bluuebunny/crossref_metadata_2025_split huggingface dataset:
EPCO-02. GERMLINE SINGLE NUCLEOTIDE POLYMORPHISM rs55705857 AT 8q24 INTERACTS WITH SOMATIC IDH1 MUTATION TO ENHANCE HUMAN GLIOMA FORMATION
Authors: Drucker Kristen , Yanchus Connor , Kollmeyer Thomas , Ali Asma , Paul Decker , Kosel Matthew , Panda Arijit , Caron Alissa , Giannini Caterina , Burns Terence , Abyzov Alexej , Wang Liguo , Wrensch Margaret , Wiencke John , Eckel-Passow Jeanette , Lachance Daniel , Schramek Daniel , Jenkins Robert 
Published: November 2021

Abstract

BACKGROUND
Determination of the causation of germline single nucleotide polymorphisms (SNPs) located in non-coding regions of the genome is challenging. The genomic region of 8q24 has been identified as important in many kinds of cancer, linked to a topologically associated domain (TAD) encompassing MYC; this TAD contains a GWAS SNP (rs55705857) associated with IDH-mutant glioma.

METHODS
Germline genotyping data from 622 IDH-mutant glioma and 668 controls were used to fine map the rs55705857 locus by detailed haplotype analysis. Chromatin immunoprecipitation sequencing (ChIP-seq) of histone markers H3K4me1, H3K4me3, H3K27ac and H3K36me3 was performed on normal brain samples (n=8) and human glioma samples (n=11 IDH-wt and 52 IDH-mut). RNAseq from 9 normal and 83 brain tumors (n=26 IDH-wt and 55 IDH-mut) were used to assess differential gene expression.

RESULTS
Fine-mapping identified rs55705857 SNP as the most likely causative allele (OR=8.69; p&lt;0.001) within 8q24 for the development of IDH-mutant glioma. At rs55705857, both H3K27ac and H3K4me1 in IDH-mutant vs IDH-wt tumors were increased 3.05- and 1.58-fold, respectively (DiffBind; p=5.81×10-7 and p=2.31×10-3). ChromHMM analysis of the marks indicated that promoter and enhancer functions were significantly increased, and the activity broadened at rs55705857 in IDH-mut gliomas compared to IDH-wt tumors and normal brain samples. This enhancement correlated with significant increased MYC expression in IDH-mut gliomas (p=3.1×10-13), as well as alterations of Myc signaling targets. Publicly available ATACseq, ChIPseq and long-range DNA interaction data demonstrated that the rs55705857 locus is open and interacts with the MYC promoter.

CONCLUSIONS
Fine-mapping of the 8q24 locus provided strong evidence that rs55705857 is the causative 8q24 locus associated with IDH-mut glioma. Functional experiments suggest that IDH mutation facilitates rs55705857 interaction with MYC to alter downstream MYC targets.
EPCO-06. AGE- AND REGION-SPECIFIC MULTI-OMIC CHARACTERIZATION OF H3-K27M MUTANT DIFFUSE MIDLINE GLIOMA
Authors: Liu Ilon , Li Jiang , Jeong Daeun , Hack Olivia A , Shaw McKenzie , Englinger Bernhard , Avihai Byron , Ernst Kati , Resnick Adam , Diaz Aaron , Jones David , Koschmann Carl , Kleinman Claudia , Jabado Nada , Cotter Jennifer , Ligon Keith , Alexandrescu Sanda , Yung W K Alfred , Czech Thomas , Arrillaga-Romany Isabel , Gojo Johannes , Slavc Irene , Monje Michelle , Filbin Mariella G 
Published: November 2021

