipid building block. High-density porphyrin packing in the nanoparticle bilayer enables light absorption andconversion to heat with extremely high efficiency, making porphysomes ideal candidates for photothermal therapy
and photoacoustic imaging. Upon nanostructure dissociation during cell uptake, the fluorescence and photodynamic
activity of the porphyrin monomers is restored. In addition, metal ions can be directly incorporated into theporphyrin building blocks, thus unlocking porphysome’s potential for PET and MRI. To translate porphysomesfrom bench to bedside, we have embarked a 10-year journey where we first validated porphysome’s utilities
preclinically in different tumor types, models and animal species. We then, with purely academic resources,
completed the scale up manufacturing of both porphyrin-lipid building blocks and nanoparticle formulation,
optimized the cGMP methods and processes. In parallel, we completed the metabolism and phototoxicity studies
and demonstrated porphysome safety profiles in rodents, dogs and non-human primates. In addition,
pharmacokinetics and disposition of porphysomes were established in preclinical animals and subsequently
extrapolated to human. We anticipate the launch of the first-in-human porphysome study shortly. The simple yet
intrinsic multimodal nature of porphysome nanotechnology popularize the one-for-all nanomedicine and confers its
high clinical translation potential.

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IL-7.3.2
Biological models for in vitro and in vivo imaging, PDT, drug or siRNA delivery
Magali GARY-BOBO1
1) IBMM, Univ Montpellier, CNRS, ENSCM, Montpellier, France
Currently, the development of in vitro et in vivo models in accordance with the 3Rs rules (reduction, replacement,
refinement) is of great importance. In fact, research activities need such robust and easy-to-use technics todemonstrate the imaging and therapeutic potential of innovative anticancer molecules or nanoparticles. We are abiologists team specialized in the study of photoactivable nanoparticles for fluorescent imaging, photodynamic
therapy and drug or siRNA delivery for anticancer application. In our group, the biological effects of thesenanoparticles are studied first on human cancer cell lines in culture (in 2 dimensions or in spheroids) and could also
be analyzed on zebrafish embryos. The robustness of these models allows us to quickly carry out proof of concept
studies of nanoparticles exhibiting a wide range of biomedical properties.

A

2D

3D

In vivo

B

Fig. 1 – (A) In vitro and in vivo biological models. Cell culture in 2 dimensions (2D), in spheroids (3D) and zebrafish embryos.
(B) Scheme of human cancer cells injection in zebrafish embryo.

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IL-7.3.3

Fe3+-Driven Assembly of Catalase-Like Supramolecular Photosensitizing Nanozymes for
Combating Hypoxic Tumor
Yongxin Li1, Pan Sun2, Luyang Zhao3, Xuehai Yan3, Dennis K. P. Ng4, and Pui-Chi Lo*,1
1) Department of Biomedical Sciences, City University of Hong Kong, Tat Chee Avenue, Kowloon, Hong Kong, China;
2) CAS Key Laboratory of Green Process and Engineering, Institute of Process Engineering, Chinese Academy of
Sciences, Beijing 100190, China; 3) National Key Laboratory of Biochemical Engineering, Institute of Process
Engineering, Chinese Academy of Sciences, Beijing 100190, China; 4) Department of Chemistry, The Chinese Universityof Hong Kong, Shatin, N.T., Hong Kong, China.
Photodynamic therapy (PDT) has emerged as a compelling treatment modality for a range of superficial and localized
cancer.1 It involves excitation of a photosensitizer through light irradiation followed by the generation of reactive oxygen
species (ROS), in particular singlet oxygen, to cause cellular damage at the tumor site. However, PDT still suffersfrom two major hurdles that limit its clinical application, namely the low selectivity of the photosensitizers
toward cancer cells and the insufficient oxygen concentration at hypoxic tumor. Being inspired by the naturally

occurring catalases which can convert H2O2 into water and oxygen via the Fenton reaction, we report herein a facile
approach to assemble catalase-like photosensitizing nanozymes with a self-oxygen-supplying ability.2 The process
involved Fe3+-driven self-assembly of fluorenylmethyloxycarbonyl-protected cysteine (Fmoc-Cys) in the presence of azinc(II) phthalocyanine-based photosensitizer (ZnPc) and the hypoxia-inducible factor 1 (HIF-1) inhibitor acriflavine(ACF). Molecular self-assembly provides a facile and environmental green approach to encapsulate photosensitizers andother therapeutic components into nanoparticles that can promote passive tumor targeting via the enhanced permeability
and retention effect. The nanovesicles Fmoc-Cys/Fe@Pc and Fmoc-Cys/Fe@Pc/ACF were prepared, which could be
disassembled intracellularly. The released Fe3+ could catalyze the transformation of H2O2 enriched in cancer cells tooxygen efficiently, thereby ameliorating the hypoxic condition and promoting the photosensitizing activity of thereleased ZnPc. With an additional therapeutic component, Fmoc-Cys/Fe@Pc/ACF exhibited higher in vitro and in vivo
photodynamic activities than Fmoc-Cys/Fe@Pc, demonstrating the synergistic effect of ZnPc and ACF.

References
1) D. E. J. G. J. Dolmans, D. Fukumura, R. K. Jain, Nat. Rev. Cancer 2003, 3, 380-387.
2) Y. Li, P. Sun, L. Zhao, X. Yan, D. K. P. Ng, P.-C. Lo, Angew. Chem. Int. Ed. 2020, 59, 23228-23238.
Acknowledgements. This work was supported by a grant from the Research Grants Council of the Hong Kong Special
Administrative Region (Ref. No. 11300719).

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OC-7.3.4

Covalently cross-linked tetrafunctionalized m-THPC chitosan hydrogels as drugdelivery platforms in the treatment of melanoma.
Piotr Gierlich1,2, Claire Donohoe1,2, Kevin Behan4, Daniel Kelly4, Lígia C. Gomes-da-Silva2, Mathias O. Senge1,31) Medicinal Chemistry, Trinity Translational Medicine Institute, Trinity Centre for Health Sciences, Trinity College
Dublin, The University of Dublin, St. James's Hospital, Dublin 8, Ireland; 2) Department of Chemistry, Universityof Coimbra, Rua Larga, 3004-535 Coimbra, Portugal; 3) School of Chemistry, SFI Tetrapyrrole Laboratory, Trinity
Biomedical Sciences Institute, 152-160 Pearse Street, Dublin 2, Ireland; 4) Trinity Centre for Biomedical
Engineering, Trinity Biomedical Sciences Institute, Trinity College Dublin, 152-160 Pearse Street, Dublin 2, Ireland.
Photodynamic therapy (PDT) is an anti-cancer treatment method, which uses the combined effect of aphotosensitizing drug (as pro-drug activating agent), light, and oxygen to cause selective damage to target tissue.1
The second generation photosensitizer (PS) 5,10,15,20-tetrakis(m-hydroxyphenyl)chlorin (m-THPC) is a widely
characterized, clinically tested, and commercially available drug with the market formulation known as Foscan.2 In
order to develop advanced treatment modalities there is a need for improved drug delivery platforms. Hydrogels,
which have been investigated as effective drug delivery systems, can prevent PSs aggregation and offer significant
potential as drug carries due to the ability to swell in aqueous media.3 Chitosan (CS), a natural polysaccharide, is asuitable biodegradable material for hydrogel formulation and has been used in pharmaceutical applications on
account of its lack of toxicity and good biocompatibility.4

Fig. 1 – Schematic representation of the hydrogel synthesis.

In the present work, m-THPC was used as a starting point to obtain a library of compounds aimed at overcoming PS
limitations while maintaining its photophysical and clinical properties. Substitution, esterification and Sonogashiracoupling reactions were employed to modify the m-THPC skeleton providing aldehyde and carboxylic acid moieties
used as a suitable synthetic handle for covalent cross-linking in the formation of CS hydrogels. Injectable, selfhealing properties of hydrogels were confirmed macroscopically and by the rheological analysis. Next,
tetrafunctionalized m-THPC derivatives were tested in vitro against melanoma (B16F10) cancer cells. In vivo
biodistribution of the released PSs was studied using fluorescence imaging technique. Prepared hydrogel
formulations are expected to allow for a local, injectable administration towards melanoma tumors, while preventing
from systemic side effects related to the m-THPC treatment and are under ongoing in vivo evaluation.
References
1) 1. T. J. Dougherty, C. J. Gomer, B. W. Henderson, G. Jori, D. Kessel, M. Korbelik, J. Moan, Q. Peng, J. Nat.
Cancer Inst. 90, 889–905, 1998; 2) M. O. Senge, J. C. Brandt, Photochem. Photobiol. 87, 1240–1296, 2011; 3) B.
Khurana, P. Gierlich, A. Meindl, L. C. Gomes-da-Silva, M. O. Senge, Photochem. Photobiol. Sci. 18, 2613–2656,
2019; 4) M. Dash, F. Chiellini, R. M. Ottenbrite, E. Chiellini, Prog. Polym. Sci. 36, 981–1014, 2011.
Acknowledgements. This project has received funding from the European Union’s Horizon 2020 research andinnovation programme under the Marie Sklodowska-Curie grant agreement n°764837 and supported by grants from
Science Foundation Ireland (IvP 13/IA/1894).
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IL-7.3.5

Recent Progress on Activatable Photosensitizers and Heavy Atom Free
Photosensitizers
Juyoung Yoon
Department of Chemistry and Nanoscience, Ewha Womans University, Seoul 03760, Korea
Switchable phototheranostic nanomaterials are of particular interest for specific biosensing, high-quality imaging,and targeted therapy in the field of precision nanomedicine.1 Here, we develop a “one-for-all” nanomaterial(NanoPcTBs) that self-assembles from flexible and versatile phthalocyanine building blocks.2 Fluorescence andreactive oxygen species (ROS) generation could be triggered depending on a targeted, protein-induced, partial
disassembly mechanism, which creates opportunities for low-background fluorescence imaging and activatable
photodynamic therapy (PDT). We also reported a facile strategy to directly assemble a phthalocyanine
photosensitizer (PcS) with an anticancer drug mitoxantrone (MA) to form uniform nanostructures (PcS-MA), whichhave the capability of undergoing nucleic acid-responsive disassembly.3 On the other hand, the in vivo specificbinding between albumin and PcS, arising from the disassembly of injected NanoPcS, was recently confirmed using
an inducible transgenic mouse system.4 In a recent investigation, we devised a novel molecular design approach tocreate heavy-atom-free photosensitizers for enhanced photodynamic therapy under hypoxia conditions.5 The
thionaphthalimides display dramatically enhanced quantum yields for photosensitized singlet oxygen formation (Φ∆~ 1.00, in air-saturated acetonitrile).6

References
1) H.-B. Cheng, B. Qiao, H. Li, J. Cao, Y. Luo, K. M. K. Swamy, J. Zhao, Z. Wang, J. Y. Lee, X.-J. Liang, J. Yoon,
Protein-Activatable Diarylethene Monomer as a Smart Trigger of Noninvasive Control Over Reversible Generationof Singlet Oxygen: A Facile, Switchable, Theranostic Strategy for Photodynamic-Immunotherapy. J. Am. Chem.
Soc., 143, 2413-2422, 2021.
2) X. Li, C-y. Kim, S. Lee, D. Lee, H.-M. Chung, G. Kim, S.-H. Heo, C. Kim, K.-S. Hong, J. Yoon, Nanostructured
Phthalocyanine Assemblies with Protein-Driven Switchable Photoactivities for Biophotonic Imaging and Therapy.
J. Am. Chem. Soc., 139, 10880-10886, 2017.
3) X. Li, S. Yu, D. Lee, G. Kim, B. Lee, Y. Cho, B.-Y. Zheng, M.-R. Ke, J.-D. Huang, K. T. Nam, X. Chen, J. Yoon,
Facile Supramolecular Approach to Nucleic-Acid-Driven Activatable Nanotheranostics That Overcome Drawbacksof Photodynamic Therapy. ACS Nano, 12, 681-688, 2018.
4) X. Li, S. Yu, Y. Lee, T. Guo, N. Kwon, D. Lee, S. C. Yeom, Y. Cho, G. Kim, J.-D. Huang, S. Choi, K. T. Nam,
J. Yoon, In Vivo Albumin Traps Photosensitizer Monomers from Self-Assembled Phthalocyanine Nanovesicles: A
Facile and Switchable Theranostic Approach. J. Am. Chem. Soc., 141, 1366-1372, 2019.
5) V.-N. Nguyen, Y. Yim, S. Kim, B. Ryu, K. M. K. Swamy, G. Kim, N. Kwon, C-Y. Kim, S. Park, J. Yoon,
Molecular Design of Highly Efficient Heavy-Atom-Free Triplet BODIPYs for Photodynamic Therapy andBioimaging. Angew. Chem. Int. Ed., 59, 8957-8962, 2020.
6) V.-N. Nguyen, S. Qi, S. Kim, N. Kwon, G. Kim, Y. Yim, S. Park, J. Yoon, An Emerging Molecular Design
Approach to Heavy-Atom-Free Photosensitizers for Enhanced Photodynamic Therapy under Hypoxia. J. Am. Chem.
Soc., 141, 16243-16248, 2019.
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IL-7.3.6

Improving the photodynamic efficiency of phthalocyanine with nanotechnology
Fabienne Dumoulin
Acıbadem Mehmet AliAydınlar University, Department of Medical Engineering,Kayışdağı Cad. 34, Ataşehir, Istanbul, Turkey

The maximum absorption of phthalocyanines is centered around
700 nm, which is a significant advantage for photosensitisers ofanti-cancer photodynamic therapy.1 In addition, phthalocyanines
can be easily functionalized and their photoproperties can be
tailored with an appropriate substitution and metalation pattern.2
As nanotechnology offers the opportunity to benefit from the
Enhanced Permeation and Retention Effet, and also to fabricate
multi-functionalized platforms, incorporating phthalocyanines
into nanoparticles is an aim intensively pursued by researchers in
the last decade.
We have used covalent and non-covalent methods toincorporate/encapsulate/embed/ photosensitising phthalocyanines
into nanoparticles.
Polyacrylamide3, micellar formulation4, phthalocyanine-polyglutamic acid polymeric constructs5,6, amongst othernanoparticles have been produced, each of them exhibiting specific advantages and challenges that will be
discussed.
References
1) P.-C. Lo, M. S. Rodriguez-Morgade, D. K. P. Ng, T. Torres, R. Pandey and F. Dumoulin, Chemical Society
Reviews, 2020, 49, 1041-1056
2) S. Z. Topal, Ü. İşci, U. Kumru, D. Atilla, A. G. Gürek, C. Hirel, M. Durmuş, J.-B. Tommasino, D. Luneau, S.
Berber, F. Dumoulin and V. Ahsen, Dalton Trans. 2014, 43, 6897-6908
3) F. Giuntini, F. Dumoulin, R. Daly, V. Ahsen, E. M. Scanlan, A. S. P. Lavado, J. W. Aylott, G. A. Rosser, A.
Beebye and R. W. Boyle, Nanoscale 2012, 4, 2034-2045
4) B. Pucelik, I. Gürol, V. Ahsen, F. Dumoulin, J. M. Dabrowski, Eur. J. Med. Chem. 2016, 124, 284-298
5) L.V. Kiew, H. Y. Cheah, S. H. Voon, E. Gallon, J. Movellan, K. H. Ng, S. Alpugan, H. B. Lee, F. Dumoulin, M.
J. Vicent, Lip Yong Chung, Nanomedicine: NBM, 2017, 13, 1447-1458.
6) H. Y. Cheah, E. Gallon, F. Dumoulin, S. Z. Hoe, N. Japundžić-Žigon, S. Glumac, H. B. Lee, P. Anand, L. Y.
Chung, M. J. Vicent, L. V. Kiew, Mol. Pharmaceutics 2018, 15, 2594-2605.

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IL-7.3.7

Multifunctional nanomaterials for detection and photoinactivation of microbial cells
Anzhela Galstyan
University of Münster, Center for Soft Nanoscience, Münster, Germany.
With tunable structures, remarkable photophysical properties and the ability to produce reactive oxygen species
under red light irradiation, phthalocyanine derivatives have been investigated for many years as photosensitizers for
antimicrobial PDT.1 Nanotechnology has created new opportunities in aPDT by providing efficient nanoscale
photoactive agents that can be formed via a bottom-up or top-down approach.
Self-association is the spontaneous organization of molecular units and is widely used bottom-up approach tofabricate nano-PDT agents.2 Novel nanostructured Zn(II)phthalocyanine assemblies able to reduce bacterial
viability in planktonic cultures and biofilms will be presented. How the degree of ordered molecular arrangement
affects the photophysical properties and how high antimicrobial efficacy could be achieved by tuning the ratio
between solubility and cell penetration will be discussed.
Another approach towards photoactive nanomaterials is electrospinning, which is considered a simple top-down
approach for obtaining nanofibres of high quality. Though there are many reports showing that such nanoporous
surfaces exhibit very strong antibacterial and antiviral effect upon irradiation, major regulators of activity are stilllargely unverified and the requirements for effective antimicrobial action remain controversial. Design andsynthesis of photoactive compounds that could be used as structural components of nanoscale materials3,4 as well as
SAR of such materials will be presented.

Fig. 1 – Schematic view of the formation of nano-PDT-agents through bottom-up and top-down approach.

References

1) A. Galstyan, Turning photons into drugs: phthalocyanine-based photosensitizers as efficient photoantimicrobials, Chem. – A
Eur. J., (2021), 27(6): 1903–1920. 2) A. Galstyan, A. Ricker, H. Nüsse, J. Klingauf, U. Dobrindt Exploring the Impact of
Coordination-Driven Self Assembly on the Antibacterial Activity of Low-Symmetry Phthalocyanines. ACS Appl. Bio Mater.
(2020), 3, 400–411. 3) K. Strokov, A. Galstyan, Chitosan‐Silicon Phthalocyanine Conjugate as Effective Photo‐Functional
Hydrogel for Tracking and Killing of Bacteria, Eur. J. Org. Chem. (2020), 47, 7327-7332. 4) K. Strokov, A. H. Schäfer, U.
Dobrindt, A. Galstyan, Facile fabrication of silicon(IV)phthalocyanine embedded poly(vinyl alcohol)-based antibacterial andantifouling interfaces, ACS Appl. Bio Mater. (2020), 3(6), 3751–3760.

Acknowledgements. This work was supported by Deutsche Forschungsgemeinschaft (GA 2365/2-1), Fonds derChemischen Industrie and WWU Münster.

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OC-7.3.8
Porphysome nanoparticles are effective photosensitizers for photodynamic therapy
treatment for cancer
Keegan Guidolin1,2,3, Lili Ding3, Juan Chen3, Gang Zheng2,31) Division of General Surgery, Department of Surgery, University of Toronto, Toronto, Canada; 2) Institute of
Biomedical Engineering, University of Toronto, Toronto, Canada; 3) Princess Margaret Cancer Center, University
Health Network, Toronto, Canada
Background: PDT has been investigated as a cancer therapeutic pre-clinically using many photosensitizers and formany applications. One particular photosensitizer, Photofrin (porfimer sodium, PHO), is a mixture of oligomers
containing porphyrins. PHO benefits from some of the inherent tumouritropic properties of porphyrins, but there isconcern about its off-target effects [1]. Nanomedicines are designed and engineered materials with healthapplications that measure between 1 nm and 100 nm in at least one dimension. Their size confers tumouritopic
properties through a mechanism called the enhanced permeability and retention (EPR) effect [2]. One suchnanomedicine is called the “porphysome” (PS) and is composed of porphyrin-lipid subunits which self-assemble into
spherical liposome-like structures ~100 nm in size. These porphysomes have demonstrated both diagnostic andtherapeutic applications in pre-clinical studies conducted over the past ten years, including fluorescence imaging,
photoacoustic imaging, photothermal therapy, and drug delivery; however, they have never been investigated as
photosensitizers for PDT on their own [3]. The aim of this study was to determine whether or not porphysomes are
viable photosensitizers for PDT applications.
Methods: Female athymic nude mice, underwent subcutaneous injection of 5 x 10 6 A549 lung cancer cells. Whentumours reached 5 mm, animals were randomized into treatment groups as per the specific aim protocol. After
treatment, animals were followed with tumour measurement three times per week for a total of 30 days posttreatment. An a priori tumour size endpoint of 500 mm3 was established for ethical purposes to represent terminal
tumour growth.
Aim 1 – establishment of laser dose. To establish that no photothermal effects are present at the proposed light dose,
three animals were administered 10 mg/kg of PS, and after a drug-light interval (DLI) of 24 hours, tumours were
subjected to localized external laser delivery at a dose of 135 J/cm2 (100 mW, 10 mm diameter beam, 671 nm, 22.5
minutes). Throughout this period, the surface temperature of the tumour was measured by thermal imaging. An apriori endpoint of a 5°C temperature increase was established to indicate photothermal heating.
Aim 2 – dose optimization. To establish the optimal dose and treatment plan for PS PDT, five animals each received:
5 mg/kg PS, 24h DLI; 7.5 mg/kg PS, 24h DLI; 10 mg/kg PS, 24h DLI; and 5 mg/kg PHO, no laser treatment. In
addition, the best performing dose of PS from these groups was administered with a DLI of 48h, and with no laser
treatment with five animals in each group.
Aim 3 – head-to-head comparison with Photofrin. To measure the effectiveness of PS PDT against the current clinicalstandard, twenty animals each received treatment with 5 mg/kg PHO, 24h DLI, and with the optimized dose and DLIof PS.
Results: Tumour temperatures measured throughout treatment with 100 mW/cm2 for 22.5 minutes did not rise above
the a priori threshold of 5°C above baseline. All PS doses demonstrated a significant tumour ablative effect comparedto groups not receiving laser treatment; however, the greatest effect was seen in the 10 mg/kg PS group at a DLI of
24 hours. In the dose-finding group, the 10 mg/kg treatment and 7.5 mg/kg treatment demonstrated 67% and 60%
cure respectively, with partial response and tumour growth suppression in the remaining tumours for both. The 5
mg/kg treatment showed partial response and tumour growth suppression without cure. Negative control groups (bothPS and PHO drug-only, and no drug no light groups) demonstrated uncontrolled tumour growth requiring sacrificeprior to the 30-day endpoint as tumours reached an a priori maximum volume. Direct comparison of PHO 5 mg/kg
and PS 10 mg/kg demonstrated similar tumour growth suppression (MANOVA p>0.05), which was significantly
different from the uncontrolled tumour growth seen in the untreated control groups. Tumour growth reached a nadir
12-15 days after treatment; after this point, tumours began to regrow at a suppressed rate (Figure 1). Complete curewas not different between PHO and PS treatment groups (15% vs. 25%, p=0.52).

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Figure 1. Comparison of mean relative tumour volume in animals treated with PS-PDT (n=20) and PHO-PDT (n=20).

Discussion: We identified an optimal dose of 10 mg/kg and DLI of 24 hours. This corroborates previous findingsthat tumour PS concentration is maximized at 24 hours following administration [3-5]. The optimized dose is also
identical to the dose used in studies of other therapeutic applications of PS, namely photothermal therapy (PTT) [4,6]. This study PS PDT was compared to a clinically approved PDT photosensitizer, PHO, as a positive control. Our
results suggest that not only can the PS be used as a photosensitizer for PDT, but that it ablates tumours at a similarrate to PHO.
When delivered to a tumour, porphysomes initially exist as intact nanovesicles, but as time passes, porphysomes aretaken into tumour cells, at which time they dissociate into their component porphyrin-lipids. Intact porphysomes
absorb light energy and convert it into vibrational energy, released as heat (PTT) [4, 6-9]. In their dissociated state,porphyrin-lipids absorb energy which can be released as fluorescence or can produce reactive oxygen species.
Previous studies have confirmed the fluorescence of porphysomes at 24 hours and have used fluorescence imaging
to guide PTT [8-10]. The results of this study suggest that PDT and PTT could be used in combination as a single
agent multimodality photosensitizer. Future studies will directly investigate this combined multimodality therapy
investigating for additive or synergistic tumour ablation potential.
References
1.
Dougherty, T.J., et al., Photodynamic therapy. J Natl Cancer Inst, 1998. 90(12): p. 889-905.
2.
Kalyane, D., et al., Employment of enhanced permeability and retention effect (EPR): Nanoparticle-based
precision tools for targeting of therapeutic and diagnostic agent in cancer. Mater Sci Eng C Mater Biol
Appl, 2019. 98: p. 1252-1276.
3.
Lovell, J.F., et al., Porphysome nanovesicles generated by porphyrin bilayers for use as multimodal
biophotonic contrast agents. Nat Mater, 2011. 10(4): p. 324-32.
4.
Jin, C.S., et al., Ablation of hypoxic tumors with dose-equivalent photothermal, but not photodynamic,
therapy using a nanostructured porphyrin assembly. ACS Nano, 2013. 7(3): p. 2541-50.
5.
Lovell, J.F., et al., Enzymatic regioselection for the synthesis and biodegradation of porphysome
nanovesicles. Angew Chem Int Ed Engl, 2012. 51(10): p. 2429-33.
6.
Jin, C.S., J.F. Lovell, and G. Zheng, One minute, sub-one-watt photothermal tumor ablation using
porphysomes, intrinsic multifunctional nanovesicles. J Vis Exp, 2013(79): p. e50536.
7.
MacDonald, T.D., T.W. Liu, and G. Zheng, An MRI-sensitive, non-photobleachable porphysome
photothermal agent. Angew Chem Int Ed Engl, 2014. 53(27): p. 6956-9.
8.
Kinoshita, T., et al., Evaluation of Novel Imaging Devices for Nanoparticle-Mediated Fluorescence-Guided
Lung Tumor Therapy. Ann Thorac Surg, 2019. 107(6): p. 1613-1620.
9.
Ng, K.K., et al., Self-sensing porphysomes for fluorescence-guided photothermal therapy. Bioconjug Chem,
2015. 26(2): p. 345-51.
10.
Philp, L., et al., Use of Porphysomes to detect primary tumour, lymph node metastases, intra-abdominal
metastases and as a tool for image-guided lymphadenectomy: proof of concept in endometrial cancer.
Theranostics, 2019. 9(9): p. 2727-2738.
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P-7.3.9
Heme Biosynthesis and Degradation changes after 5-ALA application: A proteomic
study
Sara Sansaloni Pastor1, Emmanuel Varesio, and Norbert Lange1
1) Department of Pharmaceutical Technology, School of Pharmaceutical Sciences, University of Geneva, Institut desSciences Pharmaceutiques de Suisse Occidentale (ISPSO), Geneva, Switzerland
Photodynamic therapy (PDT) has been applied to treat neoplastic and non-neoplastic diseases. It has been shown thatafter exogenous application of 5-aminolevulinic acid (5-ALA) or its derivatives induces is a preferential accumulationof protoporphyrin IX (PpIX). However, the mechanisms for this accumulation remain undiscovered.
To better understand the effect of PpIX accumulation in cancer cells, we studied how the different enzymes in heme
biosynthesis and degradation enzymes are affected by 5-ALA and porphyrins. To this end, we used T24 cell line (bladdercancer cell line) treated for 24 h by 5-ALA (1 mM), succinylacetone (SA) (1 mM), 5-ALA+SA (1mM) and compared to
non-treated control. All analyses were performed by mass quantification using a LC-ESI-MS/MS on an Orbitrap Fusion
Lumos Tribrid mass spectrometer. Statistical analysis was carried out using two ways ANOVA and individual t-test.
We identified approximately 5000 proteins per sample. In addition, our technical replicates showed a good correlation(Pearson correlation > 0.91). Our results showed a change in the of heme enzymes’ expression not only on the synthesis,but also on the degradation of porphyrins. A deregulation effect was observed in the biosynthetic pathway caused by theporphyrins, with a statistically significant upregulation of HMBS, UROD and PPOX and a downregulation of CPOX.
5-ALA also showed a statistically significant downregulation effect on HMBS, PPOX and FECH. Additionally, thedegradation pathway is highly upregulated by the porphyrins, which affects HMOX1 and BLVRA enzymes.
Our findings shed a light in the proteomic role of 5-ALA and porphyrins and its applications in PDT therapeutic options.
Thus, leading research efforts in improving novel and better treatment approaches.
References:
1) Sansaloni-Pastor S, Bouilloux J, Lange N. The Dark Side: Photosensitizer Prodrugs. Pharmaceuticals (Basel). 2019Oct 4;12(4):148. doi: 10.3390/ph12040148. PMID: 31590223; PMCID: PMC6958472.
Acknowledgements. This work was supported by the Swiss National Science Foundation (SNF)

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P-7.3.10

Targeted bioorthoganal prodrug activation enabled by pH responsive classicalpolymer photocatalysts
Rong Li, Thomas Kuckhoff, Katharina Landfester and Calum T. J. Ferguson
Max Planck Institute for Polymer Research, Ackermannweg 10, 55128 Mainz, Germany
The use of photocatalytic in situ prodrug activation and against cancer has received increasing attention over recent
years. However, the application of the currently approved photocatalysts is limited by their poor aqueous solubility,
aggregation, photobleaching, off target effects and poor tumour uptake. One approach to overcome these limitations
is through the use of a delivery vector for the photoactive species. Recently we have reported a new class of polymer
photocatalysts formed by combining small molecular photocatalysts/ with classical polymer chemistry. 1-3 Creating
photocatalytic polymer constructs that contain the beneficial properties from both components.
We have designed pH responsive classical polymer catalysts that respond to the well reported pH changes between
healthy and cancerous tissue. Here, we have used RAFT polymerisation to produce diblock copolymers consistingof PEG114-b-(PAEMA40-s-PPH2BT2). At pH >7 crosslinking of primary amines in the PAEMA block withterephthalaldehyde occurs, resulting in the formation of nanoparticles (NPs). In this particulate form the polymer isphotocatalytically inactive as the active centres are imbedded within the core of the particle. These non-active NPs
can freely circulate around the body and will accumulate in tumour tissue through the EPR effect. Within the tumour
cell environment a pH drop is observed to pH 6.4. This change stimulates the disassembly of the NP, where thepolymer once again becomes photocatalytically active. We have used this photocatalytic material for the in situ
activation of a cancer prodrug based on fluorouracil (5-FU). Where the prodrug version of 5-FU shows limited
toxicity but can be easily activated in the presence of light and the photoactive polymer into the toxic form. We havedemonstrated by both In vitro and cellular studies that this approach has the potential to enable targeted prodrug
activation for cancer treatment.
References[1]

C. T. Ferguson, N. Huber, T. Kuckhoff, et al., J. Mater. Chem. A, 2020.

[2]

C. T. Ferguson, N. Huber, K. Landfester, et al., Angew. Chem. Int. Ed., 2019.

[3]

N. Huber, R. Li, C. T. Ferguson, et al., Catal. Sci. Technol., 2020.

Acknowledgements. This work was supported by grant 42 from the Ministry of Truth.

