bluuebunny/crossref_metadata_2025_split huggingface dataset:
EPCO-11. BIASES IN HEALTHY BRAIN SINGLE-CELL DATASETS: FINDING THE BEST COMPARISON FOR GLIOBLASTOMA STUDIES
Authors: Carcanholo Felipe , Simões Renan , Santos Emerson , Malta Tathiane 
Published: November 2024

Abstract
Tumor cells undergo metabolic alterations that support their proliferation and simultaneously modify immune populations, resulting in an immunosuppressive tumor microenvironment. This study aims to compare the metabolic expression of immune cells within the tumor microenvironment to those in a healthy environment. However, there is a lack of single-cell transcriptomic data for normal brain cells, necessitating an analysis of different methods to compare immune cells in tumors with those in a healthy brain. To address this, we explored cell-to-cell metabolic variability using public single-cell RNA-seq (scRNA-seq) data of glioblastoma (n=338564), and from immune cells in the brains of post-mortem individuals (n=14177), fetuses (n=40000), and epilepsy patients (n=466), as well as from immune blood cells (n=329762). We employed ssGSEA to generate activity scores for 70 key metabolic pathways from KEGG for each individual cell. Known marker genes were utilized to identify normal cell populations (macrophages, T cells, oligodendrocytes) in datasets lacking cell type annotations. Our results revealed that blood cells exhibited high metabolic activity in macrophages but showed moderate activity in other cell types. Immune blood cells also demonstrated high oxidative phosphorylation (OXPHOS) but low fatty acid activity. Fetal brain cells displayed a lack of heterogeneity and showed high fatty acid activity with low OXPHOS activity. Brain cells from post-mortem individuals displayed heterogeneity but exhibited low metabolic activity across nearly all cells and pathways. Additionally, the hierarchy of pathway activity differed from the other datasets. Epilepsy data revealed a metabolic activity pattern similar to tumor-infiltrating immune cells, with comparable activity hierarchy and heterogeneity, although the sample size was limited for detailed comparison. Understanding the differences between types of datasets is important to choose the most appropriate comparison, allowing us to observe the metabolic alterations in immune cells caused by glioblastoma and to develop better immunotherapy strategies.
EPCO-20. MUTANT IDH INHIBITORS INDUCE LINEAGE DIFFERENTIATION IN IDH-MUTANT OLIGODENDROGLIOMA
Authors: Spitzer Avishay , Gritsch Simon , Nomura Masashi , Jucht Alexander , Fortin Jerome , Raviram Ramya , Weisman Hannah R , Castro L Nicolas Gonzalez , Druck Nicholas , Chanoch-Myers Rony , Lee John J Y , Mylvaganam Ravindra , Servis Rachel Lee , Fung Jeremy Man , Lee Christine K , Nagashima Hiroaki , Miller Julie J , Arrillaga-Romany Isabel , Louis David N , Wakimoto Hiroaki , Pisano Will , Wen Patrick Y , Mak Tak W , Sanson Marc , Touat Mehdi , Landau Dan A , Ligon Keith L , Cahill Daniel P , Suvà Mario L , Tirosh Itay 
Published: November 2024

Abstract
IDH-mutant gliomas exhibit variable responses to mutant-IDH inhibitor (IDHi) therapy; yet, the molecular mechanisms underlying these responses remain poorly understood. Here, we investigate the cellular underpinnings of response by leveraging single cell/nuclei RNA-sequencing of on-treatment IDH-mutant oligodendroglioma tumor samples resected from three patients who benefited clinically from treatment. We integrate these findings with single cell and bulk RNA-seq data from independent cohorts and experimental models. We find that IDHi treatment promotes robust differentiation towards the astrocytic lineage, accompanied by stem-like cell depletion and reduced proliferation. Notably, NOTCH1 mutations are associated with impaired astrocytic differentiation, potentially negatively impacting response to IDHi. This study unveils the differentiating effects of IDHi on oligodendroglioma cellular hierarchies and identifies a potential genetic marker for optimizing patient selection. Paper was recently published in Cancer Cell (10.1016/j.ccell.2024.03.008).
EPCO-25. TRANSPOSABLE ELEMENTS INDUCE ONCO-EXAPTATION EVENTS IN MALIGNANT ATRX-DEFICIENT GLIOMAS
Authors: Malgulwar Prit Benny , Johnson William E , Dharmaiah Sharvari , Machado Annette A , Barathi Bhargavi Brahmendra , Lu Lina , Singh Anand , Saw Ajay , Danussi Carla , Irwin David , Rai Kunal , Gao Ruli , Huse Jason T 
Published: November 2024