Abstract
Diffuse midline gliomas driven by lysine27-to-methionine mutations in histone 3 (H3-K27M DMGs) are among the most fatal brain tumors. Molecular studies including single cell RNA-sequencing (scRNA-seq) of pediatric and predominantly pontine H3-K27M DMGs have shown that the H3-K27M oncohistone keeps glioma cells locked in a stem-like oligodendrocyte precursor cell (OPC) state that is capable of self-renewal and tumor-initiation. However, a comprehensive dissection of the cellular architecture of H3-K27M DMGs across different midline regions and age groups is required to better understand the cell-intrinsic and contextual regulation of H3-K27M DMG cell identities. In particular, the more recently described group of adult H3-K27M DMGs remains understudied. Here, we have collected and characterized 45 H3-K27M mutant patient tumors, spanning pontine (n=26), thalamic (n=17), and spinal (n=2) locations. Median age at surgery was 12 (2-68) years, encompassing 21 early childhood (0-10 years), 12 adolescent (11-20 years), and 12 adult (≥ 21 years) tumors. The majority of samples were obtained pre-treatment (n=28), as opposed to post-treatment or at autopsy (n=17). We profiled all 45 tumors by single cell/single nucleus RNA-seq and selected tumors were further characterized by the single cell assay for transposase-accessible chromatin (scATAC-seq). Our integrated analyses highlight the predominance of transcriptionally and epigenetically defined OPC-like tumor cells as the main cell population of H3-K27M DMGs across all age groups and locations. We further identify distinct age- and location-specific OPC-like cell subpopulations. Comparison of pediatric and adult tumors further demonstrates a significant increase of mesenchymal cell states in adult H3-K27M DMGs, which we link to differences in glioma-associated immune cell compartments between age groups. Together, this study sheds light on the effects of age- and region-dependent microenvironments in shaping cellular identities in H3-K27M DMGs.
EPCO-09. LONGITUDINAL ANALYSIS OF DIFFUSE GLIOMA REVEALS CELL STATE DYNAMICS AT RECURRENCE ASSOCIATED WITH CHANGES IN GENETICS AND THE MICROENVIRONMENT
Authors: Varn Frederick , Johnson Kevin , Wade Taylor , Malta Tathiane , Sabedot Thais , Barthel Floris , Kim Hoon , Ahmed Nazia , Datta Indrani , Barnholtz-Sloan Jill , Bakas Spyridon , D'Angelo Fulvio , Gan Hui , Garofano Luciano , Huse Jason , Khasraw Mustafa , Kocakavuk Emre , Migliozzi Simona , Ormond David , Paek Sun Ha , Van Meir Erwin , Walenkamp Annemiek , Watts Colin , Weller Michael , Weiss Tobias , Wesseling Pieter , Stead Lucy , Poisson Laila , Noushmehr Houtan , Iavarone Antonio , Verhaak Roel 
Published: November 2021

Abstract
Diffuse glioma is characterized by a poor prognosis and a universal resistance to therapy, though the evolutionary processes behind this resistance remain unclear. The Glioma Longitudinal Analysis (GLASS) Consortium has previously demonstrated that therapy-induced selective pressures shape the genetic evolution of glioma in a stochastic manner. However, single-cell studies have revealed that malignant glioma cells are highly plastic and transition their cell state in response to diverse challenges, including changes in the microenvironment and the administration of standard-of-care therapy. To interrogate the factors driving therapy resistance in diffuse glioma, we collected and analyzed RNA- and/or DNA-sequencing data from temporally separated tumor pairs of over 300 adult patients with IDH-wild-type or IDH-mutant glioma. In a subset of these tumor pairs, we complemented these data with single-nucleus RNAseq and multiplex imaging mass cytometry at each time point. Recurrent tumors exhibited diverse changes that were attributable to changes in histological features, somatic alterations, and microenvironment interactions. IDH-wild-type tumors overall were more invasive at recurrence and exhibited increased expression of neuronal signaling programs that reflected a possible role for neuronal interactions in promoting glioma progression. In contrast, recurrent IDH-mutant tumors exhibited a significant increase in proliferative expression programs that correlated with discrete genetic changes. Hypermutation and acquired CDKN2A homozygous deletions associated with an increase in proliferating stem-like malignant cells at recurrence in both glioma subtypes, reflecting active tumor expansion. A transition to the mesenchymal phenotype was associated with the presence of a specific myeloid cell state defined by unique ligand-receptor interactions with malignant cells, providing opportunities to target this transition through therapy. Collectively, our results uncover recurrence-associated changes in genetics and the microenvironment that can be targeted to shape disease progression following initial diagnosis.
EPCO-27. SINGLE-CELL ANALYSIS OF ETMR PATIENT SAMPLES LINKS TRANSCRIPTIONAL PHENOTYPES TO GENETIC DRIVER ALTERATIONS AND INFORMS NOVEL THERAPEUTIC STRATEGIES
Authors: Hovestadt Volker , Beck Alexander , Lambo Sander , Shaw McKenzie , Hack Olivia A , Englinger Bernhard , Gojo Johannes , Madlener Sibylle , Alexandrescu Sanda , Pfister Stefan , Kool Marcel , Filbin Mariella G 
Published: November 2021