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P-7.3.11
Radioluminescent nanoparticles and deep-tissue photodynamic therapy to enhanceradiotherapy efficacy
Sarah Stelse-Masson1, Thibault Jacquet1, Mans Broekgaarden1,2, Frédéric Chaput3, Kseniya Popovich4, Anamarija
Abu el Rub1, Anthony Nomezine1, Lucie Sancey2, Eva Mihokova5, Vaclav Cuba4, Hélène Elleaume1,
Anne-Laure Bulin1
1) Synchrotron Radiation for Biomedicine, Grenoble Alpes University, INSERM UA7, Grenoble, France.
2) Institute for Advanced Bioscience, Grenoble-Alpes University, INSERM U1209, 38000 Grenoble, France.
3) École Normale Supérieure de Lyon, CNRS - Université Claude Bernard Lyon 1, France.
4) Department of Nuclear Chemistry, FNSPE, Czech Technical University in Prague, Czech Republic.
5) Institute of Physics, Czech Academy of Sciences, Prague, Czech Republic.
Radioluminescent nanoparticles, also known as nanoscintillators, are gaining
increasing interest as radiotherapeutics. Because they down-convert ionizing
radiations into visible light, they can act as internal light sources remotely activatedby penetrating X-ray 1 . They can therefore induce photodynamic therapy (PDT)
during radiation therapy in deep-tissue and potentially within large tumor
volumes 2,3. Proof-of-concept studies have been published by us and others, yet theexact therapeutic mechanisms that are activated remain to be fully understood 4,5.
When using nanoscintillators in combination with radiotherapy, mechanisms otherthan PDT could also be activated. As nanoscintillators are typically composed ofhigh-Z elements, we hypothesized the existence of another therapeutic effect thatis currently under-explored for nanoscintillators: the purely physical radiation
Figure 1: Radiotherapeutic
dose-enhancement effect. This effect is initiated by a higher photoelectric contributions that can be activatedand potentially synergize whenabsorption of orthovoltage X-rays by high-Z elements compared to soft tissues thatnanoscintillators are used toleads to a higher production of photoelectrons and Auger electrons that can
activate deep-tissue PDT during
enhance the damage to cancer tissues 6. Investigating each individual
radiotherapy.
radiotherapeutic effect induced by the nanoscintillator-photosensitizer conjugates,
as well as their potential synergy, is crucial to better understand this novel
modality.We will present the results obtained with various nanoscintillators made of high-Z elements and we will
describe their ability to enhance the efficacy of radiotherapy through different mechanisms. Techniques ranging fromphysics to preclinical studies were employed to achieve these results.
References
1) C. Dujardin et al. Needs, Trends, and Advances in Inorganic Scintillators, IEEE T Nucl Sci 65, 2018
2) W. Sun et al. Nanoscintillator-mediated X-ray induced photodynamic therapy for deep-seated tumors: fromconcept to biomedical applications, Theranostics 10, 20203) J. Klein et al. Radioluminescence in biomedicine: physics, applications, and models, Phys Med Biol 64, 20194) A-L. Bulin et al. X-ray induced singlet oxygen activation with nanoscintillator-coupled Porphyrins, J. Phys Chem
C 117, 20135) H. Chen et al., Nanoscintillator-mediated X-ray inducible photodynamic therapy for in vivo cancer treatment,
Nano Lett. 15 20156) A-L. Bulin et al., Radiation dose-enhancement is a potent radiotherapeutic effect of rare-earth composite
nanoscintillators in preclinical models of glioblastoma, Advanced Science, 7 2020
Acknowledgements. The work was supported by grants from the French Medical Research Foundation (Fondation
pour la Recherche Médicale, ARF20170938739), the Auvergne Rhône-Alpes Region (NEPTUNE project), INSERM
Cancer, the French Ligue Contre le Cancer Foundation and the LABEX PRIMES of the Université of Lyon (ANR11-LABX-0063). A-L. B. also received additional funding from the “Rotary Club” Association through the program“Jetons Cancer” and from INSERM.
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P-7.3.12
Rational Design and Development of a New Class of Metal-Based Photosensitizers for
Photodynamic Therapy
Houston D. Cole1, John Roque III1,2, Ge Shi1, Elam Ramasamy1, Colin G. Cameron1, Sherri A. McFarland1
1) University of Texas at Arlington, Department of Chemistry and Biochemistry, Arlington, TX, United States; 2)
University of North Carolina at Greensboro, Department of Chemistry and Biochemistry, Greensboro, NC, UnitedStatesPhotodynamic therapy (PDT) is a unique approach to cancer treatment that remains underutilized despite
its intrinsic selectivity towards cancerous tissue. PDT utilizes a non-toxic photosensitizer (PS), light, andoxygen to initiate localized cell death (and elicit an antitumor immune response in some cases). Currently,
the only FDA-approved PS for cancer therapy is Photofrin, a porphyrin-based oligomer. With improved
PSs, we propose that PDT could become a more important adjuvant or alternative therapy for certain
cancers. Metal-based PSs are of particular interest in this regard due to their tuneable photophysics andanti-cancer activity. Ruthenium- and osmium-centered PSs are emerging as examples, with our own
TLD1433 currently undergoing Phase 2 clinical trials for the treatment of bladder cancer (Clinicaltrials.gov
identifier NCT03945162). Herein, we describe the rational design and development of a new class of metalbased PSs to further optimize their light-driven PDT effects.

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P-7.3.13

Remote-controlled drug release with photons: From light to X-rays
Tristan le Clainche1, Nazareth Milagros Carigga Gutierrez1, Eaazhisai Kandiah2, Xavier Le Guevel1, Lucie Sancey1,
Hélène Elleaume3, Anne-Laure Bulin3, Jean-Luc Coll1, Mans Broekgaarden1.
1) Institute for Advanced Bioscience, Grenoble-Alpes University, INSERM U1209, 38000 Grenoble, France.
2) European Synchrotron Radiation Facility, 38000, Grenoble, France3) Synchrotron Radiation for Biomedicine, Grenoble-Alpes University, INSERM UA7, 38026 Grenoble, France
Liposomes have been the most successful type of nanomedicine for cancer patients, playing a leading role in
improving the tolerability of chemotherapeutics. However, to advance the success of liposomal drug delivery andcancer treatment, new approaches to physically trigger drug release in cancer tissues and increase the permeabilityof the protective cancer stroma are needed 1. Therefore, we investigate whether photosensitizers and radiocatalytic
nanomaterials can be integrated in liposomes for remote-controlled drug release through excessive production ofreactive oxygen species (ROS) upon exposure to light and radiotherapy.
When embedded in the lipid bilayer of liposomes, photosensitizers can be used for the light-controlled release of
encapsulated drugs 2. The oxidation of unsaturated phospholipids through the photodynamic effect triggers
conformational changes in their acyl chains, resulting in lipid packing defects, loss of membrane integrity, and therelease of liposomal cargoes 3. To optimize this effect for 690 nm light-controlled drug release, we developedoxidation-responsive liposomes supplemented with benzoporphyrin derivative (BPD). We discovered that distinct
lipids and phospholipids act synergistically to promote drug release, and developed a novel formulation with which
complete drug release could be achieved at low radiant exposures (<10 J/cm2).
Radiation controlled liposomal drug delivery would be a substantial improvement over photo-triggered drug release,as X-rays penetrate deeply into tissues and radiotherapy is more broadly involved in the standard-of-care for cancer
patients. We thus further supplemented the oxidation-responsive liposomes with radiocatalytic gold nanoclusters,i.e., 2 nm nanoparticles that generate ROS upon exposure to X-rays
through the radiation dose-enhancement effect 4. Using
monochromatic synchrotron radiation, we discovered that liposomes
containing both BPD and gold nanoclusters generated significantly
elevated levels of ROS. The ROS generation occurred in a golddependent manner, as it was highest with X-ray energies tuned above
the absorption K-edge of gold (82 keV), suggesting a synergistic
interaction between gold and BPD. This interaction was successfully
leveraged to induce X-ray-controlled drug release.
In conclusion, this work provides important new insights in the roleof specific lipids in achieving photodynamic drug release. Moreover,
we provide compelling proof-of-concept for the feasibility ofradiotherapy-triggered drug release, which is a ground-breaking newconcept. Ongoing research focuses on investigating the biologicaleffects of these novel oxidation-responsive liposomes on 3D culture
Fig. 1 – Remote-controlled drug release frommodels of cancer 5. Especially the increased permeability of tumor
oxidation-responsive liposomes, induced either
tissues following exposure to photo- and/or X-ray-generated ROS by the excitation of photosensitizers by light, orthe excitation of radiocatalytic gold
may significantly advance the standard-of-care for many cancer
nanoclusters by X-rays.
patients.
References
1) Crommelin et al., J Control Release 2019. 2) Luo et al., Small, 2016. 3) Massiot et al., J Mater Chem B, 2019.
4) Broekgaarden et al., Nanoscale 2020. 5) Broekgaarden et al., Biomaterials 2019.
Acknowledgements. This work was supported by the Phospholipid Research Center (Heidelberg, Germany, MAB2020-080/1-1), and the French National Institute for Health and Medical Research (INSERM).

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P-7.3.14

Squalene-NIR dye nanoassemblies targeting mitochondria with photosensitizing
properties for the detection and treatment of cancer by phototherapy
Souad Adriouach1,2 and Eric Allémann1,2
1School of Pharmaceutical Sciences, University of Geneva, 1211 Geneva, Switzerland; 2Institute of Pharmaceutical
Sciences of Western Switzerland, University of Geneva, 1211 Geneva 4, Switzerland,
Squalene (Sq) is a natural precursor of cholesterol (1). The conjugation of Sq to a drug produces bioconjugates that
self-assemble in water to give nanoassemblies (NAs) (2).
Near-infrared (NIR) fluorescence has great potential for in vivo tumour imaging. NIR dyes such as the recently
synthesized SSA-15 (3) are lipophilic cations with preferential accumulation in the mitochondria of cancer cells.
They also have photosensitizing properties inducing tumour cell death after NIR light irradiation.
This study aims to develop Sq-SSA-15 NAs targeting mitochondria, with imaging and photosensitizing properties
and to assess their capacity to induce tumour cell death by phototherapy after NIR light irradiation. Here, we report:
 in vitro fluorescence imaging of Sq-SSA-15 NAs and specific mitochondrial localization
 antitumour effect of Sq-SSA-15 NAs after NIR light irradiation
Material and methods:
For this study, we used MCF7 human breast cancer cells and MCF-10 non-tumorigenic human breast cells to assess
the specific localization of Sq-SSA-15 NAs in the mitochondria of cancer cells. MCF7 and MCF10 were incubatedwith different concentrations of SSA-15 or the Sq-SSA-15 NAs for 1h. Mitochondria were stained with
MitoTracker® Orange and nuclei with Hoechst®. Cells were observed by live-cell imaging using the automated
IXM-XL microscope (Molecular devices, 1 x 40). To study the photosensitizing properties of our product, we
incubated MCF7 cells with 20 µM of SSA-15 and Sq-SSA-15 for 1 h. The cells were washed and irradiated with a
NIR LED lamp at a wavelength of 630 nm, at 100mW/cm2 for 10 min. We performed the cytotoxicity assay after 24
h using WST-1 cell proliferation assay.
Results: Fluorescence imaging of MCF-7 cells demonstrated effective localization of both SSA-15 and Sq-ssa-15
NAs in mitochondria of cancer cells by the co-staining with MitoTracker® Orange. The comparison with MCF10
non-tumorigenic breast cells demonstrated the preferential accumulation of our product in cancer cells. The WST-1
cell proliferation assay indicated that both SSA-15 and Sq-SSA-15 induced tumour cell death after light irradiationcompared to the control (dark condition).
Conclusion: This study demonstrated the preferential accumulation of Sq-SSA-15 in the mitochondria of cancer
cells by comparison to healthy cells. Sq-SSA-15 induced cancer cell death after NIR light irradiation. The
fluorescence and photosensitizing properties of the Sq-SSA-15 NAs suggest their potential use as a nanotheranostic
agent for imaging and treatment of cancer by phototherapy.
References:1.
Desmaële D, Gref R, Couvreur P. Squalenoylation: A generic platform for nanoparticular drug delivery. J
Control Release. 2012;161(2):609-18.
2.
Adriouach S, Vorobiev V, Trefalt G, Allémann E, Lange N, Babič A. Squalene-PEG: Pyropheophorbide-a
nanoconstructs for tumor theranostics. Nanomedicine: Nanotechnology, Biology and Medicine. 2019;15(1):243-51.
3.
Vorobiev V, Adriouach S, Crowe LA, Lenglet S, Thomas A, Chauvin A-S, et al. Vascular-targeted
micelles as a specific MRI contrast agent for molecular imaging of fibrin clots and cancer cells. European Journalof Pharmaceutics and Biopharmaceutics. 2021;158:347-58.

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P-7.3.15
Metallo-surfactant mediated nanocolloids for photodynamic therapyLluïsa Pérez-García1,2, Bunty Sharma2,3, Ganga Ram Chaudhary3, and Gurpreet Kaur3
1) Universitat de Barcelona, Department of Pharmacology, Toxicology and Therapeutic Chemistry, Institut deNanociència i Nanotecnologia UB (IN2UB), 08028 Barcelona, Spain; 2) University of Nottingham, School of
Pharmacy, Nottingham, UK; 3) Panjab University, Department of Chemistry, Chandigarh, 160014, India.
Metallosurfactant (MS) aggregates have grasped great attention from researchers worldwide due to the dual
properties of both metals and surfactants. On complexing surfactants with metal ions, a decrease in the critical micelleconcentration (CMC) is usually observed compared to those of the parent innocent surfactants.1 Catanionic vesicles
are synthesized by mixing cationic metallosurfactant and anionic surfactant in non-stoichiometric ratios which leads
to spontaneous vesicle formation. The size and surface charge of these vesicles can be controled by varying thecationic/anionic components ratio. Photodynamic therapy results from combination of photosensitizers (PSs), lightof specific wavelength, and molecular oxygen; conversion of molecular oxygen into phototoxic singlet oxygen is
greatly improved by incorporation of PSs into nanomaterials.2
We have formulated metallocatanionic vesicles (MCVs) from a combination of a double-and single-chain copperand iron-based cationic metallosurfactant (CuCPCI, and FeCPCI) and an anionic surfactant sodium bis(2ethylhexyl)sulfosuccinate (AOT). We have prepared mixtures with different ratios, from 10:90 to 90:10, in PBS of
7.4 pH. In this approach, two of the fractions, one each from a cationic rich and anionic rich side, were selected toencapsulate anionic rose bengal (RB). The vesicles were characterized by SAXS, AFM, FE-SEM, cryo-TEM, and
Zeta-sizer measurements. These studies reveal that the MCVs have dual functionality i.e. encapsulate PSs andenhance the singlet oxygen yield of RB, and the nanomaterials show antibacterial properties against S. Aureus, E.
Coli.3,4 Recently, we have applied these PS-loaded MCV against U-251 Glioblastoma cell lines. These experiments
showed MCVs biocompatible nature in dark and high phototoxicity against cancer cell lines which were confirmed
by WST-8 assay. Further, SOSG assay and differential nuclear staining assay (DNS), confirmed the intracellular
singlet oxygen generation and live/dead cell after PDT. Caspase assay confirmed the apoptotic pathway of cell killing.
This work provides a new metal hybrid smart biocompatible material that possesses dual functionality and is preparedby an easy, fast, and feasible procedure which resulted in enhanced PDT against a drug-resistant bacterium and cancer
cell lines.
References
1) Kaur, N.; Kaur, S.; Kaur, G.; Bhalla, A.; Srinivasan, S.; Chaudhary, G. R. Metallovesicles as smart nanoreactors
for green catalytic synthesis of benzimidazole derivatives in water, J. Mater. Chem. A, 2019, 7, 17306-17314.
2) Samperi, M.; Limón, D.; Amabilino, D. B.; Pérez-García, L. Enhancing singlet oxygen generation by selfassembly of a porphyrin entrapped in supramolecular fibers, Cell Reports Physical Science, 2020, 1, 100030.
3) Sharma, B.; Kaur, G.; Chaudhary, G. R.; Gawali, S. L.; Hassan, P. A. High antimicrobial photodynamic activityof photosensitizer encapsulated dual-functional metallocatanionic vesicles against drug resistant bacteria S aureus,
Biomater. Sci. 2020, 8, 2905−2920.
4) Sharma, B.; Kaur, G.; Chaudhary, G. R. Optimization and utilization of single chain metallocatanionic vesicles
for antibacterial photodynamic therapy (aPDT) against E. Coli, J. Mater. Chem. B, 2020, 8, 9304-9313.
Acknowledgements. This work was supported by the Commonwealth scholarship (INCN-2019-409), the European
Regional Development Fund (FEDER), Spanish government grant PID2020-115663GB-C32, and the University of
Nottingham.

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IL-7.4.1

Illuminating the plant calendar
Joshua M. Gendron
1) Yale University, Department of Molecular, Cellular, and Developmental Biology, New Haven, USA
Plants have served as a preeminent study system for photoperiodism due to their propensity to flower in concordancewith the seasons. A nearly singular focus on understanding photoperiodic flowering has prevented discovery of otherphotoperiod measuring systems necessary for vegetative health. Here we use bioinformatics to identify photoperiodinduced genes in Arabidopsis. We show that one, PP2-A13, is expressed exclusively in, and required for plant fitness
in, short winter-like photoperiods. We create a real-time photoperiod reporter, using the PP2-A13 promoter driving
luciferase, and show that photoperiodic regulation is independent of the canonical CO/FT mechanism forphotoperiodic flowering. We then reveal that photosynthesis combines with circadian clock-controlled starch
production to regulate cellular sucrose levels to control photoperiodic expression of PP2-A13. This work
demonstrates the existence of a photoperiod measuring system housed in the metabolic network of plants thatfunctions to control seasonal cellular health.
Acknowledgements. This work was supported by grants from the National Institutes of Health and National Science
Foundation

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IL-7.4.2
The molecular basis for day/night signaling in the cyanobacterial circadian clock
Carrie L. Partch1,2, Jeffrey A. Swan1, Colby R. Sandate3, Joel Heisler4, Alfred Freeberg1, Dustin Ernst5, Archana
Chavan4, Susan S. Golden2,5, Andy LiWang2,4, Gabriel C. Lander3
1) University of California Santa Cruz, Department of Chemistry & Biochemistry, Santa Cruz, USA; 2) Universityof California San Diego, Center for Circadian Biology, La Jolla, USA; 3) Scripps Research, Department ofIntegrative and Computational Structural Biology, La Jolla, USA; 4) University of California Merced, Departmentof Chemistry & Biochemistry, Merced, USA; 5) University of California San Diego, Division of Biological
Sciences, La Jolla, USAIn cyanobacteria, three Kai (or ‘cycle’) genes, KaiA, B and C, work together to generate circadian rhythms thatsynchronize its physiology with Earth’s day/night cycle. KaiC is the enzymatic driver of the clock with two tandem
AAA+ ATPase domains that assemble into two hexameric ATPase rings. During the day, KaiA stimulates
autophosphorylation of KaiC at two sites in the C-terminal ATPase domain, while KaiB binds at night to the Nterminal ATPase domain to sequester KaiA and allow KaiC to autodephosphorylate, completing one cycle of theclock1. KaiB association with KaiC is restricted to the evening by the phosphorylation state of KaiC, although themechanism by which this information is communicated ~70 angstroms through KaiC to the KaiB binding interface
is not known. Here we present structures of KaiC representing its daytime and nighttime states, obtained by cryoelectron microscopy, that reveal the structural basis of KaiB selectivity for the nighttime state. Mutants that disrupt
the allosteric transmission of information through KaiC reduce affinity for KaiB and link cooperative recruitmentof KaiB to ATP hydrolysis by KaiC, abrogating circadian rhythms in vitro and in vivo. This work highlights how
ultrasensitive signaling mechanisms contribute to day/night transitions in the cyanobacterial circadian clock.

Fig. 1 – The central timekeeping mechanism of the cyanobacterial circadian clock.

References
1) Swan, J.A., Golden, S.S., LiWang, A., and Partch C.L. Structure, function, and mechanism of the core circadianclock in cyanobacteria. Journal of Biological Chemistry, 2018.
Acknowledgements. This work was supported by grants GM121507 and GM141849 from the National Institutesof Health.

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IL-7.4.3
Superoxide is a metabolic signal that acts on the Arabidopsis circadian clock in theevening
Mike Haydon1
1) University of Melbourne, School of BioSciences, Parkville, Australia
Plants must coordinate photosynthetic metabolism with the daily environment and adapt rhythmic physiology anddevelopment to match carbon availability. Products of photosynthetic metabolism, including sugars and reactive
oxygen species (ROS), are closely associated with the plant circadian clock and sugars have been shown to provide
metabolic feedback to the circadian oscillator. However, distinguishing the effects of light and sugars inphotoautotrophic cells is challenging. We have used transcriptomics and chemical screens to explore regulation of
the circadian clock by sugars. From a sugar-regulated transcriptome of Arabidopsis we identified genes associatedwith redox and ROS processes. We found that sucrose increased levels of superoxide and identified circadianrhythms of superoxide-regulated transcripts which are phased around dusk, including several oscillator genes. Ourdata suggest a new role for superoxide as a rhythmic sugar signal which acts in the evening and affects circadiangene expression and growth.
Acknowledgements. This research was funded by BBSRC grant (BB/L021188/1), Royal Society Research Grant(RG150144) to MJH and by The University of Melbourne through the Research Grants Support Scheme to MJH
and a Melbourne Research Scholarship to XL.

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OC-7.4.4

Diurnal Rhythmicity of Cucumber Root Iron Deficiency Response Eliminates Rapidly
Upon Nanohaematite Iron Resupply
Amarjeet Singh1, Viktória Kovács Kis2 Áron Keresztes3, Fruzsina Pankaczi1, Brigitta Müller1, Ferenc Fodor1 Zoltán
Klencsár2, Ádám Solti 11) Department of Plant Physiology and Molecular Plant Biology, ELTE Eötvös Loránd University, Pázmány Péter
sétány 1/C, Budapest H-1117, Hungary
2) Centre for Energy Research, Konkoly-Thege Miklós út. 29-33, Budapest, H-1121, Hungary3) Department of Plant Anatomy, ELTE Eötvös Loránd University, Pázmány Péter sétány 1/C, Budapest H-1117,
Hungary
Iron (Fe) is an essential cofactor of redox enzymes in photosynthesis, respiration and antioxidative protection. Plants
take up Fe from the soil. Under alkaline conditions the oxidised ferrous Fe forms insoluble ferric oxide/hydroxide
precipitates that limits Fe acquisition of plants. Fe shortage leads to various dysfunctions, primarily impaired
photosynthetic functions, growth and development. To overcome Fe limitation, the majority of angiosperm plants
operate a reduction based Fe uptake strategy that also involves the acidification of the rhizosphere and the excretionof plant metabolites that contribute in the chelation and/or reduction of Fe. This reduction based Strategy I thus
primarily rely on the operation of Ferric Reductase Oxidase (FRO) family enzymes that perform a transmembraneelectron transport from cytoplasmic reductant NADH to ferric compounds. As a Strategy I plant, cucumber (Cucumis
sativus) expresses three root plasma membrane localised FROs viz. CsFRO1, CsFRO2 and CsFRO3 associated withroot ferric chelate reductase (FCR) activity. Cucumber belongs to the flavin excreting dicot plants: as response for
Fe deficiency, it excretes riboflavin derivative 5’-ketoriboflavin compounds that were previously suggested to be
involved in the reduction of insoluble ferric precipitates. Once Fe taken up into the roots, the root-to-shoot xylem
translocation requires citrate as for complexing compound. Taken together, both generating reducing power, and thebiosynthesis of flavins and organic acids requires a metabolic input. Previous studies on Populus sp. indicated that
Fe uptake of dicots is a diurnal activity. Here we applied an Fe deficient plant model system performing intensive Fe
uptake and nanohaematite suspension as for Fe source to establish a quasi-natural condition for Fe uptake and to
study the circadian rhythm dependent relaxing of the reduction based Fe acquisition system.
As for model, Fe deficient cucumber plants were cultivated in hydroponics with a complete retraction of Fe from themedium for three weeks from germination. Nanocrystalline haematite colloid suspension with a particle size ranging
from 10-20 nm was applied in a concentration equivalent to 20 µM nominal concentration of Fe as for Fe deficiency
recovery treatment starting at 9 am. Expression profile of CsFRO1, CsFRO2, CsFRO3 and CsRIBA1 (GTP
cyclohydrolase II) was analysed using quantitative real time PCR. The ferric chelate reductase (FCR) activity was
analysed using a bathophenanthroline disulfonate based ferrous Fe trapping method. Utilisation of the nanohaematite
particles was validated by microscopy energy-dispersive X-ray spectroscopy (EDS) on ultrathin cuts of root
segments. Expression of the genes of interest showed two peak circadian expression arrangement with markedly
increased expression at noon and evening for all genes exception of CsFRO3 in Fe deficient model plants. Utilizationof nanohaematite resulted in decrease in expression of CsFRO1, CsFRO3 and CsRIBA1 within 10 hours of treatment
but total decline to control level took 24 h. CsRIBA1 strongly responded to Fe deficiency and Fe supply. On the other
hand, decline in expression of CsFRO2 and activity of FCR took approximately 48 hours demonstrating a biphasic
suppression of Fe deficiency response. Besides nanohaematite particles were able to penetrate the root apoplast andintercellular space confirmed by EDS, transmission electron microscopy analysis verified the nanoparticles as for Fe
source with the reduction in their particle size. The rapid decline in the rhythmic expression of CsFRO1, CsFRO3
and CsRIBA1 underlines the effective reciprocal feedback effect of cellular Fe sensing mechanisms that overwrite
the rhythmicity in the transcript amounts.
Acknowledgements: This work was supported by the grants financed by the National Research, Development andInnovation Office, Hungary (NKFIH K-124159 and VEKOP-2.3.3-15-2016-00008). This work was completed aspart of the ELTE Thematic Excellence Programme 2020 supported by the National Research, Development andInnovation Office (TKP2020-IKA-05).

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IL-7.4.5
Evaluation of plant circadian rhythms based on the cellular circadian behaviour
Tokitaka Oyama1
1) Kyoto University, Department of Botany, Kyoto, JapanThe circadian clock gives time information to various physiological processes that would be involved in the daynight response. In plants, many genes are under control of the clock and their gene expression shows circadianrhythms even under constant conditions. The self-sustained oscillation is generated by transcription-translation
feedback loops composed of a number of clock genes in each cell. Hence the circadian clock system of plant is basedon the cell-autonomous machinery. Circadian rhythms of plants have been effectively analysed at thetissue/organ/body levels; circadian properties such as phase/period preciseness and entrainability of individual cells
are largely unknown.
In my laboratory, we developed the bioluminescence monitoring system that enabled us to observe circadianrhythms of individual cells in the plant1. A circadian bioluminescent reporter, AtCCA1:LUC was used for genetransfection to cells in a duckweed with a particle bombardment method. CCA1 is a clock gene of which expression
peaks in the morning. Using an EM-CCD camera system, the bioluminescence of individual cells was automaticallymonitored under various light/dark schedules1,2. Duckweed plants are suitable for imaging the entire plant becauseof their tiny, flat, floating bodies. Cellular circadian behaviour under constant conditions were heterogeneous amongcells in the same frond (the plant unit of duckweed) of Lemna gibba. The heterogeneous period lengths affected theheterogeneous entrainment among individual cells in the frond under non-24 h light/dark cycles. Even though
individual cells showed a high independency in the circadian behaviour, spatial phase patterns that were a sign ofcell-cell interaction (coupling between cellular rhythms) were observed in the fronds.
The CCA1:LUC reporter has been widely utilized to study circadian rhythms of various plants. This reporter
represents the oscillation of a clock gene, that is, the behaviour of the circadian clock. Recently, we have found thata luciferase gene under the control of a “constitutive” promoter (e.g. CaMV35S:LUC) showed a circadianbioluminescence rhythm when introduced into duckweed plants. A dual-colour bioluminescence monitoring system
at a single-cell level (simultaneous monitoring of cellular bioluminescence from two different-colour reporters)
revealed that the CaMV35S:LUC bioluminescence rhythm and the AtCCA1:LUC one showed different period lengths
and phase-relationship among cells in the same frond3. The CaMV35S:LUC bioluminescence rhythm of a cell looked
to be associated with the synchrony around it without the direct control of its cellular clock. Since the synchrony ofcellular clocks is highly associated with the coupling phenomena, the cellular bioluminescence behaviour of
CaMV35S:LUC will be a tool to analyse the organization of circadian behaviour in tissues/organs.
Cellular circadian behaviour in a tissue/organ is influenced by other cells/tissues; the cellular nature ofcircadian clock would be inaccessible. Then, we have approached it by observing bioluminescence rhythms ofindividual protoplast-derived cells isolated from the Arabidopsis CCA1:LUC transgenic plant. Even under the highly
artificial conditions, leaf- and root-derived cells showed typical properties of the circadian rhythm. The differencesin the properties between these two cell-types seemed to relate to their adaptiveness in the totally different local
environments for leaves and roots.
In this paper, I will present our recent results of cellular circadian rhythms of plants and discuss thecharacteristics of the plant circadian system as a cell-based organization.
References
1) Muranaka T, Oyama T, Heterogeneity of cellular circadian clocks in intact plants and its correction under lightdark cycles. Sci. Adv., 2, e1600500., 2016.
2) Okada M, Muranaka T, et al., Synchrony of plant cellular circadian clocks with heterogeneous properties underlight/dark cycles. Sci. Rep., 7, 317., 20173) Watanabe E, Isoda M, et al., Detection of uncoupled circadian rhythms in individual cells of Lemna minor using
a dual-color bioluminescence monitoring system. Plant Cell Physiol., in press, 2021.
Acknowledgements. This work was supported in part by the Japan Society for the Promotion of Science KAKENHI(Grant numbers 25650098, 17KT0022, and 19H03245).
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for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-7.4.6
Integration of circadian and environmental signals that regulate chloroplast gene
expression
Antony N. Dodd1
1) John Innes Centre, Norwich, UK.
Photosynthesis within chloroplasts is essential for virtually all plants, including the crops that provide our food.
Photosynthesis is regulated by both circadian rhythms and a variety of environmental cues. Understanding themechanisms through which environmental cues regulate photosynthesis is important to predict the effects upon
photosynthesis of future climates, and to identify potential targets for crop improvement. We are investigating thenature of the signalling mechanisms in plants that integrate circadian and environmental cues, and communicate
this information to chloroplasts (1, 2). We reason that plants must integrate circadian timing cues with information
concerning unpredictable changes in their environment in order to respond to predictable (24 h) and unpredictablechanges in the environmental conditions. We investigated this idea within the context of signalling from thenucleus to chloroplasts, by examining the integration of circadian, light and temperature signals by a group ofproteins called sigma factors. In plants, sigma factors are nuclear-encoded regulators of chloroplast transcription. I
will show new results identifying a key role for a sigma factor in the integration of circadian and low temperature
signals, and position this within the upstream and downstream components of the signalling pathway, and its
impacts upon plant physiology.
References
1) Belbin FE, Noordally ZB, Wetherill SJ, Atkins KA, Franklin KA, Dodd AN. Integration of light and circadiansignals that regulate chloroplast transcription by a nuclear-encoded sigma factor. New Phytol. 213, 727-738 (2017).
2) Noordally ZB, Ishii K, Atkins KA, Wetherill SJ, Kusakina J, Walton EJ, Kato M, Azuma M, Tanaka K,
Hanaoka M (2013). Circadian control of chloroplast transcription by a nuclear-encoded timing signal. Science 339,
1316-1319. (2013).
Acknowledgements. This research was funded by BBSRC (UK) (BB/I005811/2, BB/J014400/1; studentship
1518540; Institute Strategic Programme GEN BB/P013511/1), The Leverhulme Trust (RPG-2018-216), NAGASE
Science Technology Foundation (Japan), the Ministry of Education, Culture Sports, Science and Technology(Japan) (Grants-in-Aid 17K07438), Tokyo Institute of Technology World Research Hub Initiative (WRHI)
Program of Institute of Innovative Research and the Consejo Nacional de Ciencia y Tecnología (Mexico).

