tbuckley/cabot-search huggingface dataset:
P26.10.B OPTIMIZING DELIVERY OF CHEMOTHERAPEUTIC AGENTS BY INTRA-ARTERIAL INFUSION IN GLIOBLASTOMA
Authors: J Latulippe, L Roy, F J Gobeil, D Fortin
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND Being the most aggressive malignant primary brain tumor, glioblastoma (GBM) is a difficult disease to treat. The first line treatment consists in maximal resection followed by concomitant radiotherapy and temozolomide. However, most patients will relapse during the after treatment. The overall survival is only 14,6 months while the progression-free survival is 6,9 months. Even if the first-line treatment is well established, there is no consensus for the second line. At our institution, Dr. Fortin prioritize the intra-arterial (IA) administration of chemotherapeutic agents (CTA), which conveys a higher drug concentration to the tumor site. Carboplatin is the most used chemotherapeutic, but certain patients will not respond or will even develop chemoresistance to this treatment. Therefore, there is a strong need for bringing different CTAs into clinical care to offer more options. Topotecan, cytarabine and paclitaxel are molecules evaluated in this study since they are all already used intravenously (IV) in the treatment of GBMs. MATERIAL AND METHODS Since these agents have never been used for IA, we need to determine their maximal tolerable dose (MTD) to avoid any adverse effects. As such, healthy rats were given the CTA by retrograde infusion in the external carotid. They were euthanized five days following that surgery. After determining the right dosages, the survival study was initiated. Indeed, every CTAs were administered IA but also IV to compare both administrations. We used the Fischer-F98 glioma rat model. For the efficacy study, we implanted 10 000 F98 cells into the brain of the Fischer rats. At day 10, they received the treatment associated to their group. The animals were followed every day following the surgeries and were euthanized when their condition was too deteriorated. The IV doses used (found in the literature) were topotecan: 4 mg/kg, cytarabine: 40 mg/kg and paclitaxel: 10 mg/kg. RESULTS The doses found in the MTD study for topotecan and cytarabine were 4 mg/kg and 8,75 mg/kg respectively. However, paclitaxel was toxic when administered via IA, probably because of its lipidic vehicle. As for the efficacy study, the IA group of topotecan showed a promising outcome with a higher survival rate (29 days) compared to IV (23,5 days). As for cytarabine groups, IA was detrimental, with a 22 days median survival, compared to 26,5 days for the IV group. CONCLUSION While topotecan and cytarabine could be given intra-arterially, only topotecan showed conclusive results with a higher survival rate. More animals will be added to this group to validate this outcome. Cytarabine does not appear to be a good option since the MTD IA dose yielded a worse survival rate than the dose administered via IV. As for paclitaxel, considering it cannot be used intra-arterially, intra-tumoral infusion will be tested to see if we can get positive results.
P12.08.B DUAL-CAR T CELLS AGAINST NKG2D LIGANDS AND VEGFR2 ACCUMULATE IN ORTHOTOPICALLY IMPLANTED GLIOMA AND PROLONG SURVIVAL OF IMMUNOCOMPETENT GLIOMA-BEARING MICE
Authors: M Mastall, G Dunkel, T Weiss, N Okada, B Weigelin, M Weller, P Roth
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND Glioblastoma is the most common malignant brain tumor with a devastating outcome. Novel therapeutic strategies are currently explored including the use of chimeric antigen receptor (CAR) T cells. However, in glioblastoma an impaired tumor infiltration by CAR T cells, antigen heterogeneity and an immunosuppressive tumor microenvironment remain significant obstacles. NK group 2D (NKG2D)-CAR T cells efficiently lyse glioma cells and confer a survival benefit of glioma-bearing immunocompetent mice upon intratumoral administration. This effect is less prominent after intravenous injection, suggesting an insufficient trafficking of CAR T cells into the tumor. Vascular endothelial growth factor receptor 2 (VEGFR2)-CAR T cells were shown to accumulate in murine glioma upon intravenous administration and to have anti-glioma and anti-angiogenic activity. However, VEGFR2 expression in glioblastoma is heterogenous. Here, we generated Dual-CAR T cells targeting VEGFR2 and NKG2D ligands. METHODS Murine