bluuebunny/crossref_metadata_2025 huggingface dataset:
P11.03 Next generation sequencing-identified IDH-1, ATRX, TP53 mutation and MYCN amplification in glioblastoma with primitive neuroectodermal tumor-like feature and extensive subarachnoid spread: a case report
Authors: Lee K , Gheeyoung C
Published: September 2019

BACKGROUND

In recent years, there have been numerous molecular studies in the glioblastoma. With regard to glioblastoma showing primarily extensive subarachnoid spreading, however, the pathogenesis remains to be determined due to its low incidence. Present report is a rare case of glioblastoma with primitive neuroectodermal tumor (PNET)-like feature, showing extensive subarachnoid spread, as well as the distinguishing result of its molecular analysis using next generation sequencing (NGS).

DESCRIPTION

A 37-year-old male presented with a new onset, severe headache. Magnetic resonance imaging (MRI) test revealed a diffuse infiltrative mass in the right frontal lobe. The tumor was also spread broadly in subarachnoid space. The patient underwent craniotomy and tumorectomy. Microscopically, glioblastoma showed PNET-like feature. Tumor cells had high nuclear/cytoplasmic ratios, oval, slightly elongated or angulated hyperchromatic nuclei, markedly increased mitotic activity, and obvious apoptotic bodies. Immunohistochemical analysis showed partly positivity of GFAP. Also this tumor demonstrated CD56 and synaptophysin immunoreactivity. IDH-1 immunohistochemistry was negative, which initially led to the diagnosis of glioblastoma, IDH wild-type. However, IDH-1 (p.Arg132His) mutation was found in subsequent NGS test. Additionally, two frame shift genetic alteration was found including ATRX (p.Tyr2208fs) and TP53 (p.Pro301fs). Interestingly, also MYCN (2p24.3) amplification was identified by both NGS and fluorescence in situ hybridization (FISH) test. Estimated MYCN copy number was 18. Despite the post-operative therapy including temozolomide and radiotherapy was performed, the tumor aggressively progressed and the patient expired nine months after the first symptom onset.

CONCLUSION

Although there have been several previous reports of MYCN amplification in glioblastoma with PNET-like feature, this is the first case report which finding in IDH-mutant type. IDH-1 mutation and younger age are usually associated with a better prognosis in glioblastoma. However, this case may suggest that when MYCN amplification coexists in IDH-mutant type, the tumor behavior could be contrarily aggressive.
P11.06 Non epigenetic effect of vorinostat in glioblastoma cells
Authors: Perez T , Berges R , Maccario H , Braguer D , Honoré S
Published: September 2019

BACKGROUND

Glioblastoma multiform (GBM) is the most frequent primitive brain tumor. GBM has a high recurrence and mortality. Histone deacetylase (HDAC) inhibitors have evoked great interest because they are able to change transcriptomic profiles to promote tumor cell death but also show undesirable side effects due to the lack of selectivity.We show new properties of low dose vorinostat, which inhibits cytoplasmic HDAC6 and display interesting non-epigenetics effects, especially on the microtubular cytoskeleton.

MATERIAL AND METHODS

We used murine (GL261) and human (U87 and GBM6 stem cells) cellular models. The cell proliferation was assessed by MTT tests, the migration by the 24 hours Transwell technic and by wound/healing tests. The expression levels of proteins of interest were assessed by Western Blot. Microtubules dynamics were assessed by time-lapse videomicroscopy. The synergy between drugs was tested by Loewe model.

RESULTS

Vorinostat inhibited the proliferation and the migration of the three cell lines mentioned above at level below the EC50.Vorinostat induces tubulin acetylation and alpha-tubulin c-terminal detyrosination that signed microtubular stabilization and these effects are independent of histone acetylation (HADC3). Interestingly, vorinostat decreases EB1 expression (a bad prognostic factor in GBM) and decreases microtubule dynamics. Moreover, vorinostat decreases neural markers such as GFAP, beta3-tubulin and CNPase and increases mural markers expression such as SMA/EGFR and PDGFR. Finally, it showed a synergy combined with erlotinib.

CONCLUSION

Low dose vorinostat, which do not affect histone désacetylase, has antitumor effect on glioblastoma cells by a new mechanism involving microtubule cytoskeleton. Interestingly, combination of low doses vorinostatand erlotinibshowed a strong synergy. Low dose, vorinostat could therefore represent an interesting therapeutic option and fewer side effects and that could be used to increased GBM patient sensitivity to erlotinib.
P11.10 The IFNγ pathway mediates brain metastasis formation of breast cancer
Authors: Pedrosa R , Kros J M , Schrijver B , Marques R , Leenen P , Dik W , van Eijck C , Mustafa D
Published: September 2019

BACKGROUND

In previous work we showed the prominence of the T-cell response in the formation of brain metastases of primary ER negative breast cancers (Mustafa et al, Acta Neuropathol 2018). We also showed that breast cancer cells co-cultured with stimulated T lymphocytes overexpress Guanylate-binding protein 1 (GBP1) accompanying increased trespassing ability through an in vitro blood-brain barrier (BBB) model. In addition, we demonstrated a predilection for metastasizing to brain of breast cancer cells that were co-cultured with activated T cells in a mouse model. We now scrutinize the importance of the IFNγ pathway for tresspassing of the tumor cells through the BBB following T cell contact.

MATERIAL AND METHODS

Anti-hIFN-γ-IgA antibodies were used to neutralize the IFNγ effects on the tumor cells. The effects on the tumor cells is only due to native IFNγ produced by activated T cells, not by recombinant IFNγ. Since the IFNγ expression itself enhances its expression by the T cells, we blocked IFNγ receptors prior to adding CD3+ T cell conditioned media to the breast cancer cells. The receptor blocking was achieved by antibodies to the IFNγα and IFNγβ subunits. Activation of the STAT1 pathway was monitored by GBP1 expression. For functional read-out the in vitro BBB model was used.