Abstract
Deficiency of ATRX (Alpha Thalassemia/Mental Retardation Syndrome X-linked), a core member of SWI/SNF family chromatin regulator is altered in diffuse gliomas and coexist with IDH1/2 mutations in adults and histone G34R/V alterations in pediatric populations. Loss of ATRX has been associated with global alterations in chromatin accessibility and epigenomics landscape, however, the detail mechanism behind the oncogenic transformation remains elusive. Integrating transcriptomics and in-depth positional epigenome analysis over repetitive DNA sequences in progenitor and tumors models of murine and human gliomas, we describe ATRX-deficient gliomas is fundamentally a disease of altered Transposable Elements (TE’s) and regulates neuronal differentiation and cell motility in gliomas, as validated pharmacologically and genetically. Epigenome assessment reported de-repression of LINE-1 (Long interspersed nuclear elements-1), specifically at Lamina-Associated Domains (LADs) with gain of enhancer histone marks, with association in chromatin loops/topology in ATRX-deficient glioma cells. Moreover, our single cell-RNA seq analysis focusing on TE’s and molecular docking studies in human glioma stem cells demonstrated LINE-1 regulates oncogenic alternative splicing events and neuro-developmental signatures. Expanding the analysis, we identified a subset of evolutionary young regulatory TE’s to play critical role in ATRX-deficient gliomas involved in neuronal differentiation phenotype. To summarize, our cross-species analysis establishes tangible links between ATRX deficiency and multiscale dysregulated 3D-epigenomic architecture that hijack regulates onco-exaptation programs in gliomas. We further discover that enhancer-like TE’s fine-tune the transcriptional program with important clinical implications, that could help in developing treatment options in malignant gliomas.
EPCO-26. LONGITUDINAL SINGLE-CELL ANALYSES IDENTIFY DRIVERS OF GENETIC, EPIGENOMIC, AND CELLULAR EVOLUTION IN IDH-MUTANT GLIOMA
Authors: Johnson Kevin , Spitzer Avishay , Varn Frederick , Nomura Masashi , Garofano Luciano , Chowdhury Tamrin , Anderson Kevin , D’Angelo Fulvio , Bussema Lillian , Gritsch Simon , Oh Young Taek , Moon Hyo-Eun , Paek Sun Ha , Bielle Franck , Laurenge Alice , Stefano Anna-Luisa Di , Mathon Bertrand , Picca Alberto , Sanson Marc , Lipsa Anuja , Hertel Frank , Zhao Zheng , Wang Qianghu , Jiang Tao , Hermes Beth , Sanai Nader , Golebiewska Anna , Niclou Simone , Huse Jason , Yung W K Alfred , Lasorella Anna , Suvà Mario , Iavarone Antonio , Tirosh Itay , Verhaak Roel 
Published: November 2024

