bluuebunny/crossref_metadata_2025_split huggingface dataset:
TMIC-58. ALTERATIONS IN GLIOBLASTOMA TUMOR MICROENVIRONMENT TRANSCRIPTOME AFTER TREATMENT WITH PEMBROLIZUMAB
Authors: Majd Nazanin , Yang Liu , Williford Garret , Alfaro-Munoz Kristin , Dasgupta Pushan , Heimberger Amy , Kannan Kasthuri , Huse Jason , Bhat Krishna , de Groot John 
Published: November 2023

Abstract

BACKGROUND
Immune checkpoint inhibitors (CPI) have failed to show survival benefit in glioblastoma. Understanding the alterations in the tumor microenvironment after treatment with CPI is crucial for development of immunotherapy in glioblastoma. We previously reported our window-of-opportunity trial of pembrolizumab in GBM (NCT02337686). In the current study, we obtained 8 paired glioblastoma tumors pre- and post-treatment with pembrolizumab and conducted RNA sequencing analysis to understand the alterations in the tumor microenvironment transcriptome in response to pembrolizumab. Method: We conducted bulk RNA-sequencing profiling and differential gene expression analysis to compare the expression levels of patient-derived glioblastoma pre- and post-treatment with pembrolizumab, while controlling for paired information. We performed gene enrichment analysis on the differentially expressed genes. We employed the single sample mww-GST method to assess the enrichment of various immune cell types that were previously identified, across the two groups.

RESULTS
Our analysis revealed 294 down-regulated and 259 up-regulated genes in the post treatment samples. The gene enrichment analysis demonstrated that the up-regulated genes were enriched in pathways related to the immune system. The single sample mwwGST analysis indicated a consistent overall up-regulation of expression in immune cell infiltration. Notably, T cell-related signatures, such as CD4, CD8, and NKT exhibited significantly higher infiltration in the post-treatment group. Interestingly, gene ontology analyses showed upregulation of phagocytosis and antigen presentation gene modules in the post-treatment samples. Additionally, we found neuronal components and genes related to hyperexcitability and synaptic connectivity with an overrepresentation of AMPA and DLG4 pathways in post-treatment tissues.

CONCLUSION
Our data show that in addition to the anticipated infiltration of myeloid and lymphoid cell populations, checkpoint blockade can also unexpectedly influence glioma-neuron interactions, which may represent additional immunotherapy targets in GBM. Further proteomic validation of infiltration of different cell types and their interactome in pre- and post-CPI treated tissue is warranted.
TMIC-67. GLIOBLASTOMA AUGMENTS SYSTEMIC IMMUNUNOSUPPRESSION UPON LOSS OF TESTOSTERONE VIA ALTERATION OF GLUCOCORTICOIDS PRODUCTION
Authors: Lee Juyeun , Lathia Justin , Chung Yoon-Mi , Sharifi Nima , Hao Yue , Berens Michael 
Published: November 2023

Abstract
While sex differences in incidence, genetics, and immune changes are established in glioblastoma (GBM), the specific contribution of sex hormones to these differences has not been fully explored. Testosterone has been shown to have a pro-tumorigenic effect on human and mouse GBM tumor cells. However, the systemic effect of testosterone in an immune competent model has not been tested. To test this, we performed castration or sham surgery on C57BL/6 male mice, followed by intracranial implantation of syngeneic glioma cells. Surprisingly, castrated mice exhibited more aggressive tumor growth compared to sham surgery controls, and this castration-induced aggressiveness could be reversed by administering exogenous testosterone. This finding was further confirmed by chemically blocking androgen receptor signaling using enzalutamide. We observed a significant increase in adrenocorticotropic hormone (ACTH) levels in the serum from castrated mice compared to the sham group after tumor implantation. Additionally, mass spectrometry analysis detected elevated levels of circulating glucocorticoids in castrated mice, and pharmacological blockade of glucocorticoid receptor using mifepristone reversed the shorted survival in castrated mice, but not in sham control. Flow analysis revealed decreased production of anti-tumor cytokines such as IFN-γ and TNF-α in CD4+ and CD8+ T cells from tumors and lymph nodes in castrated mice, suggesting systemic immunosuppression in these animals. Indeed, castration of immune-compromised mice, specifically NSG and RAG1KO strains, did not result in reduced survival. Moreover, when GBM cells were subcutaneously implanted into the flank, tumor growth in castrated mice was decelerated compared to sham controls. We also observed significantly lower ACTH levels in the flank tumor model compared to the brain tumor model, indicating a brain-specific effect of steroid hormones on tumor control. Taken together, these findings demonstrate that testosterone regulates T cell function through glucocorticoids, ultimately affecting GBM growth in a site-specific manner.
TMIC-83. THE MOLECULAR AND SPATIAL ARCHITECTURE OF AN IMMUNE-RICH GLIOMA SUBTYPE
Authors: Kumar Anupam , Wang Lin , Solomon David , Hervey-Jumper Shawn , Berger Mitchel , Diaz Aaron , Phillips Joanna 
Published: November 2023