RESULTS

The presence of T-lymphocyte-secreted IFNγ in the primary breast cancer microenvironment activates the STAT1-dependent IFNγ pathway in breast cancer cells, endowing them with an increased ability to trespass the in vitro BBB. Moreover, direct inhibition of soluble IFNγ, or blocking of the IFNγ-specific receptor in breast cancer cells significantly decreases their ability to cross the BBB.

CONCLUSION

The results illustrate the specific action of T lymphocytes in the formation of cerebral metastasis involves the IFNγ signaling pathway as one of the crucial entangled pathways Subsequent studies should aim at the interference with the IFNγ pathway to develop preventive strategies against the formation of cerebral metastases of breast cancer.
P11.19 Glioblastoma cancer stem cells proliferation and cell cycle arrest under TTFields influence
Authors: Mazzanti M , Verduci I , Cannavale G , Pyromali I , Poloni R , Rampini D
Published: September 2019

BACKGROUND

Glioblastoma (GBM) is the most aggressive and fastest growing-type among brain tumours. The current standard of care is a combination of surgery, radio- and chemotherapy. However, GBM is characterized by a poor prognosis and an average life expectancy of 15 months under current treatment. Tumor treating fields (TTFields) are low-intensity electric fields alternating at an intermediate frequency (200kHz), which have been demonstrated to block cell division and interfere with organelle assembly. It has been evaluated in randomized phase 3 trials in GBM and demonstrated to prolong progression-free survival and overall survival when administered together with standard maintenance temozolomide (TMZ) chemotherapy in patients. This technology represents the biggest step forward against GBM in more than a decade.

METHODS

Primary cultures of human glioblastoma enriched in cancer stem cells (GSCs) were stimulated using the InovitroTM System (NOVOCURE) for 72 hours. Remaining cells were collected for further experiments.

RESULTS

Our investigation has targeted GSCs, a subset of cells that are considered responsible for tumour recurrence and confer resistance to chemotherapy and radiotherapy. Our data show that after 72 hours of TTFields exposure, more than 60% of the cellular population are dead. The residual GSCs were able to recover proliferation as soon as TTFields application had ended. We used different combinations of TTFields exposure times and pharmacological treatments with the main objective to impair cell proliferation relapse. Prolong TTF stimulation kills more cells but survivor cells are still able to resume proliferation. The addition of 10 mM TMX increase drastically the efficacy of the TTFields treatment even if the few cells left were able to slowly form colonies in a single cell cloning assay.

CONCLUSION

GCSs are sensitive to TTFields and to the combining pharmacological treatment with TMZ. The strategy to optimize the antiproliferative action on GCSs relapse will be to find a best combination of anticancer drugs concentration, together with TTFields exposure.
P11.21 Treating numerous secondary tumors in the brain with tumor treating fields: Simulation study to identify optimal transducer array layout
Authors: Kinzel A , Yesharim O , Naveh A , Bomzon Z
Published: September 2019

BACKGROUND

Tumor Treating Fields (TTFields) use alternating electric fields for the treatment of solid tumors. The therapy is approved for glioblastoma multiforme (GBM), and a phase III trial in 1–10 brain metastases from non-small cell lung cancer (METIS) is currently enrolling patients. In GBM, the layout of the transducer arrays delivering the TTFields to the tumor is optimized for high field intensity in the tumor, while the dose in other regions is decreased. In the setting of secondary brain tumors, as they manifest as brain metastases in 10–30 % of adult cancer patients - especially in melanoma, lung, breast, colon, and kidney cancer - a high TTFields dose in the entire brain would be beneficial. Thus, numerous tumors instead of only one lesion should receive therapeutic TTFields doses. In this study, transducer array layouts aiming for a homogeneous TTFields distribution in the whole brain were investigated.

MATERIAL AND METHODS

We used computer simulations in a realistic computational head model of a 40+ years old man, constructed in-house from a T1 MRI series, to compute the field distributions obtained with various transducer array layouts. The distribution of TTFields delivered by pairs of transducer arrays at different positions on the head and neck was simulated using Sim4Life v3.0 (ZMT Zürich). For each layout, we determined and compared the mean and median field intensities in five pre-determined sections of the brain: (1) the cerebellum and brain stem together with other infra-tentorial anatomical regions; and (2–5) the four cerebral quadrants.

RESULTS

One array layout could be identified yielding median intensities between 1.5 V/cm to 1.7 V/cm in all areas and a homogeneous distribution within the brain. This layout is composed of one pair of arrays positioned on the right temple and left scapula, and the other pair positioned on the left temple and right scapula.

CONCLUSION

This study was able to determine a novel TTFields transducer array layout that might be used for treatment of the entire brain with therapeutic intensities, as would be beneficial in patients with brain metastases.
P11.22 Radiotherapy effects in GBM Rat model CNS1
Authors: Furman O , Daniels D , Guez D , Last D , Sharabi S , Talianski A , Mardor Y , Zach L
Published: September 2019

BACKGROUND

CNS1 is a syngeneic glioma model in Lewis Rats. It is an aggressive infiltrating tumor cell line that mimics important aspects of human GBM such as rapid growth, dispersal along blood vessels and white matter, pseudopallisading cells with features of hemorrhage and necrosis. CNS1 tumors are infiltrated with macrophages and T-cells, and were studied in the context of immunotherapy and gene therapy, extracellular matrix and invasion, but CNS1 response to radiation has not yet been described. If we wish to combine novel immune-based therapies with existing GBM protocols that include radiation and chemotherapy, we will need models that respond to these protocols. As a first step in this direction, we sought to describe CNS1 response to radiation in vitro and in vivo.