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for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-7.4.7
TOC1 phosphorylation controls formation and function of an NF-TOC1 complex
regulating hypocotyl growthDavid E Somers1, Jiapei Yan1
1) Ohio State University, Department of Molecular Genetics, Columbus, USA
Plant photoperiodic growth is coordinated by interactions between circadian clock and light signaling networks. How
post-translational modifications of clock proteins affect these interactions to mediate rhythmic growth remains
unclear. Here we identify five phosphorylation sites in the core clock protein TIMING OF CAB EXPRESSION 1(TOC1) which when mutated to alanine (5X) eliminate detectable phosphorylation. 5X is unable to fully rescue theclock, growth and flowering phenotypes of the toc1 mutant. 5X exhibits advanced phase, a faster degradation rate,
reduced interactions with PHYTOCHROME INTERACTING FACTOR3 (PIF3) and HISTONE DEACETYLASE
15 (HDA15), and poor binding at pre-dawn hypocotyl-growth-related genes (PHGs), leading to de-repression of
hypocotyl growth in the middle of the night. NUCLEAR FACTOR Y subunits B and C (NF-YB/C) stabilize TOC1
at target promoters and this novel trimeric complex (NF-TOC1) acts as a transcriptional co-repressor with HDA15
to inhibit hypocotyl elongation. Collectively, we identify a molecular mechanism of how TOC1 phosphorylation
alters its phase, stability and physical interactions with co-regulators to precisely phase PHG expression to control
photoperiodic hypocotyl growth.

Fig. 1 -- Proposed mechanism for the role of TOC1 phosphorylation in the control of hypocotyl growth. At early night, wild-type(left) and 5X (right) TOC1 are equally present and recruit transcriptional repressors NF-YB/C and HDA15 to inhibit expressionof PHGs. As night progresses, PIF3 levels rise, and the advanced phase of 5X, more rapid degradation and poorer PIF3
interaction result in lesser presence of 5X at target promoters, relative to wild-type TOC1, leading to attenuated inhibition of
PHGs and enhanced hypocotyl growth. At late night, low TOC1 levels reduce NF-YB/C and HDA15 recruitment, resulting in
PIF3 release to activate downstream PHGs leading to maximal pre-dawn hypocotyl growth.

Acknowledgements. This work was supported by the National Institutes of Health (R01GM093285 and to D.E.S.),
the Next-Generation BioGreen21 Program (PJ01327305), the Rural Development Administration, Republic of Korea(to D.E.S.) and the World Class University Program of South Korea (No. R31-2008-000-10105-0), NRF, MEST (toD.E.S.).

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-7.4.8

Interplay between circadian clock and unfolded protein response in Arabidopsis
thaliana
Ozlem GUMUS1, Buse PINAR1, Baris UZILDAY1, Ismail TURKAN1, Rengin OZGUR1
1) Ege University, Faculty of Science, Department of Biology, Izmir, 35100, Turkey
The movement of Earth generates light and temperature change in environment and organisms such as plants havedeveloped molecular oscillators to adapt these conditions1. Circadian clocks are internal timekeeping systems thatdrive daily rhythm in physiology and metabolism due to changeable environmental conditions 2. Abiotic stress suchas high temperature or salt stress are the example of those conditions, and these environmental fluctuations cancause endoplasmic reticulum stress (ER stress) by increasing the misfolded or unfolded proteins in the ER 3. ER
stress induces a specific mechanism within the cell called unfolded protein response (UPR) which stimulates
transcription of a series genes that increase the protein folding capacity in the ER4 Aim of this work was toinvestigate interaction among circadian clock and protein folding capacity and ER stress response. For this aim,
Arabidopsis thaliana plants were trained to 12/12 h (LD) and to 24 h constant light (LL) to determine their
response to tunicamycin induced ER stress. Moreover, expressions of genes related to UPR were measured at 4,8,
12, 16 and 24 h time points after onset of ER stress. In addition, we also investigated if ER stress can change period
or amplitude of A. thaliana molecular clock. As results, Tm enhanced the expressions of ER stresssensor/transducer genes (bZIP17, bZIP28, bZIP60, IRE1A, IRE1B), ER chaperones and folding helper genes (BiP1,
BiP3, ERO1, CNX), ER-associated degradation (ERAD) genes (DER1, SEL, HRD1), and ER stress associatedapoptosis genes (AGB1, NAC089) in shoots. LL and LD treatments have different effects on UPR of A. thaliana
whereas LL grown plants showed decreased ER stress response as compared to LD grown plants.
References
1) Ding, Z., Millar, A. J., Davis, A. M., & Davis, S. J. (2007). TIME FOR COFFEE encodes a nuclear regulator in
the Arabidopsis thaliana circadian clock. The Plant Cell, 19(5), 1522-1536.
2) Haydon, M. J., Mielczarek, O., Robertson, F. C., Hubbard, K. E., & Webb, A. A. (2013). Photosynthetic
entrainment of the Arabidopsis thaliana circadian clock. Nature, 502(7473), 689-692.
3) Moreno, A. A., & Orellana, A. (2011). The physiological role of the unfolded protein response inplants. Biological research, 44(1), 75-80.
4) Ozgur, R., Uzilday, B., Iwata, Y., Koizumi, N., & Turkan, I. (2018). Interplay between the unfolded protein
response and reactive oxygen species: a dynamic duo. Journal of experimental botany, 69(14), 3333-3345.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-8.1.1
COVID-19 through a UV lensShelley Gorman1
1) Telethon Kids Institute, University of Western Australia, Perth, Australia
In response to the COVID-19 (coronavirus disease of 2019) pandemic, researchers have been seeking low-cost andaccessible means of providing protection from its harms, particularly for at-risk individuals such as those withcardiovascular disease, diabetes and obesity. One possible way is via safe sun exposure, and/or dietary
supplementation with induced beneficial mediators, such as vitamin D and nitric oxide. Potential benefits could be
mediated through the disinfectant properties of UV radiation, and its capacity to modulate immunity and limit
cardiometabolic dysfunction. Anti-inflammatory effects of UV-induced mediators might limit the extent of the
COVID-19 cytokine storm to reduce morbidity. There are also likely important UV dose considerations. While
there has been significant commentary on potential benefits for vitamin D, the direct effects of exposure to UV
light on COVID-19 has received less attention. Data collected to-date suggests that ambient levels of both UVA
and UVB may be beneficial for reducing severity or mortality due to COVID-19. Currently unresolved are thenature of the associations between blood 25(OH)D and COVID-19, with more prospective data needed that better
considers lifestyle factors, such as physical activity and personal sun exposure levels. Indeed, there has been
limited data collected on the associations between personal sun exposure and COVID-19-related outcomes. Many
clinical trials are underway testing the potential for vitamin D supplementation or inhaled nitric oxide in treating
COVID-19 and related outcomes. While some protective effects are hypothesized for UV light (and induced
mediators) for improving respiratory and cardiometabolic health during COVID-19, more evidence is needed todemonstrate causality1.
References
1) S. Gorman, R. B. Weller, Investigating the potential for ultraviolet light to modulate morbidity and mortality fromCOVID-19: A narrative review and update. Frontiers in Cardiovascular Medicine, 7, Article 616527, December 2020.

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for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-8.1.2

UV, blood pressure and cardiovascular disease
Richard B Weller,
1)Centre for Inflammation Research, University of Edinburgh, UK.; 2) Department of Dermatology, Royal
Infirmary of Edinburgh, UK
Excess sunlight is a risk factor for skin cancer in white skinned individuals, but there are no data linking it withincreased all-cause mortality. By contrast, high blood pressure (BP) is the leading global cause of disability
adjusted life years lost. A 5 mmHg fall in BP, even from a ‘healthy’ level produces a 10% reduction in major
cardiovascular events1. Population BP at national level correlates with latitude, and in temperate countries
population winter BP is 2-3 mmHg lower than summer. Prospective data from Sweden show that increased sunlight
exposure independently correlates with reduced cardiovascular and all-cause mortality after 25 years2.
The well-known seasonal variation in BP and incident cardiovascular events has historically been ascribed totemperature and vitamin D synthesis.
I performed an observational study of 340,000 patients on renal dialysis, in >2,000 centres in the USA. BP wasmeasured thrice weekly for 3 years in these patients, and cross referenced against satellite derived UV irradiation
data at each location throughout this time, as was temperature. BP correlated inversely with UV and this effect was
independent of temperature, although temperature had an independent additive effect. BP was higher in the Black
American than White American patients and the UV related fall was attenuated3.
Physiological formation of biologically active 1,25 dihydroxyvitamin D3 requires UVB exposure. Measured
vitamin D levels correlate inversely with incident cardiovascular disease, yet meta-analyses of extensive studies oforal vitamin D supplementation show that vitamin D plays no part in maintaining cardiovascular health 4.
Mendelian randomisation studies confirm this. Serum vitamin D levels are also a marker of sunlight exposure.
UVA mobilises nitric oxide from stores in the skin to the systemic circulation where it is an arterial vasodilator5.
This vitamin D independent mechanism probably mediates UV’s action on cardiovascular disease.
I have recently completed a randomised cross-over, sham controlled, single blinded clinical trial of the use of UVA
lamps to treat patients with mild hypertension. Treatment was whole body irradiation with 5J/cm2 UVA from ahome phototherapy lamp, daily for two weeks. After a 2-4 week washout period, sham irradiation with theidentical lamps screened to block wavelengths <500nm. The sequence of treatments was randomised. Clinic BP
was measured at the start and end of active and sham irradiation periods. 14 patients were recruited. Mean arterial
BP fell significantly more in the active (-8 ± 2.9mmHg) than control (+1 ± 3 mmHg p=0.033) arm of the trial. Thechange in BP was more marked for patients recruited in the darker (autumnal equinox to vernal equinox) thanlighter half of the year.
Deaths from hypertension related disease in the UK are around 60 times more common in the UK than skin cancer
deaths. Epidemiological, mechanistic and now interventional data all show an effect of sunlight or ultraviolet
radiation in lowering BP. Advice on sun exposure needs to consider benefits as well as hazards.
References

1. Rahimi K, Bidel Z, Nazarzadeh M, et al. Pharmacological blood pressure lowering for primary and secondary

prevention of cardiovascular disease..... The Lancet 2021
2. Lindqvist PG, Landin-Olsson M. The relationship between sun exposure and all-cause mortality. Photochem
Photobiol Sci 2017;
3. Weller RB, et al. Does Incident Solar Ultraviolet Radiation Lower BP? J Am Heart Assoc 2020;
4. Weller RB. Beneficial Effects of Sunlight May Account... JAMA Cardiol 2020;
5. Liu D, Fernandez BO, Hamilton A, et al. UVA irradiation of human skin vasodilates arterial vasculature andlowers blood pressure independently of nitric oxide synthase. J Invest Dermatol 2014;
Acknowledgements. This work was part supported by project grant PG/15/23/31362from the British Heart
Foundation.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-8.1.3
Myopia progression during COVID-19 lockdown and UV deprivation
Seyhan Yazar1,2
1) Garvan Institute of Medical Research, Garvan-Weizmann Centre for Cellular Genomics, Sydney, Australia; 2)
Lions Eye Institute, Genetics and Epidemiology, Perth, Australia
Staying at home has become the new norm as a result of COVID-19. All of us had to reduce our world to the four
walls of our houses for weeks if not for months. Most school children are learning at home, online. This newenvironment has major implications on eye health. Myopia or short-sightedness is linked to spending lots of timelooking at near objects and very little time looking at distant objects. In the pre COVID-19 era, myopia prevalence
was on the rise and this increase has been linked to children spending too much time inside and studies have shownthat encouraging children to spend more time outside can prevent myopia.1,2 There are multiple hypotheses explaining
this protective effect including exposure to the bright sunlight and UV. In this talk, the most recent myopia rates andeffect of UV deprivation on these rates will be discussed.
References
1) He M, Xiang F, et al. Effect of Time Spent Outdoors at School on the Development of Myopia Among Children
in China: A Randomized Clinical Trial. JAMA, 2015.
2) Wu Pei-Chang, Chueh-Tan Chen MS et al. Myopia Prevention and Outdoor Light Intensity in a School-Based
Cluster Randomized Trial. Ophthalmology, 2018.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-8.1.4

Evaluation of blue light protection of the skin
Terje Christensen1,2, Bjørn J. Johnsen 2, Ellen M. Bruzell 31) University of South-Eastern Norway, Drammen, Norway; 2) Norwegian Radiation and Nuclear Safety Authority(DSA), Østerås, Norway; 3) Nordic Institute of Dental Materials (NIOM), Oslo, Norway
Visible-light effects on the skin have been regarded less hazardous than effects of ultraviolet radiation (UV).
Uncertainties exist about the limit values for visible exposure, types of effects, and the action spectrum ofchromophores that may give rise to unwanted effects. DNA is assumed to be the most important chromophore for
UV, while several others absorbing visible light are distributed in the tissues, some of which are naturally occurringand some exogenous, resulting from e.g. pollution, use of cosmetics or photoactive medication. In the absence ofexact knowledge, industry markets various protective products. The main aim of this presentation is to evaluate
whether protection against blue light is needed and if so, in what situations and for which groups.
We have measured the irradiance values for several typical general lighting sources and electronic screens, such asof mobile phones and personal computers. The lowest dose of blue light reported to cause biological effects innormal skin was estimated from a literature study. The estimated dose was used to compute the minimum exposure
times to light from various sources necessary for obtaining reported skin effects (Tthres). Corresponding exposure
times were calculated for outdoor conditions and a selection of medical devices.
The Tthres for a range of biological effects on human skin and skin cells was between 5 and 200 Jcm-2, and the only
consistently reported effect, pigmentation, was observed after an average exposure of 65 Jcm-2. This dose will bereached by at least one month of continuous exposure to electronic screens or two weeks in a 500 lx LED-lit officeenvironment. In comparison, Tthres is reached in 2 h at noon mid-summer in Oslo (at UV-index 6, which requires
protection against UV-damage to the skin). Medical sources intended for light curing of dental materials emit blue
light which is 100 times stronger than the maximum outdoor values in Oslo; however, the exposure conditions
cannot be compared1.
We conclude that for the normal, healthy population, UV protection of the skin is essential, compared with thesuggested use of protection against blue light. Exposure to indoor general lighting and electronic screens will not
exceed doses that can induce skin effects within practical exposure times. Exceptions may exist for e.g.
photosensitizing drug use or exposure to exogenous substances.
Reference
1) Christensen, T., Johnsen, B.J. & Bruzell, E.M. Violet-blue light exposure of the skin: is there need forprotection? Photochem Photobiol Sci 20, 615–625 https://doi.org/10.1007/s43630-021-00043-9, 2021.

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for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-8.1.5
Solar ultraviolet radiation slows lung function decline in premenopausal women in alongitudinal and population-based cohort (ECRHS)
Kai Triebner (PhD)1, 2, Steinar Hustad (MD)1, 2, Jose Barrera-Gómez (MSc)3, 4, 5, Xavier Basagaña (PhD)3, 4,
5,Bryndís Benediktsdóttir (MD)6,7, Shyamali Dharmage (MD)8, Karl A Franklin (MD)9, Elaine Fuertes (PhD)10,
Vanessa Garcia-Larsen (PhD)11, Jose Gullón-Blanco (MD)12, Rain Jõgi (MD)13, Bénédicte Leynaert (PhD)14,15, Eva
Lindberg16, Jesús Martínez-Moratalla (MD)17, 18, Nerea Muniozguren Agirre (MD)19, Isabelle Pin (MD)20, 21,
Chantal Raherison (MD)22, Antonio Pereira-Vega (MD)23, Vivi Schlünssen (PhD)24, 25, Antonia Valentin (PhD)3, 4, 5,
Francisco Gómez Real (MD)1, 26* and Payam Dadvand (PhD)3, 4, 5** The authors contributed equally.
1

University of Bergen, Jonas Liesveg 92, 5021 Bergen, Norway; 2Core Facility for Metabolomics, Jonas Liesveg
92, 5021 Bergen, Norway; 3ISGlobal, Carrer del Dr. Aiguader, 88, 08003 Barcelona, Spain; 4Universitat Pompeu
Fabra, Plaça de la Mercè, 10, 08002 Barcelona, Spain; 5CIBER Epidemiología y Salud Pública, Avenida Monforte
de Lemos 3-5, 28029 Madrid, Spain; 6Faculty of Medicine, University of Iceland, Reykjavik, Iceland; 7Department
of Sleep, Landspitali University Hospital, Hringbraut 101, Reykjavík, Iceland; 8Allergy & Lung Health Unit,
Melbourne School of Population Health, Melbourne University, 207 Bouverie Street, 3053 Carlton, Australia;
9
Department of Surgical and Perioperative Sciences, Surgery, Umea University, 901 87 Umeå, Sweden;
10
Department of Respiratory Epidemiology, Occupational Medicine and Public Health, Imperial College, South
Kensington, London, UK; 11Department of International Health, Johns Hopkins University, Bloomberg School of
Public Health, 615 N Wolfe Street, Baltimore, USA; 12Department of Pneumology, Hospital San Agustin de Aviles,
Camino de Heros 6, 33401 Avilés, Asturias, Spain; 13Department of Lung Medicine, Tartu University Clinic,
Ludvig Puusepa 1a, 50406 Tartu, Estonia; 14Inserm-U1168, VIMA, 16 Avenue Paul Vaillant Couturier, 94807
Villejuif, France; 15UMR-S 1168, UVSQ, Université Versailles St-Quentin-en-Yvelines, 55 Avenue de Paris, 78000
Versailles, France; 16Department of Medical Sciences, Uppsala University, Akademiska sjukhuset, 75185 Uppsala,Sweden; 17Pulmonology Service, Albacete University Hospital, Health Service Castilla - La Mancha, C. Hermanos
Falco 37, 02008 Albacete, Spain; 18Medicine School of Albacete, University of Castilla-La Mancha, Calle
Almansa, 14, 02008 Albacete, Spain; 19Unit of Epidemiology and Public Health, Department of Health, Basque
Government, Gran Vía 85, 48011 Bilbao, Spain; 20Department of Pediatrics, University Hospital Grenoble Alpes,
Inserm, Avenue Maquis du Grésivaudan, 38700 La Tronche, France; 21Institute for Advanced Biosciences,
Grenoble; University Grenoble Alpes, CS 10217, 38043 Grenoble cedex 9, France; 22U1219, Bordeaux Population
Health Research, Bordeaux University, 146 rue Leo Saignat, 33076 Bordeaux, France, 23Service of Pneumology
and Allergy, University Hospital Juan Ramón Jiménez, Ronda Exterior, Paseo Nte., 21005 Huelva, Spain;
24
Department of Public Health, Environment, Occupation and Health, Danish Ramazzini centre, Aarhus
University, Palle Juul-Jensens Boulevard 99, 8200 Aarhus, Denmark; 25National Research Center for the Working
Environment, Lersø Parkallé 105, 2100 Copenhagen, Denmark; 26Department of Gynecology and Obstetrics,
Haukeland University Hospital, Jonas Liesveg 92, 5021 Bergen, Norway;
Introduction: During the mid-twenties a woman’s lung function starts declining and this decline accelerates after
menopause.1 The slope of decline may be influenced by exposure to solar ultraviolet radiation through its effects on
inflammatory status and endogenous vitamin D concentrations. 2,3
Objectives: To explore associations between exposure to solar ultraviolet radiation and lung function decline, before
and after menopause, and potential effect modification by vitamin D intake through dietary sources.
Methods: Using 10-year follow-up data from the international European Community Respiratory Health Survey andsatelite data on the daily effective ultraviolet irradiance during this period, we modelled individual exposure to solar
ultraviolet radiation. Forced vital capacity (FVC) and forced expiratory volume in one second (FEV1) were measured
at baseline and follow-up. Linear mixed effects regression models with lung function change as outcome and solar
ultraviolet radiation as exposure were fitted and adjusted for potential confounders.
Results: An increase of one interquartile range in solar ultraviolet radiation exposure was associated with a reduced
FVC decline of -3.3mL/y (95%CI: -6.5 to -0.1mL/y) in the whole study population. This association was driven byBook of Abstracts ����������������������������������������������������������������������������������������������� 353

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austriapremenopausal women (-8.1mL/y, -14.7 to -1.5mL/y) and was independent of dietary vitamin D intake. No
statistically significant associations were observed for FEV1.
Conclusions: Exposure to solar ultraviolet radiation is associated with reduced lung function decline in women. This
is of public health interest and a previously unknown health benefit of solar ultraviolet radiation.

No. of subjects

Estimate (95% CI)

p-Value

202
80
234
152

−8.1 (−14.7 to −1.5)−4.5 (−11.9 to 2.9)
−1.9 (−7.3 to 3.4)
−0.9 (−6.1 to 4.4)

0.022
0.277
0.497
0.751

202
80
234
152

−0.9 (−5.0 to 3.1)
−2.0 (−7.2 to 3.2)
−2.2 (−6.6 to 2.9)
−0.1 (−3.9 to 3.7)

0.668
0.487
0.356
0.975

202
80
234
152

−1.2 (−2.4 to −0.1)−0.1 (−1.8 to 1.6)
0.3 (−0.7 to 1.4)
−0.1 (−1.0 to 0.8)

0.048
0.911
0.540
0.838

Annual FVC decline [mL/year]
Premenopause
Menopausal transition
Early postmenopause
Late postmenopause
Annual FEV1 decline [mL/year]
Premenopause
Menopausal transition
Early postmenopause
Late postmenopause
Annual change of FEV1/FVC [%]Premenopause
Menopausal transition
Early postmenopause
Late postmenopause

−10

−5

0

5

10

Fig. 1 – Forest plot of the results for FVC decline, FEV1 decline and change of FEV1/FVC over 10 years of follow-up, associatedwith one interquartile range increase in UVR exposure; adjusted for age, weight, height, skin type, dietary vitamin D intake, age
at completed full-time education (as socio-economic proxy), difference in weight, pack-years, population sample and spirometer,
including 95% confidence interval (negative values represent reduced decline);

References

1.
Triebner K, Matulonga B, Johannessen A, et al. Menopause Is Associated with Accelerated Lung
Function Decline. American journal of respiratory and critical care medicine 2017; 195(8): 1058-65.
2.
Brumpton BM, Langhammer A, Henriksen AH, et al. Vitamin D and Lung Function Decline in
Adults With Asthma: The HUNT Study. American Journal of Epidemiology 2016; 183(8): 739-46.
3.
Thuesen BH, Skaaby T, Husemoen LLN, Fenger M, Jørgensen T, Linneberg A. The association ofserum 25-OH vitamin D with atopy, asthma, and lung function in a prospective study of Danish adults.
Clinical & Experimental Allergy 2015; 45(1): 265-72.
Acknowledgements. Kai Triebner received a postdoctoral fellowship from the University of Bergen. Payam
Dadvand is funded by a Ramón y Cajal fellowship (RYC-2012-10995) awarded by the Spanish Ministry of Economy
and Finance. The present analyses are part of a project funded by the Norwegian Research Council (Project No.
228174). The coordination of ECRHS 2 was supported by the European Commission, as part of their Quality of Life
program, (Grant code: QLK4-CT-1999-01237). The coordination of ECRHS 3 was supported by the Medical
Research Council (Grant No. 92091). Bodies funding the local studies are listed in the online data supplement
sections 1 and 2.

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IL-8.1.6
Melanin, DNA photodamage and vitamin D synthesis
Antony R Young
King’s College London, St John’s Institute of Dermatology, London, UK
Exposure to solar ultraviolet radiation (UVR) has beneficial as well as adverse effects, and it is important to optimise
the risk benefit ratios of these effects under different conditions. Human epidermis contains melanin, the quantity ofwhich depends on Fitzpatrick skin type (FST). Light skinned FST I/II individuals have very low levels of constitutive
melanin whereas very high levels are present in FST VI with black skin. Irrespective of FST, melanin shows agradient with the highest concentration in the basal layer that contains melanocytes and keratinocyte stem cells.
Epidermal DNA photodamage with mutagenic and carcinogenic potential, specifically cyclobutane pyrimidine
dimers (CPD), and vitamin D synthesis are acute responses to solar UVR. These responses are very dependent on
melanin as determined in studies comparing FST I/II with FST VI. In the case of CPD1, the previously unexposed
buttock skin of healthy young volunteers was exposed to single comparably erythemal doses of solar simulated
radiation (SSR) and CPD were assessed immediately after exposure in three different epidermal zones; basal, middle
and upper. Protection factors by melanin against CPD were estimated in each zone and shown to be ~60 for the basal
layer, ~17 for the middle layer and ~5 for the upper layer, and the level of protection was related to melaninconcentration. The very high level of DNA protection afforded by melanin in the basal layer is the likely reason forthe large difference in skin cancer incidence in black and white skins.
The chromophore for vitamin D synthesis is epidermal 7-dehydrocholesterol (7-DHC). The effect of melanin on
vitamin D synthesis was compared in black and white skins after 5 serial exposures (with 3-4 day intervals) of wholebody (85% body surface area) fluorescent SSR or TL01 (monochromatic UVB at ~311nm)2. The exposure doses
were the same for all skin types and were sub-erythemal for FST I/II. Blood samples were taken before, during andafter the exposure protocol and assessed for 25(OH)D3 (gold standard for vitamin D synthesis) by HPLC tandem
mass spectrometry. These data were used to construct linear dose response curves and the ratio of the black vs whiteslopes was used to give an index of melanin inhibition of vitamin D synthesis. With both UVR spectra the melanininhibition factor was <1.5. The most likely reason for this very low value is that there is ample 7-DHC above themelanin rich basal layer in black skin to allow good vitamin D synthesis.
Taken together, these studies shown that the inhibitory role of melanin in deeply pigmented skin depends on its
location and quantity within the epidermis, and how it interacts with different chromophores. In the case of CPD, thechromophore is nuclear DNA and melanin is highly protective in the melanin rich basal layer. In the case of vitaminD, there is clearly sufficient chromophore (i.e., 7-DHC) distributed above the basal layer to allow good vitamin D
synthesis, such that the inhibitory effect of melanin is very small compared to its effect on nuclear DNA. This meansthat any discussion on the effects of melanin on photobiological outcomes must be specific not only for outcome, butalso the location in the epidermis at which that outcome occurs.
Finally, these results have public health implications. People with deeply pigmented skins, who often have suboptimal vitamin D status, may be encouraged to improve their vitamin D status by spending more time in the sun
because their risk of skin cancer is very low. The high incidence of skin cancer in those with light skins can only be
reduced to that of black skins if their level of routine photoprotection is very high, though small sub-erythemal dosesof solar UVR are necessary for vitamin D synthesis.
References
1) D. Fajuyigbe, S. M. Lwin, B. L. Diffey, R. Baker, D. J. Tobin, R. P. E. Sarkany and A. R. Young, Melanin
distribution in human epidermis affords localized protection against DNA photodamage and concurs with skin cancer
incidence difference in extreme phototypes, FASEB J, 32, 3700-3706, 2018.
2) A. R. Young, K. A. Morgan, T. W. Ho, N. Ojimba, G. I. Harrison, K. P. Lawrence, N. Jakharia-Shah, H. C. Wulf,
J. K. Cruickshank and P. A. Philipsen, Melanin has a small inhibitory effect on cutaneous vitamin D synthesis: A
comparison of extreme phenotypes, J Invest Dermatol, 140, 1418-1426 e1411, 2020.