Abstract
Diffuse glioma exhibit an immunosuppressive microenvironment, yet glioma subsets driven by RAF-alterations exhibit an increased immune signature. In pleomorphic xanthoastrocytoma (PXA), we identified CXCL14 as an important factor promoting a cytotoxic T cell response. The molecular and spatial relationships within this tumor type, however, are unknown. To address this, we use single nucleus RNA sequencing (snRNAseq), spatial transcriptomics, and spatial molecular imaging to construct a high-resolution molecular landscape of PXA. SnRNAseq was performed on 36,781 cells from 10 tumors, including initial and recurrent pairs, spatial transcriptomes were performed on a total of 91 FOVs comprised of 17 PXAs, and spatial molecular imaging (SMI) of 1,000 RNA probes at subcellular resolution was performed on 48,518 cells from 6 tumors, including two PXA, two pilocytic astrocytoma, and two ganglioglioma. For snRNAseq, neoplastic cells were separated from non-malignant cells based on analysis of inferred copy number alterations. CellChat was used to infer cell-cell communication. For spatial transcriptomics, data from 200-300µm FOVs were normalized and divided into CD3-high versus CD3-low groups. For SMI, cells with fewer than 20 transcripts were eliminated, data were normalized using simple total counts, cells were segmented using the Nanostring pipeline, and semi-supervised clustering was performed. Dual CXCL14 RNAscope and Iba1/CD3 immunostaining confirmed spatial relationships within the tumor. In PXA with high-CD3, differential gene expression analysis demonstrated broad immune-enrichment and identified upregulation of several ligand-receptor pairs. Integrating these data with snRNAseq and cell-cell communication analysis inferred paracrine interactions between tumor cells and T cells. One such factor, CXCL14, was expressed widely by tumor cells but spatial analysis revealed cohesive clusters of CXCL14-high expressing tumor cells. In anaplastic (CNS WHO grade 3) tumors, however, both CXCL14 expression and microglia decreased. These high-resolution molecular analyses of PXA identify dynamic tumor-immune crosstalk regulating the immune microenvironment.
MODL-02. DEVELOPMENT OF RODENT GLIOBLASTOMA MODELS OF MGMT-MEDIATED TEMOZOLOMIDE RESISTANCE
Authors: Lee Teresa , Bhatt Deepti , Sundaram Ranjini , Saltzman W Mark , Bindra Ranjit S , Vasquez Juan C 
Published: November 2023