MATERIAL AND METHODS

In vitro, survival of irradiated CNS1 cells was assessed with clonogenic assay. Radiation varied in dose from 0 to 10 Gy and was delivered via Kimtron Polaris X-ray generator. In vivo, male Lewis rats were intra-cranially inoculated with 0.5*106 CNS1 tumor cells and monitored for survival. Treated rats (N=6) were subjected to a single 20Gy whole-head radiation treatment under full anesthesia, delivered five days post-inoculation. Control rats (N=5) were anesthetized but not irradiated. Tumor size was monitored using contrast enhanced T1-weighted MRI in both treated and control rats at several time points (4, 6, 11, 18 and 32 days post tumor inoculation).

RESULTS

CNS1 cells are sensitive to radiation in vitro, as cell survival decreased after exposure to increasing amounts of radiation. In vivo, while initial tumor size did not significantly differ between groups, rats treated with radiation survived significantly longer than control rats (23.8 ± 5.0 days vs. 11 ± 4.1 days, p<0.005). Growth arrest following irradiation in vivo was not detected 1d after treatment but was observed 6d post-irradiation. Growth arrest was recorded in half of the treated rats, showing no increase in tumor size (N=2) or reduction in tumor volume (N=1) relative to 1d post-irradiation. Tumor growth rates were lower in all irradiated rats relative to control rats. Survival time was negatively correlated with initial tumor size in the control group but not in the treatment group.

CONCLUSION

CNS1 rat model of GBM is a valid model of radiotherapy effects on GBM tumors. Further studies combining radiation and chemotherapy are the next step. Support for this work was provided by Israel Cancer Association.
P11.28 Alteration of blood brain barrier (BBB) permeability by Tumor Treating Fields (TTFields) in vivo
Authors: Keßler A F , Salvador E , Domröse D , Burek M , Tempel Brami C , Voloshin Sela T , Giladi M , Ernestus R , Löhr M , Förster C , Hagemann C
Published: September 2019

BACKGROUND

Alternating electric fields with intermediate frequency (100 - 300 kHz) and low intensity (1 - 3 V/cm), known as Tumor Treating Fields (TTFields), have been established as a novel adjuvant therapy for glioblastoma (GBM) patients. The blood brain barrier (BBB) tightly controls the influx of the majority of compounds from blood to brain. Due to this regulation, the BBB may block delivery of drugs for treatment of brain tumors, in particular GBM. In this study, we investigated the influence of TTFields on BBB permeability in vivo.

MATERIAL AND METHODS

For determination of BBB permeability, rats were treated with 100 kHz TTFields for 72 h. At the end of treatment, rats were i.v. injected with Evan′s Blue (EB), which binds Albumin (~70 kDa) upon injection to the blood. EB was extracted after brain homogenization and quantified at 610 nm. In addition, cryosections of rat brains were prepared following TTFields application at 100 kHz for 72 h, and sections were stained for Claudin 5, Occludin and immunoglobulin G (IgG) to assess vessel structure. Moreover, serial dynamic contrast-enhanced DCE-MRI with Gadolinium contrast agent (Gd) was performed before and after TTFields application.

RESULTS

In vivo, the EB accumulation in the brain was significantly increased by application of TTFields to the rat head. Claudin 5 and Occludin staining was visible in vessel endothelial cells and localized at the cells’ edges in control cryosections of rat brains. In TTFields-treated rats, the vessel structure became diffuse; Claudin 5 and Occludin were delocalized and IgG was found throughout the brain tissue and not solely inside the vessels, as it is normally the case. Serial DCE-MRI demonstrated significantly increased accumulation of Gd in the brain, detected directly after 72 h of TTFields application. 96 h after end of TTFields treatment the effect on the BBB disappeared and no difference in contrast enhancement between controls and TTFields treated animals was observable.

CONCLUSION

Application of TTFields at 100 kHz could have the potential to deliver drugs to the brain, which normally are unable to cross the BBB by altering BBB integrity and permeability. Utilizing TTFields to open the BBB and its subsequent recovery, as demonstrated by the data presented herein, could lead to a clinical approach of drug delivery for treatment of malignant brain tumors and other diseases of the central nervous system. These results will be further validated in clinical trials.
P11.37 Evaluating water content and electrical properties at 200 kHz in brain and GBM tumor tissue of three TTFields patients with conventional imaging
Authors: Bomzon Z , Wenger C , Hershkovich H K , Tempel Brami C , Giladi M
Published: September 2019

BACKGROUND

Electrical properties (EPs) of brain tissue, specifically brain tumors, crucially determine the field distribution of Tumor Treating Fields (TTFields), an anti-mitotic treatment approved for glioblastoma multiforme (GBM). Due to the correlation of TTFields efficacy and field intensity at the tumor region, the knowledge of EPs in each patient is of great importance for patient-specific planning of treatment. Water content electrical properties tomography (wEPT) is a non-invasive imaging technique using water content (WC) maps obtained from rapidly acquired and processed conventional sequences to estimate the EPs of brain tissue at 128 MHz. The WC maps of this approach are constructed from two spin echo sequences similar to a T1 and a PD image. Following previous studies in rat tumor models demonstrating promising wEPT mapping of EPs in the brain at 200, this study examines the feasibility of this approach in human GBM patients.

MATERIAL AND METHODS

For three patients of the EF-14 trial population, we divided T1 and PD images pixel-by-pixel to obtain the image ratio. Using a transfer function, WC maps were generated and maps of the electrical conductivity σ and the relative permittivity ε r at 200 kHz were calculated with two different equations.