VEGFR2, NKG2D, Mock or Dual-CAR T were generated by retroviral transduction. In vitro, their activity was explored in co-culture assays. Glioma cells were orthotopically implanted into brains of immunocompetent mice and the infiltration of intravenously administered CAR T cells was assessed by in vivo fluorescence molecular tomography (FMT), ex vivo flow cytometry as well as 3D light-sheet microscopy (3D-LSFM). RESULTS Co-culture experiments demonstrated increased lysis of glioma cells by Dual-CAR T cells compared to VEGFR2-CAR T cells. By ex vivo 3D-FLSM, we observed substantial trafficking of systemically administered VEGFR2-CAR T cell into gliomas in vivo. In two different glioma models, FMT imaging as well as ex vivo flow cytometry revealed that Dual-CAR T cells accumulated to a similar extent at the tumor site as VEGFR2-CAR T cells and around 10-fold more Dual-CAR T cells were detected compared to CAR T cells expressing the NKG2D-CAR alone. Finally, Dual-CAR T cell treatment resulted in reduced growth of orthotopically growing gliomas and significantly prolonged the survival of glioma-bearing mice compared to Mock or NKG2D-CAR or VEGFR2-CAR T cell treatment. CONCLUSION Recent advances in CAR T cell optimization are mostly focused on CAR design, CAR generation and improving metabolic fitness, while improvements in CAR T cell trafficking are limited. VEGFR2-CAR T cells exhibited enhanced infiltration into the tumor site in the brain and arming of VEGFR2-CAR T cells with a NKG2D-CAR resulted in prolonged survival of glioma-bearing mice. Therefore, Dual-CAR T cells offer a versatile approach for the treatment of glioblastoma addressing two main hindrances, tumor infiltration and antigen heterogeneity.
P26.05.B DOSE-DEPENDENT DISTRIBUTION OF METHOTREXATE IN THE JUVENILE PIG BRAIN FOLLOWING INTRAVENOUS HIGH-DOSE METHOTREXATE INFUSION
Authors: T Rasmussen, M Guscott, Z Bascal, S Ward, S Howard, R K Buddington, J Palmfeldt, T S Mikkelsen
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND High-dose methotrexate (HD-MTX) is an essential element in the treatment of numerous cancers, including pediatric acute lymphoblastic leukemia with central nervous system involvement, and malignant brain tumors. However, our understanding of MTX distribution and elimination within the brain is incomplete. This limits the development of optimal dosing strategies. Additionally, while HD-MTX is effective, it is also known to cause acute central nervous system (CNS) toxicity in some patients. The exact mechanisms behind this toxicity are still unknown but is assumed by many to be linked to MTX mediated changes in brain metabolism. This calls for further investigation into MTX distribution and elimination within the brain, alongside a deeper understanding of MTX-induced changes in brain metabolism. MATERIAL AND METHODS This study assessed MTX levels in the brain of healthy, anesthetized juvenile pigs following a four-hour intravenous infusion of either standard 5 g/m2 HD-MTX or an increased dose of 20 g/m2 very high-dose MTX (VHD-MTX). A microdialysis probe was inserted into the cerebral frontal lobe to assess extracellular fluid MTX concentration. Blood plasma and urine samples were also collected. MTX concentrations were measured across samples using a highly sensitive liquid chromatography-mass spectrometry (LC-MS). RESULTS Serial samples of microdialysate, plasma, and urine were successfully collected from sedated pigs and MTX was detectable across all samples. The median MTX exposure represented as area under the drug concentration curve (AUC) in plasma and microdialysate was increased 3.6-fold and 3.1-fold respectively at the end of the four-hour infusion of VHD-MTX compared to HD-MTX. This increase was 4.9-fold and 3.1-fold after an additional four hours of observation post MTX-infusion (p &amp;lt; 0.05, n=12). CONCLUSION This study is part of a larger investigation exploring the use of VHD-MTX in treating CNS cancers. The results demonstrate a dose dependent increase in MTX exposure in extracellular fluid of the cerebral frontal lobe as demonstrated by microdialysis. Further studies using this model will investigate the effects of combining VHD-MTX with strategies to manage systemic toxicity. Additionally, we will employ explorative LC-MS metabolomics to assess changes in the metabolomic profile in CNS samples to address possible changes that could explain the phenomenon of CNS toxicity.