Abstract
Embryonal tumor with multilayered rosettes (ETMR) is a malignant brain tumor that typically occurs in children under the age of three. Most patients die within two years of diagnosis, and more effective, targeted therapies are urgently needed. To better characterize the oncogenic mechanisms of key driver alterations and identify novel therapeutic targets, we studied the cellular heterogeneity of ETMR using single-cell RNA sequencing. Analyses conducted on &gt;3,000 high-quality cells collected from ten primary and relapse specimens revealed a common cellular hierarchy across all tumors: A highly proliferative neural stem cell-like population (SOX2+) gives rise to intermediate progenitors (ASCL1+) and more differentiated neuron-like cells (STMN2/4+). These malignant populations closely match histological patterns of ETMR (i.e. rosettes, neuropil), as observed by immunofluorescence microscopy. Comparison to single-cell datasets from human embryos indicates resemblance to cell populations of the developing brain, but also reveals key ETMR-specific differences, including expression of the chromosome 19 miRNA cluster (C19MC, the presumed genetic driver of most ETMRs), which is restricted to the stem cell-like population. We next investigated if targeting C19MC is a viable strategy to disrupt the cellular hierarchy of ETMR. Silencing with antisense oligonucleotides shows pronounced reduction of cell line growth for a specific subset of the 46 members of C19MC. These miRNAs share seed sequences with evolutionary conserved miRNAs that have been shown to regulate pluripotency and self-renewal of embryonic stem cells. We hypothesize that select C19MC members play similar roles in ETMR and represent bona fide targets for therapeutic targeting using antisense technology.
EPCO-28. SINGLE-CELL RNA SEQUENCING IDENTIFIES INTRATUMORAL HETEROGENEITY AND ACTIVATION OF PRO-INVASIVE PATHWAYS IN NON-FUNCTIONING PITUITARY ADENOMAS
Authors: Jain Saket , Wang Elaina , Babikir Husam , Shamardani Karin , Diaz Aaron , Aghi Manish K 
Published: November 2021

Abstract
Pituitary adenomas (PA) are one of the most common primary brain tumors and comprise 15% of brain neoplasms. Most PAs are histologically benign but can cause significant morbidity. The genetic profile of PAs is poorly understood. We used single-cell RNA sequencing using the 10X genomic platform to investigate cellular heterogeneity in twelve non-functioning pituitary adenoma samples from nine patients including site-specific (core vs edge) samples from three patients. Our analysis identified discrete clusters of cells associated with activation of specific functional pathways including lipid metabolism, angiogenic, and antigen presentation and processing pathways regardless of location within the tumor. MALT1, a lncRNA associated with increased proliferation and metastasis was ubiquitously expressed amongst these samples. Analysis of the core vs edge samples showed two specific clusters with activated invasion-promoting pathways including PI3k/AKT signaling, Wnt signaling (Wnt6 and FZD4), and epithelial-mesenchymal transition (TGFB1, SMAD1, ZEB1, and SNAI2) in the edge of the tumors. The activated Wnt signaling cascade drove a proinflammatory tumor microenvironment induced by the expression of IL-1, IL-17, and Toll-like receptors (TLR6 and TLR7/8) resulting in suppression of Tregs. Copy number variation analysis using the CONICS-CNV algorithm highlighted distinct chromosomal alterations within our samples that led to insight into clonal variations within each tumor with loss of chromosome 2 an early event in tumorigenesis and gain/loss of chromosome 19 as late events. Mapping the copy number variation analysis with the somatic variant analysis using the Vartrix algorithm identified novel driver mutations within these tumors. These findings help define the molecular fingerprint of pituitary adenomas and provide insights which could be utilized for better management of these tumors.
BIOM-07. QUANTITATIVE MGMT PROMOTER METHYLATION INDEX INDICATES A NON-LINEAR PROGNOSTIC EFFECT IN GLIOBLASTOMA, SUGGESTING THAT USE OF OPTIMAL CUTOFF POINTS MAY BE CLINICALLY DISADVANTAGEOUS
Authors: Gibson David , Ravi Akshay , Almaraz Eduardo Rodriguez , Chang Susan , Oberheim-Bush Nancy Ann , Taylor Jennie , Clarke Jennifer , Solomon David , Scheffler Aaron , Witte John , Lambing Hannah , Berger Mitchel , Okada Hideho , Chehab Farid , Butowski Nicholas 
Published: November 2021

Abstract

BACKGROUND
Epigenetic inhibition of the O6-methylguanine-DNA-methyltransferase (MGMT) gene has emerged as a clinically relevant prognostic marker in glioblastoma (GBM). Methylation of the MGMT promoter has been shown to increase chemotherapy efficacy. While traditionally reported as a binary marker, recent methodological advancements have led to quantitative approaches that measure methylation, providing clearer insights into methylation’s functional relationship with survival.