RESULTS

The median value of estimated WC remains similar in healthy brain tissues among all patients, ~73.5% in the white matter, ~82% in the gray matter. The median values of wEPT-estimated σ at 200 kHz in the white matter is ~0.09 S/m and in the gray matter ~0.18 S/m, corresponding median values of ε r at 200 kHz are ~2100 and ~3000 in white and gray matter respectively. Contrary, in the tumor the spread between the median values of WC and EPs is much higher. Stating the most important findings, in the necrosis median WC are 90.3%, 92.3%, 85.2% in patients 1–3 respectively with corresponding median σ values of 0.494, 0.657, 0.25 S/m. In the enhancing tumor the spread of median WC is even higher (67.2%, 83.6%, 85.5%), yet lower spread but also very heterogeneous median σ values of 0.075 S/m, 0.208, 0.259 S/m are estimated with wEPT.

CONCLUSION

Our results demonstrate the adaption of wEPT for mapping of WC and EPs at 200 kHz in three human GBM patients. In contrast to the vastly irregular tumor tissue, our estimations in healthy brain tissue are similar between patients and in accordance with EPs experimentally measured during our animal experiments and consistent with reported values in the literature. Hence, wEPT is a promising, fast technique based on regular MRI that might help patient-specific treatment planning of TTFields therapy, although the mapping of tumor tissue needs further confirmation in a greater population and investigations of EPs of excised tumor tissue samples should be conducted.
P11.44 Conception of a promising future therapy: Drug loaded-microbubbles against glioblastoma
Authors: Schulz E , Mawamba V , Sturm V , Ernestus R , Schatzschneider U , Löhr M , Hagemann C
Published: September 2019

BACKGROUND

Major obstacles for an effective chemotherapy of glioblastomas (GBM) are the blood-brain-barrier (BBB) and serious systemic side effects of the cytotoxic drugs. A new promising strategy could be the delivery of microbubbles, encapsulating the chemotherapeutics, across the BBB to the tumor site. This will shield the drug from detrimental systemic effects. Low intensity focused ultrasound (LIFU) is able to open the BBB and triggers targeted release of the drugs within the tumor. First data on the synthesis of microbubbles, specifically designed new drugs and the targeted rupture of microbubbles by LIFU are presented.

MATERIAL AND METHODS

Thin-film hydration of lipids was utilized to prepare microbubbles, which were tested for toxicity on the GBM cell lines GaMG, U87, U138 and U343. In addition these cells were treated with 6 platinum(II) and palladium(II) complexes conjugated to lipophilic side chains of different length (C1, C8, C10) for 72h. To evaluate cell viability and calculate EC50 values MTT assays and a real-time proliferation assay using the impedance-based xCELLigence DP-System were executed.

RESULTS

Microbubbles ≤ 2µm in diameter were synthesized and could be disintegrated by applying LIFU. Neither the intact bubbles nor the lipids alone had any toxic effects on the GBM cells. In contrast, all six drugs were highly effective with EC50 values far below those of Temozolomide (67µM) and in the range of the reference drug cisplatin (3µM). Especially the palladium(II) compound with the C1-chain displayed a very low EC50 value (<10µM), while the longer chains and the platinum(II) compounds were less effective (EC50 10–40µM). An early and concentration-dependent onset of the cytotoxic effect of drugs with C1 and C8 side chains was revealed in the real time proliferation assay.

CONCLUSION

All components for a new microbubble-based therapeutic strategy are in place. Microbubbles were synthesized without having toxic effects in cell culture. New highly potent palladium(II) and platinum(II) compounds with low EC50 values were developed. The next step will be their encapsulation into the microbubbles via their lipophilic side chains to develop an effective drug-delivery system for the treatment of GBM in combination with LIFU. This will allow increasing the local concentration of chemotherapeutic agents at the tumor site, irrespectively of their molecular size and BBB penetration capacity.
P11.55 Thymoquinone enhances the sensitivity of B16-F10 melanoma cell line to Gamma Knife radiosurgery
Authors: Hatiboglu M A , Kocyigit A , Guler E M , Akdur K , Khan I , Nalli A , Karatas E , Tuzgen S
Published: September 2019

BACKGROUND

Melanoma patients with brain metastasis have poor prognosis. Gamma Knife radiosurgery (GKR) is an effective treatment to control brain metastasis from melanoma. Thymoquinone (TQ) has gained attention as a potential therapeutic option due to its anti-tumor effects on various cancers. The aim of the study was to invetsigate the effect of GKR in B16-F10 melanoma cells in vitro and in established intracerebral melanoma model in mice in vivo, and the sensitizing efficacy of TQ towards GKR.

MATERIAL AND METHODS

Effects of GKR and combination treatment of GKR and TQ were studied on B16-F10 melanoma cells via analyzing cytotoxicty by adenosine triphosphate (ATP) assay; apoptosis induction by acridine orange staining and genotoxicity by comet assay. Expression of STAT3, p-STAT3 (Tyr705) and apoptotic pathway proteins was investigated by western blot analysis. Enzyme Linked Immunosorbent Assay (ELISA) was performed to assess the expression of inflammatory cytokines. Also survival was evaluated in mice with brain tumor after GKR and its combination treatment with TQ.