P26.07.A FGFR1 INHIBITION SENSITIZES GLIOBLASTOMA STEM CELLS TO TUMOR TREATING FIELDS
Authors: V Gouaze-Andersson, P Deshors, E Cohen-Jonathan Moyal
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND Glioblastoma (GBM) is the most lethal primary brain tumor in adult and is known to be particularly aggressive and resistant despite a treatment associating surgery and radio-chemotherapy. Over the last few decades, only few therapeutic advances have been achieved on GBM, but Tumor Treating Fields (TTFields) have recently improved outcomes of clinical trials. TTFields are a non-invasive, physical, local therapy relying on the delivery of alternating electrical fields with low intensity and intermediate frequency and are now a part of the first-line clinical treatment for GBM. Although TTFields increase patient’s survival, GBM is still an incurable disease with recurrences due to treatment resistance. Therefore, there is a need to understand mechanisms of GBM resistance to TTFields and to identify new therapeutic targets. One of the causes of resistance is the presence of Glioblastoma Stem Cells (GSC) known to be radio- and chemo-resistant and responsible for tumor regrowth. We have previously shown that tyrosine kinase receptor FGFR1 controls GBM agressiveness as well as GSC invasion and radioresistance and that its inhibition leads to radiosensitization through increasing mitotic cell death and microenvironment modulation. Because one of the main mechanisms of action of TTFields is mitotic disturbance and because TTFields act synergistically in vitro with irradiation, we hypothesize that targeting FGFR1 could sensitize GSC to TTFields. MATERIAL AND METHODS In this work we used a panel of primary GSC established from patient’s surgical GBM sample to test the potential role of FGFR1 in GSC response to TTFields. For this purpose, GSC were treated for 72 h with TTFields at 200 kHz which is the same frequency delivered to the patient receiving this therapy. TTFields treated GCS were also treated with siRNA targeting FGFR1 or pemigatinib, which is an inhibitor of FGFR1-3. RESULTS TTFields treatment increases expression and activation of FGFR1. Treatment of GSC with pemigatinib decreases FGFR1 expression and inhibits FGFR1 signaling pathway even when TTFields are applied. We demonstrated that pemigatinib, as well as FGFR1 silencing with RNA interference, sensitizes all GSC used in our experiments to TTFields by decreasing their survival. Finally, the adjunction of pemigatinib to treatment combining irradiation (IR) and TTFields also sensitizes GSC by inducing a further decrease in the repair of IR-induced DNA damages. CONCLUSION These results demonstrated that FGFR1 inhibition in concomittance with TTFields application as standard GBM treatment would improve patient prognosis.
P26.11.B PIWIL2 AS A POTENTIAL REGULATOR OF STEM-CELL PHENOTYPE IN GLIOBLASTOMA
Authors: E Garcia-Borja, R Mateu, F Siegl, J Šána, A Šedo, P Bušek
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND PIWI proteins are highly conserved RNA-binding proteins that constitute a germline-specific subfamily of Argonaute proteins. They bind to short non-coding RNAs known as piRNAs forming a piRISC (piRNA-induced silencing complex). The PIWI/piRNA complexes are pivotal in repressing transposable elements and regulating gene expression in stem cells. Recent findings reveal their implication in the progression of several cancer types, including glioblastoma (GBM). This study analyzed the expression profiles of the four human PIWI proteins (PIWIL1, PIWIL2, PIWIL3 and PIWIL4) in gliomas and in glioblastoma stem-like cells (GSCs), a critical subpopulation of malignant cells involved in GBM progression, treatment resistance, and recurrence. MATERIAL AND METHODS A panel of 160 GBMs (grade 4, IDHwt), 20 low-grade gliomas (grade 2 astrocytomas and oligodendrogliomas), and 12 non-tumorous brain tissues (pharmacoresistant epilepsy) was selected to examine the expression of PIWIL1-4 by qRT-PCR. 24 paired GSC and non-GSC cultures were derived from freshly resected human GBMs to analyze the expression of PIWIL1-4 by RT-qPCR. RESULTS PIWIL3 expression was not detected in an initial screening of 30 GBM tissues, leading to its exclusion from further analyses. PIWIL1 was detected in only half of the 160 GBM tissues, exhibiting very low expression. In contrast, PIWIL2 and PIWIL4 were present in the majority of tissues, with their expression being lower in GBMs compared to low-grade gliomas and non-tumorous brain tissues. In agreement with our results in tissues, PIWIL1 and PIWIL3 were either undetectable or expressed at very low levels in GSCs and non-GSCs. PIWIL4 was detected in both cell subpopulations displaying similar expression in GSCs and non-GSCs. In contrast, PIWIL2 expression was significantly higher in GSCs compared to non-GSCs and declined in GSCs upon serum-induced differentiation. Moreover, only PIWIL4 and PIWIL2, were detected in GSC-derived orthotopic tumors in immunodeficient mice. CONCLUSION Our findings challenge the previously reported upregulation of PIWIL1 in GBMs and its presumed relevance in GSCs. Instead, our data indicate potential contribution of PIWIL2 in maintaining the stem cell characteristics of glioma cells. Experiments aimed at silencing PIWIL2 with siRNAs in GSCs are currently ongoing. This work was supported by the Ministry of Health of the Czech Republic (NV19-03-00501), the National Institute for Cancer Research (Programme EXCELES, LX22NPO5102), the European Union - Next Generation EU, EATRIS-CZ (LM2023053), and Charles University Cooperatio Program, research area”Oncology and Haematology”.