METHODS
A CLIA assay and bisulfite sequencing was utilized to develop a quantitative, 17-point MGMT promoter methylation index derived from the number of methylated CpG sites. Retrospective review of 240 newly diagnosed GBM patients was performed in order to discern how risk for mortality transforms as promoter methylation increases. Non-linearities were captured by fitting splines to Cox proportional hazard models, plotting smoothed residuals, and creating survival plots. Covariates included age, KPS, IDH1 mutation, and extent of resection.

RESULTS
Median follow-up time and progression free survival were 16 and 9 months, respectively. 176 subjects experienced death. A one-unit increase in CpG methylation on a scale of 1-17 resulted in a 4% reduction in hazard (95% CI 0.93–0.99, P&lt; 0.005). Moreover, GBM patients with low-levels of methylation (1-6 CpG sites) fared markedly worse (HR=1.62, 95% CI 1.03-2.54, P&lt; 0.036) than individuals who were unmethylated (reference group). Subjects with medium-levels of methylation (7-12 CpG sites) had the greatest reduction in hazard (HR=0.48, 95% CI 0.29-0.80, P&lt; 0.004), followed by individuals in the highest methylation tertile (HR=0.62, 95% CI 0.40-0.97, P&lt; 0.035).

CONCLUSION
This novel approach offers greater bisulfite conversion efficiency when compared to alternative methods, reducing the likelihood of false positives. Analysis of the resulting methylation index scores demonstrates a non-linear relationship between MGMT methylation and survival, suggesting conformation of the marker’s protective effect. These findings challenge the current understanding of MGMT’s functional form and underline why implementing an “optimal cutoff point” may be disadvantageous.
BIOM-13. DEVELOPMENT AND VALIDATION OF AN LC-MS/MS ASSAY TO MEASURE GLUTATHIONE, GLUTATHIONE DISULFIDE, CYSTEINE, AND CYSTINE IN HUMAN BRAIN AND HUMAN GLIOBLASTOMA
Authors: Liu Miao , Margaryan Tigran , Sanai Nader , Tovmasyan Artak 
Published: November 2021

Abstract

BACKGROUND
Oxidative stress is implicated in many pathological conditions. Herein, we report on our development and validation of a sensitive and rapid LC-MS/MS method for the determination of oxidative stress biomarkers glutathione (GSH), glutathione disulfide (GSSG), cysteine (Cys) and cystine (CySS) in human brain and glioblastoma tissue.

METHODS
Freshly-acquired human glioblastoma tissue was homogenized with N-ethylmaleimide solution to prevent thiol oxidation. Analytes were then extracted from homogenate samples by protein precipitation with 2% sulfosalicylic acid (SSA). Stable isotope-labeled analytes were used as internal standards. Independent calibration curves for thiols and disulfides were prepared in analytical solutions. Three levels of quality controls were prepared in human brain homogenate. The detection was performed on Sciex QTRAP 6500+ mass spectrometer in positive electrospray ionization mode.

RESULTS
Linear regression model was used to cover a concentration ranging 0.4-100 µmol/L for GSSG/CySS and 1-400 µmol/L for GSH/Cys. Chromatographic separation was optimized on Intrada Amino Acid column with total run time of 5 min using gradient elution. For all analytes the maximum coefficient of variation for intra- and inter-day precision was 11.4% and the accuracy was within 80.9-113.7% in analytical solution and matrix. The analytes were stable in brain homogenate for 1 hour and 3 hours at room temperature (RT) and 4 °C, respectively. Stability at -80°C was demonstrated for at least 35 days in human brain homogenate. Stability of stock and working solutions was demonstrated for at least 4 hours (RT) and 25 days (-20°C).

CONCLUSIONS
A bioanalytical method to quantify GSH, GSSG, Cys, and CySS is successfully developed and validated. The method is currently applied to measure thiols and related disulfides in human glioblastoma tissue undergoing 5-aminolevulinic acid sonodynamic therapy (NCT04559685).
BIOM-25. ADULT GANGLIOGLIOMA: A 4-PATIENT CASE SERIES WITH MOLECULAR PROFILING
Authors: Shen Erica , Shams Alireza , Song Xianyuan , Daher Ahmad 
Published: November 2021