RESULTS

The effects of GKR on cytotoxicity, apoptosis and genotxicity were significantly enhanced by TQ on B16-F10 melanoma cells. GKR induced apoptosis through inhibition of p-STAT3 expression, which subsequently regulated pro- and anti-apoptotic proteins such as caspase-3, Bax, Bcl-2, survivin. Combination TQ with GKR irradiation enhanced the apoptotic effect of GKR irradiation. Furthermore, GKR decreased the levels of tumor-related inflammatory cytokines on B16-F10 melanoma cells. This effect was more pronounced when TQ was added to GKR irradiation. GKR with 15 Gy improved the survival of mice with intracerebral melanoma compared to untreated mice. Although, the additive effect of TQ in combination with GKR irradiation on B16-F10 melanoma cells in vitro, TQ did not add any significant survival benefit to GKR treatment in mice with intracerebral melanoma.

CONCLUSION

Our findings indicate that TQ would be a potential therapeutic and sensitizing agent in addition to GKR to enhance the anti-tumor effect of irradiation. Further studies are required to support our findings.
P11.58 Astrocytic Environment Aid the Evolution of Immunosuppressive Environment in Glioblastoma
Authors: Heiland D H , Ravi V M , Behringer S P , Schnell O
Published: September 2019

BACKGROUND

Glioblastomas are referred to as immunologically “cold” tumors since their cellular environment promotes an anti-inflammatory environment leading to an unresolved barrier to immunotherapy. Although the role of reactive astrocytes in other inflammatory diseases has been investigated in several studies, the immunoregulatory functions of astrocytes in the tumor environment remains poorly understood.

MATERIAL AND METHODS

We purified reactive astrocytes from de-novo glioblastoma and non-infiltrated cortex specimens by immunoprecipitation and analyzed the transcriptional phenotype by RNA sequencing. In order to investigate the origin of astrocytic transformation, we used a microglia loss-of-function model in human organotypic slices. Microglia was depleted by clodronat stimulation. We injected tumor cells and analyzed gene expression of the astrocytes after 7d incubation by RNA sequencing. Environmental cytokines were analyzed by multi-ELISA. Immunostainings of slices were processed by confocal microscopy and 3D reconstruction.

RESULTS

Here we address the immunological impact of tumor-associated astrocytes, we were able to identify a novel reactive subtype marked by JAK/STAT pathway activation and CD274 expression. Our results show a distinct astrocytic transcriptional phenotype that mutually arises from both microglia and astrocytes of the tumor environment. This interaction leads to a large release of anti-inflammatory cytokines such as TGFß and IL10. The reactive subtype switch of astrocytes was recovered by inhibition of the JAK pathway, which caused an increase of pro-inflammatory environment.

CONCLUSION

Our results can form the basis for a novel therapeutic approach by directly targeting tumor-associated astrocytes in order to transformimmunologically “cold” into “warm” tumors.
P11.62 Brain distribution models to select polymer-delivered drugs for the intra-cavity treatment of malignant glioma
Authors: Rowlinson J , McCrorie P , Smith S , Barrett D , Kim D , Grundy R , Scurr D , Rahman R
Published: September 2019

BACKGROUND

Conventional oral or intravenous chemotherapy distributes drugs to the whole body whereby systemic toxicity to healthy parts of the body (e.g. bone marrow failure) limits the maximum dose that can be achieved in the brain. This presents a particular concern for CNS tumours where the blood-brain-barrier (BBB) restricts drug influx from the circulation. The ability to deliver chemotherapy locally at the tumour site offers the opportunity to target residual cancer cells post-surgery whilst minimising systemic toxicity. We have developed a poly(lactic-co-glycolic acid)/poly(ethylene glycol) (PLGA/PEG) polymer matrix that forms a porous paste at room temperature when mixed with chemotherapy-containing saline, solidifying only at body temperature, with close apposition to the irregular surgical cavity. It is important that we can observe whether the drugs released from PLGA/PEG can penetrate brain parenchyma beyond the surgical resection margin at therapeutic doses. Currently the only way to measure the distribution of drugs in the body is to inject radioactive drugs into an animal. We aim to establish drug distribution parameters using label-free mass spectrometry imaging methods, prior to selection of drug formulations for clinically-relevant in vivo models. Drugs that penetrate the brain the furthest will be identified as good candidates for localised brain cancer drug delivery using PLGA/PEG paste.

MATERIAL AND METHODS

Diffusion rates were measured by examining the proportion of olaparib, dasatnib, carboplatin, etoposide, paclitaxel and gemcitabine at 2mg/ml concentration, which passes through 1mm slices of rat brain tissue within Franz cell chambers over a 6 hour period. The spatio-temporal distribution of label-free olaparib and dasatinib within mouse brain homogenate was quantitatively measured using innovative 3D OrbiSIMS, a hybrid time-of-flight / OrbitrapTM secondary ion mass spectrometer.

RESULTS

Within the Franz cell model, carboplatin and gemcitabine showed the highest diffusion rate diffusion at 16.4 and 6.53 µg/cm2/h respectively whereas olaparib, etoposide and paclitaxel were relatively poorly diffused at 1.87, 3.82 and 2.27 µg/cm2/h respectively. The minimum threshold of OrbiSIMS detection for label-free olaparib and dasatinib ions was 0.025 mg/ml and 0.2 mg/ml respectively throughout brain homogenate.

CONCLUSION

This study demonstrates different diffusion rates through brain tissue, between label-free chemotherapy drugs of distinct chemistries, with highest diffusion rates observed for carboplatin and gemcitabine. We also demonstrate label-free detection of olaparib and dasatinib using the innovative 3D OrbiSIMS method. These models will facilitate the rapid identification of agents most amenable for localised biomaterial-based chemotherapy delivery with high brain penetrance.
P12.06 Clinicopathological evaluation of PD-L1 expression and cytotoxic T-lymphocyte infiltrates across intracranial molecular subgroups of ependymomas
Authors: Sharma A
Published: September 2019

BACKGROUND

Immune check-point blockade targeting programmed cell death ligand-1 (PD-L1)/programmed death-1 (PD-1) axis has created paradigm shift in the treatment of advanced cancer. The prognosis of the ‘ST-RELA’ and ‘PF-A’ molecular subgroups of ependymomas (EPN) remains poor. This study aims to understand the potential candidature of EPNs for ICB.