Abstract
O6-methylguanine-DNA methyltransferase (MGMT) is an enzyme that repairs O6-methylguanine DNA damage lesions generated by alkylators such as temozolomide (TMZ), the standard chemotherapeutic for glioblastoma (GBM) treatment. High MGMT expression leads to TMZ resistance and is correlated with lower survival, making it an important prognostic biomarker and potential therapeutic target. Xenograft models with human GBM cell lines require implantation into immunodeficient animals, precluding the study of MGMT-dependent alkylator sensitivity in the setting of an intact tumor immune microenvironment. In this study, we developed rat and mouse glioma models of MGMT overexpression and characterized response to alkylator therapy. RG2 rat glioma and SB28 mouse glioma cells, which have low endogenous MGMT levels, were transduced with cDNA expressing FLAG-tagged MGMT to generate an isogenic pair. RG2 MGMT+ and SB28 MGMT+ cells showed robust expression of MGMT, which was effectively abrogated by MGMT inhibitors. In vitro cell viability assays showed that MGMT overexpression resulted in increased resistance to TMZ compared to parental cells, and this resistance was reversed when cells were concurrently treated with the MGMT inhibitor lomeguatrib. To model MGMT overexpression in vivo, luciferase-expressing RG2 MGMT+ cells were implanted into the brains of syngeneic Fischer 344 rats. Rats were treated with TMZ at 50mg/kg daily x 5 days via intraperitoneal injection. Tumor growth was measured by bioluminescence imaging and rats were monitored for survival and toxicity. RG2 MGMT+ tumors were highly resistant to TMZ, as treatment did not improve survival or delay tumor growth. Overall, our newly developed isogenic rat and mouse glioma models recapitulate MGMT-dependent TMZ sensitivity and provide useful preclinical tools for studying MGMT as a biomarker and potential therapeutic target in GBM. In addition, these syngeneic models facilitate studies on the immune modulation of chemotherapeutic agents in the context of MGMT expression and allows for testing of chemoimmunotherapy combinations.
MODL-07. NF1 MUTATION IN OLIGODENDROCYTE PRECURSOR CELLS INDUCES PRENEOPLASTIC LESIONS IN THE BRAIN
Authors: Pan Yuan , Hysinger Jared , Yalçın Belgin , Lennon James , Raghavan Preethi , Schindler Nicole , Anastasaki Corina , Chatterjee Jit , Mount Christopher , Nagaraja Surya , Scheaffer Suzanne , Attardi Laura , Gutmann David , Monje Michelle 
Published: November 2023

Abstract
Oligodendrocyte precursor cells (OPCs) are progenitor cells that give rise to oligodendrocytes in the central nervous system throughout the lifespan. Transcriptomic and lineage tracing studies suggest that OPCs may be the putative cell of origin for many gliomas in children and adults. While children with the neurofibromatosis type 1 (NF1) cancer predisposition syndrome typically develop optic pathway and brainstem gliomas, as many as 50% of children develop gliomas in other brain regions using radiologic criteria. Relevant to these gliomas, discrete patches of hyperdense OPCs were found in the brains of Nf1+/- mice. The OPCs within the lesions display a transient increase in proliferation and hypercellularity. When the brains of mice carrying different heterozygous NF1 patient derived germline Nf1 gene mutations were examined, hyperdense OPC lesions were only seen in mice whose Nf1 mutations also predispose to optic gliomas. Mechanistically, monoallelic KRAS hyperactivation also results in similar OPC hyperdense lesions. Moreover, OPC-specific Nf1-heterozygous mice display similar lesions, while OPC-specific Nf1-knockout results in generalized OPC hyperdensity throughout the entire brain. Taken together, these data suggest the existence of a preneoplastic state conferred by Nf1 mutation in OPCs and establish a tractable model system to explore the transition to glioma in children with NF1.
MODL-13. DEVELOPMENT OF TUMOR TREATING FIELDS (TTFIELDS) ARRAYS FOR TREATMENT OF HEAD TUMORS IN MICE MODELS
Authors: Zbidat Sewar , Blat Roni , Sellevoll Mariell , Gabay Martin , Schlachet Inbar , Cahal Shay , Davidi Shiri , Tzchori Itai , Haber Adi , Giladi Moshe , Palti Yoram 
Published: November 2023

Abstract

INTRODUCTION
Glioblastoma (GBM) is the most common malignant brain tumor. Tumor Treating Fields (TTFields) therapy is an approved treatment for GBM, delivered continuously by two array pairs placed on the skin surrounding the tumor region. Currently, most GBM animal models are based on mice. Since arrays for TTFields application to the mouse head are lacking, GBM-TTFields in vivo studies are limited. The small dimensions and specific geometries of the mouse head make the development of such arrays challenging.

METHODS
We tested different array layouts to identify one that will optimally accommodate the geometries of the mouse head while minimally restricting head movement. Additionally, we investigated adhesive tapes for array attachment to the skin, to allow good adherence and hence provide efficient treatment delivery. To validate that the selected array layout delivers therapeutic field intensity to the desired region, we performed simulations and fields measurements.

RESULTS
To overcome the issue of small head size we developed a layout in which the arrays on the head were divided into two smaller disks, with the opposing arrays situated on the mouse torso. We also identified a thin and transparent adhesive tape that facilitated the correct positioning of the arrays on the mouse head, offered good tackiness, and allowed for easy removal without leaving residual adhesive on the skin. The arrays met the minimum treatment requirements: field intensity ≥ 1 V/cm RMS, current ≥ 50 mA, usage ≥ 75%.