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IL-8.1.7

UVR, B cells and regulation of the development of multiple sclerosis
Prue Hart1, Stephanie Trend1,2, Jonatan Leffler1, Scott N Byrne3,4, Allan G Kermode2,5, Martyn A French6
1)Telethon Kids Institute, University of Western Australia, Perth, WA, Australia; 2)Centre for Neuromuscular and
Neurological Disorders, Perron Institute for Neurological and Translational Science, University of Western
Australia, Perth, WA, Australia; 3)The University of Sydney, School of Medical Sciences, Faculty of Medicine and
Health, NSW, Australia; 4)Westmead Institute for Medical Research, Centre for Immunology and Allergy
Research, Westmead, NSW Australia; 5)Institute for Immunology and Infectious Disease, Murdoch University,
Perth, WA, Australia; 6)UWA Medical School and School of Biomedical Sciences, University of Western Australia,
Perth, WA, Australia
Reduced sun exposure is a risk factor for development of multiple sclerosis (MS). To investigate the beneficial
effect of UV radiation on MS, the PhoCIS trial (Phototherapy for Clinically Isolated Syndrome) was establishedwhereby participants with the earliest form of MS, namely Clinically Isolated Syndrome (CIS), were given a courseof suberythemal narrowband UVB (311-312 nm) phototherapy over 8 weeks (3 sessions/week). Both thosereceiving phototherapy and those in the control group (no phototherapy) were followed for 12 months by magnetic
resonance imaging (MRI) and by venesection at regular intervals. When the primary outcome of MRI changes was
examined, there was a non-significant reduction (100% to 70%) in the rate of conversion from CIS to MS within 12
months in participants who received phototherapy. UVB-induced changes in the circulating immune cells of theparticipants were investigated to illuminate the mechanisms of action of narrowband UVB, and whether they
contributed to reducing CIS to MS progression.
The most substantial changes over the first 2 months of phototherapy involved B cells, a cell type whose
importance has been highlighted by the successful use of anti-CD20 monoclonal antibodies to treat MS. There wasa significant decrease in CD27+/IgD- memory B cells (MBC) and a significant increase in naïve B cells(CD27-/IgD+) (both as a % of B cells) in participants that received phototherapy compared with those who did not.
In a study of the function of circulating B cells following phototherapy, responses to an innate B cell stimulus,
resiquimod (R848) via the production of TNF and IL-10 were investigated. TNF is not only a pro-inflammatory
cytokine but is important as a co-factor supporting B cell-T cell and B cell-myeloid cell interactions. IL-10, in
contrast, is the prototypic anti-inflammatory cytokine and has been shown to be important for identification ofhuman B cells with immune regulatory properties.
Using multi-parametric flow cytometry and regression analysis, significantly fewer B cells produced TNF in
response to R848, whilst there were no significant changes detected in the capability of B cells to produce IL-10 in
response to a TLR7 ligand. Thus, narrowband UVB phototherapy reduced B cell responses in vitro to a TLR7
ligand. This study highlights that narrowband UVB phototherapy exerts systemic immunological effects beyond theskin. We propose that the B cell function curtailed by phototherapy is pathogenically important and may reflect
priming effects of phototherapy that allow for reduced responses by B cells to polyclonal activation.

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OC-8.1.8

Determining the efficacy of a popular natural homemade sunscreen promoted bywellness bloggers on social media.
Furkan A. Ince1, James McClelland1, Katie M. Dixon1.
1) School of Medical Sciences, Faculty of Medicine and Health, The University of Sydney, NSW 2006

Australia.

In Australia, the use of commercially available sunscreens is strongly recommended by health authorities to protectfrom the harmful effects of ultraviolet (UV) radiation, including keratinocyte cancers and melanoma. However,these commercial sunscreens contain some ingredients associated with negative impacts on health and theenvironment. For example, oxybenzone has been shown to have adverse effects on endocrinological function infish and rats and additionally can contribute to coral bleaching. Furthermore, some ingredients in commercial
sunscreens may cause contact dermatitis in humans. Subsequently, this has led to rising popularity in natural
homemade sunscreens. Numerous natural sunscreen recipes have been published online by different wellness
bloggers with a large following. However, these natural sunscreens lack the backing of scientific evidence for theireffectiveness. We tested the efficacy of a natural sunscreen published online by a wellness blogger with 694,000
followers on Facebook and 32,000 followers on Twitter, with the aim of determining whether this natural sunscreen
is indeed as photoprotective as commercial SPF50+ sunscreens. The sunscreen contained (v/v) almond oil 39 %,coconut oil 19 %, shea butter 10%, beeswax 19 %, red raspberry seed oil 1.6 %, carrot seed oil 1.6 % and zincoxide 5 %. Ex vivo skin samples were obtained with consent from patients undergoing elective surgery. Skinsamples were cut into equal pieces and were then treated with either a commercial SPF50+ sunscreen, natural
sunscreen or base lotion (2 mg/cm2). Treatments were applied 20 minutes prior to irradiation with a solar simulator(20 J/cm2) and kept on during irradiation. The natural sunscreen was prepared either one day or three weeks before
irradiation and stored at room temperature. Some of the skin samples were fixed three hours following UVirradiation and assessed for the levels of UV-induced DNA damage in the form of cyclobutane pyrimidine dimers(CPDs) using immunohistochemistry. The remaining skin samples were fixed 24 hours following UV-irradiationand stained with H&E to assess for sunburn cells and epidermal thickness. The red raspberry seed oil and carrot
seed oil, which are claimed to have SPFs of 25-50 and 35-40 respectively by the wellness blogger, were further
studied in vitro in human skin cells to determine if they could protect against UV-induced cell death and DNA
damage. With the trend toward social media for health advice, consumers choosing to formulate homemade
sunscreens lacking scientific testing for efficacy and shelf-life risk unknowingly exposing themselves to theharmful effects of UV including skin cancers. On the other hand, the protective effects of these natural homemade
sunscreens may help to overcome sceptical attitudes towards commercial sunscreen use, and may further avoid thenegative health and environmental impacts associated with commercial sunscreens.

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IL-8.2.1

PDT Enhancement of Anti-tumor Immunity: Mechanisms and Usage
Sandra O. Gollnick1,2, Riddhi Falk-Mahaptara2, Kimberley Ramsey1
Roswell Park Comprehensive Cancer Center, PDT Center, Department of Cell Stress Biology, Buffalo, NY, USA;
2) Roswell Park Comprehensive Cancer Center Department of Tumor Immunology, Buffalo, NY, USA Department
of Cell Stress Biology, Buffalo, NY, USA
Over the last decades the interest in PDT as an enhancer of anti-tumor immunity and our understanding of themechanisms by which PDT enhances anti-tumor immunity have dramatically increased. We and others have shownthat oxygen conservation is critical to the ability of PDT to enhance anti-tumor immunity and that PDT-induced
inflammation, which is defined by increased neutrophil migration and cytokine production, drives induction of antitumor immunity1. These results suggest that PDT regimens that augment anti-tumor immunity (imPDT) have thepotential to act as adjuvants that in combination with other ablative anti-cancer therapies, potentially leading toincreased efficacy against distant disease.

References
1) Falk-Mahapatra R and Gollnick SO, Photodynamic Therapy and Immunity: An Update. Photochemistry andPhotobiology 96 (3):550-559, 2020.
Acknowledgements. This work was supported in part by the National Cancer Institute of the National Institute of
Health under Award 5P01CA98156 (SG) and the Roswell Park Alliance Foundation. The study used shared resources
supported by Roswell Park Cancer Comprehensive Cancer Center Support Grant (P30CA016056). Riddhi FalkMahapatra was supported by the Department of Immunology Training Grant (5T32CA085183-15).

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IL-8.2.2
Immunogenic cell death induced by PDT: a new approach for cancer therapy
Dmitri V. Krysko1,2,3,, Victoria D. Turubanova3, Tatiana A .Mishchenko3, Irina V. Balalaeva3, Iuliia Efimova1,2,
Olga Krysko4 , Maria V. Vedunova3
1) Cell Death Investigation and Therapy Laboratory, Department of Human Structure and Repair, Ghent
University, Ghent, Belgium; 2) Cancer Research Institute Ghent, Ghent, Belgium; 3) Institute of Biology andBiomedicine, National Research Lobachevsky State University of Nizhni Novgorod, Nizhni Novgorod, RussianFederation; 4) Upper Airways Research Laboratory, Department of Head and Skin, Ghent University, Ghent,
Belgium
Immunotherapy has become an important part in cancer treatment during the last decade. One of the major factors
for induction of immune response during therapy is immunogenic cell death (ICD)1. Immunogenicity of dying cancer
cells is mediated by their adjuvanticity and antigenicity. Damage-associated molecular patters (DAMPs) areendogenous molecules located inside the cells and in normal conditions contribute to different physiologicalprocesses while they are released (or exposed on the outer surface of plasma membrane) when a cell is damaged ordying. Once released, DAMPs acquire immunostimulatory properties and increase adjuvanticity of dying cancer
cells. Of course, DAMPs are not the only factors released during cell death which are involved in their adjuvanticity.
Antigenicity of dying cancer cells is another major determinant of ICD which is required for an efficient targeted
induction of anti-tumor immunity. ICD can be induced by different stimuli and anticancer treatment modalities,
including chemotherapy with anthracyclines and oxaliplatin, radiotherapy, UVC irradiation, oncolytic viruses andphotodynamic therapy (PDT)2. The ICD induced by various stimuli can differ in the DAMPs’ profile and has also
been linked to different cell death modalities such as apoptosis, necroptosis and ferroptosis3. Thus, in this lecture, wefirst discuss the role of PDT in the induction of ICD 4 and then assess the advantages and disadvantages of PDT in
the induction of ICD. Finally, we will discuss a possible synergistic action between PDT and ferroptotic cell death5,
a novel, iron-dependent form of regulated cell death6. PDT can act as a source of reactive oxygen species for the
Fenton reaction, which may reinforce ferroptosis induction and increase PDT efficacy in anticancer therapy.
References
1) Galluzzi, Vitale, et al. Consensus guidelines for the definition, detection and interpretation of immunogenic cell
death. J Immunother Cancer. 2020.
2) Alzeibak, Mishchenko et al. Targeting immunogenic cancer cell death by photodynamic therapy: past, present
and future. J Immunother Cancer. 2021.
3) Efimova, Catanzaro et al. Vaccination with early ferroptotic cancer cells induces efficient antitumor immunity.
J Immunother Cancer. 2020.
4) Turubanova, Balalaeva et al. Immunogenic cell death induced by a new photodynamic therapy based on
photosens and photodithazine. J Immunother Cancer 2019.
5) Mishchenko, Balalaeva et al. Ferroptosis and Photodynamic Therapy Synergism: Enhancing Anticancer
Treatment. Trends Cancer. 2021.
6) Friedmann Angeli, Krysko et al. Ferroptosis at the crossroads of cancer-acquired drug resistance and immune
evasion. Nat Rev Cancer. 2019.
Acknowledgements
This study was supported by the Russian Science Foundation (RSF) https://rscf.ru/en/project/18-15-00279/ and by
FWO-Flanders projects FWO G043219N and FWO G016221N and by UGent BOF 01/O3618.

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IL-8.2.3

Immune responses triggered by nanobody-targeted photodynamic therapy
Sabrina Oliveira
Utrecht University, Department of Biology and Department of Pharmaceutical Sciences, Utrecht, The Netherlands
Nanobodies are the variable domain of heavy chain antibodies that exist in Camelids1. The relatively small
dimensions of nanobodies combined with high binding affinities to their targets, lead to rapid tumor accumulation,
homogenous distribution, and rapid clearance of unbound fractions. We took advantage of these properties andhave developed an alternative approach for targeted photodynamic therapy (PDT) by conjugating photosensitizers
to nanobodies2. Several of our studies are targeting the epidermal growth factor receptor (EGFR) for head and neckcancer. As photosensitizer, we have employed the IRDye700DX, that is a silicon phthalocyanine derivative,
currently being tested in clinical trials conjugated to the monoclonal antibody cetuximab, and already approved forclinical use in Japan. Generally, with nanobody-targeted PDT, cytotoxicity is selectively induced in cells whichhave high target expression level. Preclinical studies have shown selective tumor necrosis 3 and significant tumor
regression after a single treatment session4.
More recently, we have explored the immunomodulatory potential of nanobody-targeted PDT, using high andmoderate EGFR-expressing cells. After nanobody-targeted PDT, the cytoplasmic damage-associated molecularpattern (DAMP) HSP70 was detected on the cell membrane of tumor cells and the nuclear DAMP HMGB1 wasfound in the cell cytoplasm. Furthermore, nanobody-targeted PDT induced the release of the DAMPs HSP70 andATP, as well as the pro-inflammatory cytokines IL- 1β and IL-6. Conditioned medium from high EGFR-expressing
tumor cells treated with nanobody-targeted PDT led to the maturation of human monocyte-derived dendritic cells,
as indicated by the upregulation of CD86 and MHC II on their cell surface. Subsequently, these dendritic cells
induced CD4+ T cell proliferation, accompanied by IFNγ release. Together, these results suggest nanobodytargeted PDT can stimulate the immune system5. This presentation will summarize these studies and discuss current
efforts to further investigate the systemic reach of this selective and localized treatment.
References
1) Hamers-Casterman, C.; Atarhouch, T.; et al., Naturally occurring antibodies devoid of light chains. Nature
1993, 363 (6428), 446-8.
2) Heukers, R.; van Bergen en Henegouwen, et al., Nanobody-photosensitizer conjugates for targeted
photodynamic therapy. Nanomedicine : nanotechnology, biology, and medicine 2014, 10 (7), 1441-51.
3) van Driel, P.; Boonstra, M. C.; et al., EGFR targeted nanobody-photosensitizer conjugates forphotodynamic therapy in a pre-clinical model of head and neck cancer. Journal of controlled release :official journal of the Controlled Release Society 2016, 229, 93-105.
4) Deken, M. M.; Kijanka, M. M.; et al., Nanobody-targeted photodynamic therapy induces significant tumor
regression of trastuzumab-resistant HER2-positive breast cancer, after a single treatment session. Journal ofcontrolled release : official journal of the Controlled Release Society 2020, 323, 269-281
5) Beltrán Hernández I, Angelier ML, et al., The Potential of Nanobody-Targeted Photodynamic Therapy to
Trigger Immune Responses. Cancers (Basel). 2020 Apr 15;12(4):978
Acknowledgements. This work was supported by the European Research Council (ERC, Starting Grant 677582).

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OC-8.2.4

Ablation of the cancer-associated fibroblast in pancreatic cancer using FAP-targeted
photodynamic therapy
Daphne Dorst1, Esther Smeets1, Anne Freimoser-Grundschober, Christian Klein2, Marija Trajkovic-Arsic3, Martin
Gotthardt1, Jens Siveke3, Erik Aarntzen1 and Sanne A.M. van Lith1
1)Radboud university medical center, Medical Imaging, Nijmegen, The Netherlands; 2) Innovation Center Zurich,
Roche Pharmaceutical Research and Early Development, Zurich, Switzerland; 3) University Hospital Essen,
Translational Oncology, Germany
Pancreatic ductal adenocarcinoma (PDAC) patients have a dismal 5-year survival rate of 8%. PDAC tumors arecharacterized by a dense extracellular matrix that is produced by the cancer-associated fibroblast (CAF). This
extracellular matrix causes increased interstitial pressure and vascular collapse, thereby impeding perfusion anddelivery of systemic agents, and mediating exclusion of CD8+ T cells. Secondly, the CAFs create an immunesuppressive tumor-microenvironment by secretion of immune-cell inhibiting chemokines. Altogether, the CAFs
create a pro-tumorigenic microenvironment, causing currently used chemotherapies and novel immunotherapies tobe largely ineffective. An elegant solution to modulate the microenvironment in PDAC is to remove the cellorchestrating it, the CAF. In this way, both the physical barrier posed by the hyaluronic acid,- and collagen-rich
extracellular matrix as well as the other factors through which these CAFs mediate the pro-tumorigenic
microenvironment are eliminated. As CAFs in PDAC overexpress fibroblast activation protein (FAP), we here
developed an anti-FAP monoclonal antibody conjugated to a phthalocyanine-photosensitizer, for targeted depletionof the CAF using photodynamic therapy. We characterized efficiency in vitro and in a mouse model for PDAC.
The anti-FAP monoclonal antibody 28H1 and control monoclonal antibody DP47GS were conjugated withthe chelator diethylenetriaminepentaacetic acid (DTPA) and the silicon-phthalocyanine photosensitizer
IRDye700DX. Light-induced toxicity was investigated by incubation of NIH-3T3 murine fibroblasts and their FAPoverexpressing counterpart (3T3-FAP) with increasing concentrations of the conjugates and illumination with 50
J/cm2 280 mW/cm2 690 nm light. Subsequently, CKP (Ptf1awt/Cre;Kraswt/LSL-G12D;p53fl/fl) mice, which develop
spontaneous PDAC tumors, were injected with 0.3 nmol 111In-labeled 28H1-IRDye700DX (n=4) or 0.3 nmol 111Inlabeled DP47GS-IRDye700DX (n=3). Tumor targeting was determined by ex vivo biodistribution studies at 24 hours
post injection. Six mice were subjected to photodynamic therapy by exposing the tumor to 50 J/cm2 280 mW/cm2
690 nm light at 24 hours post injection with 0.3 nmol 28H1-IRDye700DX (n=3) or 0.3 nmol DP47GS-IRDye700DX(n=3). One hour after light exposure, tumor tissue was collected and IRDye700DX-derived fluorescence was imaged.
Tissues were formalin fixed and paraffin embedded, and presence of apoptotic cells was assessed using quantificationof cleaved-caspase-3 immunohistochemistry.
Both 28H1 and DP47GS were conjugated successfully to DTPA and IRDye700DX, reaching substitution
ratios of 2-2.5 IRDye700DX dyes per antibody. Only 28H1-IRDye700DX induced cell death of 3T3-FAP cells upon
illumination, while no effect on 3T3 was observed. Both 111In-labeled 28H1-IRDye700DX and 111In-labeled
DP47GS-IRDye700DX showed uptake in the pancreatic lesion of CKP mice at 24 hours post injection (13.1 ± 3.6%ID/g and 11.4 ± 4.4 %ID/g, respectively). Residual blood activity of 28H1-IRDye700DX was however lower
compared to that of DP47GS-700DX (8.2 ± 4.2 %IA/g and 21.2 ± 8.1 %IA/g, respectively), leading to significantly
higher tumor-to-blood ratios for 28H1-IRDye700DX when compared to DP47GS-IRDye700DX (1.76 ± 0.45 and0.51 ± 0.10, respectively (p = 0.006)). After illumination of the pancreatic tumors with 690 nm light, ex vivo
fluorescence imaging illustrated that the IRDye700DX derived fluorescent signal was quenched in the light-exposedparts of the tumor. In the tumors treated with 28H1-IRDye700DX, both manual and automated analyses revealed
increased expression of cleaved-caspase-3 when compared to DP47GS-treated tumors and when compared to the
non-illuminated region of the same tumor.
The anti-FAP antibody 28H1-IRDye700DX targets PDAC tumors in mice with good signal-to-backgroundratios. Furthermore, it induces apoptosis upon illumination, and co-localization studies with fibroblast markers will
reveal specificity of this cell death. Furthermore, in future experiments, the effect of depletion of FAP-expressing
CAFs on the tumor micro-environment and efficacy of other systemic therapies such as chemo- and immunotherapies
in pancreatic cancer models will be determined.

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IL-8.2.5
Size-Transformable Antigen-Presenting Cell–Mimicking Nanovesicle PDT
Potentiates Effective Cancer Immunotherapy
Xiaoyuan Chen1
1) National University of Singapore, Departments of Diagnostic Radiology, Surgery, Chemical and BiomolecularEngineering, Biomedical Engineering, Yong Loo Lin School of Medicine and Faculty of Engineering, Singapore,
Singapore
Artificial antigen-presenting cells (aAPCs) provide three key signaling components: (i) major histocompatibility
complex I/T cell receptor (MHC I/TCR) stimulatory signal, (ii) cluster of differentiation 80/cluster of differentiation
28 (CD80/CD28) costimulatory signal, and (iii) cytokine release [e.g., interleukin-2 (IL-2)]. aAPCs can stimulate
CD8+ T cell activation. While nanosized aAPCs (naAPCs) have a better safety profile than microsized (maAPCs),
they generally induce a weaker T cell response. Treatment with aAPCs alone is insufficient due to the lack of
autologous antigen-specific CD8+ T cells. Here, we devised a nanovaccine for antigen-specific CD8+ T cell
preactivation in vivo, followed by reactivation of CD8+ T cells via size-transformable naAPCs. naAPCs can beconverted to maAPCs in tumor tissue when encountering preactivated CD8+ T cells with high surface redox potential.
In vivo study revealed that naAPC's combination with photodynamic therapy had an impressive antitumor efficacy.
Our findings provide a generalizable approach for using size-transformable naAPCs in vivo for immunotherapy in
combination with nanotechnologies that can activate CD8+ T cells.
Acknowledgements. This work was supported by the start-up fund of the National University of Singapore(NUHSRO/2020/133/Startup/08) and NUS School of Medicine Nanomedicine Translational Research Programme(NUHSRO/2021/034/TRP/09/Nanomedicine)Book of Abstracts ����������������������������������������������������������������������������������������������� 362

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IL-8.2.6

Induction of long-lasting anti-tumor immunity following PDT treatment using an
investigational VLP-drug conjugate
Rhonda C. Kines1, Cynthia D. Thompson2, Sean Spring1, Zhenyu Li1, Elisabet de los Pinos1, Stephen Monks1, John
T. Schiller2
1) Aura Biosciences, Cambridge, MA, USA; 2) Laboratory of Cellular Oncology, National Cancer Institute, National
Institutes of Health, Bethesda, MD, USA
The papillomavirus virus-like particle (VLP) preferentially targets tumor cells via cell surface modified heparinsulfate proteoglycans1. We have demonstrated in vivo tumor cytotoxicity using an investigational VLP-IR700DX
drug conjugate, belzupacap sarotalocan (AU-011), upon activation with near-infrared light (nIR) in murine models2.
Upon in vitro activation with nIR light AU-011-mediated cell killing was pro-immunogenic in nature, resulting in
the release of damage associated molecular patterns such as DNA, ATP and HMGB-1, activation of caspase-1, andsurface re-localization of calreticulin and HSP70 on killed tumor cells. Using immunocompetent murine tumor
models, TC-1 and MB49, we investigated the ability of AU-011 to eradicate subcutaneous tumors and to subsequently
elicit long-term anti-tumor immunity alone or in combination with checkpoint inhibitor antibodies, anti-PD-1 or antiCTLA-4. A single in vivo administration of AU-011 followed by nIR caused rapid cell death leading to complete
tumor regression in ~50% of all animals out to 100 days and within hours of treatment, calreticulin surface expression,
caspase-1 activation and depletion of immunosuppressive leukocytes were observed in tumors. Combination of AU011 with immune checkpoint inhibitor antibodies, anti-CTLA-4 or anti-PD-1, improved therapeutic activity, resulting
in a 70-100% complete response rate that was durable up to 100 days post-treatment. In both AU-011 monotherapy
and AU-011/checkpoint inhibitor combination groups, 50-80% of complete responders displayed protection fromsecondary tumor re-challenge up to 100 days after the initial treatment. In the TC-1 model, depletion of CD4+ orCD8+ T-cells, either at the time of AU-011 treatment or secondary tumor re-challenge of complete responders,
indicated that both cell populations were vital to AU-011’s ability to eradicate primary tumors and to induce longlasting anti-tumor protection. Furthermore, tumor-specific CD8+ T-cell responses could be observed in circulating
PBMCs within three weeks of AU-011 treatment. These data, taken together, support the conclusion that AU-011
had a direct cytotoxic effect on tumor cells and induced long-term anti-tumor immunity in the absence of specificallytargeted tumor antigens, and this activity was enhanced when combined with checkpoint inhibitor antibodies. AU011 is currently being investigated in a Phase 2 clinical trial for the indication of choroidal melanoma.
References
1) Kines RC, Thompson CD, et al. Human papillomavirus capsids preferentially bind and infect tumor cells. Int J
Cancer. 2016 Feb 15;138(4):901-11.
2) Kines RC, Varsavsky I, et al. An Infrared Dye-Conjugated Virus-like Particle for the Treatment of Primary Uveal
Melanoma. Mol Cancer Ther. 2018 Feb;17(2):565-574.
3) Kines RC, Cerio RJ, et al. Virus-Like Particle-Drug Conjugates Induce Protective, Long-lasting Adaptive
Antitumor Immunity in the Absence of Specifically Targeted Tumor Antigens. Cancer Immunol Res. 2021 Apr
14.
Acknowledgements. This work was supported by Aura Biosciences and the National Institutes of Health intramuralresearch program.

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IL-8.2.7

Combining PDT and immunotherapy
Ferry Ossendorp
Leiden University Medical Center, Tumor Immunology Group, Department Immunology, The Netherlands
Photodynamic therapy (PDT) has shown encouraging clinical results when used as an ablative treatment of solid
tumors. Several studies have reported the photodynamic cancer treatment is associated with immunogenic cell
death. Combining PDT with immunotherapy is a promising option to enhance the immunogenic effects andovercome the immunosuppressive state observed in large, advanced tumors. In our study, we employ strategies inwhich we use a combination of chlorinE6-based PDT with either specific therapeutic vaccination strategies 1 or
immunotherapy with immune checkpoint blocking antibodies to PD-1 or CTLA-4 in preclinical murine tumor
models2. These have resulted in improved induction of systemic cancer-specific CD8 T cell responses whichshowed significant abscopal effects to secondary, untreated tumors in a T cell dependent fashion. Currently ouraims are to deliver defined immune modulating compounds into the tumor microenvironment (TME) using
biodegradable poly(lactic-co-glycolic) acid (PLGA)/PEG nanoparticles (NPs). Intravenous injection of NPs
showed accumulation in myeloid cells in the TME after PDT3. Recently we have used immunostimulatory NPs
consisting of PLGA/PEG particles loaded with stimulatory combined Toll-like receptor ligands and a dendritic-cell(DC)-attracting chemokine for intra-tumoral treatment. The combination provoked strong anti-tumor responses,
including abscopal effects, in three clinically relevant murine models of cancer: MC-38 (colorectal), CT26
(colorectal) and TC-1 (HPV16-induced). We could show that the local and distal anti-tumor effects depended on
the presence of CD8+ T cells. The combination elicited tumor-specific, oncoviral or neoepitope directed CD8+ Tcell immune responses against the respective tumors. In these tumor models we studied the TME in detail,whichshowed higher levels of inflammatory myeloid and CD8+ T cells compared to untreated mice. The rationale for the
combination of PDT with local or systemic immunotherapy treatment of solid tumors will be discussed.
References
1) Kleinovink et al. Combination of photodynamic therapy and specific immunotherapy efficiently eradicatesestablished tumors. Clinical Cancer Research 22, 1459-1468. 2016
2) Kleinovink et al. Photodynamic-immune checkpoint therapy eradicates local and distant tumors by CD8+ T
cells. Cancer immunology research 5, 832-838. 20173) Huis in ‘t Veld, R et al. Photodynamic cancer therapy enhances accumulation of nanoparticles in tumorassociated myeloid cells. Journal of Controlled Release 320, 19-31. 2020

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OC-8.2.8

Low-dose PDT induces a rapid recruitment of CD14+ cells in the skin of mice
Katarzyna Kuczkowska1, Tord Hompland1, Qian Peng2, Ane Sofie Viset Fremstedal1, Pål Kristian Selbo1, KristianBerg11) Oslo University Hospital – The Norwegian Radium Hospital, Department of Radiation Biology, Oslo, Norway;
2) Oslo University Hospital – The Norwegian Radium Hospital, Department of Pathology, Oslo, Norway;
There is a need for better strategies that improve the efficacy of therapeutic cancer vaccines including robust
activation of antigen presenting cells (APCs). Pre-conditioning the vaccination site by inducing mild, transient
inflammation may provide enhanced recruitment of APCs. Photodynamic therapy (PDT) is a clinically approved
treatment for non-melanoma skin cancers and pre-malignant lesions of the skin. PDT generates inflammation inlight exposed tissues and seems an excellent candidate for pre-conditioning the skin for vaccination. In the present
study, we have developed a PDT-based strategy to attract APCs to the site of vaccination in normal mouse skin by
low-dose PDT. We tested various PDT doses using two clinically relevant photosensitizers: 1, TPCS2a/Fimaporfin(used in the drug delivery method photochemical internalization (PCI)) and 2, mTHPC/Temoporfin (trade name
Foscan®). We first evaluated inflammatory responses post PDT at 24 h by quantifying cytokines involved in bothinflammation and chemotaxis of innate immune cells. The PDT dose consisting of 5 μg photosensitizer and 3 minblue light exposure (λmax~420 nm, 13.5 mW/cm2, 2.4 J/cm2) was established as the optimal for inducing a mild andtransient inflammation in normal mouse skin. The time point for maximal CD14+ influx post PDT was investigatedby analysing cytokine kinetics and recruitment of immune cells in excised skin biopsies by ELISA and IHC,
respectively. The results revealed elevated levels of the pro-inflammatory cytokines IL-6, CCL3 and IL-12,
suppression of the anti-inflammatory cytokine IL 10 and influx of CD14+ and CD45+ cell populations to PDTtreated skin area 6 h and 24 h after PDT, suggesting these two time points as most relevant for post PDT
vaccination. The present study comprises a novel strategy to use low-dose PDT as a pre-conditioning and an
adjuvant strategy with a potential to improve therapeutic cancer vaccines.

Acknowledgements. This work has been supported by grants from EuroNanoMed II (Nanovax).