MATERIAL AND METHODS

All supratentorial (ST) Grade II/III EPNs were classified into ST-RELA, ST-YAP and ST-not otherwise specified (NOS), based on L1CAM protein expression and/or presence of RELA/YAP1 fusion transcripts by RT-PCR. All posterior fossa (PF) BEPNs were classified into PF-A and PF-B based on H3K27me3 protein expression and chromosome 1q gain. Immunohistochemistry for PD-L1 (SP263, Ventana) and CD8 was performed. RelA protein enrichment at PDL1 promoter site was analysed by chromatin immunoprecipitation-qPCR (ChIP-qPCR) in three ST-RELA samples.

RESULTS

A total of 83 intracranial EPNs were included as follows: ST-RELA (n=32), ST-YAP (n=1), ST-NOS (n=11), ST-subependymoma (n=1), PF-A (n=25) and PF-B (n=13). Median tumor infiltrating CD8+ cytotoxic T-lymphocyte (CTL) density was 6/mm2, and was higher in ST EPNs (median 10/mm2) as compared to PF EPNs (median 3/mm2). PD-L1 protein expression in ≥1% tumor cells was noted in 17/83 (20%) of all EPNs, including 12/32 (37%) of ST-RELA and rare ST-NOS (2/11), PF-A (2/25) and PF-B (1/13) tumors. Twelve intracranial EPNs (14%, 12/83) showed high CTL density and concurrent PD-L1 positivity, of which majority (83%, 10/12) were ST-RELA EPNs. Enrichment of RelA protein was seen at PD-L1 promoter.

CONCLUSION

Increased CTL densities and upregulation of PD-L1 in ST-RELA ependymomas suggests potential candidature for immunotherapy.
P12.10 Treatment of pharmacoresistent epilepsy by combining stereotactic destruction and authologous mesenchymal stem cells
Authors: Slobina E , Dokukina T , Khlebokazov F , Misyuk N , Makhrov M , Korolevich P , Shamruk I , Marchuk S
Published: September 2019

BACKGROUND

Existing anti-epileptic drugs (AED) have limited efficiency in many patients, necessitating the search for alternative approaches such as stereotactic radiosurgery (SRS) and followed by stem cell therapy to improve results of previous treatment.

MATERIAL AND METHODS

From 2016 to 2017, a prospective study included 6 patients with a combined pathology, a median of 56 years of age, in the past TBI - 3 patients, a solitary metastatic focus in the brain (GM) - 2 patients. The indications for stereotactic intervention were both the presence of a metastatic lesion of the GM, and the bitemporal epilepsy with generalized convulsive, complex and simple partial attacks, regardless of the presence of an organic focus on CT or MRI GM. SRS was undertaken in patients undergoing a course of anticonvulsant drug therapy in combination with other types of symptomatic therapy in the absence of therapeutic effect. The frequency of generalized convulsive seizures ranged from 2–3 to 7–10 per month, complex partial from daily to 2–3 per month. SRS was performed on a stereotactic irradiation system with conical “BrainLab” collimators mounted on a linear CLINAC accelerator and allowing the session of SRS arc arches. Total dose was 12–30 Gy in isocenter, 1 fraction. After 2–3 monthes from SRS the patients received AED supplemented with single intravenous administration of undifferentiated autologous MSC (target dose of 1 × 106 cells/kg), followed by a single intrathecal injection of neurally induced autologous MSC (target dose of 0.1 × 106 cells/kg).

RESULTS

duration of epilepsy at the time of surgery ranged from 3 to 42 years. Cessation of attacks after 2–3 months after SRS with followed MSC therapy in 2 out of 6 patients was achieved remission (no seizures for one year and more), and 3 patients became responders to AEDs additionally, reduction of the pathological EEG pattern (disappearance of beta focusing, reduction of theta2-index to 23.6). Persistent clinical onco- and neurological remission 18 months. Reduction of the dose of enkorata-chrono - 3 patients, complete abolition of anticonvulsant therapy - 2 patients. MSC injections were well tolerated and did not cause any severe adverse effects.

CONCLUSION

MSC possess unique immunomodulatory properties and are a safe and promising candidate for cell therapy in AED resistant epilepsy patients in the frame of combined treatment.
P13.15 DNA methylation abnormalities in non-promoter regulatory regions are associated with invasive behavior in pituitary tumors
Authors: Mosella M Q S , Sabedot T S , Malta T M , Rock J , Felicella M , Aho T , Noushmehr H , Castro A V
Published: September 2019

BACKGROUND

Despite histologically benign, pituitary tumors (PT) may invade important adjacent neurovascular structures which can incur in significant comorbidities preventing a complete surgical resection and contributing to resistance to medical treatment. DNA methylation clearly stratified PT based on their functional status i.e. nonfunctioning PTs (NFPTs) from functioning PT (FPTs). However associations of methylation aberrations with invasive behavior is less clear.

MATERIAL AND METHODS

In order to evaluate whether DNA methylation alterations in regulatory regions other than promoter and coding regions are associated with invasive behavior we performed a meta-analysis of the genome-wide methylome of three public available PT cohorts plus our own (Illumina HumanMethylation platforms- 450K/EPIC). Pituitary specimens comprised of 43 invasive pituitary tumors (InvPT) and 37 noninvasive (NInvPT); 12 FPT and 68 NFPTs, in addition to 20 non-tumor pituitaries. RNA-seq data were available for one cohort (n=23, 12 InvPT,11NInvPT) and integrated with DNA methylation. Invasiveness criteria was based on Knosp grade >= 2 and/or sphenoid or dural invasion.