Abstract
Despite standard treatment, IDH-mutant gliomas inevitably recur. Previous studies suggest that therapeutic resistance may result from a combination of intratumoral cellular heterogeneity, epigenetic evolution, and acquired genetic alterations. However, how these multilayered molecular features interact to influence the evolutionary paths of IDH-mutant gliomas remains incompletely understood. To chart this evolution, 75 glioma samples were longitudinally collected from 35 patients (n = 13 oligodendroglioma, n = 22 astrocytoma) and profiled using single nucleus RNA sequencing, ATAC sequencing, and bulk DNA sequencing. Analysis of 331,016 nuclei (snRNA) revealed eight tumor microenvironment cell types and five pan-IDH-mutant malignant cellular states. The malignant states were distributed along a cellular hierarchy of stem-like populations (neural progenitor cell-like, oligodendrocyte progenitor cell-like, and undifferentiated) and more differentiated populations (mesenchymal-like and astrocyte-like) with cycling cells enriched in the stem-like populations. Joint open chromatin accessibility data (snATAC) was available for a subset of malignant cells (71,088 nuclei) and state-specific differentially accessible peaks supported that these states are epigenetically encoded. Across both subtypes, higher tumor grade was associated with reduced malignant cell differentiation and increased cycling populations. Between the two time points, we identified that there was a longitudinal increase in the cycling, undifferentiated, and mesenchymal-like populations with a corresponding decrease in the astrocyte-like population. Longitudinal genetic analysis revealed that 19 of 35 tumors acquired at least one of the following genetic alterations: treatment-associated hypermutation, cell cycle alteration, or large changes in copy number alterations. Importantly, tumors that acquired these key genetic alterations demonstrated significant shifts towards reduced differentiation and increased cycling populations while those tumors that did not had more stable malignant profiles. Collectively, our results suggest a common cellular hierarchy across IDH-mutant gliomas with a shift towards reduced differentiation, increased cycling populations during disease progression that is driven by acquired genetic alterations.
EPCO-28. UNVEILING INVASIVE MECHANISMS OF GLIOBLASTOMA CELLS THROUGH MULTIMODAL SINGLE-CELL SEQUENCING
Authors: Wu Yiyan , Wu Benson , Ellenbogen Yosef , Mansouri Sheila , Kant Joan , Li Xuyao , Singh Olivia , Habibi Parnian , Kossinna Pathum , Rodriguez Sandra Ruth Lau , Gao Andrew , Zadeh Gelareh , Gaiti Federico 
Published: November 2024

Abstract
Glioblastoma (GBM) is the most common malignancy of the central nervous system, characterized by a dismal prognosis and inevitable recurrence despite aggressive standard-of-care therapy. Extensive colonization of the surrounding brain parenchyma precludes complete surgical resection, posing a significant therapeutic challenge. While well-studied in model systems, the molecular features and epigenetic regulators of invasive GBM cells remain under-explored directly in clinical samples. To address this gap, we performed a multimodal single-cell sequencing characterization of GBM tumor samples from anatomically distinct regions collected using MRI guidance. Our results demonstrate an enrichment of progenitor-like (neural- and oligodendrocyte-like) malignant states and neurons at the tumor margins. In contrast, differentiated-like states and myeloid cells were more abundant in the tumor core. Peri-tumoral progenitor-like malignant states expressed a unique neuronal signature associated with synaptic signaling, neurogenesis, and Notch signaling. Further, the expression of this neuronal signature was correlated with increased invasiveness. Analysis of matched primary-recurrent GBM patient cohorts revealed an expansion of this neuronal activity program, which was associated with worse overall survival. Motifs of proneural transcription factors implicated in neuronal lineage differentiation were also found to be differentially accessible in progenitor-like malignant states marked by the neuronal invasive signature, potentially contributing to the remodeling of cell states at the invasive margin. Further, cell-cell interaction analysis predicted greater communication between neurons and peri-tumoral progenitor-like malignant states, mediated predominantly through neurexin-neuroligin trans-synaptic signaling, facilitating synaptogenesis and the integration of tumor cells into neural circuits. Altogether, these findings suggest a model in which GBM invasive cells communicate with normal brain neurons in peri-tumoral regions, exploiting neurodevelopmental pathways to facilitate invasion and potentially seed recurrence. Our characterization of invasive GBM cells provides insight into the mechanisms of brain invasion and highlights potential therapeutic vulnerabilities of malignant invasive cells. Understanding these mechanisms opens new avenues for targeted therapies aimed at curtailing the invasive and adaptive capabilities of GBM.
EPCO-32. PROSPECTIVE GERMLINE SEQUENCING OF PATIENTS WITH GLIOMAS, GLIONEURONAL OR NEURONAL TUMORS
Authors: Lin Andrew , Nandakumar Subhiksha , Mehine Miika , Kemel Yelena , Mandelker Diana , Karajannis Matthias , Sait Sameer Farouk , Gavrilovic Igor , Pentsova Elena , Schaff Lauren , Stone Jacqueline , Nolan Craig , Boire Adrienne , Grommes Christian , Santomasso Bianca , Diamond Eli , Wilcox Jessica , Piotrowski Anna , Kaley Thomas , Young Robert , Imber Brandon , Brennan Cameron , Moss Nelson , Tabar Viviane , Rosenblum Marc , Bale Tejus , DeAngelis Lisa , Mellinghoff Ingo , Berger Michael , Stadler Zsofia , Schultz Nicholas 
Published: November 2024