OS09.5.A UNEXPECTED FUNCTIONAL ROLE OF GLYCOCALYX ENCAPSULATION DURING GBM METABOLIC ADAPTATION OPENS AVENUES FOR NEW TREATMENT CONCEPTS
Authors: A Bång-Rudenstam, M Cerezo-Magaña, K Gonçalves de Oliveira, V Governa, H Talbot, L Kjellen, E Tykesson, A Malmström, T van Kuppevelt, K Forsberg-Nilsson, J Bengzon, M Belting
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND The inherent adaptive capacity of GBM to metabolic stress represents a key clinical challenge, and the underlying mechanisms remain ill-defined. Our lab focuses on the emerging role of lipid metabolism in cancer cell stress adaptation. We have previously found increased scavenging of lipid particles, including extracellular vesicles (EVs) and lipoproteins, during acute stress, further resulting in a lipid droplet (LD) storage phenotype. Here, we propose a new mechanistic link between chondrotin sulfate proteoglycan (CSPG) remodelling, extracellular lipid scavenging, and LD formation that together serve to fine tune intracellular lipid metabolism to escape ferroptosis. MATERIAL AND METHODS We have comprehensively profiled GBM tissues and cells during different acidic stress adaptation conditions, including freshly resected GBM specimens, primary GBM cultures, and established GBM cell-lines. Methods/analyses include: Multiomics (RNA, proteome, glycome), laser capture microdissection for spatial profiling, snRNA-seq, immunophenotyping by FACS and imaging, and in vitro treatment studies (in 2D and 3D) with live imaging of cytotoxicity and ferroptosis. Mouse GBM models were employed for in vivo treatment studies. RESULTS We provide first evidence of a functional role of cell-surface CSPGs during GBM adaptation to metabolic acidosis. Substantial CSPG induction coincides with LD formation in a process dependent on extracellular lipids and acidification. Specifically, CSPG encapsulation of stressed GBM cells acts as a shield against excessive lipid particle scavenging while LDs consitute an intracellular sink for lipid storage. Based on these findings, we introduce combined targeting of CSPGs and LDs as a strategy to challenge the stressed tumor niche. Concomitant disruption of CSPG and LD formation unleashes a metabolic vulnerability in acidic cells driven by uncontrolled lipid uptake and peroxidation, mitochondrial disintegration, and ferroptotic death. CONCLUSION In conclusion, this study identifies a potentially targetable adaptative mechanism in the treatment resistant, acidic niche of GBM. We propose that cancer cell escape from acidic stress involves the induction of CSPGs - providing a shield against the lipid rich tumor microenvironment - and LDs - forming an intracellular lipid sink. Combined targeting of CSPG and LD formation to induce ferroptosis may offer new therapeutic opportunities in GBM and other solid tumors.