CONCLUSIONS
Our newly developed mouse head arrays are a flexible construct, that adheres strongly to the skin to enable efficient electric field delivery. By facilitating TTFields delivery to the heads of mice, we will expand the scope of GBM treatment research and will contribute to further advance this field.
MODL-17. OBESOGENIC DIET INCREASES NF1-OPTIC PATHWAY GLIOMA FORMATION THROUGH EFFECTS ON THE TUMOR CELL OF ORIGIN
Authors: Chan Ambrose , Zhang Kailong , Martin Gemma , Bano Sabiha , Avery Wang , Sarvika Mahto , Gutmann David , Brossier Nicole 
Published: November 2023

Abstract
Rates of both pediatric low-grade glioma and obesity have been rising in the U.S., and children of obese mothers have increased rates of several tumor types, including brain tumors. Importantly, obesity in the U.S. is largely dietary-driven, given the prevalence of high fat and high sugar (HFHS) diets. High-fat diet exposure can promote proliferation of embryonic neuroglial progenitor cells (NPCs), which animal modeling suggests are potential cells of origin for low-grade glioma. Together, these data suggested that in utero exposure to an obesogenic diet might affect the formation of pediatric brain tumors. Using preclinical models of the Neurofibromatosis type 1 (NF1) cancer predisposition syndrome, in which optic pathway gliomas (Nf1-OPGs) arise from NPCs in the developing third ventricular zone (TVZ), we sought to determine whether maternal obesogenic exposure would increase glioma formation through intrinsic effects on the tumor cell of origin. We demonstrated that progeny from obese dams exposed to HFHS diet during and preceding gestation had increased proliferation and glial differentiation of WT and Nf1-mutant TVZ NPCs. Progeny of non-obese dams exposed to this diet during gestation demonstrate a similar phenotype, suggesting these effects are related to maternal diet rather than weight. Obesogenic diet exposure (Ob) also increased glioma penetrance in two low-penetrance murine models of Nf1-OPG. Moreover, we demonstrated that Ob mice developed earlier tumor onset with elevated Erk activity in a high-penetrance Nf1-OPG model. In contrast, when obesogenic diet was introduced postnatally, only 11% of mice showed accelerated tumor formation. Ob-exposed nerves also displayed elevated levels of activated Stat3, consistent with upregulation of the IL6/Jak/Stat3 pathway with HFHS diet exposure. These findings establish maternal obesogenic diet as a risk for pediatric glioma formation, in part through in utero effects on the tumor cell of origin, and identify the MAPK and IL6/Jak/Stat3 pathways as targets for further investigation.
MODL-18. ESTABLISHMENT OF A NOVEL HUMANIZED PERIPHERAL BLOOD MONONUCLEAR CELL (HU-PBMC) MOUSE MODEL TO EVALUATE LYMPHOCYTE INVOLVEMENT IN GLIOBLASTOMA AND ITS TREATMENT
Authors: Deshpande Krutika , Kwang Denna , Cadiente Marvin , Delany Jasmine , Ahmed Taufeeq , McDonald Bailey , Wang Xi , Barrette Anne Marie , Sun Dongxu 
Published: November 2023

