# Multimodal nanovectors with a theranostic potential for cancer treatment

Ambre Carrouée, Christophe Alric, Stephanie David, Katel Hervé-Aubert, Nicolas Aubrey, Hervé Marchais, Sandra Même, Emilie Allard-Vannier, Jean-Claude Beloeil, Igor Chourpa

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Ambre Carrouée, Christophe Alric, Stephanie David, Katel Hervé-Aubert, Nicolas Aubrey, et al. Multimodal nanovectors with a theranostic potential for cancer treatment. Journées RITS 2015, Mar 2015, Dourdan, France. p14-15. inserm-01143905

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Submitted on 20 Apr 2015

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# Multimodal nanovectors with a theranostic potential for cancer treatment

Ambre Carrouée<fnref n="1" />,<fnref n="2" />, Christophe Alric<fnref n="1" />, Stephanie David<fnref n="1" />, Katel Hervé-Aubert<fnref n="1" />, Nicolas Aubrey<fnref n="3" />, Hervé Marchais<fnref n="1" />, Sandra Même<fnref n="2" />, Emilie Allard-Vannier<fnref n="2" />, Jean-Claude Beloeil<fnref n="2" /> and Igor Chourpa<fnref n="1" />*.

**Abstract.** Targeted theranosis (diagnosis combined to therapy) strategies are intended to improve cancer treatment efficacy while decreasing side effects. Nanovectors of hybrid (organic/inorganic) structure allow to combine multiple physical and biological functions and thus are being developed as promising contrast agents for multimodal diagnosis and drug nanocarriers for therapy of cancers. We present here three types of theranostic nanovectors our group develops, all based on combination of magnetic and optical properties with biological targeting and anticancer activity. Development of these nanovectors is being made in the frame of a multidisciplinary research, including biotechnology, chemical synthesis, pharmaceutical technology, physico-chemical and biological analysis in vitro and in vivo. The rational design of our nanovectors has been confirmed by complementary techniques as leading to promising biocompatibility and theranostic potential.

**Index terms** - Biomedical sensors, Nano medicine

## I. INTRODUCTION

Diagnosis of cancers and the image-monitored treatment are the basis of the so called theranostic strategies which are being actively developed. For more efficient and accurate diagnosis, magnetic resonance imaging (MRI), computed tomography (CT), ultrasound (US) and nuclear imaging of positron emission tomography (PET) [1] are being combined in multi-modal approaches [2]. Indeed, spatially resolved 3D detection of tumors with MRI can be advantageously combined with endoscopic detection of superficial tumors by means of optical probes operating in the tissue optical window in Very Near InfraRed (VNIR). Sensitive real-time VNIR detection of small tumors can be achieved using fluorescence emission and/or Raman scattering, in particular if one used surface-enhanced resonance Raman scattering (SERRS) [3]. SERS is as sensitive as fluorescence but provides even higher molecular specificity due to a unique vibrational signature of molecules. Moreover, SERS allows simultaneous (multiplex) detection of several signals within complex biological environments [4].

Hybrid, organic-inorganic nanostructures can be used for multimodal diagnosis since they allow combining multiple nanomaterials (magnetic NPs, plasmonic NPs, drugs, biological targeting ligands) and functionalities [5]. This is the case of nanovectors we developed for bi-modal (MRI-fluorescence) and tri-modal (MRI-SERRS-fluorescence) detection of cancer cells as described below.

The core of our nanovectors is composed of superparamagnetic iron oxide nanoparticles (SPIONs) alone or combined with plasmonic AgNPs. The SPIONs generate a negative contrast on T2*-weighted MRI and are well known to improve the in vivo cancer detection [1]. The plasmonic surface of AgNPs provides enhanced Raman response of adsorbed molecules. For optical detection, we labelled the nanovectors with VNIR fluorophores and/or to Raman reporter molecules attached to the inorganic cores and coated them with biocompatible polymers. The biological ligands like scFv antibody fragments of Herceptin® and folic acid molecules are bound to the external polymeric surface of the nanovectors. These ligands enhance the specificity and the efficiency of the vector delivery to HER-2 overexpressing breast cancers which are the most aggressive breast tumors. Therapeutic agents like chemotherapy drugs and siRNA molecules are also hidden within the polymeric shell of the nanovectors. This structural organization insures the drug/label protection and good biocompatibility of the nanovectors, and corresponds to the rational design strategy.