RESULTS

Wilcoxon Rank-sum test; Δβ=0.15; p-value <0.001 identified 58 differentially methylated CpG sites in InvPT that were mainly hypomethylated (95%) in relation to NInvPT. NInvPT methylation profile was similar to non-tumor specimens, despite its heterogeneity. Thirty-four percent (n=20) of the differentially methylated CpG sites were located within predicted enhancer regions distributed in intronic (40%), intergenic (40%) and promoter (20%) regions. Predicted enhancer-target genes were enriched for actin filament cell movement, response to starvation, growth factor stimulus and protein autophosporilation pathways. Among them, ZNF625 and INO80E were found mostly negative correlated among methylation and expression data (-0.50 and -0.48, respectively), besides DOC2A found to be one potentially differentially expressed gene under enhancer control (log2FC > 0.2, pvalue <0.05).

CONCLUSION

Our results suggest that methylation alterations in predicted regulatory regions, such as enhancers, annotated in non-promoter regions (introns and intergenic) may contribute to the invasive behavior of PT.
P14.01 Differential diagnosis of IDH mutant/IDH wildtype of glioma by using 11C-methionine, 11C-choline, and18F-fluorodeoxyglucose positron emission tomography
Authors: Takei H , Shinoda J , Ikuta S , Maruyama T , Muragaki Y , Kawasaki T , Ikegame Y , Okada M , Ito T , Asano Y , Yokoyama K , Nakayama N , Yano H , Iwama T
Published: September 2019

BACKGROUND

Positron emission tomography (PET) is important in noninvasive diagnostic imaging of gliomas. There are many PET studies on glioma diagnosis based on the 2007 World Health Organization (WHO) classification; however, there are no studies on glioma diagnosis using the new classification (the 2016 WHO classification).Here we investigated the relationship between PET imaging using 11C-methionine (MET), 11C-choline (CHO), and 18F-fluorodeoxyglucose (FDG) and wildtype isocitrate dehydrogenase (IDH) (IDH-wt)/mutant IDH (IDH-mut) in astrocytic and oligodendroglial tumors according to the 2016 WHO classification.

MATERIAL AND METHODS

In total, 105 patients with newly diagnosed cerebral gliomas (six diffuse astrocytomas [DAs] with IDH-wt, six DAs with IDH-mut, seven anaplastic astrocytomas [AAs] with IDH-wt, 24 AAs with IDH-mut, 26 glioblastomas [GBMs] with IDH-wt, five GBMs with IDH-mut, 19 oligodendrogliomas [ODs], and 12 anaplastic oligodendrogliomas [AOs]) were included. All OD and AO patients had both IDH-mut and 1p/19q codeletion. The maximum standardized uptake values (SUVs) of the tumor/normal cortex mean SUV ratios (T/N ratios) for MET, CHO, and FDG were calculated; the mean T/N ratios of DA, AA, and GBM with IDH-wt/IDH-mut were compared. The diagnostic accuracy for distinguishing gliomas with IDH-wt from those with IDH-mut was assessed using receiver operating characteristic (ROC) curve analysis of the mean T/N ratios for the three PET tracers.

RESULTS

There were significant differences in the mean T/N ratios for all three PET tracers between the IDH-wt and IDH-mut groups including all histological classifications (p<0.001). Among the 27 gliomas with mean T/N ratios higher than the cutoff values for all three PET tracers, 23 (85.2%) were classified into the IDH-wt group using ROC analysis. In DA, there were no significant differences in the T/N ratios for MET, CHO, and FDG between the IDH-wt and IDH-mut groups. In AA, the mean T/N ratios of all three PET tracers in the IDH-wt group were significantly higher than those in the IDH-mut group (p<0.001). In GBM, the mean T/N ratio in the IDH-wt group was significantly higher than that of the IDH-mut group for both MET (p=0.034) and CHO (p=0.01). However, there was no significant difference in the ratio for FDG.

CONCLUSIONS

PET imaging using MET, CHO, and FDG was confirmed to be informative for preoperatively differentiating gliomas according to the 2016 WHO classification, particularly for differentiating IDH-wt and IDH-mut tumors.
P14.05 Treatment of relapsing patients with intracranial germ cell tumors: the French experience
Authors: Callec L , Patte C , Lardy-Cleaud A , Vignon L , Alapetite C , Chastagner P , Faure Conter C , Frappaz D
Published: September 2019

BACKGROUND

Optimal strategy for treatment of relapsing intracranial malignant germ cell tumors remains ill-defined. Authors describe the French experience over a 25-year period.

MATERIAL AND METHODS

Relapsing patients were retrieved by their participation in the SFOP-TGM 90-92 or GCT 96 protocols or from National Childhood Solid Tumour Registry.

RESULTS

The cohort included Group A: documented germinomas (n=14), Group B: patients treated as germinomas without histopathological proof (n=5) and Group C: secreting germ cell tumours (n=25).

Patients all received standard dose chemotherapy, and some high dose chemotherapy (VP16 Thiotepa with stem cell rescue) and/or various type of radiation.

The 5 year EFS and OS post relapse are: 79%[47 to 93%] and 86% [54 to 96%] respectively in group A, 20% [1 to 58%] and 80% [20 to 97%] respectively in group B, 56% [35 to 73%] and 60% [38 to 76%] respectively in group C.

Among 14 Group A patients, 7/12 receiving high dose versus 3/3 receiving standard dose chemotherapy, and 9/11 re-irradiated versus 1/3 not re-irradiated are CR2.