Abstract

BACKGROUND
Several genetic syndromes have been linked to the development of central nervous system (CNS) tumors; however, the prevalence and clinical significance of germline pathogenic variants in this population remain unclear.

METHODS
2,367 patients with a glioma, glioneuronal, or neuronal tumor (WHO grade 1-4) ages 0.2-93.8 years underwent tumor and matched normal sequencing with MSK-IMPACT. Germline variants and copy-number variants affecting 90 well-established cancer predisposing genes were analyzed; the presence of biallelic inactivation was determined using FACETS.

RESULTS
Eleven percent harbored germline pathogenic mutations in high-, moderate-, and low-penetrance genes. High- and moderate-penetrance genes included CHEK2 (n=32, 1.3%), BRCA2 (n=10; 0.4%), TP53 (n=9; 0.4%), NF1 (n=6; 0.2%), and mismatch repair (MMR) genes (n=20, 0.9%). Low-penetrance genes included monoallelic MUTYH (n=42; 1.8%) and the APC I1307K variant (n=28, 1.2%). Biallelic inactivation was found in all tumors from patients with germline TP53 and NF1 mutations, with lower rates in tumors from patients with a germline MMR alteration (53%). Biallelic inactivation was rare among patients with a germline mutation in homologous recombination pathway (HRD) genes (27%), and in the aforementioned low-penetrance APC (15.3%) and MUTYH (10.2%) variants. All patients with biallelic inactivation of an MMR gene were hypermutated. Biallelic inactivation of MMR genes and TP53 were observed in IDH-mutant astrocytoma and IDH-WT gliomas, while biallelic inactivation of NF1 and somatic IDH1/2 mutation were mutually exclusive. When examining the age at diagnosis among patients with germline alterations, tumors with biallelic inactivation were diagnosed at a younger age compared to tumors with monoallelic alterations (37.7 vs 51.6 years, p=0.002).

CONCLUSION
Clinical germline sequencing identifies a germline mutation in a high proportion of patients with CNS tumors. Biallelic inactivation was most commonly identified in tumors from patients with germline TP53 or NF1 mutations and were less common in patients with a germline MMR alteration.
EPCO-39. MULTIOMIC AND CLINICAL ANALYSIS OF MULTIPLY RECURRENT MENINGIOMAS REVEALS RISK FACTORS, UNDERLYING BIOLOGY, AND INSIGHTS INTO EVOLUTION
Authors: Pugazenthi Sangami , Patel Bhuvic , English Collin , McGeehan Kyle , McCornack Colin , Mok David , Anzaldua-Campos Markus , Nouri Shervin Hosseingholi , Yano Hiroko , Klisch Tiemo , Harmanci Akdes , Patel Akash , Kim Albert 
Published: November 2024