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P-8.2.9

A photoinmunoconjugate for the treatment of breast cancer
Mireia Jordà-Redondo1,2, Jaume Nos1, Mª Lluïsa Sagristá2, Lydia Blay3, Joan Francesc Julián3 and Santi Nonell1
1) IQS School of Engineering, Universitat Ramon Llull, Via Augusta 390, 08017, Barcelona, Spain
2) Departament de Bioquímica i Biomedicina Molecular, Facultat de Biologia, Universitat de Barcelona, Avinguda
Diagonal 645, 08028, Barcelona, Spain
3) Departments of General and Gastrointestinal Surgery, Hospital Universitari Germans Trias i Pujol, Carreterade Canyet, s/n, 08916 Badalona, Barcelona, Spain
Finding biomarkers that allow the detection of neoplasms by illuminating them in real time in the operating roomand combining it with photodynamic therapy (PDT) can be an advantage for the successful resection of the tumor.
In this study, we chemically conjugated a water-soluble phtalocyanine with a monoclonal antibody to assess theselectivity for a cancer cell type. Photoinmunotherapy combine the antibody selectivity and the PDT properties ofthe photosensitizier.1,2,3,4
One photoinmunoconjugate is presented, the antibody employed is Trastuzumab which has selectivity for HER-2
receptor and the moiety is IRDye® 700DX NHS. The in vitro assays were performed on two tumoral cells: BT-474
as a positive control and HeLa as a negative control. The biological in vitro assays show that this conjugate is able
to photoinactivate cancerous mammalian cells that presents the receptor HER-2 at sub-micromolar concentrations.
Furthermore, it is possible to differentiate the targeted cells by means of using a real-time fluorescence guided
device adapted for this photoinmunoconjugate.
References
1) Zhang, R. R.; Schroeder, A. B.; Grudzinski, J. J.; Rosenthal, E. L.; Warram, J. M.; Pinchuk, A. N.; Eliceri,
K. W.; Kuo, J. S.; Weichert, J. P. Beyond the Margins: Real-Time Detection of Cancer Using Targeted
Fluorophores. Nat. Rev. Clin. Oncol. 2017, 14 (6), 347–364.
2) Rosenthal, E. L.; Warram, J. M.; Boer, E. De; Basilion, J. P.; Biel, M. A.; Bogyo, M.; Bouvet, M.;
Brigman, B. E.; Colson, Y. L.; DeMeester, S. R.; Gurtner, G. C.; Ishizawa, T.; Jacobs, P. M.; Keereweer,
S.; Liao, J. C.; Nguyen, Q. T.; Olson, J.; Paulsen, K. D.; Rieves, D.; Sumer, B. D.; Tweedle, M. F.;
Vahrmeijer, A. L.; Weichert, J. P.; Wilsopn, B. C.; Zenn, M. R.; Zinn, K. R.; van Dam, G. M. Successful
Translation of Fluorescence Navigation During Oncologic Surgery: A Consensus Report. J. Nucl. Med.
2016, 57 (1), 144–150.
3) Photodynamic Medicine: From bench to clinic, edited by Herwig Kostron and Tayyaba Hasan, The Royal
Society of Chemistry, London, 20164) Mitsunaga, M.; Ogawa, M.; Kosaka, N.; Rosenblum, L. T.; Choyke, P. L.; Kobayashi, H. Cancer CellSelective in Vivo near Infrared Photoimmunotherapy Targeting Specific Membrane Molecules. Nat. Med.
2011, 17 (12).
Acknowledgements. This work was supported by Asociación Española Contra el Cáncer (IDEAS19135NONE).

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IL-8.3.1

Role of nitric oxide in tumor response to PDT: Perspectives from the early studies
Mladen Korbelik
BC Cancer, Integrative Oncology Department, Vancouver BC, Canada

Potential of nitric oxide (NO) to markedly influence the outcome of tumor treatment by photodynamic
therapy (PDT) was first reported in 1996, which is within several years of the recognition that thisunstable gas has a major impact on tumor progression. Early studies of NO effects on tumor PDT
response, conducted from mid to late 1990’s, have conclusively established the endogenously-produced
NO in tumors as an important determinant of sensitivity to PDT and shown that this critical role can be
exploited for therapeutic gain. The effects of NO revealed in these studies to have fundamental roles in
the outcome of tumor PDT include the vasodilatation (pre-PDT tumor oxygenation and post-PDT tumor
blood-flow), interaction with PDT-induced superoxide, participation in oxidative stress signaling, andinfluence on inflammatory events in vascular endothelium of treated tumors. While a number of gained
insights have been capitalized upon during the following decades, some of the valuable leads still remain
unexploited. Particularly promising are profits attained by controlling the integral role of NO in tumor
immunosuppression.

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IL-8.3.2

Hyper-aggressiveness of tumor cells that survive a photodynamic challenge: role ofendogenous nitric oxide
Albert W. Girotti1, Jonathan M. Fahey2, Witold Korytowski3
1) Department of Biochemistry, Medical College of Wisconsin, Milwaukee, WI, USA; 2) Department of Pathology,
University of Colorado, Aurora, CO, USA; 3) Department of Biophysics, Jagiellonian University, Krakow, Poland
Anti-tumor photodynamic therapy (PDT) has an advantage over conventional chemotherapy and radiotherapy inbeing predominantly tumor site-specific. Like chemo/radiotherapy, however, PDT can be limited by pre-existing or
acquired resistance, studies ca. 2000 showing that tumor-derived nitric oxide (NO) is substantially involved. Morerecently, our work revealed that NO generated by inducible NO synthase (iNOS) not only stimulated resistance, butalso proliferative, migratory, and invasive aggressiveness of cells that could withstand a PDT challenge. This wasdemonstrated for a variety of human cancer cell lines in vitro, including prostate PC3, breast MDA-MB-231, andglioblastoma U87, each of which was photosensitized in mitochondria with 5-aminolevulinic acid (ALA)-induced
protoporphyrin IX. The indicated post-PDT responses were accompanied by a substantial and long-lasting
upregulation of iNOS/NO, whereas other NOS isoforms were typically unaffected. Hyper-resistance andaggressiveness are attributed to NO from PDT-upregulated iNOS rather than that from basal enzyme, a finding thatis virtually unprecedented for anti-tumor therapeutics. Elevated iNOS/NO-dependent resistance to ALA-PDT has
also been demonstrated for an MDA-MB-231 mouse xenograft model. Key upstream signaling events leading toiNOS/NO upregulation by PDT will be discussed for one cancer line (glioblastoma), along with promising
pharmacologic approaches for suppressing this negative (pro-tumor) response.
Acknowledgements. This work was supported by NIH Grant CA70823 and MCW Cancer Center Grant FP12605

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IL-8.3.3

Nitric Oxide-Mediated Bystander Effects in Photodynamic Therapy
Witold Korytowski1,2, Jerzy Bazak1, and Albert W. Girotti2
1) Dept. of Biophysics, Jagiellonian University, Krakow, Poland; 2) Dept. of Biochemistry, Medical College of
Wisconsin, Milwaukee, WI, USA
We have recently uniquely shown that human breast cancer cells (adenocarcinoma MDA-MB231), exploit
endogenous nitric oxide (NO) to resist elimination by photodynamic therapy (PDT), cisplatin (CDDP)
chemotherapy or combination of both. Inducible nitric oxide synthase (iNOS) was rapidly and persistently
upregulated after the treatment challenge, and the resulting NO signaled not only for greater resistance but also
growth, migration, and invasion/aggressiveness of not only cells surviving the therapeutic challenge, but also
bystander cells, which were not challenged. Ionizing radiation of specifically targeted cells in a given population is
known to elicit pro-death or pro-survival responses in non-targeted bystander cells, but far less is known about such
effects in non-ionizing PDT. We are testing the hypothesis that photodynamically or pharmacologically stressedbreast adenocarcinoma cells can elicit NO-mediated pro-growth/migration aggressive responses not only insurviving target cells but also in non-stressed bystanders. Switching to a more aggressive phenotype during clinicaltherapeutic treatment would be an alarming prospect because it might lead to greater cancer metastatic spread. We
will also report on our attempts to overcome/minimize such effects by using classical inhibitors of iNOS activity(e.g. 1400W) and recently introduced bromodomain and extra-terminal domain (BET) transcriptional inhibitors(e.g. JQ1).
(Supported by NCN grant 2017/27/B/NZ5/02620).

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OC-8.3.4

A Novel Fluorescent Generator of Peroxynitrite Activatable with Red Light
Cristina Parisi1, Mariacristina Failla2, Aurore Fraix1, Luca Menilli3, Francesca Moret3, Elena Reddi3, Barbara
Rolando2, Loretta Lazzarato2, Alberto Gasco2, Salvatore Sortino1.
1) University of Catania, Department of Drug and Health Sciences, Catania, Italy; 2) University of Torino,
Department of Science and Drug Technology, Torino, Italy. 3) University of Padova, Department of Biology,
Padova, Italy.
The alarmingly low turnover of new clinically approved anticancer drugs and the Multi Drug Resistance (MDR)
phenomena emerging for drugs actually used, call for an urgent shift of attention to other “unconventional” andunderexplored therapeutic modalities. Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are
emerging as suitable alternative agents. In particular, peroxynitrite (ONOO‒) has encountered an increasing interestby virtue of its potent oxidative and cytotoxic activity and of its nitrating activity against tyrosine of proteins, 1 aprocess at the basis of the inhibition of the cell’s efflux pumps mainly responsible for MDR. However, due to thelack of selectivity of ONOO‒ for bio-substrates, the control of its delivery in terms of space, time and dosage
represents a desirable requisite. Light gives the possibility to reach this purpose with high accuracy by using
appropriate photoprecursors.2

Fig. 1 – Structure of the novel fluorescent generator of peroxynitrite activatable with red light.

We report a novel molecular hybrid able to generate ONOO‒ under activation with the highly biocompatible red
light (Fig. 1). The hybrid consists of a benzophenothiazine moiety which act as light-harvesting antenna, joined toan N-nitroso appendage through a flexible spacer. The red fluorescence of the antenna represents an additional
advantage permitting the localization of the hybrid within the cellular environment, an important requisite in viewof image-guided phototherapies. The biological activity of the hybrid has been evaluated against two different
cancer cell lines. The hybrid appears well-tolerated in the dark and induces a significant cell mortality underirradiation in a very low concentration range and with very low light doses ( ca. 1 J cm -2).3
References
1) S. Bartesaghi, R. Radi, Fundamentals on the biochemistry of peroxynitrite and protein tyrosine nitration.
Redox Biology, 2018.
2) S. Sortino, Light- Responsive Nanostructured Systems for Applications in Nanomedicine. Springer, 2016.
3) C. Parisi, M. Failla, A. Fraix, L. Menilli, F. Moret, E. Reddi, B. Rolando, F. Spyrakis, L. Lazzarato, R.
Fruttero, A. Gasco, S. Sortino, A generator of peroxynitrite activatable with red light. Chemical Science,
2021.
Acknowledgements. We thank AIRC – Italian Association for Cancer Research (IG-198559) for financial support.

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IL-8.3.5

Strategies towards the use of ruthenium phthalocyanine compounds for theenhancement of photodynamic therapy
Roberto Santana da Silva, Francisco Rinaldi Neto and Amanda Blanque Becceneri
Departament of Biomolecular Sciences, School of Pharmaceutical Sciences, University of Sao Paulo, Ribeirão Preto,
SP-Brazil.
Photodynamic therapy (PDT) is currently being used as an alternative treatment for the control of several malignant
diseases including cancer. Essentially PDT uses light irradiation, oxygen and photosensitizers that have been largely
synthesized and studied. Recently, several new strategies have emerged to enhance PDT but its use may also affectearly and late onset side effects under normal cells. We have found that nitrosyl phthalocyanine ruthenium-based
compounds may be an interesting approach to improve PDT combined with light irradiation therapy. They are singlet
oxygen and nitric oxide (NO) producer agents, beyond what they can also protect normal cells from the cytotoxicity.
In this context, combination of reactive oxygen and nitrogen species (RONS) and photobiomodulation (PBM), a kindof near infrared light irradiation therapy, could be an important tools for a beneficial cancer treatment option. We
have used two types of ruthenium compounds, ([Ru(Pc)], Pc = phthalocyanine) and trans-[Ru(NO)(NO2)(Pc)]. The
UV-Vis spectra of both complexes displayed a band in the 660 nm region. In the case of 0.5 µM trans[Ru(NO)(NO2)(Pc)], light irradiation at the Q-band reduced the percentage of viable human melanoma (A375) cells
to around 50% more as compared to [Ru(Pc)]. We hypothesized that these results were due to a synergistic effectbetween singlet oxygen and nitric oxide. Similar experiments performed with PDT (660 nm) combined with PBM(850 nm) induced more photocytotoxicity using both [Ru(Pc)] and trans-[Ru(NO)(NO2)(Pc)]. This was interpreted
as PBM increasing cell metabolism (ATP production) and the consequent higher uptake of the ruthenium
phthalocyanine compounds and more efficient apoptosis. Interestingly, the cytotoxicity described above seems to be
restrict to cancer cells once no cytotoxicity was observed for the normal cell 3T3, when NO donor agent was
associated to the ROS production. Aim to better understand this effect we have evaluated the cytotoxicity of thosecomplexes against the triple-negative malignant breast cells MDA-MB-231, an extremely aggressive and difficult totreat subtype. The complexes were cytotoxic, changed cell morphology and inhibited cell migration in the presenceof light irradiation. Preliminary tests with trans-[Ru(NO)(NO2)(Pc)] complex demonstrated cytotoxic potential in 3D
cell culture, a biologically relevant model that better represents the tumor microenvironment observed in vivo.
Besides, this combination can also reduce 30 % the cytotoxicity effect under normal MCF-10 cells. The use of metalbased photosensitizers that produce RONS combined with light therapy may represent an advance in the field ofphotodynamic therapy.
Acknowledgments. This work was supported by the Brazilian agencies CNPq, CAPES, and FAPESP.

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IL-8.3.6
Nitric Oxide Photodynamic Therapy with Molecular and Supramolecular Constructs
Salvatore Sortino
Department of Drug and Health Chemical Sciences, University of Catania. Email: ssortino@unict.it
The multiple role nitric oxide (NO) plays in a number of physiological and pathophysiological processes has
stimulated a massive interest in the development of new strategies and methods for generating NO in a controlled
fashion, with the exciting prospect to tackle important diseases. In this context, the therapeutic outcome of any NObased drug is strictly dictated by three main parameters: i) concentration, ii) delivery site and iii) dosage. In somecases, this creates a complex role for the molecule in opposing beneficial and deleterious events. This dichotomy has
made the development of new strategies and methods for generating NO with precise spatiotemporal control a hot
topic in the burgeoning field of nanomedicine. 1 Light represents a minimally invasive “micosyringe” to provide ahighly localized “burst” of NO in biological systems with a superb spatiotemporal control through the aid of suitable
NO photoprecursors. In fact, photoexcitation permits not only to confine site of action of NO at the illuminated areawith high precision but also to define its dosage with great accuracy by tuning the light intensity and/or duration. 2
These unique features make the NO photoreleasing compounds a powerful therapeutic arsenal much more appealing
than those based on either thermal, enzymatic or pH stimuli.
In this contribution, an overview of the most significant examples of NO photodelivering molecular andsupramolecular constructs developed in our laboratories over the last few years is presented, emphasizing the logicaldesign and the potential applications in anticancer and antibacterial therapy alone or in combination with typicalphotosensitizers for photodynamic therapy.3-8
References
1) S. Sortino, Light-controlled nitric oxide delivering molecular assemblies. Chem. Soc. Rev. 2010, 39, 2903-2913.
2) S. Sortino Ed. Light- Responsive Nanostructured Systems for Applications in Nanomedicine. Springer, 2016.
3) A. Fraix and S. Sortino, Combination of PDT photosensitizers with NO photodonors, Photochem. Photobiol. Sci..
2018, 17, 1709-1727.
4) A. Fraix, V. Kirejev, M. Malanga, E. Fenivesi, S. Beni, M. B. Ericson and S. Sortino, A three-color fluorescentsupramolecular nanoassembly of phototherapeutics activatable by two-photon excitation with near infrared light,
Chem. Eur. J. 2019, 23, 7091-7095.
5) M. Malanga, M. Seggio, V. Kirejev, A. Fraix, I. Di Bari, E. Fenyvesi, M. B. Ericson and S. Sortino, A
phototherapeutic fluorescent b-cyclodextrin branched polymer derivering nitric oxide, Biomater. Sci. 2019, 7,
2272-2276.
6) F. Sodano, R. J. Cavanagh, A. K. Pearce, L. Lazzarato, B. Rolando, A. Fraix, T. F. Abelha, C. E. Vasey, C.
Alexander, V. Taresco and S. Sortino, Enhancing Doxorubicin Anticancer Activity with a Novel Polymeric
Platform Photoreleasing Nitric Oxide, Biomater. Sci. 2020, 8, 1329-1344.
7) C. Parisi, M. Failla, A. Fraix, L. Menilli, F. Moret, E. Reddi, F. Spyrakis, B. Rolando, L. Lazzarato, R. Fruttero,
A. Gasco and S. Sortino, A Generator of Peroxynitrite Activatable with Red Light, Chemical Science 2021, 12,4770-4776.
8) A. Fraix, C. Parisi, M. Failla, K, Chegaev, F. Spyrakis, L. Lazzarato, R. Fruttero, A. Gasco and S. Sortino, NO
Release Regulated by Doxorubicin as Green Light-Harvesting Antenna, Chem. Commun. 2020, 56, 6332-6335.
Acknowledgements. We thank AIRC – Italian Association for Cancer Research (IG-198559) for financial support.

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OC-8.3.7
Can NO induced by photooxidative stressed tumour cells influence tumour
microenviroment?
Mariachiara Gani1, Luigi E. Xodo1, Valentina Rapozzi1
1

University of Udine, Department of Medicine, Udine, Italy,

Our previous studies have shown that PC3 prostate cancer cells exposed to chronic photooxidative stress upregulate
inducible nitric oxide synthase (iNOS), stimulating the development of a more aggressive tumor population
characterized by greater growth, invasiveness, and resistance1.
Since NO is a gaseous signaling molecule, it can act as a "messenger" that can communicate to the neighbouring
cells, determining their fate. We therefore wondered whether and how NO, induced by low doses of PDT, might
affect the tumor microenvironment.
To investigate this, we took the supernatant of Pheophorbide-α (Pbα)/PDT-treated PC3 cells and replaced it to theculture medium of untreated PC3 tumor cells, after irradiation. We indeed found a stimulation of PC3 untreated cell
growth, confirming the data reported by Girotti et al. 2,3.
Based on these results, we are investigating what happens in PNT1A, non-prostate cancer cells. Preliminary results
suggest that their growth is stimulated, an aspect that could influence tumour progression and affect treatment
efficacy.

Fig.1 – Representation of PC3 tumour cells (yellow) and PC3 “bystander” untreated cells (pink). The PC3 “bystander” cells
are the cells that are not directly treated.

References
1) F. D’Este F, E. Della Pietra, G.V. Badillo Pazmay, L.E. Xodo, V. Rapozzi. Role of nitric oxide in the
response to photooxidative stress in prostate cancer cells. Biochem. Pharmacol. 2020.
2) J. Bazak, J.M. Fahey, K. Wawak, W. Kortytowski, A.W. Girotti, Enhanced aggressiveness of bystander cells
in an anti-tumor photodynamic therapy model: Role of nitric oxide produced by targeted cells. Free Radic.
Biol. Med. 2017.
3) J. Bazak, W. Kortytowski, A.W. Girotti, Bystander effects of nitric oxide in cellular models of anti-tumor
photodynamic therapy. Cancers, 2019.
Acknowledgements. This work was supported by the National FFABR,VQ3DAME.

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OC-8.3.8

Synthesis and Characterization of Os(II) Oligothienyl Complexes for Photodynamic
Therapy Applications
Elamparuthi Ramasamy, Houston Cole, John Roque, III, Colin G. Cameron and Sherri A. McFarland*
Department of Chemistry and Biochemistry, The University of Texas at Arlington, Texas, 76019, USA
Photodynamic therapy (PDT) is an emerging new technique to treat various kinds of cancer, either alone or as an
adjuvant to conventional treatments such as chemotherapy, radiotherapy, and surgery. In PDT, the photosensitizer(PS) illuminated in presence of oxygen to produce reactive oxygen species (ROS) to destroy cancerous tumor cells.
Being activated by light, one of the key benefits of PDT is its selectivity, confining treatment to the affected area.
Photofrin, an organic compound based on porphyrins, was the first FDA-approved PDT agent and is the basis ofseveral organic derivatives. Our approach uses metal complexes; we synthesized the Ru-based complex TLD 1433
that is in Phase II clinical trials. Here, we have synthesized trifluoromethyl substitution of bipyridine with series ofoligothiophene pi-expansive ligands heteroleptic Os(II) complexes and would expect to alter the excited state
photophysical properties for efficient PDT. We strongly believe that the electron withdrawing trifluoromethylgroups tend to impart longer wavelength absorption and longer excited state lifetimes. The longer wavelength
absorption helps to deep tissue penetration and will help to sensitize efficient singlet oxygen production.
Acknowledgements. This work was supported by grant R01CA222227 from the National Cancer Institute.

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for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-8.4.1

Plant responses to UV light mediated by UVR8
Gareth I. Jenkins
University of Glasgow, Institute of Molecular Cell and Systems Biology, Glasgow G12 8QQ, UK.
Exposure of plants to UV wavelengths modifies numerous aspects of metabolism, morphology and physiology.
These regulatory responses are underpinned by the differential expression of hundreds of genes. UVRESISTANCE LOCUS8 (UVR8) is the only photoreceptor known to mediate regulatory transcriptional responses
to UV-B and shortwavelength UV-A. UVR8 exists as a homodimer in the absence of UV light and photoreception
causes rapid dissociation of the dimer into monomers to initiate signaling. Under photoperiodic illumination withwhite light supplemented with UV-B a dimer/monomer photoequilibrium is established, where approximately 70%
of UVR8 is in the dimeric form. Regulation of the photoequilibrium and initiation of transcription by the UVR8
monomer is mediated by interaction with other proteins: CONSTITUTIVELY PHOTOMORPHOGENIC 1(COP1), REPRESSOR OF UV-B PHOTOMORPHOGENESIS (RUP) proteins, and a number of transcription
factors. However, the regulation of UVR8 action in the initiation of transcriptional responses is not fully
understood.

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-8.4.2

UVR8 Signalling and Plant Development
Ashutosh Sharma1, Gareth I. Jenkins2 and Keara A. Franklin1,

1) University of Bristol, School of Biological Sciences, Bristol, UK, 2) University of Glasgow, Institute of
Molecular Cell and Systems Biology, Glasgow, UK.

Plants perceive different wavelengths of light using specialised photoreceptors. This information is integrated
with temperature signals to control the abundance and activity of the PHYTOCHROME INTERACTING
FACTOR (PIF) family of transcription factors. PIFs regulate multiple developmental processes throughout
plant development and perform a central role in controlling stem elongation. In this talk, I will discuss how
UV-B, perceived by the photoreceptor UV RESISTANCE LOCUS 8 (UVR8), regulates auxin and gibberellin
signalling to control plant architecture in different environments.

Acknowledgements: This work was supported by the Biotechnology and Biological Sciences ResearchCouncilBook of Abstracts ����������������������������������������������������������������������������������������������� 376

19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-8.4.3

Interplay among Nitric Oxide and UVR8 in plants exposed to UV-B radiation
Fernández María Belén1*, Latorre Lucas1, Cassia Raúl1
1) Instituto de Investigaciones Biológicas, UNMdP-CONICET. Mar del Plata, Argentina. E-mail:mbfernan@mdp.edu.ar
The ultraviolet- B (UV-B) resistance locus 8 (UVR8) is the only known UV-B photoreceptor in plants1 and isconserved from Chlorophytes to angiosperms2. After UV-B irradiation, UVR8 turns from an inactive homodimer toan active monomer regulating the expression of several genes involved in tolerance and acclimation3. Nitric oxide(NO) is an UV-B downstream gaseous signal that increases in response to UV-B and attenuates its damaging
effects on plants4. The aim of this work was to analyse the interplay between NO and UVR8 in the UVR8
signalling pathway of plants exposed to UV-B. To this end, we first analysed the expression of NIA1 (nitrate
reductase 1, a NO enzymatic source) in three-week-old control and UV-B irradiated (3.47 m−2 s−1, for 2h)
Arabidopsis Col 0 (WT) and uvr8-1 null mutant plants using RT-qPCR. Results obtained show that NIA1 wasupregulated after UV-B radiation in WT plants, however, this response was impaired in mutants. This indicates that
expression of NIA1 after UV-B treatment depends on the UVR8 signalling pathway, and thus, NO increase throughthis enzyme may be UVR8 dependent. We also analysed if the expression of PHR1 (cyclobutane pirimidine dimers
photolyase involved in DNA repair), IAA19 (an auxin responsive gene) and UVR8 are regulated by UV-B andUVR8. RT-qPCR results demonstrated that PHR1 was upregulated whereas IAA19 and UVR8 were downregulated
after UV-B treatment, and all responses were abolished in mutants. This confirms that regulation of these genes
occurs through the UVR8- dependent pathway. IAA19 expression is repressed under UV-B due to MYB77, BIM1
and BES1 transcription factors (TFs) sequestration by UVR85. In order to determine if similar regulation occurs for
the UVR8 gene, in silico analysis of the predicted UVR8 promoter using PlantPAN database revealed the presenceof binding motifs for these TFs, supporting its negative regulation after UV-B. We also analysed possible UV-B
and NO co-regulation of the UVR8 signalling pathway components, using publicly available microarray and RNAseq databases from Arabidopsis plants treated with NO. This analysis revealed co- regulation by NO and UV-B ofseveral genes involved in this pathway. We found that CHS, IAA2 and GA2OX (GA2-oxidase) are upregulated in
Arabidopsis leaves both by UV-B and exogenous application of NO. On the other hand, other genes were
negatively co- regulated by UV-B and NO, such as the growth promoting genes PRE1 (paclobutrazol resistance 1)
and EXP1 (expansin b1). Finally, prediction analysis of S-nitrosylated proteins from the UVR8 signalling pathway
and cysteine conservation using multiple sequence alignment revealed that COP1, PHR1, IAA14 and NIA1 could
be targets of this post-translational modification. In conclusion, we report an interplay between UVR8 and NO in
the UVR8 signalling pathway as: 1) NO increment depends on UVR8 2) and NO regulates the components fromthe UVR8 signalling pathway at the gene transcription and post-translational levels under UV-B irradiation.
References
1) Brown BA, et al., A UV-B specific signaling component orchestrates plant UV protection. PNAS,
102 (50) 18225-18230, 2005.
2) Fernández MB et al., A Comprehensive Phylogeny Reveals Functional Conservation of the UV-B Photoreceptor
UVR8 from Green Algae to Higher Plants. Frontiers in Plant Science, 7:1698, 2016.
3) Favory JJ et al., Interaction of COP1 and UVR8 regulates UV-B induced photomorphogenesis and stressacclimation in Arabidopsis. The EMBO Journal 28:591-601. https://doi.org/10.1038/emboj.2009.4
4) Cassia R, et al., The role of nitric oxide in the antioxidant defense of plants exposed to UV-B radiation. React.
Oxygen, Nitrogen Sulfur Spec. Plants, https://doi.org/10.1002/9781119468677.ch22 2019.
5) Yang Y, et al., UV-B photoreceptor UVR8 interacts with MYB73/MYB77 to regulate auxin responses andlateral root development. EMBO J, 39(2):e101928 ,2020.
Acknowledgements. This work was supported by grant number 3589 from Agencia Nacional de Promoción deCiencia y Técnica and EXA 929/19 from Universidad Nacional de Mar del Plata. FMB and CR are permanent
researchers of CONICET, Argentina. LL is doctoral fellowship of CONICET.
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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-8.4.4

Broad and narrow band (311 nm) ultraviolet-B radiation activate distinct defensive
antioxidant pathways
Arnold Rácz, Éva Hideg
University of Pécs, Department of Plant Biology, Pécs, Hungary
Ultraviolet radiation (UV, 280-400 nm) has the highest energy among the radiation components reaching thesurface of the Earth. UV-B (280-315 nm) is a potential stressor, and damaging effects of high UV-B doses onplants have been studied extensively. However, over the past decades, several studies have shown thatenvironmental UV-B by itself does not act as a stressor, but rather acts as a regulating factor. Nevertheless, evenambient UV-B can have a damaging effect, when combined with other abiotic factors, stimulating the reactive
oxygen species (ROS) production. When ROS concentrations are increasing, oxidative damage can be avoided byadjusting the ROS antioxidant balance.
A well-documented effect of acclimation to UV-B is the biosynthesis of UV absorbing phenolic compounds. Basedon their high ROS reactivities in vitro1, these compounds are also assumed to act as antioxidants in planta, although
this role is only supported by indirect evidence so far. A third, confirmed defence function of phenolic compounds
is substrate to class III peroxidase enzymes2 (POD). A selective increase in POD activity has a central role inacclimation to supplemental UV-B in model experiments3, as one of the major components of the H2O2 controllingsystem. We have also shown that POD isoforms are selectively activated by UV-B4, and that in tobacco thedominant POD isoform is supported by chlorogenic acid (CGA), a phenolic acid present at increased
concentrations in UV-treated leaves5.
In the present study, we used tobacco plants (Nicotiana tabacum L., cv. Petit Havanna) grown in a growth chamber
under photosynthetically active radiation (PAR) only, then exposed to supplemental UV-B from a narrowband(311.5  2.5 nm) source. Depending on the applied photon flux density (2.9–9.9 μmol m−2 s−1), this treatment
resulted in 0–10% decrease in leaf photochemical activity. Narrow band UV-B resulted in distinct antioxidant
enzyme responses as compared to the above effects of broadband UV-B. The 311 nm treatment decreased CGAPOD and increased SOD activity. Both changes were linear with the applied dose. In addition, an increase in nonenzymatic antioxidant capacities was observed as a non-linear response, appearing under high doses only. Thesefindings may be relevant to UVR8-independent UV-response pathways, such as the recently suggested
photoreceptor with maximum absorbance at 310-311 nm6, and also indicate the necessity of studying thecontribution of various UV-B wavelengths to complex biochemical responses leading to UV-tolerance.
References
1) Csepregi K., Hideg, É., Phenolic compound diversity explored in the context of photo-oxidative stressprotection. Phytochem. Anal. 29, 129-136, 2018.
2) Jansen, M.A.K., van den Noort, R.E., Adillah Tan, M.Y., Prinsen, E., Lagrimini, M.L., Thorneley, R.N.F.,
Phenol-oxidizing peroxidases contribute to the protection of plants from ultraviolet radiation stress. Plant Physiol.
126, 1012-1023, 2001.
3) Majer, P., Czégény, Gy., Sándor, Gy., Dix, P.J., Hideg, É., (2014) Antioxidant defence in UV-irradiated tobacco
leaves is centred on hydrogen-peroxide neutralization. Plant Physiol. Biochem. 82, 239-243, 2014.
4) Rácz, A., Hideg, É., Czégény, Gy., Selective responses of class III plant peroxidase isoforms to environmentally
relevant UV-B doses. J.Plant Physiol. 221, 101-106, 2018.
5) Rácz, A., Czégény, Gy., Csepregi, K., Hideg, É., Ultraviolet‑B acclimation is supported by functionallyheterogeneous phenolic peroxidases. Sci. Rep. 10, 16303, 2020.
6) O'Hara, A., Headland, L.R., Díaz-Ramos, L.A., Morales, L.O., Strid, Å., Jenkins, G.I., Regulation of
Arabidopsis gene expression by low fluence rate UV-B independently of UVR8 and stress signalling. Photochem.
Photobiol. Sci. 18, 1675-1684, 2019.
Acknowledgements. This work was supported by the Higher Education Institutional Excellence Programme of the
Ministry of Human Capacities in Hungary (20765-3/2018/FEKUTSTRAT).