Abstract
Glioblastoma (GBM) has traditionally been classified as an ‘immune cold’ tumor, characterized by strong immunosuppression, low mutational burden, and few presentable antigens. However, lack of standard-of-care improvement and the discovery of targetable immune-modulatory biomarkers like PD-1/PD-L1 and CTLA4 in the GBM microenvironment has prompted clinical trials to explore immune checkpoint inhibition (ICI) efficacy in patients, with moderate responses in specific patient subsets. To better characterize ICI in GBM, and to match prospective patients to suitable therapeutics, it is necessary to establish preclinical GBM mouse models that incorporate the lymphocytic milieu as xenograft and syngeneic models are unable to replicate the immune responses seen in humans. We have established a novel humanized PBMC mouse model that recapitulates human GBM pathology in the brain, supports and maintains systemic and intracranial human T cell engraftment in the presence of GBM tumors, and allows for a suitable timeline for preclinical testing of therapeutics prior to onset of GVHD. In our study, orthotopic GBM-bearing NSG mice showed successful human CD45+ engraftment in the blood (~20%) and brain (~5%) 2 weeks post intravenous implantation of hu-PBMCs. Restricted tumor growth was observed, indicating initial responsiveness of human immune cells to GBM. Engraftment remained unaffected upon treatment with temozolomide (GBM standard of care), and persisted over 6 weeks (brain ~40%, blood ~70%) before GVHD onset (week 7). ~90% of the hCD45+ population was CD3+, indicating robust T cell engraftment, which was further characterized into CD4+ and CD8+ subsets and by activation status (HLA-DR, CD45RA/RO, PD-1, FOXP3 etc) within the tumor-bearing brain and blood. Anti-PD-1 inhibitor efficacy was also tested in engrafted animals to observe ICI potential in GBM. This humanized mouse provides a powerful translational model to evaluate T cell responses in tumor pathology and immunotherapy in GBM and potentially other primary or secondary brain malignancies.
MODL-36. EX VIVO GSC CULTURES RETAIN PATIENT-UNIQUE HALLMARK GENE EXPRESSIONS AND PREDICT PATIENT RESPONSE TO TMZ MORE SPECIFICALLY THAN MGMT PROMOTER STATUS
Authors: Ntafoulis Ioannis , Kleijn Anne , Ju Jie , Jimenez-Cowell Kevin , Fabro Federica , Klein Michelle , Yen Romain Tching Chi , Balvers Rutger , Kers Trisha , Kros Johan , Lawler Sean , Kremer Andreas , Li Yunlei , Stubbs Andrew , van den Bent Martin , Dirven Clemens , Leenstra Sieger , Lamfers Martine ,   GLIOTRAIN Consortium
Published: November 2023

Abstract

BACKGROUND
Patient-derived glioma stem-like cells (GSCs) have become the gold-standard in neuro-oncological research; however, it remains to be established whether loss of in situ microenvironment affects the clinically-predictive value of this model. We implemented a GSC monolayer system to investigate in situ-in vitro molecular correspondence and the relationship between in vitro and patient response to temozolomide (TMZ).

METHODS
DNA and RNA-sequencing was performed on 56 glioblastoma tissues and 19 derived GSC cultures. Sensitivity to TMZ was screened across 66 GSC cultures. Drug response readouts were related to clinical parameters of corresponding patients and whole-transcriptome data.

RESULTS
Tumor DNA and RNA sequences revealed strong similarity to corresponding GSCs despite loss of neuronal and immune interactions. Patient-specific hallmark gene expression levels were retained in vitro. Functional screening for TMZ sensitivity yielded three response categories (responders, intermediates and non-responders) which significantly correlated with patient survival, therewith providing a more specific response prediction than the binary MGMT marker. Transcriptome analysis of GSCs and parental tissues identified 121 genes related to TMZ sensitivity and overall survival, of which 21, including MGMT, could be validated in external datasets.

CONCLUSION
GSCs retain patient-unique hallmark gene expressions despite loss of their natural environment. Drug screening using GSCs predicted patient response to TMZ more specifically than MGMT status, while transcriptome analysis identified potential biomarkers for this response. GSC drug screening therefore provides a tool to improve drug development and precision medicine for glioblastoma patients.
MODL-37. MODELING REVERSIBLE TUMORIGENESIS IN CEREBRAL ORGANOIDS
Authors: Graham Maya , Codega Paolo , Campos Carl , Nandakumar Subhiksha , Rosenblum Marc , Antonescu Cristina , Grogan Meaghan , Benhamida Jamal , Bale Tejus , von Deimling Andreas , Socci Nicholas , Mellinghoff Ingo 
Published: November 2023

Abstract
Human cerebral organoids have emerged as a new approach to modeling the role of cancer-associated genes in tumorigenesis ex vivo. Here, we asked whether high-grade tumorigenesis in cerebral organoids is a reversible process. Using inducible cassettes carrying key oncogenic mutations histone H3K27M, dominant negative p53 and activated PDGFRA, we show that their expression in cerebral organoids reproducibly drives high grade malignant transformation in vitro and in vivo, including in orthotopic xenografts. We characterize this transformation as a fusion-negative rhabdomyosarcoma (FN-RMS) phenotype and identify a putative FN-RMS cell-of-origin in cerebral organoids. Finally, we show this transformation is dependent on persistent transgenic oncogene expression, as oncogene withdrawal causes complete tumor regression. These findings demonstrate that cerebral organoids may be used to pinpoint previously unappreciated and distinct roles of different mutations in tumor development and maintenance.