## II. RESULTS

### a. Covalently assembled magnetic nanovectors of chemotherapy agent

We generate our covalently assembled nanovectors via silane-mediated binding of native or ligand-modified PEG chains and fluorophores to the SPION surface [6]. For this, one-pot synthesis protocols proposed by our team have the advantage of reducing the number of intermediate steps. To load these nanovectors with anticancer drug doxorubicin, we developed an experimental approach using pre-formed chelate doxorubicin-Fe<fnref n="2" />⁺ [7]. The interest of this approach consists in the accelerated drug release in acidic environment as it is the case in tumors and in cell lysosomes. Such nanovectors of doxorubicin have been shown to be stealthy [8] and to possess a significant MRI-

<footnote n="1">¹ Université François Rabelais, EA6295 Nanomédicaments et Nanosondes, Tours, France.</footnote>

<footnote n="2">²CNRS-UPR 4301 - Centre de Biophysique Moléculaire - Equipe « IRM, signaux, images et expression des gènes », Orléans, France.</footnote>

<footnote n="3">³ INRA, Université François Rabelais, UMR 0483 Immunologie Parasitaire et Vaccinologie, Tours, France.</footnote>

<footnote>*Corresponding and presenting author: igor.chourpa@univ-tours.fr</footnote>
fluorescent response, anticancer activity *in vitro* [7] and *in vivo* [9], and reduced hematotoxicity.

From recently, we are working to improve the
cancer targeting efficiency of our nanovectors by
decorating them with a single-chain variable fragment
(scFv, molecular weight ca 27-28 kDa) targeting HER-2.
This strategy has several advantages over whole
antibodies: (i) the scFv are one-fifth the size of whole IgG
antibodies (thus, nanoplatforms functionalised with scFv
are much smaller) and they retain full antigen bonding
capacity; (ii) unlike whole antibodies, scFv do not contain
the Fc constant domain and therefore are not able to trigger
potentially harmful immune responses.

b. Electrostatically assembled magnetic nanovectors of siRNA

Short interfering RNAs (siRNA) are a promising tool to
treat various human diseases, including cancer. The siRNA
act via the RNA interference (RNAi) mechanism.
However, the systemic administration of siRNAs does not
allow to attain efficiently the site of action. To overcome
this difficulty, we develop magnetic siRNA nanovectors
(MSNs) based on SPIONs coated with siRNA and chitosan
polymer via layer-by layer electrostatic deposition [10]. In
addition to protecting siRNAs, chitosan is intended to
enhance the efficacy of the MSNs transfection to cancer
cells. To improve the formulation and optimize the
component quantities, we applied experimental design
strategy, respectively a Box-Behnken and a Plackett-
Burman design [10]. The method provides surface plots
helpful to optimize and predict the component quantities of
the MSNs regarding their hydrodynamic diameter (D<sub>H</sub>).
The results show that the most influent parameter was the
order of the components incorporation. The model
equations allowed to obtain MSNs with D<sub>H</sub> smaller than
100 nm which was desired for their systemic
administration.

c. Electrostatically assembled magneto-plasmonic nanoprobes

Controlled electrostatic assembly of SPIONs, AgNPs, nile blue (NB) dye and chitosan (Chi) polymer has been used to obtain stable magneto-plasmonic clusters (AgION-Chi) able to act as MRI-SERRS-active nanoprobes. The interest of the AgION-Chi nanoprobes is in that they are: (i) very rapid and easy to produce without using neither toxic reagents nor conjugation chemistry; (ii) readily biodegradable due to their electrostatic assembly; (iii) colloidally and spectrally stable for nearly one month. The nanoprobes allowed triple fluorescence-SERRS-MRI detection of HeLa cancer cells.

III. DISCUSSION – CONCLUSION

Rationally designed, the nanovectors described here show
a good compromise between their physico-chemical
characteristics (size ca. 100 nm, nearly neutral surface

charge, long-term colloidal stability, sufficient optical and
magnetic response) and biological properties
(biocompatibility *in vitro* and *in vivo*, ability to recognize
cancer cells and to deliver drugs and/or siRNA).
These nanovectors are theranostic and make possible a
complementary use of the spatially resolved *in vivo*
analysis by MRI with the real time multiplex optical
sensing by means of VNIR probing. This is expected to
provide more efficient and flexible solution for cancer
detection, localization/characterization and therapy.

ACKNOWLEDGMENTS

We acknowledge financial support from Région Centre,
INCa, and Ligue Nationale contre le Cancer.

REFERENCES

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