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30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

SP-8.4.5, P-8.4.5

Stabilizing short-lived photoproducts of phytochromes in solid state at room
temperature
Lisa Köhler, Wolfgang Gärtner, Jörg Matysik and Chen Song1
1) University of Leipzig, Faculty for Chemistry and Mineralogy, Institute for Analytical Chemistry, Leipzig,
Germany
Phytochromes (Phys) are photosensoring proteins regulating phototaxis and photomorphogenesis and were first
discovered in higher plants. Especially in bacteria, a large variety of Phys were discovered within the last decades,
habouring different tetrapyrrole chromophores and spectroscopic properties.1 However, magic-angle spinning (MAS)NMR analysis of the short-lived photoproducts comprise practical obstacles using frozen solutions even with theprobe euqipped with in situ illumination, the irradiated molecules would be photocycled through numerous
photointermediates rather than maintained in the steady photoproduct state. Moreover, the highly concentrated
samples used in NMR studies, would prevent the penetration of the actinic light beyond a few hundred µm into thesample, creating a gradient of photoproduct and intermediates within the sample. Therefore we surveyed for sample
preparation techniques, namely the lyophilization2 and the immobilization in disaccharide-protein glassy systems3,
that enable retardation of fast thermal reversion of the photoproduct.
We characterized representative Phy photoproducts of all three subfamilies with different molecular mass (19–56 kDa) with the photoproduct half-lives ranging from a few seconds to hours by UV-vis and MAS NMR
spectroscopy. Upon lyophilization, the conformational dynamics of the photoproduct is “frozen out” due to theremoval of the hydration shell and thus provides the possibility to investigate conformational distributions of theheterogeneous chromophore in various states2. The disaccharide-coating Phy proteins maintain the minimum
hydration shell thus maximally preserving the structural integrity. Herein, the dark reversion is inhibited due to thehigh viscosity of the protein-sugar-water formulations and long-range hydrogen-bonding connectivities.3,5 Bothabovementioned techniques allow the photoproduct preservation against thermal denaturation and reversion for atleast three weeks at room temperature when applied in a well-designed process, which had been evidenced by redissolving the solid samples.
In particular sugar glasses show outstanding positive properties, e.g. transparency and microcrystallinity and arehence widely applicable in different analytical methods. The precise nature of protein-disaccharide interactions
responsible for the Phy biopreservation is discussed.
References
1) N.C. Rockwell, J.C. Lagarias, Phytochrome evolution in 3D: deletion, duplication, and diversification. New
Phytologist, 2020.
2) Y. Kim, Q. Xu, K.-H. Zhao, W. Gärtner, J. Matysik, C. Song, Lyophilization Reveals a Multitude of Strucural
Conformations in the Chromophore of a Cph2-like Phytochrome. J. Phys. Chem. B, 2020.
3) M. Malferrari, A. Savitsky, W. Lubitz, K. Möbius, G. Venturoli, Protein Immobilization Capabilities of Sucros
and Trehalose Glasses: The Effect of Protein/Sugar Concentration Unraveled by High-Field EPR. J. Phys. Chem.
Lett., 2016.
4) K. Anders, L.-O. Essen, The family of ohytochrome-like photoreceptors: diverse, complex and multi-colored, butvery useful. Current Opinion in Strctural Biology, 2015.
5) S. J. Hagen, J. Hofrichter, W. Eaton, Protein reaction kinetics in a room-temperature glass. Science, 1995.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

SP-8.4.6, P-8.4.6

Molecular background of iron uptake mechanisms of non-photosynthetic plastids
Máté Sági-Kazár1, Sándor Lenk2, Éva Sárvári1, Csaba Hegedűs1, Attila Barócsi2, Katalin Solymosi3, Ádám Solti11) Department of Plant Physiology and Molecular Plant Biology, ELTE Eötvös Loránd University, Pázmány Péter
sétány 1/C, Budapest H-1117, Hungary2) Department of Atomic Physics, Budapest University of Technology and Economics, Műegyetem rkp. 3, H-1111
Budapest, Hungary3) Department of Plant Anatomy, ELTE Eötvös Loránd University, Pázmány Péter sétány 1/C, Budapest H-1117,
Hungary
Iron (Fe) is an essential micronutrient for all living organisms. In plants Fe is most important for the formation of thephotosynthetic apparatus and chlorophyll biosynthesis. For the operation of one single linear photosynthetic electrontransport chain a total of 22 Fe atoms, in the form of Fe-S clusters, haem groups and non-haem Fe ions are required,thus the majority of the mesophyll cell’s Fe content is located in the chloroplasts. According to the current model,
Arabidopsis thaliana chloroplasts operate a reduction-based Fe uptake. Transmembrane electron transport isfacilitated by Ferric Reductase Oxidase 7 (FRO7) using NADPH as electron donor produced by the photosynthetic
activity. Consequently, the chloroplast Fe uptake is dependent on photosynthetic activity. FRO7 reduces ferric Fe
complexes to free ferrous Fe which is immediately translocated into the stroma by Permease in Chloroplast 1 (PIC1).
Both chlorophyll biosynthesis and the formation of the photosynthetic apparatus is light dependent in angiosperms.
Thus, the development of the photosynthetic apparatus and thylakoid structures of chloroplasts are impaired andetioplasts form instead in the absence of light. The Fe uptake mechanism of non-photosynthetic plastids such as
etioplasts remains uncharacterized so far. Upon illumination, etioplasts rapidly convert into chloroplasts.
Photosynthetic structures develop, while their formation requires the presence of Fe. However, in the lack of afunctional photosynthetic apparatus, the reduction-based uptake system may lack sufficient reducing power. Thus,
the questions arise, which is first in the plastids: Fe or the photosynthetic apparatus. How non-photosynthetic plastids
can manage their Fe acquisition? Although multiple components of the chloroplast Fe homeostasis have been
identified so far, their exact role in the Fe acquisition is still scarcely understood.
Kale (Brassica oleracea convar. capitata var. sabauda) is a close relative of Arabidopsis and was previously shown
to express orthologues of Arabidopsis chloroplast Fe homeostasis elements. Moreover, in the kale head, natural
etiolation occurs because of the light filtering effect of the overlapping leaves. Thus, here we applied the kale model
to study the differences in Fe homeostasis between etioplasts, intermediate, and developed chloroplasts. Kale heads
were separated into layers based on photosynthetic activity and chlorophyll content. Expression profiles of known
members of the reduction-based Fe uptake (BoFRO7, BoPIC1) as well as less characterised Fe homeostasis elements
were analysed using quantitative real-time PCR. The Fe content of isolated plastids was measured with abathophenanthroline disulfonate based ferrous Fe trapping method by spectrophotometry. In etiolated leaves,
expression of BoFRO7, BoPIC1 was found to be low suggesting they can only have a limited role in the Fe uptake
system of etioplasts. Isolated etioplasts contained a low amount of Fe in parallel. Among the studied Fe homeostasis
elements, BoYSL4, an Fe-nicotianamine transporter showed, indeed, an elevated expression in etiolated leaves. The
Arabidopsis homologue AtYSL4 protein has been only detected in the proplastids of the cotyledons so far where it
was suggested to be involved in the Fe unloading processes of the seed Fe storage of the dicot plant Arabidopsis. Our
results indicate that etioplasts do not operate the reduction power dependent Fe acquisition strategy. Instead, they
may transport Fe in a pathway in which YSL4 is involved.
Acknowledgements. This work was supported by the ÚNKP-20-3 New National Excellence Program of the Ministry
for Innovation and Technology from the source of the National Research, Development and Innovation Fund. This
work was supported by the grant financed by the National Research, Development and Innovation Office, Hungary(NKFIH K-124159). Investigations were principally carried out in the frame of ELTE Thematic Excellence Program
2020 supported by National Research, Development and Innovation Office - TKP2020-IKA-05. S.L. and A.B
acknowledges the NKFIH Grant No. 2017-1.2.1-NKP-2017-00001.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-8.4.7

How plants balance development and UV-B stress tolerance
Tong Liang1,2,6, Chen Shi1,2,6, Yao Peng1,2, Huijuan Tan1,2, Peiyong Xin5, Yu Yang1,2, Fei Wang1, Xu Li1, Jinfang
Chu5, Jirong Huang3, Yanhai Yin4, Hongtao Liu1,*
1) National Key Laboratory of Plant Molecular Genetics (NKLPMG), CAS Center for Excellence in Molecular
Plant Sciences, Institute of Plant Physiology and Ecology (SIPPE), Chinese Academy of Sciences, 200032
Shanghai, P. R. China; 2) University of Chinese Academy of Sciences, Shanghai 200032, P. R. China; 3) Collegeof Life and Environmental Sciences, Shanghai Normal University, Shanghai 200234, P. R. China; 4) Departmentof Genetics, Development and Cell Biology, Iowa State University, Ames, Iowa 50011, USA; 5) National Centre for
Plant Gene Research (Beijing), Institute of Genetics and Developmental Biology, Chinese Academy of Sciences,
Beijing, China; 6) These authors contributed equally to this work
Ultraviolet B (UV-B) light is a potential stress factor in plants, but how plants coordinate growth and UV-B stressresponses is not well understood. Here we report that brassinosteroid (BR) signaling inhibits UV-B stress responses
in Arabidopsis thaliana and various crops by controlling flavonol biosynthesis. We further demonstrate that BRI1EMS-SUPPRESSOR 1 (BES1) mediates the tradeoff between plant growth and UV-B defense responses. BES1, amaster transcription factor involved in BR signaling, represses the expression of transcription factor genes MYB11,
MYB12, and MYB111, which activate flavonol biosynthesis. BES1 directly binds to the promoters of these MYBs
in a BR-enhanced manner to repress their expression, thereby reducing flavonol accumulation. However, exposure
to broad-band UV-B down-regulates BES1 expression, thus promoting flavonol accumulation. These findings
demonstrate that BR-activated BES1 not only promotes growth but also inhibits flavonoid biosynthesis. UV-B
stress suppresses the expression of BES1 to allocate energy to flavonoid biosynthesis and UV-B stress responses,
allowing plants to switch from growth to UV-B stress responses in a timely manner.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

IL-8.4.8

UV-B photoreceptor signalling and responses
Roman Ulm1
1) Department of Botany and Plant Biology, University of Geneva, Geneva, Switzerland
Plants are able to perceive ultraviolet-B radiation (UV-B) using the UV-B photoreceptor UV RESISTANCE LOCUS
8 (UVR8) which activates a molecular signalling pathway leading to UV-B acclimation1. The evolutionarily
conserved UVR8 UV-B photoreceptor exists as a homodimer that monomerises upon UV-B absorption via specificintrinsic tryptophans. The UVR8 monomer interacts with CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1),
an E3 ubiquitin ligase, initiating a molecular signalling pathway that leads to gene expression changes. This signalling
output leads to UVR8-dependent responses including UV-B-induced photomorphogenesis and the accumulation of
UV-B-absorbing metabolites that function as “sunscreens”. I will present our latest understanding of how UVR8
regulates UV-B acclimation and tolerance1.
References
1) Podolec, R., Demarsy, E., Ulm, R., Perception and signaling of ultraviolet-B radiation in plants. Annu. Rev. Plant
Biol. 72: 793–822, 2021.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

OC-8.4.9

Rapid adjustment in epidermal UV sunscreen: Comparison of optical measurement
techniques and response to changing solar UV radiation conditions
Susanne Neugart1,2, Mark A. Tobler2, Paul W. Barnes2
1) Division Quality and Sensory of Plant Products, Georg-August-Universität Göttingen, Carl-Sprengel-Weg 1, D37075 Goettingen, Germany; 2) Department of Biological Sciences and Environment Program, Loyola University
New Orleans, 6363 St. Charles Avenue, 70118 New Orleans, LA, USA
The accumulation of soluble and cell-wall bound UV-absorbing compounds (i.e. flavonoids) in the epidermis andthe mesophyll of leaves is a response of plants to UV exposure. These compounds are known to function in UVscreening, but they are also of potential value for food quality. One way to non-destructively monitor UV-screening
in leaves is by optical methods, from which UVA-PAM and Dualex instruments stand out. The degree and rapidity
to which plants can modulate UV-screening in response to fluctuating solar UV conditions is poorly understood. In
this study, okra plants were exposed to two solar radiation treatments (near-ambient UV (+UV) and attenuated UV(-UV)) and the epidermal UV transmittance (TUV; UVA-PAM) and flavonoid index (Dualex) were measured in theyoungest and second youngest mature leaves over three consecutive days and within an individual day. The day-today (measured near solar noon) and diurnal (over the course of a day) measurements of leaf optical properties
indicated that TUV decreased and flavonoid index increased in the adaxial epidermis ~50% until 15:00 CDT thenreturned close to morning values later in the day. Correlations between UV-B radiation and TUV and flavonoid index
revealed highest values 30 minutes to 1 hour prior the measurements. These findings indicate that plants can respondquickly to fluctuating solar UV conditions and underlines the importance of the harvest-time point for healthpromoting compounds in fruit and vegetables. Our findings also indicate that the UVA-PAM and the Dualex
instruments are both suitable instruments to monitor rapid changes in UV-screening in plants.
References
Acknowledgements. This study was supported by the Deutsche Forschungsgemeinschaft (DFG) - Project number
359552155 and the J.H. Mullahy Endowment for Environmental Biology at Loyola University New Orleans.

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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-9.1.1
Optical force-induced three-dimensional protein assembly growing from solution
Roger Bresolí-Obach1,2, Po-Wei Yi2,3, Wei-Hsiang Chiu2, Tetsuhiro Kudo4, Johan Hofkens1, Eri Chatani5, Ryohei
Yasukuni3, Yoichiroh Hosokawa3, Teruki Sugiyama2,3, Shuichi Toyouchi2, Hiroshi Masuhara2
1) Katholieke Universiteit Leuven, Leuven, Belgium; 2) National Yang Ming Chiao Tung University, Hsinchu,
Taiwan; 3) Nara Institute of Science and Technology, Nara, Japan; 4) Toyota Technological Institute, Nagoya,Japan; 5) Kobe University, Kobe, Japan.
Currently at the microscale, photochemistry and photobiology cover a wide range of scientific fields from single
molecule spectroscopy to several applications including solar energy utilization, optical trapping and biologicalimaging. In the 70s’, Ashkin proposed that optical forces can be exerted on small objects by tightly focusing a laser
beam onto them. In one hand, single nano- and micro-scale objects can be trapped and manipulated with an
exquisite 3D spatiotemporal control in solution, which endows optical trapping a large potential for many
biological studies. On the other hand, when many nanoscale objects are gathered together in (and/or nearby) the
focus, they mutually interact with each other through the scattered trapping light.1 Specifically, at solution surface
and interface, the trapping results in various interesting (bio)-chemical phenomena including molecular
crystallization of biomolecules,2 droplet formation,3 or dynamically fluctuating assembly formation,4 among others.
In this work, we study the formation of a large and thick concentrated protein domain by laser trapping.5
Specifically, a 1064 nm laser is tightly focused on the surface of an almost saturated hen egg white lysozyme(HEWL) solution (26 mM) in D₂O through an air-immersion 60x objective lens (NA 0.90). After switching on thetrapping laser, a single protein domain quickly appeared in the transmission image as a white ring, which expanded
up to a radius of 60 µm in 120 s. To support the HEWL gathering at and around the focus, Raman spectroscopy
experiments were performed either at the focus or at 40 µm away, using a 532 nm laser as the excitation light. We
observed that the Raman signal increased approximately 3.5 and 2.5-fold at focus and 40 µm away, respectively,
indicating that the local protein concentration increased. As well, we did not observe any shift in the Raman
spectrum, indicating that no HEWL denaturation process took place. In order to gain further experimental evidence,
we also followed this protein film formation using fluorescence imaging. A small fraction (1:10000) of
Rhodamine-labelled lysozyme was added to the protein solution. In this case, the formation of a fluorescent domain
is observed, which radially expands up to few tens µm in 60 s and disappears about 40 s after switching off thetrapping laser. Both Raman and fluorescence experiments support the formation of a highly concentrated domain of
HEWL by optical trapping. Indeed, this phenomenon is probably coupled to a liquid-liquid phase separation. The
presented results set the basis for developing a new methodology based on optical trapping to generate highly
concentrated protein domains, which cannot be easily obtained using the current state-of-the-art methodologies.
References
1) T. Sugiyama, K. Yuyama, and H. Masuhara, Laser trapping chemistry: from polymer assembly to amino acidcrystallization. Acc. Chem. Res., 45, 11, 1946–1954 (2012).
2) K. Yuyama, K.-D. Chang, J.-R. Tu, H. Masuhara, and T. Sugiyama, Rapid localized crystallization of lysozyme
by laser trapping. Phys. Chem. Chem. Phys., 20, 6034-6039 (2018).
3) I. Aibara, C.-H. Huang, T. Kudo, R. Bresolí-Obach, J. Hofkens, A. Furube, and H. Masuhara, Dynamic couplingof optically evolved assembling and swarming of gold nanoparticles with photothermal local phase separation ofpolymer solution. J. Phys. Chem. C, 124, 30, 16604–16615 (2020).
4) C.-H. Huang, T. Kudo, R. Bresolí-Obach, J. Hofkens, T. Sugiyama, and H. Masuhara, Surface plasmon
resonance effect on laser trapping and swarming of gold nanoparticles at an interface. Opt. Exp., 28, 19, 2772727735 (2020).
5) P.-W. Yi, W.-H. Chiu, T. Kudo, R. Bresolí-Obach, J. Hofkens, E. Chatani, R. Yasukuni, Y. Hosokawa, T.
Sugiyama, S. Toyouchi, and H. Masuhara, Single sub-millimeter linear assembly of polystyrene microparticles by
optical trapping at protein solution surface. Abstract of 2021 Spring Meeting of Japan Society of Applied Physics(JSAP).
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19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

P-9.1.2
Polyfluorinated aromatic porphyrin as a photoactive scaffold for peptide cyclisation
Paolo Dognini1, Talhat Chaudhry1, Kehinde Ross1, Christopher Coxon2, and Francesca Giuntini1
1) School of Pharmacy and Biomolecular Sciences, Byrom Street Campus, Liverpool John Moores University,
Liverpool L3 3AF, UK; 2) Institute of Chemical Sciences, School of Engineering and Physical Sciences, HeriotWatt University, Edinburgh AH14 4AS, UK
Polyfluoroaromatic reagents readily react with thiols via SNAr and provide excellent templates on to which peptides
can be assembled through side-chain macrocyclisation. We have previously investigated the reactivity ofhexafluorobenzene (HFB, 1) towards cysteine and related nucleophiles [1–3] and subsequently extended this study
to fluorinated porphyrins, exploiting them as photoactive scaffolds for peptide conjugation and cyclisation (Figure1). We will describe how the di-substitution of tetrakis(pentafluorophenyl)porphyrin (2) with the Skin Penetrating
And Cell Entering (SPACE) peptide [4] (ACTGSTQHQCG), presenting two cysteines in position i, i+8,
successfully afforded the macrocyclised product under peptide-compatible conditions. By repeating this reaction
using peptides with different sequences and inter-thiol distances, we evaluated whether this chemistry is sequencespecific. Moreover, the combination of these results with those from a similar methodology applied tobis(pentafluorophenyl)porphyrin (3) gave us important insights regarding the geometrical constraints of themacorcyclisation (e.g. minimal number of amino acids required to reach the para positions of trans and cis meso
phenyl groups). We will confirm the substitution pattern using 19F NMR and we anticipate that the remaining para
positions of products 5 and 6 are also available for further conjugation.
F

F
F

F

S

F

F
1

F

F

F

F

F

F

F
F

F

S

F

N

NH
N

HN

F

F

F

F

F

NH

N

N
HN
F

F

F

F

F

F

F

F

F

F

F

F

S

F
F

F
F

NH

N

F

S

F

5

F
F

F

F

F

F

F

2

F

F

F

F

F

F

N

F

S

HS

peptides

HN

NH
F

F

F

F

N

F

F

F

F

F

F

4
SH

F

F

F

S
F

N H
N

F
F

F

F

F

F

F

N
H N
F

F
F
F

S

F

F

6

N
HN

F

F

F

F

S

7
3

Fig. 1 – General reaction scheme for peptide cyclisation using hexafluorobenzene (1), tetrakis(pentafluorophenyl)porphyrin(2) and bis(pentafluorophenyl)porphyrin (3) and related products.

Preliminary biological data showed that crosslinking SPACE peptide cysteines with 2 does not affect its uptake inskin cancer cells while the presence of the porphyrin core imparts fluorescence and photodynamic properties to thiswater-soluble conjugate (Figure 2). Further steps in understanding the biological behaviour of this conjugate will be
the elucidation of its uptake mechanism in comparison with the uptake of SPACE peptide alone [4] and theinvestigation of any specific subcellular localisation.

Book of Abstracts ����������������������������������������������������������������������������������������������� 385

19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

Fig. 2 – SCCIC8 cells were treated in complete media with F20-SPACE peptide conjugate (10 µM) and incubated for 24h in
the dark. Images were captured at 40X magnification with a Leica Live cell imaging microscope. A) Bright Field image ofcells; B) Nuclei stained with DAPI; C) Uptake of F20-SPACE peptide conjugate; D) Collated image of conjugate uptake withDAPI stained nuclei.

In conclusion, this robust and accessible synthetic tool allows the conjugation of two biologically active
compounds, joining and concurrently improving their individual properties. Moreover, the possibility to modulate
the peptide sequence and differently exploit the photoactive tetrapyrrole scaffold opens the way to a wide range ofapplications in biological systems, especially as molecular probes, drugs and detecting agents.
References
[1]
[2]
[3]
[4]

A.M. Spokoyny, Y. Zou, J.J. Ling, H. Yu, Y.S. Lin, B.L. Pentelute, A perfluoroaryl-cysteine SNAr
chemistry approach to unprotected peptide stapling, J. Am. Chem. Soc. 135 (2013) 5946–5949.
doi:10.1021/ja400119t.
S.J.M. Verhoork, C.E. Jennings, N. Rozatian, J. Reeks, J. Meng, E.K. Corlett, F. Bunglawala, M.E.M.
Noble, A.G. Leach, C.R. Coxon, Tuning the Binding Affinity and Selectivity of Perfluoroaryl-Stapled
Peptides by Cysteine-Editing, Chem. - A Eur. J. 25 (2019) 177–182. doi:10.1002/chem.201804163.
P. Dognini, P.M. Killoran, G.S. Hanson, L. Halsall, T. Chaudhry, Z. Islam, F. Giuntini, C.R. Coxon, Using
19F NMR and two-level factorial design to explore thiol-fluoride substitution in hexafluorobenzene and its
application in peptide stapling and cyclisation, Pept. Sci. 113 (2021) e24182. doi:10.1002/pep2.24182.
T. Hsu, S. Mitragotri, Delivery of siRNA and other macromolecules into skin and cells using a peptide
enhancer, Proc. Natl. Acad. Sci. U. S. A. 108 (2011) 15816–15821. doi:10.1073/pnas.1016152108.

Acknowledgements. This work was supported by a University Alliance Doctoral Training Alliance CO‐
FUND/Marie Skłodowska‐Curie (European Union’s Horizon 2020 research and innovation pro‐ gramme, grant
agreement No 801604) PhD Fellowship Programme in Applied Biosciences for Health (PD).
Book of Abstracts ����������������������������������������������������������������������������������������������� 386

19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

Authors Index
Bold indicates presenting authors

A

Aarntzen, Erik: OC-8.2.4
Abboud, Myriam: IL-3.1.6
Abbruzzetti, Stefania: P-1.3.16, OC-3.4.6
Abdelgawad, Ahmed: IL-3.1.1
Abdullah Elbeheiry, Hani Mohamed:
P-2.1.9
Abohujel, Reem: IL-5.1.3
Ač, Alexander: IL-4.2.1
Acherar, Samir: IL-5.4.9
Adan-Bermudez, Sergio: OC-3.4.1
Adir, Noam: IL-1.1.1
Adriouach, Souad: P-7.3.14
Agata, Kiyokazu: P-3.2.10
Agostiano, Angela: P-1.1.10
Agut, Montserrat: P-1.3.16
Ahad, Tashmeeta: OC-4.3.8
Ahmad, Israr: IL-3.1.1
Ahmetali, Erem: OC-1.3.4
Aksu, Burak: OC-1.3.4
Aktary, Zackie: IL-5.3.1
Albanese, Paola: OC-3.4.5
Albert, Lea: IL-7.1.3
Alberto, Marta Erminia: IL-2.4.5
Alcaraz-Rocha, Paulo: OC-6.2.8, P-6.2.9,
SP-6.2.9, IL-6.2.5
Alcuin Brown, Christian Thomas: OC-4.3.4
Alejo, Bea: OC-3.1.8, OC-6.3.8
Alexander, Bruce H.: IL-3.1.5
Alibes, Ramon: IL-7.1.6
Allémann, Eric: P-7.3.14
Almeida, Adelaide: P-1.3.13, P-1.3.26,
P-2.1.15
Altamura, Emiliano: OC-3.4.5
Alves, Mariana Q.: P-5.1.16
Andersen, Roger: IL-2.3.5
Andraud, Chantal: IL-3.2.1
Andreae, Fritz: P-6.4.11
Andreeva, Elena A. : P-1.4.10, IL-1.4.1,
P-1.4.9
Andrés-Gil, Cristina De: IL-3.3.7
Andreu, Inmaculada: P-7.2.9, P-2.1.10,
P-2.1.12, IL-2.1.5
Angele-Martinez, Carlos: PL-4
Angelov, Ivan: OC-1.3.8
Araujo-Soares, Vera: IL-7.2.6
Arisi, Mariachiara: IL-4.3.3
Arjona, José Aguilera: OC-6.2.4, SP-4.1.10,
OC-2.3.4, P-6.3.10
Arlt, Jochen: OC-3.4.1
Arnaut, Luis G.: IL-2.3.6, OC-2.4.4, OC2.4.9
Arnaut, Zoe A. : IL-2.3.6
Arndt, Hans-Dieter: IL-7.1.1
Arnoux, Philippe: P-1.3.30, IL-5.4.9
Aroso, Rafael T.: IL-2.3.6
Arous, Delmon: OC-6.4.6
Asha, Haritha: IL-4.1.3
Ashraf, Shoaib: P-1.3.28
Asiimwe, Nicholas L.: OC-3.4.8
Asperud, Jonas: OC-6.4.6
Assunção, Ricardo: IL-4.4.7

Audran, Gérard: IL-5.4.9
Avlasevich, Yuri: OC-7.1.9
Aylott, Jonathan W.: IL-5.1.3

B

Bacqueville, Daniel: OC-6.3.4
Bagdonas, Saulius: P-5.1.15
Bagnato, Vanderlei S.: IL-2.4.7
Balalaeva, Irina V.: IL-8.2.2, IL-8.2.2
Balasso, Marco: IL-6.1.5, OC-6.1.8
Balukova, Andrea: OC-6.4.8
Banaszak, Anastazia T.: IL-6.2.6
Bano, Shazia: OC-3.4.12
Baptista, Mauricio: IL-2.3.2, IL-2.1.2
Baraldi, Andrea: OC-3.4.9
Barata, Joana F.B.: P-2.1.14, P-2.1.15
Barnard, Isla: OC-4.3.4
Barnes, Paul W.: OC-8.4.9
Barócsi, Attila: SP-8.4.6
Barrera-Gómez, Jose: IL-8.1.5
Bartolomeu, Maria: P-2.1.15
Basagaña, Xavier: IL-8.1.5
Basarić, Nikola: P-5.4.12
Bassani, Dario M.: IL-3.2.7
Bassi, Roberto: IL-1.4.3
Bastien, Philippe: IL-6.3.7
Batista, Victor S.: IL-1.4.6
Bäuerle, Dominik: IL-1.1.6
Bäumler, Wolfgang: OC-1.3.6, P-1.3.21
Bazak, Jerzy: IL-8.3.3
Becceneri, Amanda Blanque: IL-8.3.5
Behan, Kevin: OC-7.3.4
Beiki, Dana: P-6.4.12
Bellei, Barbara: IL-1.2.5
Benediktsdóttir, Bryndís: IL-8.1.5
Béni, Szabolcs: P-7.1.12
Benito-Peña, Elena: IL-5.1.7
Bennett, Doran I. G.: OC-4.4.4
Benson, Sam: OC-3.4.8
Berg, Kristian: IL-5.4.7, OC-6.4.6, OC-8.2.8
Berger, Bastian: P-3.2.9
Bergmann, Uwe: IL-4.4.1
Bernerd, Françoise: IL-6.3.7, SP-4.1.10,
IL-5.3.2
Berot, Vincent: OC-3.1.8
Bessou-Touya, Sandrine: OC-6.3.4
Bianchini, Paolo: P-1.3.16, OC-3.4.6
Biazzotto, Juliana C.: OC-5.4.4
Bilger, Wolfgang: IL-2.2.3
Birch-Machin, Mark: OC-1.2.8
Blasco, Maria A. : SP-4.1.10
Blay, Lydia: P-8.2.9
Boghossian, Ardemis: PL-6.1
Böhm, Cornelia: OC-6.1.8, IL-6.1.5
Bonn, Catherine: OC-1.2.8
Bonvicini, Francesca: P-3.2.12
Bornman, Janet F.: IL-6.2.3
Boros, Gábor: OC-4.1.4
Bos, Desiree: P-6.4.11
Boura, Cédric: IL-5.4.9
Braga, Susana S.: P-5.1.16
Brancini, Guilherme T.P.: IL-3.3.7