P02.16.A IRF8-DRIVEN REPROGRAMMING OF THE IMMUNE MICROENVIRONMENT ENHANCES ANTI-TUMOR ADAPTIVE IMMUNITY AND REDUCES IMMUNOSUPPRESSION IN MURINE GLIOBLASTOMA
Authors: M Montoya, S A Collins, P Chuntova, T S Patel, T Nejo, A Yamamichi, N Kasahara, H Okada
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND Glioblastoma (GBM) has a highly immunosuppressive tumor immune microenvironment (TIME), largely mediated by myeloid-derived suppressor cells (MDSCs). Here, we utilized a retroviral replicating vector (RRV) to deliver Interferon Regulatory Factor 8 (IRF8), a master regulator of type 1 conventional dendritic cell (cDC1) development, in a syngeneic murine GBM model. We hypothesized that RRV-mediated delivery of IRF8 could “reprogram” intratumoral MDSCs into antigen-presenting cells (APCs) and thereby restore anti-tumor T-cell responses. MATERIAL AND METHODS Effects of RRV-IRF8 on survival and tumor growth kinetics were examined in the SB28 murine GBM model. Immunophenotype was analyzed by flow cytometry and gene expression assays. We assayed functional immunosuppression and antigen presentation by ex vivo T-cell-myeloid co-culture. RESULTS Mice with RRV-IRF8 pre-transduced intracerebral tumors had significantly longer survival and slower tumor growth compared to controls. RRV-IRF8 treated tumors exhibited significant enrichment of cDC1s and CD8+ T-cells. Additionally, myeloid cells derived from RRV-IRF8 tumors showed decreased expression of the immunosuppressive markers Arg1 and IDO1 and demonstrated reduced suppression of naïve T-cell proliferation in ex vivo co-culture, compared to controls. Further, intra-tumoral and cervical lymph node DCs from RRV-IRF8 tumors showed increased antigen presentation compared to those from control tumors. To assess mechanism of action, mice were treated azidothymidine (AZT), a viral replication inhibitor. Untreated mice maintained a superior survival benefit, which demonstrated that viral replication through proliferating cells is necessary for tumor control. This showed that IRF8 transduction in both tumor and non-tumor cells is necessary for survival benefit, associated with a reprogrammed, cDC1- and CD8 T-cell-enriched TIME. Moreover, subset of mice with RRV-IRF8 treated tumors achieved complete tumor clearance and further rejected subcutaneous GBM tumors, indicating the presence of immunological memory. CONCLUSION Our results indicate that reprogramming of glioma-infiltrating myeloid cells by in vivo expression of IRF8 may reduce immunosuppression and enhance antigen presentation in situ, achieving improved tumor control and conferring a significant survival benefit.
P02.17.B MANIPULATING GABAERGIC MECHANISMS TO DELAY GLIOMA INVASION IN PERITUMORAL MICROENVIRONMENT
Authors: M Scalera, E De Santis, A Sturlese Verduri, N Meneghetti, N Barsotti, E Novelli, A Mazzoni, M Pasqualetti, M Costa, M Caleo, E Vannini
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND The interaction between tumor cells and the main components of the tumor microenvironment (TME) seems to guide the progression of glioma. Several recent studies have highlighted that the synaptic activity of pyramidal neurons potently drives Glioblastoma (GB) proliferation. However, the role played by other neuronal cell types is still poorly understood. Based on our previous findings showing that the optogenetic stimulation of parvalbuminergic (PV)-inhibitory interneurons negatively regulates glioma progression, here we tested whether a chemogenetic manipulation of peritumoral PV-interneurons could influence GB growth. MATERIAL AND METHODS To address this issue, AAV-hSyn-DIO-hM4D(Gi)-mCherry or AAV-hSyn-DIO-hM4D(Gq)-mCherry were injected in the motor cortex bilaterally with respect to the GL261 injection site in PV-Cre mice. In order to manipulate PV-interneurons activity, animals will be randomized to receive CNO or vehicle intraperitoneally twice a day from day 12 (i.e., the glioma symptomatic stage) to day 22 (i.e., the glioma late stage) post tumor induction. As readouts, we: i) evaluated the tumor volume using Magnetic Resonance Imaging (MRI), ii) performed a longitudinal monitoring of motor capabilities using specific motor tests (i.e., Grip Strength and Grid walk), iii) assessed the survival, iv) determined the GB proliferation index of glioma-bearing mice after different treatments. RESULTS We found that the PV-interneurons inactivation resulted in worsening motor performances with respect to the vehicle mice. Despite that, no difference was observed in animal median survival or tumor volume among the two groups. However, MRI data showed a very significative reduction in the tumor volume at 28 days after glioma induction in PV-activated glioma bearing mice (p=0,004). In addition, PV-activation exert a slight effect on the maximum survival of CNO-treated glioma bearing mice with respect to the vehicle mice (39 vs 32 days). CONCLUSION Since there is no cure for GB, it is essential to find new and effective therapeutic approaches to improve the quality of life and to extend the life expectancy in patients. These data suggest that the inhibitory circuitry in the tumor-adjacent zone could delay GB growth, protecting peritumoral tissue from GB proliferation.