Abstract
Accounting for only 2% of all primary brain tumors, ganglioglioma is a rare neoplasm that most frequently arises in the temporal lobe in children or young adults. Ganglioglioma is the most common epilepsy-associated neoplasm and is typically composed of dysplastic ganglion cells and neoplastic glial cells. Recent publications have revealed that the activating p.V600E mutation in the BRAF oncogene is seen in 20-60% of gangliogliomas. However, the genetic landscape of ganglioglioma requires further elucidation, especially in gangliogliomas that arise from outside of the temporal lobe in adults. In this case series, we describe the genetic mutations of adult gangliogliomas originating outside of the temporal lobe in four patients aged 21 to 56 years. Targeted next generation sequencing via the MSK-IMPACT panel covering ~ 500 actionable mutations was used in all cases. None of the cases had the BRAF p.V600E mutation. Case 1 describes a left cerebellar cystic ganglioglioma with NTRK2 fusion mutation (NTRK2-DST) and a tumor mutation burden (TMB) = 0 mutation/megabase (mut/Mb). Case 2 describes a right parietal lobe cystic ganglioglioma with a BRAF-CLEC2Linversion mutation and TMB = 0 mut/Mb. Case 3 describes a cerebellar vermis ganglioglioma with a BRAF fusion mutation (BRAF-KIAA1549) and TMB = 0 mut/Mb. The last case describes a right cerebellopontine angle ganglioglioma with a MAP2K1 splicing mutation and TMB = 0.9 mut/Mb. To the best of our knowledge, the mutations identified in this case series have not been previously described in adult brain gangliogliomas. The BRAF-KIAA1549 fusion mutation has been found in spinal cord gangliogliomas, but not in brain gangliogliomas. MAP2K1 splicing mutations, unlike MAP2K1 in-frame deletions, have not been found in gangliogliomas. Finally, a different NTRK2 mutation (NTRK2-TLE4) has been described in a pediatric ganglioglioma, but NTRK2-DST has not been found in any adult gangliogliomas.
BIOM-28. SERIAL PLASMA AND CSF CELL-FREE TUMOR DNA (cf-tDNA) TRACKING IN DIFFUSE MIDLINE GLIOMA PATIENTS UNDERGOING TREATMENT WITH ONC201
Authors: Cantor Evan , Wierzbicki Kyle , Tarapore Rohinton , Ravi Karthik , Wadden Jack , Babilla Clarissa , Thomas Chase , Cartaxo Rodrigo , Yadav Vivek Anand , Ravindran Ramya , Bruzek Amy K , Cummings Jessica , Kawakibi Abed Rahman , Ji Sunjong , Paul Alyssa , Wolfe Ian , Leonard Marcia , Robertson Patricia , Franson Andrea , Mody Rajen , Garton Hugh , Odia Yazmin , Kline Cassie , Vitanza Nicholas , Khatua Soumen , Allen Joshua , Mueller Sabine , Gardner Sharon , Koschmann Carl 
Published: November 2021

Abstract
Diffuse midline glioma (DMG) with H3K27M mutation is a lethal childhood brain cancer, with limited means of monitoring beyond serial MRI scans. We conducted a multi-site Phase 1 trial of the imipridone ONC201 for children with H3K27M-mutant glioma (NCT03416530). Patients on Arm D of the trial (n=24) underwent serial lumbar puncture (baseline, 2 and 6-months) for cell-free tumor DNA (cf-tDNA) analysis at time of MRI. Additionally, patients on all arms of the trial at the University of Michigan underwent serial plasma collection. We collected a total of 96 plasma-samples and 53 CSF-samples from 29 patients. We performed ddPCR analysis of cf-tDNA samples and compared variant allele fraction (VAF) to radiographic change (maximal tumor area on MRI). For our H3F3A-mutated (K27M) patients, cf-tDNA was positive in 53/62 plasma samples (sensitivity 85.4%) and 28/29 CSF samples (sensitivity 96.5%) and overall specificity of 100%. There was no direct correlation between percent-change in tumor-area and plasma (p=0.47) or CSF VAF (p=0.89), implying that VAF provided information supplemental to radiographic assessments. “Spikes” in plasma cf-tDNA VAF (increase of ≥25%) co-occurred with progression in 2/9 (22%) cases and preceded progression in 5/9 cases (55%) by an average of 1.22 months. In CSF, spikes preceded progression in 4/6 cases (66%) by an average of 1.8 months. Two patients had increases in tumor-area with no increase in plasma VAF; both were later confirmed as pseudo-progressors, suggesting additional potential utility of cf-tDNA VAF monitoring. A 14yo male with spinal cord glioma received concurrent bevacizumab with ONC201, which resulted in a decrease in tumor area but continued increase in plasma VAF, predicting radiologic progression at the next time. In summary, we present data which suggests monitoring serial CSF/plasma H3K27M tDNA is a promising clinical tool. Changes in cf-tDNAVAF over time appear to correlate with response, predict progression, and differentiate pseudo-progression.