Brash, Douglas E.: PL-4
Bravo, Isabel: OC-3.4.8
Bresolí-Obach, Roger: P-9.1.1
Breton, Nolwenn: IL-5.4.9
Briganti, Stefania: IL-1.2.5
Briza, Tomas: SP-5.4.5
Broekgaarden, Mans: P-7.3.13, P-7.3.11,
SP-5.4.6, OC-6.4.7
Brondz, Efim: OC-6.4.6
Brudvig, Gary W.: IL-1.4.6
Bruzell, Ellen M.: OC-5.2.4, P-1.3.20, IL5.2.3, OC-8.1.4
Bryant, Donald A. : IL-1.4.6
Buenfil-Chi, Tomás J.: P-1.3.17
Buglak, Andrey: P-5.1.13, P-5.1.14, P-5.1.18,
P-4.3.10
Bujaldon, Sandrine: OC-4.4.4
Bulin, Anne-Laure: P-7.3.11, PL-6.2,
P-7.3.13
Burdziński, Gotard: P-1.4.9, P-1.4.10
Burfield, Laura C.: IL-7.2.2
Burnap, Robert L.: IL-4.4.7
Buscemi, Gabriella: P-1.1.10OC-1.1.8, P-1.1.9
Busch, Loris: SP-4.1.9, OC-7.1.9
Busch, Theresa: IL-6.4.1, OC-7.1.9
Busick, Jeffery: OC-2.3.8
Busqué, Félix: IL-7.1.6
Buzzá, Hilde H.: IL-2.4.7
Byrne, Scott N.: IL-3.1.3, OC-3.1.4, IL-3.1.2,
IL-8.1.7

C

Cabré, Gisela: IL-7.1.6
Cahoon, Elizabeth K.: IL-3.1.5
Calbet, Neus: OC-6.3.8
Callaghan, Susan: P-6.4.15
Callan, John F.: OC-3.2.4
Calzavara-Pinton, Piergiacomo: IL-4.3.3
Camacho, Rafael: IL-3.2.3
Camenisch, Theodore G.: OC-5.3.8
Cameron, Colin G.: P-2.4.11, IL-2.1.7, OC8.3.8
Candau, Didier: IL-6.3.7
Canfield, Martha: IL-7.2.6
Cano, C: OC-3.1.8
Cánovas, David: IL-3.3.7
Capelo, Jose Luis: IL-5.1.6
Cardinali, Giorgia: IL-1.2.5
Cardona, Francisco: P-5.1.16
Carigga-Gutierrez, Nazareth Milagros:P-5.4.6, SP-5.4.6, P-7.3.13, OC-6.4.7
Carlos, Luís D.: IL-5.1.5
Carpenter, Christopher J.: IL-5.2.2
Carrau, Analía: IL-3.3.2
Caruso, Enrico: OC-3.4.11, P-5.4.11
Carvalho, Rogério: P-1.4.11
Cassán, Fabricio: IL-3.3.3
Cassia, Raúl: OC-8.4.3
Castagna, Antonella: OC-4.2.4
Castillejo, Isabel: SP-4.1.10
Castro, Kelly A.D.F. : OC-5.4.4
Cavaleiro, José A.S. : P-2.1.14

Book of Abstracts ����������������������������������������������������������������������������������������������� 387

19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, AustriaĆavar, Iva: P-1.3.12
Cea-Sánchez, Sara: IL-3.3.7
Ceci, Luigi: IL-1.1.7
Celli, Jonathan P.: IL-5.4.1, IL-6.4.3
Ceresini, Daniela: OC-3.3.8
Chang, Che-Wei: IL-3.2.3
Chang, Yu-Cherng: P-1.3.23
Chaput, Frédéric: P-7.3.11
Chatani, Eri: P-9.1.1
Chatterjee, Sharmistha: IL-1.1.6
Chaudhary, Ganga Ram: P-7.3.15
Chaudhry, Talhat: P-9.1.2
Chauhan, Veeren M.: IL-5.1.3
Chavan, Archana: IL-7.4.2
Chavan, Bhaven: OC-1.2.8
Chen, Juan: OC-7.3.8, IL-7.3.1
Chen, Xiaoyuan: IL-8.2.5, PL-11
Chernev, Petko: IL-4.4.5, IL-4.4.7
Chilingaryan, Tatevik: P-1.3.30
Chiu, Wei-Hsiang: P-9.1.1
Cho, Ara: TS-3
Christensen, Terje: OC-8.1.4
Chu, Jinfang: IL-8.4.7
Chuiko, Yana: P-5.1.14
Chukhutsina, Volha: OC-1.4.4
Cian, Valentina: OC-3.4.4
Cicco, Stefania Roberta: P-1.1.12
Cieplik, Fabian: IL-5.2.1
Cingolani, Matteo: IL-2.4.2
Cinotti, Elisa: OC-3.1.8
Clainche, Tristan Le: OC-6.4.7, P-7.3.13,
SP-5.4.6
Coates, Philip J.: SP-5.4.5
Coffey, Aoife: IL-4.2.1
Cole, Houston D.: P-2.4.11, IL-2.1.7, OC8.3.8
Coll, Jean-Luc: P-7.3.13, SP-5.4.6, OC-6.4.7
Colletier, Jacques-Phillipe: IL-1.4.1, P-1.4.9,
P-1.4.10
Coniglio, Anahi: IL-3.3.3
Conrad, Christina: IL-5.4.1, IL-6.4.3
Cooper, Matthew: IL-3.1.2
Corral, Inés: IL-4.1.6, IL-4.1.6
Corrochano-Luque, María: IL-3.3.7
Corrochano, Luis M.: IL-3.3.7
Corticelli, Franco: P-3.2.12
Cosma, Pinalysa: P-1.1.10
Costa, Catarina C.: P-5.1.16
Costa, Letícia D.: OC-5.4.4, P-5.4.14
Coxon, Christopher: P-9.1.2
Crespo-Hernández, Carlos E.: IL-2.1.3,
IL-4.1.5
Crestani, Gaia: IL-4.2.1
Cristofolini, Luigi: OC-3.4.9
Croce, Roberta: IL-1.4.7, OC-4.4.4, OC4.4.8
Cross, Carolyn: IL-2.3.5
Csekes, Erika: P-2.1.9
Csepregi, Kristóf: P-2.2.8, P-4.2.9, OC4.2.3
Cuba, Vaclav: P-7.3.11
Cuéllar-Zuquin, Juliana: P-4.1.11
Curak, Gabrijela: OC-3.3.4
Curti, Mariano: OC-1.1.4, IL-1.1.6
Curutchet, Carles: OC-1.1.4
Czégény, Gyula: OC-4.2.3, P-4.2.9,
P-2.2.8

DD’Este, Francesca: IL-5.4.2
da Silva, Luís Pinto: OC-7.1.4
da Silva, Raquel Nunes: P-5.1.16
da Silva, Roberto S.: OC-5.4.4
da Silva, Roberto Santana: IL-8.3.5
Daniel Cediel Becerra, José Duban:
P-6.3.11
Darvin, Maxim E.: OC-7.1.9
Dau, Holger: IL-4.4.7
Davies, Michael: IL-2.1.1
Dawe, Robert S.: IL-4.3.7
de Galbe, M. Victoria: OC-2.3.4
de Gálvez Aranda, María Victoria: OC6.2.4, P-6.3.10
de Laureto, Patrizia Polverino: OC-3.4.4
de Plas, Raf Van: IL-3.2.2
Debus, Richard J.: IL-4.4.5
Degola, Francesca: OC-3.3.8
Del Carmen, Marcela G.: IL-5.4.1, IL-6.4.3
del Valle, Carla Arnau: P-5.1.10, P-5.1.11
Delassan, Sarah: OC-6.3.4
Delcanale, Pietro: OC-3.4.6, P-1.3.16
Dell‘Accio, Alfonso: P-1.3.19
Demaria, Marco: IL-1.2.3
Demessant-Flavigny, Ann‘Laure: SP-4.1.10,
OC-6.3.8
Demirci, Utkan: IL-5.4.1, IL-6.4.3
Descalzo, Ana B.: IL-5.1.7
Desnick, Robert: IL-7.2.1
Dharmage, Shyamali: IL-8.1.5
Dias, Cristina: P-2.1.13
Diaspro, Alberto: P-1.3.16, OC-3.4.6
Dilshat, Ramile: IL-5.3.6
Ding, Lili: OC-7.3.8
Dixon, Katie M.: IL-3.1.6, OC-8.1.8
Dobbek, Holger: IL-4.4.1
Dodd, Antony: IL-7.4.6
Dognini, Paolo: P-9.1.2, OC-3.4.11
Don, Anthony: OC-3.1.4, IL-3.1.3
Donohoe, Claire: OC-2.4.4OC-7.3.4
Dormael, Romain de: IL-6.3.7
Dorst, Daphne: OC-8.2.4
Douki, Thierry: OC-6.3.4
Duarte, Maria P.: IL-5.1.6
Dumoulin, Fabienne: IL-7.3.6
Dungel, Peter: OC-5.2.8
Duplan, Hélène: OC-6.3.4
Durantini, Andrés: OC-2.1.4
Durmuş, Mahmut: OC-1.3.8
Duteil, Luc: IL-5.3.2
Džeba, Iva: P-5.4.12

E

Eadie, Ewan: P-2.3.9, P-4.3.9, OC-4.3.4
Eckl, Daniel Bernhard: P-1.3.21
Edin, Nina F. J. : OC-6.4.6
Efimova, Iuliia: IL-8.2.2, IL-8.2.2
Eggenhofer, Nicole: OC-7.2.4
Eggleston, Ian: P-6.4.16, P-6.4.12, OC-6.4.9
Eichner, Anja: P-1.3.21
Einfeldt, Sven: SP-4.1.9
el Rub, Anamarija Abu: P-7.3.11
Elias, Eduard: OC-4.4.8
Elkihel, Abdechakour: P-1.3.10, SP-1.3.10
Elleaume, Hélène: P-7.3.11, P-7.3.13
Elliot, Abigail: OC-3.4.8

Elliott, Thomas: P-6.4.16
Elmets, Craig A. : IL-3.1.1
Emri, Gabriella: OC-4.1.4
Eren, Tarık: OC-1.3.4
Ernst, Dustin: IL-7.4.2
Eulálio, Ana: P-1.1.11

F

Fahey, Jonathan M.: IL-8.3.2
Fahlstedt, Peter: IL-5.2.5
Failla, Mariacristina: OC-8.3.4
Falk-Mahaptara, Riddhi: IL-8.2.1
Fardilha, Margarida: P-5.4.13, OC-5.4.3
Farinola, Gianluca Maria: OC-1.1.8, P-1.1.9,
P-1.1.10 ,P-1.1.12
Farooq, Nabeela: P-6.4.16
Faßbender, Sonja: IL-1.2.6
Fateixa, Sara: P-5.1.12, P-2.1.14
Faustino, Maria Amparo F.: P-1.3.26,
P-1.3.13, P-1.3.27, P-4.1.13, P-5.4.13, OC5.4.3, OC-5.4.4, P-1.3.13, P-1.3.27, P-4.1.13,
P-5.4.13, OC-5.4.3, OC-5.4.4, P-2.1.15
Fefer, Michael: P-1.3.18, IL-1.3.2, P-1.3.24
Feng, Yi: OC-3.4.8
Ferguson, Calum: P-7.3.10
Fernandez-Rodriguez, Julia: IL-3.2.3
Fernandez, Antonio: OC-3.4.8
Fernández, María Belén: OC-8.4.3
Ferreira, Ana Rita: P-5.4.13, OC-5.4.3
Fiala, Johannes: P-7.1.10, OC-3.3.4
Fidrus, Eszter: OC-4.1.4
Finlayson, Louise: OC-4.3.4, P-2.3.9
Fiore, Michele: OC-3.4.5
Fischer, Reinhard: IL-3.3.6
Fischer, Sophie: IL-4.4.7
Fitsious, Eleni: IL-1.2.3
Fitzgerald, Megan E.: P-4.1.12
Flanagan, Keith J.: P-6.4.15
Flesher, David A. : IL-1.4.6
Floc‘h, Caroline Le: OC-6.3.8
Flores González, Silvia Juliana: P-6.3.12,
P-6.3.9
Flori, Enrica: IL-1.2.5
Flors, Cristina: P-1.3.11
Fodor, Ferenc: OC-7.4.4
Fongaro, Benedetta: OC-3.4.4
Fonseca, Alba: P-6.4.11
Fontaine, Margot: OC-3.1.8
Foster, Barbara A. : P-4.1.12
Foster, Lesley: IL-7.2.6
Fraix, Aurore: OC-8.3.4
Francés-Monerris, Antonio: IL-4.1.2, OC4.1.7
Franklin, Karl A. : IL-8.1.5
Franklin, Keara A. : IL-8.4.2
Freeberg, Alfred: IL-7.4.2
Freimoser-Grunschober, Anne: OC-8.2.4
French, Martyn A. : IL-3.1.2, IL-8.1.7
Frendo, Federico: P-1.3.27
Frochot, Céline: IL-5.4.9, P-1.3.30
Fuentealba, Denis: P-2.1.16
Fuentes Lorenzo, Jorge Luis: OC-5.3.4,
P-6.3.11, P-6.3.12, P-6.3.9
Fuertes, Elaine: IL-8.1.5
Fusi, Franco: P-1.3.19

Book of Abstracts ����������������������������������������������������������������������������������������������� 388

19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

G

Gabre, Pia: IL-5.2.7
Galibert, Marie-Dominique: IL-5.3.5
Galindo, Francisco: P-5.1.10, P-5.1.11
Galstyan, Anzhela: IL-7.3.7
Gani, Mariachiara: OC-8.3.7, IL-5.4.2
Garcia-Larsen, Vanessa: IL-8.1.5
Garcia-Ruiz, Alejandro: PL-4
Garcia, Cesar Vicente: P-1.1.12
Gariboldi, Marzia Bruna: OC-3.4.11, P-5.4.11
Garrido-Charles, Aida: IL-7.1.6
Garrido, José Luis: SP-6.2.9, OC-6.2.8
Gärtner, Wolfgang: IL-3.3.1, SP-8.4.5
Gary-Bobo, Magali: IL-7.3.2
Gasco, Alberto: OC-8.3.4
Gascón-Moya, Marta: IL-7.1.6
Gederaas, Odrun Arna: P-6.4.15
Gelmetti, Alessandra: IL-4.3.3
Gendron, Joshua M.: IL-7.4.1
Genovese, Damiano: IL-2.4.2
Gentili, Denis: P-3.2.12
Ghosh, Goutam: P-1.3.28
Gierlich, Piotr: OC-7.3.4
Gilaberte, Yolanda: IL-6.3.5, IL-7.2.7, OC2.3.4, IL-5.4.8, OC-5.4.10
Giotta, Livia: IL-1.1.7
Girotti, Albert W.: IL-8.3.2, IL-8.3.3
Gisriel, Christopher: IL-1.4.6
Giuntini, Francesca: OC-3.4.11, P-9.1.2
Giussani, Angelo: P-4.1.11
Giza, Aleksandra: OC-4.2.7
Glueck, Michael: OC-3.4.7, P-1.3.18, IL1.3.2, P-1.3.24
Gobin, Ivana: P-1.3.12
Golden, Susan S.: IL-7.4.2
Gollnick, Sandra O.: IL-8.2.1
Golonka, David: IL-6.1.6
Gomes-da-Silva, Lígia C.: OC-2.4.9, OC7.3.4, OC-2.4.4
Gomes, Ana T. P. C. : P-1.3.13
Gómez, Iván: IL-2.2.2
Gonçalves, Gil: P-2.1.13
Goncalves, Leticia C.P.: PL-4
González-Berdullas, Patricia: OC-7.1.4
González, Salvador: IL-5.4.8, OC-5.4.10,
P-6.3.10
Gorbushina, Anna: IL-3.3.5
Görgen, Andreas: OC-6.4.6
Gorman, Shelley: IL-8.1.1
Gorostiza, Pau: IL-7.1.6
Gotthardt, Martin: P-6.4.11, OC-8.2.4
Gourinchas, Geoffrey: IL-6.1.5, OC-6.1.8
Grade, Ana: P-1.3.26
Grant, Sydney: P-4.1.12
Grattieri, Matteo: IL-1.1.2, OC-1.1.8, P-1.1.9,
P-1.1.10, IL-1.1.7
Green, James: IL-4.1.3
Greer, Alexander: OC-2.1.4, P-1.3.28,
IL-2.1.2
Grenier, Alexe: OC-1.2.4
Griffioen, Arjan W.: IL-6.4.5
Grigalavicius, Mantas: OC-6.4.6
Grillari, Johannes: OC-5.2.8, IL-1.2.7
Grinholc, Mariusz S.: P-1.3.14, OC-1.3.7, OC1.3.9, P-1.3.29, P-6.4.13
Grønlien, Krister Gjestvang: IL-5.2.3
Group, Light4Lungs: P-1.3.19

Gruber, Florian: IL-1.2.7, IL-3.2.5, OC3.2.8
Guendulain García, Sergio D.: IL-6.2.6
Guenova, Emmanuella: TS-4
Guerzo, André Del: IL-3.2.7
Guevel, Xavier Le: P-7.3.13
Guidolin, Keegan: OC-7.3.8
Guieu, Samuel: P-5.1.16
Guío-Carrión, Ana: P-4-1-10, SP-4.1.10
Gullón-Blanco, Jose: IL-8.1.5
Gümüş, Berivan Özlem: OC-7.4.8
Gushchin, Ivan: P-3.2.11
Gustavsson, Thomas: OC-4.1.8
Gutiérrez, Gabriel: IL-3.3.7
Gutowska-Owsiak, Danuta: P-1.3.15

Huang, Huang-Chiao: IL-5.4.1, IL-6.4.3
Huang, Jirong: IL-8.4.7
Huber, Harald: P-1.3.21
Hudson, Laura: OC-1.2.8
Hughes, Jonathan: IL-6.1.1
Huguet, Etienne: IL-5.3.2
Huijbers, Elisabeth J.M.: IL-6.4.5
Hujdur, Elvira: OC-6.1.8
Hunt, Matthew: P-6.4.16
Huovinen, Pirjo: IL-2.2.2
Hüsemann, Judith: IL-1.2.6
Huss, Wendy J.: P-4.1.12
Hustad, Steinar: IL-8.1.5
Huymann, Lesley: OC-3.3.4
Hytönen, Vesa P.: IL-6.1.6

H

I

Haarmann-Stemmann, Thomas: IL-1.2.1,
OC-2.1.8
Habermeyer, Benoit: P-1.3.30
Hadi, Marym Mohammed: OC-3.2.4
Hally, Cormac: P-1.3.16
Hamblin, Michael R.: OC-5.4.4
Hamilton, Lloyd: OC-3.4.8
Hammerle, Fabian: P-7.1.10, OC-3.3.4
Hamminger, Christoph: P-1.3.18, IL-1.3.2,
OC-3.4.7
Hamzavi, Iltefat H.: IL-6.3.6
Hannecker, Anna: OC-3.3.4
Harranz, Ángeles: P-2.1.13
Hart, Prue: IL-8.1.7, IL-3.1.2
Hasan, Tayyaba: PL-5, P-1.3.28, OC-3.4.12,
IL-5.4.1, IL-6.4.3
Haschemi, Arvand: IL-3.2.5
Haumann, Michael: IL-4.4.7
Haydon, Mike: IL-7.4.3
Hegedűs, Csaba: OC-4.1.4, SP-8.4.6
Heinrich, Benedikt: IL-7.1.3
Heisler, Joel: IL-7.4.2
Helguero, Luisa: P-2.1.13
Henriksen Raabe, Tine Therese: OC-6.4.6
Henry, Nelly: IL-1.3.5
Hermanowicz, Paweł: OC-4.2.7, OC-5.1.4
Hernando, Jordi: IL-7.1.6
Herrera-Ceballos, Enrique: OC-6.2.4,
P-6.3.10
Hess, Wolfgang: IL-1.4.1
Heyne, Belinda: P-2.1.16
Hickok, Jason: IL-2.3.5
Hideg, Éva: P-4.2.9, OC-4.2.3, OC-8.4.4,
P-2.2.8
Hipfl, Teresa: OC-7.2.8
Hjeltnes, Are Kristofer: IL-5.2.6
Hoerl, Markus: OC-3.4.7
Hofer, Angelika: OC-7.2.8
Hoffmann, Anja Karen: P-1.3.21
Hoffmann, Jutta: IL-4.4.7
Hofkens, Johan: P-9.1.1
Høgset, Anders: IL-5.4.7
Holzer, Barbara: OC-5.2.8
Homayoon, Donja: OC-7.2.8
Hompland, Tord: OC-8.2.8
Hori, Atsushi: OC-2.4.8
Hori, Shinichi: OC-2.4.8
Hosokawa, Yoichiroh: P-9.1.1
Hu, Chen: OC-4.4.4
Huang, Hao-Li: IL-1.4.6

Ibbotson, Sally H.: IL-7.2.3, P-4.3.9, OC4.3.4, P-2.3.9
Iglesias, Pablo: OC-3.1.8, OC-6.3.8
Ihalainen, Janne A. : IL-6.1.6
Improta, Roberto: IL-4.1.3
Ince, Furkan A. : OC-8.1.8, IL-3.1.6
Ireland, Rachael: OC-3.1.4, IL-3.1.3
Ishikita, Hiroshi: IL-4.4.3
Ito, Shosuke: OC-5.3.8
Itri, Rosangela: IL-2.1.2
Ivanov, Daniela: P-3.2.9
Izquierdo-Serra, Mercè: IL-7.1.6

J

Jabeen, Shakeela: OC-2.1.4
Jabłońska, Izabella: P-6.4.14
Jacques-Jamin, Carine: OC-6.3.4
Jacquet, Thibault: P-7.3.11
Janka, Eszter Anna: OC-4.1.4Jansen, Marcel A.K. : IL-4.2.1, IL-4.2.2
Januskaite, Emilija: P-5.1.15
Jenkins, Gareth I.: IL-8.4.1, IL-8.4.2
Jiménez, M. Consuelo: P-2.1.10
Joaquinito, Ana S.: P-1.3.27
Jõgi, Rain: IL-8.1.5
John, Swen Malte: IL-6.3.2
Johnsen, Bjørn J.: OC-5.2.4, OC-8.1.4
Johnstone, Katherine: IL-7.2.6
Jones, Anita C.: OC-3.4.1
Jordà-Redondo, Mireia: P-8.2.9
Josso, Martin: LS-3
Jouan-Hureaux, Valérie: IL-5.4.9
Jouni, Hussein: IL-5.3.2
Juarranz, Ángeles: IL-5.4.8, OC-5.4.10
Julián, Joan Francesc: P-8.2.9

K

Kadioglu, Aras: IL-1.3.5
Kalb, Larissa: OC-1.3.6
Kalia, Sunil: OC-4.3.8
Kalnaityte, Agne: P-5.1.15
Kamei, Yasuhiro: P-3.2.10
Kandiah, Eaazhisai: P-7.3.13
Kapitonova, Marina A. : OC-5.1.8
Karikó, Katalin: OC-4.1.4
Karner, Lisa: OC-5.2.8
Kassab, Giulia: IL-2.4.7
Kaur, Gurpreet: P-7.3.15
Keck, Cornelia M.: SP-4.1.9, OC-7.1.9

Book of Abstracts ����������������������������������������������������������������������������������������������� 389

19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

Kehe, Gannon M.: IL-5.2.2
Kelly, Daniel: OC-7.3.4
Keresztes, Áron: OC-7.4.4
Kermode, Allan G.: IL-3.1.2, IL-8.1.7
Kern, Jan: IL-4.4.1
Kessel, David: OC-6.4.4
Khazova, Marina: P-4.3.9
Khurana, Bhavya: P-1.3.22
Kiening, Martin: OC-3.4.10
Kikuchi, Misato: IL-6.1.2
Kim, Christopher J.: IL-4.4.5
Kines, Rhonda C.: IL-8.2.6
Kip, Annemarie: P-6.4.11
Kirilovsky, Diana: IL-1.4.1, P-1.4.9, P-1.4.10
Kis, Viktória Kovács: OC-7.4.4
Kitahara, Cari: IL-3.1.5
Klein, Christian: OC-8.2.4
Klem, Karel: IL-4.2.2, IL-4.2.1
Klencsár, Zoltán: OC-7.4.4
Knechten, Maren: IL-1.2.6
Kneissl, Michael: SP-4.1.9
Knobler, Robert: TS-1
Kobiela, Adrian: P-1.3.15
Kochevar, Irene: P-1.3.23
Köhler, Lisa: P-8.4.5, SP-8.4.5
Kohli, Indermeet: IL-6.3.6
Kok, Lai Fong: IL-3.1.3
Komura, Yasuo: OC-2.4.8
Kononov, Alexei I.: P-5.1.13, P-5.1.14, OC5.1.8, P-5.1.18
Kopton, Ramona A : IL-7.1.2
Korbelik, Mladen: IL-2.4.1, IL-8.3.1
Kőrösi, László: P-4.2.9
Korytowski, Witold: IL-8.3.3, IL-8.3.2
Kovacs, Daniela: IL-1.2.5
Kramer, Axel: SP-4.1.9, OC-7.1.9
Kramer, Bernd: OC-1.3.6
Kranner, Ilse: OC-3.3.4
Krejcir, Radovan: P-5.4.5, SP-5.4.5
Kremslehner, Christopher: OC-3.2.8, IL3.2.5, IL-1.2.7
Kruszewska, Beata: P-1.3.14, OC-1.3.7
Krutmann, Jean: IL-1.2.6, OC-2.1.8
Krysko, Dmitri V.: IL-8.2.2, IL-8.2.2
Krysko, Olga: IL-8.2.2, IL-8.2.2
Kubenko, Varvara: P-5.1.13
Kuckhoff, Thomas: P-7.3.10
Kuczkowska, Katarzyna: OC-8.2.8
Kudo, Tetsuhiro: P-9.1.1
Kühl, Anja A. : SP-4.1.9
Kuimova, Marina: IL-5.1.2
Kulu, İrem: OC-1.3.8
Kurachi, Cristina: IL-2.4.7
Kurpińska, Martyna: P-1.3.29
Kuru, Erkin: OC-3.4.8
Kussovski, Vesselin: OC-1.3.8

L

Labarile, Rossella: OC-1.1.8, P-1.1.9, P-1.1.10,
IL-1.1.7
Łabuz, Justyna: OC-5.1.4, OC-4.2.7
Lacerda, Paula S.S.: P-2.1.15
Lackner, Michaela: OC-3.3.4
Lademann, Jürgen: P-3.2.9, OC-7.1.9
Lämmermann, Ingo: IL-1.2.7
Lander, Gabriel C.: IL-7.4.2
Landfester, Katharina: P-7.3.10, OC-7.1.9

Landgraf, Nicole: P-1.3.21
Laneri, Francesca: P-7.1.5, SP-7.1.5
Lange, Norbert: P-6.4.10, SP-6.4.10, OC3.4.10, P-7.3.9
Lapalud, Priscilla: OC-6.3.4
LaRochelle, Ethan: P-4.3.9
Larue, Lionel: IL-5.3.1
Larue, Ludivine: IL-5.4.9
Latorre, Lucas: OC-8.4.3
Lazzarato, Loretta: OC-8.3.4
Lee, Jun Yup: IL-3.1.3
Lee, Jun-Seok: OC-3.4.8
Leffler, Jonatan: IL-3.1.2, IL-8.1.7
Leister, Kai: IL-3.3.6
Lelli, Davide: OC-3.4.6
Lenk, Sándor: SP-8.4.6
León, Miguel: P-4.1.14
Levantino, Matteo: P-1.4.10
Leynaert, Bénédicte: IL-8.1.5
Lhiaubet-Vallet, Virginie: IL-4.1.1, OC-4.1.8,
OC-3.4.3, P-2.1.11
Li, Rong: P-7.3.10
Li, Xu: IL-8.4.7
Li, Yongxin: IL-7.3.3
Li, Zhenyu: IL-8.2.6
Liang, Tong: IL-8.4.7
Lichtenberg, Casper de: IL-4.4.5
Lievonen, Topias: IL-6.1.6
Liguori, Nicoletta: OC-4.4.8
Lim, Henry W.: IL-6.3.3, LS-2, IL-6.3.6
Lima, Bárbara: OC-2.4.9
Lindblad, Peter: IL-1.1.3
Lindgren, Mikael: P-6.4.15
Lineros-Rosa, Mauricio: OC-3.4.3
Little, Mark: IL-3.1.5
Liu, Hongtao: IL-8.4.7
Liu, Jun: P-1.3.18, IL-1.3.2, P-1.3.24
Liu, Xin: OC-4.4.4
LiWang, Andy: IL-7.4.2
Lizondo-Aranda, Paloma: OC-4.1.8
Lo, Pui-Chi: IL-7.3.3
Lobo, Ana Catarina S.: IL-2.3.6
Lodeiro, Carlos: IL-5.1.6
Loebel, Nicolas: IL-2.3.5
Lohan, Silke B.: P-3.2.9, SP-4.1.9
Lončarić, Martin: P-5.4.12
los Pinos, Elisabet de: IL-8.2.6
Losi, Aba: IL-3.3.1, OC-3.3.8
Louisnard, Fernand: P-4.1.11
Lourenço, Leandro M.O.: P-1.3.13, P-5.1.12
Lui, Harvey: OC-4.3.8
Luque, Eva M.: IL-3.3.7
Lyons, Alan: P-1.3.28

M

Maçôas, Ermelinda: P-2.1.13
MacRobert, Alexander J.: OC-3.2.4,
P-6.4.16, OC-6.4.8, OC-6.4.9
Maffeis, Valentin: IL-1.1.6
Magalhães, Carla: OC-7.1.4
Mai, Jim Z.: IL-3.1.5
Maisch, Tim: IL-1.3.1
Majora, Marc: IL-1.2.6
Malacarne, Miryam Chiara: OC-3.4.11,
P-5.4.11
Malanga, Milo: P-7.1.12, SP-7.1.5
Malatesti, Nela: P-1.3.12, P-5.4.12