Among 21 Group C patients. who had received radiation in first line, 10/15 receiving high dose versus 1/6 receiving standard dose chemotherapy, and 7/11 re-irradiated versus 4/10 not re-irradiated are CR2.

CONCLUSION

The outcome of relapsing germinoma is favourable and intensity of second line remains matter of debate. High dose chemotherapy with radiotherapy, when feasible, should remain the reference for treatment of a relapsed non germinoma, though more active treatments are warranted.
P14.18 Prognostic value of serial dynamic O-(2-[18F]-fluoroethyl)-L-tyrosine PET in patients with non-resectable malignant glioma undergoing chemoradiation
Authors: Rosen J , Stoffels G , Lohmann P , Bauer E K , Werner J , Rapp M , Felsberg J , Fink G R , Langen K , Galldiks N
Published: September 2019

BACKGROUND

A complete resection of high-grade gliomas (HGG) is associated with improved survival, which, however, cannot be achieved in a considerable number of patients. We here evaluated the prognostic value of serial O-(2-[18F]-fluoroethyl)-L-tyrosine (FET) PET in patients with newly diagnosed, non-resectable astrocytic HGG undergoing chemoradiation with temozolomide.

MATERIAL AND METHODS

Serial dynamic FET PET scans were performed in 18 newly diagnosed patients with molecularly defined, non-resectable HGG at baseline and after chemoradiation (8±3 weeks). Both static (tumor/brain ratios, FET tumor volumes) and dynamic FET PET parameters (time-to-peak, slope), as well as MRI changes according to RANO criteria at first follow-up after chemoradiation (8±3 weeks), were obtained. The predictive ability of FET PET parameters and RANO criteria was evaluated with regard to the progression-free survival (PFS). Using ROC analyses, threshold values for FET PET parameters were obtained. Subsequently, univariate and multivariate survival analyses were performed to assess their predictive power for PFS.

RESULTS

ROC analysis revealed that the mean tumor/brain ratio (AUC, 0.84), FET tumor volume (AUC, 0.89), and slope (AUC, 0.72) at baseline were predictive for a prolonged PFS (9.3 vs. 5.7 months, P=0.05; 10.3 vs. 5.9 months; P=0.03; 13.5 vs. 6.2 months, P=0.02, respectively). Furthermore, FET tumor volume and slope remained significant in the multivariate survival analysis (both P<0.05). In contrast, relative changes of static or dynamic FET PET parameters at follow-up and MRI changes according to RANO criteria were not significant in this albeit small series of patients.

CONCLUSION

Results suggest that before initiation of chemoradiation FET PET parameters at baseline can be used to predict PFS in patients with non-resectable HGG.
P14.30 Treatment outcomes of newly-diagnosed glioblastoma multiforme (GBM) by O6-methylguanine DNA methyltransferase promoter (MGMT) status: a multi-country study
Authors: Fu A Z , Hall J P , Bailey A , Kemp J , Thompson G J , Lee C
Published: September 2019

INTRODUCTION

This study evaluated the relationship of MGMT status with first-line (1L) treatment outcomes of patients with newly-diagnosed GBM in France, Germany, Italy, Spain, the UK (EU5), and Canada.

MATERIALS AND METHODS

Medical oncologists and neuro-oncologists across EU5 and Canada completed a point in time, cross-sectional survey for the next 8 GBM patients seen between May and July 2016 within EU5 and Canada. All results apart from time to progression (TTP) were presented for patients receiving 1L active drug treatment. TTP was calculated from initiation of 1L treatment to initiation of second-line treatment. Results presented are statistically significant (p<0.05) unless otherwise specified. Bases vary depending on data availability.

RESULTS

A total of 241 physicians reported on 1747 patients with GBM. 875 were receiving 1L active drug treatment at time of survey. Mean age was 59.7 years (median=61) and 34.6% were women. Mean life expectancy was 14.9 months (median=12) at diagnosis and mean TTP was 8.5 months (median=7.3). Surgery was performed in 62% of patients (n=546) prior to 1L drug treatment; 38% of patients (n=329) had no surgery. Patients with surgery had a higher life expectancy at diagnosis vs patients with no surgery prior to 1L (mean=16.4 vs 12.2 months; median=15.0 vs 12.0). Patients who received corticosteroids (n=524) vs no corticosteroids during radiotherapy (n=64) had a shorter life expectancy at diagnosis (mean=15.0 vs 16.8 months, p=0.07; median=12.5 vs 13.9) and were more likely to have 8 or more inpatient days due to GBM (21% vs 8%, p=0.07) in the last 3 months prior to the survey. 62% of patients (n=541) had an MGMT-status recorded (tested: methylated or unmethylated), and 38% (n=334) were untested/ awaiting results (untested) at 1L. MGMT-tested patients had better life expectancy at diagnosis (mean=16.1 vs 12.9 months; median=15.0 vs 12.0) and longer TTP (mean=8.9 vs 7.8 months; median=7.8 vs 6.4) than untested patients. Among MGMT-tested patients, 58% were methylated and 42% were unmethylated. Methylated patients had similar life expectancy at diagnosis (mean=15.9 vs 16.3 months, p=0.85; median=15.0 vs 15.0) and TTP (mean=9.0 vs 8.8 months, p=0.42; median=8.0 vs 7.5) as unmethylated patients.

CONCLUSIONS

This analysis provides valuable insights into the 1L treatment outcomes of GBM patients in EU5 and Canada. Patients who did not undergo surgery had worse treatment outcomes. Steroid use appears to be associated with worse outcomes and higher healthcare resource utilization. Patient treatment outcomes varied depending on whether they are MGMT tested.