Abstract
Although meningiomas, the most common primary brain tumor, are often effectively treated with surgery and radiation, an important subset of meningiomas behave aggressively and are characterized by treatment resistance and multiple recurrences. Prior work has yet to specifically study multiply recurrent meningiomas (MRMs), including whether they are molecularly distinct from non-recurrent meningiomas (NRMs) at initial diagnosis and whether the molecular features of MRMs evolve with subsequent recurrences. The objective of this study was to identify clinical and molecular features associated with MRMs and determine if longitudinal molecular changes occur in paired MRM samples. In this dual institution study, clinical parameters were collected for 1,186 meningioma patients. After propensity score matching, 31 MRMs were matched to 84 NRMs. Molecular sequencing including whole exome sequencing (n=44), EPIC bead chip methylation array (n=43), and RNA-sequencing (n=66) were performed. On multivariable binomial logistic regression, MRMs were significantly associated with male sex (P=0.012), subtotal resection (P=0.001), higher number of meningiomas on presentation (P=0.017), and histopathological sheeting (P=0.002). Multiomic analysis of primary meningiomas revealed MRMs exhibit greater global copy number alternations (CNA) (P=0.0113) and increased DNA methylation (P=0.0161). Integrated methylation profiling and RNA-sequencing identified candidate driver genes of MRMs. Among these genes, we demonstrate that knockdown of EDNRB, a locus with higher promoter methylation and decreased gene expression in MRMs, leads to increased meningioma cell proliferation, suggesting a tumor suppressor function. CNAs, subclonal evolution, and methylation profiles of MRMs did not significantly change from primary tumor to recurrence, even after radiation treatment, suggesting MRMs are molecularly aggressive from initial diagnosis. Overall, we identify several novel clinical and molecular risk factors associated with MRMs. MRMs harbor unique molecular features on presentation, which do not appear to change during evolution and after treatments, which has implications for development of biomarkers and therapeutic agents for these aggressive tumors.
EPCO-46. SPATIAL TRANSCRIPTOMICS ANALYSIS OF GLIOBLASTOMA REVEALS THREE DISTINCT REGIONAL PROGRAMS OF T-CELL INFILTRATION
Authors: Wang William L , Chen Yiyun , Mo Chiu Ho , Good Zinaida , Sotillo Elena , Mackall Crystal L 
Published: November 2024

Abstract
Glioblastoma (GBM) resists T-cell killing by developing an immunosuppressive tumor microenvironment. Transcriptomic technologies, such as single-cell RNA sequencing (scRNA-seq), have characterized immune cell phenotypes and putative cell-cell interactions in GBM. However, the tumor microenvironment surrounding T-cells is still not well characterized, motivating the need for spatial analyses. Two publicly-available GBM spatial transcriptomic datasets were analyzed, with one discovery cohort (N=18) and one validation cohort (N=12). Using cell deconvolution with a scRNA-seq reference dataset, areas of high T-cell infiltration were identified and clustered into regional programs based on transcriptomic signatures. To highlight which ligand-receptor interactions are most prominent in these regional programs, a novel spatial enrichment algorithm was applied. The impact of spatially enriched ligand-receptor interactions on overall survival was evaluated using bulk RNA-sequencing data from The Cancer Genome Atlas (TCGA). Three distinct T-cell regional programs (transitional, immunosuppressive, and infiltrative) were found across both the discovery and validation cohort. The infiltrative T-cell regional program resided on the periphery of the tumor with a neural progenitor phenotype (NRGN, VSNL1, SLC17A7). In contrast, the immunosuppressive T-cell region had a higher proportion of malignant cells and a greater mesenchymal tumor signature (ANXA1, CD44, complement pathway). Furthermore, ligand-receptor enrichment analysis showed that the immunosuppressive region had higher tumor and tumor-associated macrophage (TAM) signaling towards CD4 T-cells along the collagen, midkine, and osteopontin pathways. TCGA survival analysis showed that higher collagen and midkine signals were associated with worse overall and disease free survival. Our results identified three distinct regional programs in GBM and highlight a set of signaling pathways associated with T-cell suppression. Our findings suggest that the disruption of these pathways could reduce immunosuppressive signals from tumor and TAMs to T-cells. Subsequent validation with spatial proteomics data in glioblastoma is ongoing.
EPCO-52. INVESTIGATING EGFR CONTAINING EXTRACHROMOSOMAL DNA DYNAMICS IN GBM
Authors: Zhao Bo , Cho Chae Yun , Ye Lingqun , Akdemir Kadir 
Published: November 2024