Malinen, Eirik: OC-6.4.6
Mallidi, Srivalleesha: IL-6.4.2
Malova, Polina S.: OC-5.1.8
Malvehy, Josef: OC-6.3.8
Manet, Ilse G.J.: P-3.2.12
Manns, Fabrice: P-1.3.23
Mannucci, Alessia: OC-4.2.4
Manori De Silva, Warusavithana G.: IL-3.1.6
Mantareva, Vanya: OC-1.3.8
Mar Gil-Sánchez, María del: IL-3.3.7
Marat, Xavier: IL-6.3.7
Marchetti-Deschmann, Martina: IL-3.2.6,
OC-3.2.8, IL-1.2.7
Mariangeli, Matteo: OC-3.4.6
Marín, María J.: P-5.1.10, P-5.1.11
Marini, Alessandra: IL-1.2.6
Mariño-Ocampo, Nory: P-2.1.16
Marionnet, Claire: IL-5.3.2, IL-6.3.7
Mariotti, Lorenzo: OC-4.2.4
Markovitsi, Dimitra: IL-4.1.3
Marmol, Veronique Del: OC-3.1.8
Marotta, Roberto: OC-3.4.5
Marque, Sylvain: IL-5.4.9
Marsh-Wakefield, Felix: IL-3.1.3
Martasek, Pavel: SP-5.4.5
Martín, Almudena: SP-4.1.10
Martín, Natalia: OC-2.3.4
Martin, Nazario: P-2.1.13
Martínez-Abaigar, Javier: IL-2.2.7
Martinez-Fernandez, Lara: IL-4.1.3
Martinho, José: P-2.1.13
Mascaraque-Checa, Marta: OC-5.4.10
Mascio, Paolo Di: IL-2.1.6
Mason, Rebecca S.: IL-3.1.6
Masood, Hamzah: OC-3.2.4
Mastrangelopoulou, Maria: OC-6.4.6
Masuhara, Hiroshi: P-9.1.1
Matabuena, María: SP-4.1.10
Mateu, Judit: OC-6.3.8
Matysik, Jörg: SP-8.4.5
Mavelli, Fabio: OC-3.4.5
McCarthy, Allison K.: P-4.1.12
McCarthy, Bianca Y.: IL-3.1.6
McClelland, James: OC-8.1.8
McFarland, Sherri A. : IL-2.1.7, P-2.4.11,
OC-8.3.8
McGreer, Matt: IL-2.3.7
McHale, Anthony: OC-3.2.4
McKenzie, Richard: IL-6.2.1
McLay, Emily R.: IL-4.2.6
McMillan, Lewis: OC-4.3.4
Megias, Diego: SP-4.1.10
Meinke, Martina C.: SP-4.1.9, P-3.2.9,
OC-7.1.9
Melø, Thor B.: P-6.4.15
Mendes, Adélio: P-1.1.11
Menilli, Luca: OC-8.3.4
Mennucci, Benedetta: IL-1.4.2
Menon, Vijay: PL-4
Mercado-Uribe, Hilda: P-1.3.17
Mesquita, Mariana Q.: OC-5.4.3, P-5.4.13
Messinger, Johannes: IL-4.4.5, IL-4.4.1
Metzger, Magdalena: OC-5.2.8
Michalska, Klaudia: P-1.3.25, P-1.3.29
Micieli, Maria: P-1.3.19
Mihaljević, Branka: P-5.4.12
Mihokova, Eva: P-7.3.11
Milano, Francesco: OC-3.4.5, IL-1.1.7

Book of Abstracts ����������������������������������������������������������������������������������������������� 390

19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

Miller, Anne: IL-3.2.5
Miolo, Giorgia: OC-3.4.4, P-1.3.27
Miralles-Durán, Alejandro: IL-3.3.7
Miranda, Miguel Ángel: IL-2.1.5, OC-4.1.8,
OC-3.4.3, P-7.2.9, P-2.1.11, P-2.1.12
Mishchenko, Tatiana A. : IL-8.2.2, IL-8.2.2
Miyamoto, Makilo: OC-3.1.8
Mizintsev, Artem: P-5.1.18
Mochizuki, Nobuyoshi: IL-6.1.2
Moghissi, Keyvan: IL-2.3.3
Mokrzyński, Krystian: OC-5.3.8
Molina, Romina: IL-3.3.3
Moliner, Fabio de: OC-3.4.8
Monks, Stephen: IL-8.2.6
Monteiro, Ana R.: P-5.1.12, P-4.1.13
Monteiro, Carlos J.P.: P-1.3.27
Monteiro, Fátima: P-2.1.13
Mora, Verónica: IL-3.3.3
Moreno-Bondi, Maria C.: IL-5.1.7
Moreno, Ana: OC-3.4.6
Moret, Francesca: OC-8.3.4
Morgan, Myfanwy: IL-7.2.6
Morita, Akimichi: IL-4.3.2
Morozov, Dmitry: IL-6.1.6
Morsi, Rania: P-3.2.12
Moszyńska, Agata: P-1.3.29
Moura, Nuno M.M.: P-4.1.13, P-3.2.13, P-1.1.11
Mourot, Alexandre: IL-7.1.8
Moussounda Koumba, Tataye
Moussounda: IL-5.4.9
Mroginski, Maria Andrea: IL-6.1.7
Müller, Brigitta: OC-7.4.4
Muller, Petr: SP-5.4.5
Multamäki, Elina: IL-6.1.6
Mummolo, Liviana: IL-2.4.2
Muñoz Herranz, M. Paz: P-5.1.10, P-5.1.11
Muranishi, Hiromi: OC-2.4.8, P-2.4.10
Muranyi, Peter: OC-1.3.6
Mušković, Martina: P-5.4.12, P-1.3.12
Mussini, Andrea: P-1.3.16, OC-3.4.6
Muzzopappa, Fernando: IL-1.4.1, P-1.4.10

N

Nagatani, Akira: IL-6.1.2
Nair, Devatha P.: IL-5.2.2
Nájera, Laura: OC-5.4.10
Nakamura, Motoki: OC-2.1.8
Nakonieczna, Joanna: IL-1.3.3, P-1.3.15,
P-1.3.25, P-1.3.29
Nanekar, Rahul: IL-6.1.6Narzt, Marie-Sophie: IL-3.2.5, IL-1.2.7
Nath, Shubhankar: IL-5.4.1, IL-6.4.3
Navarrete-Miguel, Miriam: OC-4.1.7
Navarro-Bielsa, Alba: OC-2.3.4
Nawrocki, Wojciech J.: OC-4.4.4
Nazarenko, Vera: P-3.2.11
Neale, Patrick J.: IL-6.2.5
Neale, Rachel E.: IL-7.2.2
Neto, Francisco Rinaldi: IL-8.3.5
Neugart, Susanne: OC-8.4.9
Neves, Maria Graça P.M.S. : P-3.2.13,
P-1.3.13, P-2.1.13, P-2.1.14, P-2.1.15, P-1.1.11,
P-1.3.26, P-1.3.13, P-2.1.13, P-2.1.14, P-2.1.15,
P-1.1.11, P-1.3.26, P-1.3.27, P-4.1.13, P-5.1.12,
P-5.4.13, OC-5.4.3, OC-5.4.4, OC-5.4.4
Nievas, Sofia: IL-3.3.3
Nilsen, Ola: P-1.3.20

Nizinski, Stanislaw: P-1.4.9, IL-1.4.1, P-1.4.10
Noguchi, Takumi: IL-4.4.6
Nogueira, Ana Teresa: P-1.1.11
Nogueira, Helena I. S.: P-2.1.14
Nomezine, Anthony: P-7.3.11
Nomikou, Nikolitsa: OC-3.2.4
Nomoto, Takahiro: IL-2.4.3
Nonell, Santi: IL-1.3.5, P-1.3.16, P-8.2.9
Norton, Sam: IL-7.2.6
Nos, Jaume: P-8.2.9
Nouveau, Stéphanie: IL-5.3.2
Nowak-Sliwinska, Patrycja: IL-6.4.5
Nowak, Martyna: P-6.4.14
Ntziachristos, Vasilis: PL-10
Núñez, Luis Alberto: P-6.3.9
Nürnberg, Dennis J.: IL-4.4.7
Nyokong, Tebello: OC-1.3.4

OO‘Mahoney, Paul: P-4.3.9, P-2.3.9
Obaid, Girgis: OC-3.4.12, OC-6.4.4
Oberdanner, Christian: OC-7.2.4
Obukhov, Yurii: P-4.3.10
Ogonowska, Patrycja: P-1.3.15
Oliveira, Elisabete: IL-5.1.6
Oliveira, Reginaldo: P-1.4.11
Oliveira, Sabrina: IL-5.1.6, IL-8.2.3
Oliver, Nicholas: IL-4.4.7
Öllinger, Karin: IL-3.1.7, IL-5.3.3
Olsen, Cathrine Elisabeth: IL-5.4.7
Orellana, Guillermo: IL-5.1.7
Orellano, Elena G.: IL-3.3.2
Orozova, Petya: OC-1.3.8
Orsi, Davide: OC-3.4.9
Ortega-Zambrano, Alberto D.: P-1.3.17
Ortega, Ingrid: P-1.3.11
Ortiz, Maria: IL-5.3.2
Ossendorp, Ferry: IL-8.2.7
Ouardi, Meryem El: P-7.2.9, P-2.1.12
Ouk, Tan Sothéa: SP-1.3.10, P-1.3.22
Oyama, Tokitaka: IL-7.4.5
Özbaşak, Hande: P-2.1.9
Ozgur, Rengin: OC-7.4.8

P

P. Ng, Dennis K.: IL-7.3.3
Pallecchi, Lucia: P-1.3.19
Palomares, Emilio: IL-1.3.5
Pankaczi, Fruzsina: OC-7.4.4
Pantazis, Dimitrios: IL-4.4.2, OC-1.4.8
Papaccio, Federica: IL-1.2.5
Papkovsky, Dmitri: IL-5.1.1
Paragh, Gyorgy: P-4.1.12
Parel, Jean-Marie: P-1.3.23
Parisi, Cristina: OC-8.3.4
Pariyar, Shyam: OC-4.2.8
Partch, Carrie L.: IL-7.4.2
Pastor, Sara Sansaloni: P-7.3.9, SP-6.4.10
Patel, Shiv: OC-3.2.4
Patentreger, Andrew D.: P-4.1.12
Patra, Vijaykumar: IL-4.3.5
Patzelt, Alexa: OC-7.1.9
Pedersen, Mona Elisabeth: IL-5.2.3
Pedraza Barrera, Carlos Adolfo: OC-5.3.4,
P-6.3.12
Peintner, Ursula: P-7.1.10, OC-3.3.4
Peng, Qian: OC-8.2.8

Peng, Yao: IL-8.4.7
Pereira, Ana M.V.M.: P-1.1.11
Pereira, Mariette M.: OC-2.4.4, OC-2.4.9,
IL-2.3.6
Pérez-Anker, Javiera: OC-3.1.8, OC-6.3.8
Pérez-García, Lluïsa: P-7.3.15
Pérez-Gilaberte, Juan B.: OC-2.3.4
Pescheck, Frauke: IL-2.2.3
Peterson, Jeffrey: P-1.3.23
Petrocelli, Silvana: IL-3.3.2
Picardo, Mauro: IL-1.2.5
Pierański, Michał K.: OC-1.3.9
Piffaut, Virginie: IL-5.3.2
Pils, Vera: IL-1.2.7
Pinar, Buse: OC-7.4.8
Pinnola, Alberta: IL-1.4.3
Pinto, Sara M.A.: IL-2.3.6
Planel, Emilie: IL-5.3.2, IL-6.3.7
Plätzer, Kristjan: IL-1.3.2, P-1.3.18, P-1.3.24,
OC-5.2.8, OC-3.4.7, OC-7.2.4
Plíhalová, Lucie: P-4.3.11
Pogue, Brian: OC-3.4.12, P-4.3.9
Pollet, Marius: IL-1.2.6
Pontaroli, Ana Clara: IL-4.2.6
Popovich, Kseniya: P-7.3.11
Pouliot, Roxane: OC-1.2.4
Pourzand, Charareh: P-6.4.12, OC-6.4.9
Prandini, Juliana A. : OC-5.4.4
Premi, Sanjay: PL-4
Price, Janet: OC-2.3.8
Prieto, Leonor: SP-4.1.10, OC-6.3.8
Prodi, Luca: IL-2.4.2
Puig-Fábregas, Julia: IL-6.2.5, SP-6.2.9,
OC-6.2.8
Puig, Susana: OC-6.3.8, OC-3.1.8
Pujante, Alejandro Mateos: P-2.1.10

Q

Quartacci, Mike Frank: OC-4.2.4
Quirós-Guerrero, Luis: P-7.1.10

R

Raber, Bailey: OC-4.4.4
Račková, Lucia: P-2.1.9
Rácz, Arnold: OC-8.4.4, P-4.2.9, OC-4.2.3,
P-2.2.8
Radolf, Thorsten: P-6.4.11
Raffy, Guillaume: IL-3.2.7
Ragni, Roberta: P-1.1.12
Raman, Chander: IL-3.1.1
Ramasamy, Elamparuthi: OC-8.3.8
Ramesh, Bala S.: OC-3.2.4
Ramos, Catarina I.V.: P-4.1.13, P-5.1.12
Ramos, Paula: P-1.3.26
Rampazzo, Enrico: IL-2.4.2
Ramsey, Kimberley: IL-8.2.1
Ranieri, Annamaria: OC-4.2.4
Rapacka-Zdończyk, Aleksandra: OC-1.3.7
Rapozzi, Valentina: IL-5.4.2, OC-8.3.7
Rashdan, Eyman: OC-1.2.8
Rastelli, Alessandra N.S.: P-1.3.28
Ratkaj, Ivana: P-5.4.12
Reddi, Elena: IL-1.3.5, OC-8.3.4
Redoulès, Daniel: OC-6.3.4
Reis, Melani J.A.: P-1.1.11
Reiss, Krystle: IL-1.4.6
Remeeva, Alina: P-3.2.11

Book of Abstracts ������������������������������������������������������������������������������������������������ 391

19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

Remenyik, Éva: OC-4.1.4
Reveguk, Zakhar: P-5.1.13, P-5.1.14
Rhodes, Lesley E.: IL-7.2.2
Ricci, Ada: OC-3.3.8
Riccobono, Eleonora: P-1.3.19
Ripa, Maria Belén: IL-3.3.2
Rizvi, Imran: IL-5.4.1, IL-6.4.3, OC-6.4.4
Robinson, Sharon A. : IL-2.2.6, IL-6.2.2
Roca-Sanjuán, Daniel: PL-3, P-4.1.11, OC4.1.7
Rocha, João: P-5.1.16
Rochette, Patrick J: OC-1.2.4
Rodrigues-Santos, Paulo: IL-2.3.6
Rodrigues, Joana: P-5.1.12
Rodriguez-Luna, Azahara: P-6.3.10
Rodríguez-Pérez, Laura: P-2.1.13
Rodriguez, María Victoria: IL-3.3.2
Rogowska, Melania: P-1.3.20
Röhlen-Schmittgen, Simone: OC-4.2.8
Rolando, Barbara: OC-8.3.4
Rolfes, Katharina Maria: OC-2.1.8
Romano, Giovanni: P-1.3.19, IL-1.3.5
Romero, Elisabet: IL-1.1.6, OC-1.1.4
Romo, Cristina: IL-2.3.5
Roque III, John A. : P-2.4.11, IL-2.1.7, OC8.3.8
Rosas, Susana: IL-3.3.3
Ross, Kehinde: P-9.1.2
Rossi, Jari: IL-6.1.6
Rößler, Linda: IL-3.1.7, IL-3.1.7
Rossolini, Gian Maria: P-1.3.19
Roudot, Angelina: IL-6.3.7
Roullet, Nicolas: OC-6.3.4
Rovira, Xavier: IL-7.1.7
Rozpądek, Piotr: OC-4.2.7
Ruban, Alexander: IL-1.4.5
Rugiero, Matteo: P-5.4.11
Ruhi, Mustafa K.: IL-5.4.1, IL-6.4.3
Rutter, Kirsty J.: IL-7.2.2
Rutz, Adriano: P-7.1.10
Rybchyn, Mark S.: IL-3.1.6
Ryšavá, Alena: P-4.3.11SSæterbø, Kristin G.: P-6.4.15
Saga, Yoshitaka: OC-4.4.8
Sági-Kazár, Máté: P-8.4.6, SP-8.4.6
Sagristá, Mª Lluïsa: P-8.2.9
Sainsbury, Kirby: IL-7.2.6
Samkoe, Kimberly: OC-3.4.12
Sancey, Lucie: P-7.3.11, P-7.3.13, SP-5.4.6
Sandate, Colby R.: IL-7.4.2
Santaniello, Antonietta: OC-4.2.4
Santiago, Juan Luis: OC-5.4.10
Santin, Marco: OC-4.2.4
Santos, Carla: P-2.1.13
Santos, Maria João G.: P-5.1.16
Santos, Patrícia: P-1.3.13
Sargen, Michael R.: IL-3.1.5
Sarkany, Robert P.E.: IL-7.2.6
Sarmadi, Roxana: IL-5.2.7
Sarna, Michał: OC-5.3.8
Sarna, Tadeusz: OC-5.3.8
Sárvári, Éva: SP-8.4.6
Sasai, Jun: IL-5.3.2
Sauter, Nicholas K.: IL-4.4.1
Scartazza, Andrea: OC-4.2.4

Schäfers, Judith: OC-4.4.8
Schalka, Sérgio: LS-1
Schamberger, Barbara: OC-3.4.7
Scharbele, Fábio: OC-2.4.4
Schicho, Kurt: OC-5.2.8
Schiller, John T.: IL-8.2.6
Schinagl, Christoph: OC-3.3.4
Schiphorst, Christo: OC-3.4.2
Schirò, Giorgio: P-1.4.10
Schlau-Cohen, Gabriela: IL-1.4.3
Schleusener, Johannes: SP-4.1.9
Schmidt, Marius: IL-6.1.3
Schneider-Brachert, Wulf: OC-1.3.6
Schneider-Warme, Franziska: IL-7.1.2
Schneider, Sabine: IL-1.2.6
Schöbel, Harald: OC-3.3.4
Schosserer, Markus: OC-3.2.8, IL-1.2.7
Schüler, Nadine: P-3.2.9
Schulz, Martin: P-2.1.9
Schumacher, Julia: IL-3.3.5
Schurr, Michael J.: IL-5.2.2
Schwender, Holger: IL-1.2.6
Sciscione, Fabiola: OC-3.2.4
Seggio, Mimimorena: P-7.1.12
Seiger, Dirk: OC-3.4.8
Selbo, Pål Kristian: IL-5.4.7, OC-8.2.8
Semenov, Oleg: P-3.2.11
Sen, Pinar: OC-1.3.4
Şener, M. Kasım: OC-1.3.4
Senge, Mathias O.: PL-8, P-1.3.22, OC7.3.4, OC-2.4.4, P-6.4.15
Sensen, Christoph Wilhelm: OC-6.1.8
Sereno, Jeremy: OC-6.3.4
Serra, Sofia G.: P-3.2.13
Serra, Vanda V.: P-3.2.13
Shareef, Saeed: IL-1.1.6
Shariev, Artur: IL-3.1.6
Sharma, Ashutosh: IL-8.4.2
Sharma, Bunty: P-7.3.15
Shary, Nico Nico: P-4.1.12
Sherwani, Mohammad Asif: IL-3.1.1
Shi, Chen: IL-8.4.7
Shi, Ge: P-2.4.11, IL-2.1.7
Shinohara, Kanako: IL-6.1.2
Sicher, Claudia: SP-4.1.9
Siem, Sunniva: OC-6.4.6
Sierra Prada, Lady Johanna: P-6.3.12
Siewert, Bianka: OC-3.3.4, P-7.1.10
Silva, Artur M. S. : P-5.1.16
Simoncik, Oliver: SP-5.4.5
Singh, Amarjeet: OC-7.4.4
Sinha, Rajeshwar: IL-2.2.1
Sirohiwal, Abhishek: OC-1.4.8
Siveke, Jens: OC-8.2.4
Škařupová, Denisa: P-4.3.11
Sliney, David H.: IL-2.3.1
Sliwa, Michel: IL-1.4.1, P-1.4.9, P-1.4.10
Smeets, Esther: OC-8.2.4
Smolentseva, Anastasia: P-3.2.11
Sobrino, Cristina: IL-6.2.5, SP-6.2.9, OC6.2.8
Sochorova, Michaela: OC-3.2.8
Sol, Vincent: SP-1.3.10, P-1.3.22
Soler, Juan Antonio: P-2.1.11
Solhaug, Knut Asbjørn: IL-2.2.5
Solti, Ádám: OC-7.4.4, SP-8.4.6
Solymosi, Katalin: SP-8.4.6
Somers, David E.: IL-7.4.7

Son, Minjung: IL-1.4.3
Sondermann, Natalie C.: OC-2.1.8
Song, Chen: SP-8.4.5
Sortino, Salvatore: IL-8.3.6, P-7.1.12, SP7.1.5, OC-8.3.4
Spring, Sean: IL-8.2.6
Stano, Pasquale: OC-3.4.5
Staring, Koen: OC-3.4.2
Stashenko, Elena E.: OC-5.3.4, P-6.3.12,
P-6.3.9
Stavrinidou, Eleni: PL-9
Steglich, Claudia: IL-1.4.1
Steingrímsson, Eiríkur: IL-5.3.6
Stelse-Masson, Sarah: P-7.3.11
Sterba, Martin: SP-5.4.5
Stojković, Emina A. : IL-6.1.3
Strassert, Cristian A. : IL-2.4.6
Streng, Christian: IL-3.3.6
Stucki-Buchli, Brigitte: IL-6.1.6
Süer, Naime Ceren: OC-1.3.4
Suescún Sepúlveda, Jhon Alexander:
P-6.3.11
Sugiyama, Teruki: P-9.1.1
Sun, Pan: IL-7.3.3
Suzuki, Tomomi: IL-6.1.2
Svobodová, Alena Rajnochová: P-4.3.11
Swan, Jeffrey A. : IL-7.4.2
Sych, Tomash: P-5.1.18
Szpadel, Krzysztof: P-6.4.14

T

Takala, Heikki: IL-6.1.6
Tamarit, Lorena: P-2.1.12, P-7.2.9
Tan, Huijuan: IL-8.4.7
Tanew, Adrian: IL-4.3.1
Tano, Maria Josefina: IL-3.3.2
Tão, Helder: OC-2.4.9
Taylor, Glyn: IL-1.3.5
Telegina, Taisiya: P-4.3.10
Tell, Gemma: OC-6.3.8
Tenhaken, Raimund: P-1.3.18
Terrén, Samuel: OC-5.4.10
Teszlák, Péter: P-4.2.9
Theodossiou, Theodossis: OC-6.4.6
Thomas, Andrés: IL-2.1.2
Thomas, Paul: P-5.1.10, P-5.1.11
Thompson, Cynthia D.: IL-8.2.6
Thor, Andreas: IL-5.2.7
Thor, Jasper van: OC-1.4.4
Tichauer, Kenneth: OC-3.4.12
Tobler, Mark A. : OC-8.4.9
Todorovic, Nikolina: IL-6.1.5, OC-6.1.8
Tomé, Augusto C.: OC-5.4.4
Tønnesen, Hanne Hjorth: IL-5.2.3
Tonon, Caroline Coradi: P-1.3.28
Tornier, Carine: IL-6.3.7
Torra, Joaquim: P-1.3.11
Torres, Tomas: P-6.4.11
Tovar, Johan S.D.: IL-2.4.7
Toyouchi, Shuichi: P-9.1.1
Trajkovic-Arsic, Marija: OC-8.2.4
Tran, Martin: P-1.3.20
Trautinger, Franz: PL-1
Treghini, Chiara: P-1.3.19
Trend, Stephanie: IL-3.1.2, IL-8.1.7
Triebner, Kai: IL-8.1.5
Trindade, Tito: P-4.1.13, P-5.1.12, P-2.1.14

Book of Abstracts ����������������������������������������������������������������������������������������������� 392

19th Congress of the European Society
for Photobiology

30 August – 3 September 2021 | World Wide Web and Salzburg, Austria

Trotta, Massimo: IL-1.1.7, OC-3.4.5, OC1.1.8, P-1.1.9 ,P-1.1.10
Tse, Benita C.Y.: OC-3.1.4, IL-3.1.3
Tucker, Margaret A. : IL-3.1.5
Turkan, Ismail: OC-7.4.8
Turnbull, Johanna: IL-2.2.6
Turubanova, Victoria D.: IL-8.2.2, IL-8.2.2

Vojtesek, Borivoj: SP-5.4.5
Volpicella, Maria Teresa: IL-1.1.7
Vona, Danilo: OC-2.2.4, OC-1.1.8, P-1.1.9,
P-1.1.10 ,P-1.1.12
Vostálová, Jitka: P-4.3.11
Vrabl, Pamela: OC-3.3.4
Vu, Hong Nhung: IL-5.3.6

UW

Ulm, Roman: IL-8.4.8
Ulrichová, Jitka: P-4.3.11
Urban, Otmar: IL-4.2.2, IL-4.2.1
Uriati, Eleonora: P-1.3.16, OC-3.4.6
Urraca, Javier L.: IL-5.1.7
Uzilday, Baris: OC-7.4.8

V

Vaccari, Marco: OC-3.4.9
Vainikka, Linda: IL-3.1.7, IL-3.1.7
Valen, Håkon: P-1.3.20, IL-5.2.3
Valic, Michael: IL-7.3.1
Vallejo, Mariana C. S. : P-3.2.13
Valzano, Felice: P-1.3.19
van Beijnum, Judy R.: IL-6.4.5
van Lith, Sanne A.M. : OC-8.2.4, P-6.4.11
Varesio, Emmanuel: P-7.3.9, SP-6.4.10
Vargas, Franklin: P-4.1.14
Varsalona, Maria: P-1.1.9, OC-1.1.8, IL-1.1.7
Vayá, Ignacio: P-7.2.9, P-2.1.12, IL-2.1.5
Vázquez, Olalla: IL-7.1.3
Vechtomova, Yuliya: P-4.3.10
Vedunova, Maria V.: IL-8.2.2, IL-8.2.2
Vendrell, Marc: OC-3.4.8
Ventura, Barbara: P-3.2.12
Venturini, Marina: IL-4.3.3
Viappiani, Cristiano: P-1.3.16, OC-3.4.6
Vieira, Sandra I.: P-5.1.16, P-2.1.13
Vieyra-Garcia, Pablo: TS-2
Vignoni, Mariana: IL-2.1.2
Vileno, Bertrand: IL-5.4.9
Villalba, María: P-6.3.10
Villamizar Mantilla, Diego Armando:
P-6.3.9
Vilutis, Osvaldas: P-5.1.15
Vindstad, Benedikte E.: P-6.4.15
Viset Fremstedal, Ane Sofie: OC-8.2.8
Vitry, Catherine de: OC-4.4.4
Vogeley, Christian: OC-2.1.8

Wagner, Stephan: IL-5.3.7
Wahlgren, Weixiao Yuan: IL-6.1.6
Wakamatsu, Kazumasa: OC-5.3.8
Walburn, Jessica: IL-7.2.6
Waleron, Krzysztof: OC-1.3.7
Wang, Dong: OC-6.4.9
Wang, Fei: IL-8.4.7
Wang, Jimin: IL-1.4.6
Wargent, Jason J.: IL-4.2.6
Wäster, Petra: IL-3.1.7, IL-3.1.7
Waterman, Melinda: IL-2.2.6
Watson, Georgia: IL-6.2.2
Ważny, Rafał: OC-4.2.7
Wei, Lei: P-4.1.12
Weinman, John: IL-7.2.6
Weller, Richard: IL-8.1.2
Wen Wong, Judith Jing: IL-5.4.7
Westenhoff, Sebastian: IL-6.1.6, IL-6.1.3
Weyergang, Anette: IL-5.4.7
White, Christopher: OC-6.2.7
White, David: P-6.4.16
Wiebeler, Christian: OC-6.1.4
Wientjes, Emilie: OC-3.4.2
Wilson, Adjele: IL-1.4.1, P-1.4.9, P-1.4.10
Wimmer, Annette: P-1.3.24, IL-1.3.2
Winkler, Andreas: IL-6.1.5, OC-6.1.8
Witzel, Christian: SP-4.1.9
Wolf, Peter: IL-4.3.6, IL-6.3.1, IL-7.2.5,
PL-7
Wolfender, Jean-Luc: P-7.1.10
Wondrak, Georg: IL-1.2.2
Wood, Kenneth: OC-4.3.4, P-2.3.9
Woźniak, Agata: P-6.4.13, P-1.3.14, OC1.3.7, P-1.3.29
Woźniak, Marta: P-6.4.14
Wudarski, Jakub: P-3.2.10
Wulf, Hans Christian: IL-7.2.6

X

Xin, Peiyong: IL-8.4.7
Xodo, Luigi Emilio: IL-5.4.2, OC-8.3.7
Xu, QianFeng: P-1.3.28

Y

Yachandra, Vittal K.: IL-4.4.1
Yacoby, Iftach: IL-1.1.5
Yaghini, Elnaz: OC-6.4.8
Yan, Jiapei: IL-7.4.7
Yan, Xuehai: IL-7.3.3
Yang, Chen: IL-3.1.6
Yang, Kevin: IL-3.1.1
Yang, Yu: IL-8.4.7
Yano, Junko: IL-4.4.1
Yasukuni, Ryohei: P-9.1.1
Yazar, Seyhan: IL-8.1.3
Yi, Po-Wei: P-9.1.1
Yin, Yanhai: IL-8.4.7
Yoon, Juyoung: IL-7.3.5
Yoshikawa, Yukiko: IL-6.1.2
Young, Antony: IL-8.1.6
Yu, Boram: IL-4.4.7
Yudenko, Anna: P-3.2.11
Yusuf, Nabiha: IL-3.1.1

Z

Zaccheroni, Nelsi: IL-2.4.2
Zaharieva, Ivelina: IL-4.4.7
Zala, Ninon: P-1.4.10
Zálešák, Bohumil: P-4.3.11
Zatloukalova, Pavlina: SP-5.4.5
Zavala, Jorge A. : IL-4.2.5
Zawada, Jeff: OC-5.2.4
Zhan, Jiao: IL-1.4.1
Zhang, Haijiang: IL-3.2.3
Zhang, Rui: IL-3.1.5
Zhao, Luyang: IL-7.3.3
Zheng, Gang: IL-7.3.1, OC-7.3.8
Ziółkowski, Piotr: P-6.4.14
Zitter, Elke De: P-1.4.10
Zoratto, Samuele: OC-3.2.8, IL-1.2.7
Zouni, Athina: IL-4.4.1
Zurbriggen, Matias: PL-2
Zwicker, Paula: SP-4.1.9, OC-7.1.9
Zwiesler-Vollick, Julie: OC-2.3.8
Zybała, Izabela: P-6.4.13

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