Abstract
The epidermal growth factor receptor (EGFR) is the most frequently altered gene in GBM, with EGFR amplification occurring in approximately 40% of all glioblastomas, half of which exhibit the EGFRvIII deletion variant. EGFR amplification often occurs through the formation of extrachromosomal DNA (ecDNA), where EGFR typically co-amplifies with upstream enhancers. ecDNA is commonly found in a wide range of cancer types, particularly Glioblastoma (GBM), but is rare in normal cells. The uneven segregation of ecDNAs results in oncogene copy number variations in subclones, producing highly heterogeneous tumor cells and providing a selective advantage to cancer cells in response to changing environmental conditions. Meanwhile, ecDNAs keep evolving through fusion, deletion, or integration of new fragments, resulting in alterations to their structure and sequence. Some ecDNAs can reintegrate into chromosomal DNA, disrupting the integrity of chromosomal genes or rewiring chromosomal gene expression via ecDNA resident enhancers. However, the dynamics of EGFR-containing ecDNA (ecEGFR) have not been thoroughly investigated across primary and recurrent GBM. In this study, we profiled 47 glioblastoma specimens (both primary and recurrent) using a multi-omic approach, including WGS, RNA-seq, and ATAC-seq, identifying ecDNA in 14 samples, 10 of which carried EGFR. EGFR deletions were observed in 4 ecEGFR-positive samples, with 2 being the vIII variant. Interestingly, EGFR deletions predominantly occurred in the primary GBM. These ecEGFR samples exhibited significantly high EGFR copy number, gene expression, and chromatin accessibility. Additionally, there were samples with ecEGFR in the primary tumor but not in the recurrent tumor, or ecEGFR appears only in recurrent tumor. One patient showed an ecEGFR deletion in the primary tumor, while the recurrent tumor exhibited wild type ecEGFR.
EPCO-56. INHERITED PREDISPOSITION TO ADULT GLIOMAS: OVERVIEW OF THE CURRENT CLINICAL PRACTICE AND FUTURE RECOMMENDATIONS
Authors: Patel Saumya , Balay Lara , Maga Tara , Maraka Stefania 
Published: November 2024

Abstract
Gliomas are the most common adult primary malignant brain tumor and occur in ages 45-65 years old. While most gliomas are sporadic in nature, a small portion is related to cancer predisposition syndromes (CPS), such as Li Fraumeni or Lynch syndromes. Given that 5–10% of patients will have a family history of glioma, there is currently an effort in identifying germline variants associated with gliomas. Additionally, with the advent of somatic tumor testing, it is possible to pick up suspected germline mutations in genes that cause CPS. Our experience includes a 32-year-old patient diagnosed with astrocytoma IDH mutant, Grade 2 who was found to have a BRAC1 mutation on somatic brain tumor testing. 10 years later, the patient was found with metastatic ovarian cancer and a germline BRAC1 mutation. Early identification of CPS can lead to early detection or prevention of malignancies in affected patients and relatives. However, gliomas are rarely the primary or presenting tumor of CPS, and guidance of CPS screening for these individuals is still lacking. Due to the increasing interest and importance of CPS testing in patients with glioma we have developed a collaborative protocol between our neuro-oncologists and genetic counselors aso that patients have access to germline testing. We propose that all patients with glioma at age of 50 or under, patients with strong family history of cancer including gliomas, and patients with suspected germline mutations based on tumoral somatic mutational profile should be referred to the genetic counselors. Our preliminary findings include 8 patients who completed germline genetic testing. Of these, one patient tested positive for a BRCA1 pathogenic variant. We are planning to continue to collect this data to better inform clinical practice.