bluuebunny/crossref_metadata_2025_split huggingface dataset:
DIPG-46. TARGETING THE METABOLIC ERK5-PFKFB3 AXIS IN PEDIATRIC DIFFUSE MIDLINE GLIOMA
Authors: Casillo Stephanie , Gatesman Taylor , Chilukuri Akanksha , Jane Esther , Premkumar Daniel , Barber Callie , Halbert Matthew , Cruz Andrea , Mullett Steven , Michel Joshua , Chang Yue-Fang , MichaelRaj Antony , Lieberman Frank , Felker James , Shiva Sruti , Abdullah Kalil , Zinn Pascal , Friedlander Robert , Abel Taylor , Venneti Sriram , Mack Stephen , Wendell Stacy , Pollack Ian , Agnihotri Sameer 
Published: June 2023

Abstract
Diffuse midline glioma is an aggressive brain tumor with a median age at diagnosis of 6.3 years and 5-year survival rate of 2.2%.The defining histone mutation H3K27M precludes docking of a protein responsible for epigenetic modification resulting in erroneously accessible chromatin and subsequent transcription of oncogenic pathways, including the RAS-MERK5-ERK5 signaling cascade. To determine which oncogenic processes are regulated by extracellular signal-regulated kinase 5 (ERK5), gene-set enrichment analysis of patient-derived datasets demonstrated gene networks involved in glycolysis to be enriched with ERK5 expression. In confirmation, loss of ERK5 via shRNA interference reduced cell proliferation and glycolysis in DIPG IV and SF8628 cells. Reintroduction of ERK5 wildtype (WT) into ERK5 knockdown lines rescued cell proliferation and glycolysis, while the addition of ERK5 kinase dead domain (KDD) only partially rescued these survival and metabolic defects. Targeted evaluation of glycolysis enzyme expression via qRT-PCR revealed a direct relationship between ERK5 and proglycolytic enzyme 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3). Mechanistically, ERK5 activation of PFKFB3 was mediated by activation of transcriptional factor myocyte enhancer factor 2A (MEF2A) as demonstrated by coimmunoprecipitation and luciferase promoter assays. Expression of PFKFB3 was elevated at both the mRNA and protein level in DMG patient-derived samples. Genetic knockdown of PFKFB3 via shRNA interference resulted in reduced proliferation and glycolysis in DIPG IV and SF8628 cells. Similarly, pharmacologic inhibition of PFKFB3 with small molecule inhibitor PFK-158 capitulated these results. This inhibitor demonstrated blood brain barrier penetrance in silico and extended survival of in vivo mouse models. Multitargeted drug therapy against both ERK5 and PFKFB3 produced a synergetic in vitro response with increased sensitivity of these cells to apoptosis compared to single treatment alone. In conclusion, these results support ERK5 regulation of glycolysis through the critical metabolic effector PFKFB3. Multitargeted drug therapy against this axis represents a therapeutic vulnerability in pediatric diffuse midline glioma.
DIPG-51. ABDOMINAL METASTASES OF DIFFUSE MIDLINE GLIOMA, H3K27-ALTERED WITH A VENTRICULOPERITONEAL SHUNT
Authors: Lazow Margot , Schieffer Kathleen , Palmer Joshua , Salloum Ralph , Widemeyer Michelle , Sribnick Eric , Kumar Shiva Senthil , Umaru Banlanjo , Drissi Rachid , Mardis Elaine , Boué Daniel , Fouladi Maryam , Thomas Diana 
Published: June 2023

Abstract
Extraneural metastases are extremely rare in diffuse midline glioma (DMG), H3K27-altered, despite a tendency toward aggressive local growth and potential neuroaxis dissemination at progression. Ventriculoperitoneal shunt (VPS) placement for cerebrospinal fluid diversion is commonly performed in patients with thalamic tumors and increasingly considered in brainstem disease with developing hydrocephalus. Herein, we present a case of DMG seeding the abdomen presumably via the VPS. This patient was diagnosed at 22 years of age with a bithalamic tumor with extension into the brainstem and cerebellum as well as non-contiguous periventricular spread to the bilateral frontal lobes, with associated hydrocephalus. The patient underwent VPS insertion and biopsy, with pathology consistent with DMG, H3K27-altered. Whole exome sequencing revealed H3-3A and TP53 alterations as well as PDGFRA amplification. Treatment consisted of upfront craniospinal photon irradiation, with subsequent reduction in thalamic disease burden. Adjuvant therapy with an oral PDGFRA inhibitor (avapritinib) was initiated, though discontinued after two weeks due to worsening neurocognitive deficits. The patient presented three months post-completion of radiation, five months from diagnosis, and six weeks following VPS revision with rapidly progressive abdominal distension and altered mental status, with imaging demonstrating omental caking and significant malignant ascites, resulting in shunt malfunction and increased intracranial pressure. With limited therapeutic options, the family decided not to pursue further intervention and the patient died five days later. Autopsy confirmed metastatic disease throughout omentum, in addition to diffuse leptomeningeal dissemination. Comprehensive molecular profiling of diagnostic and post-mortem thalamic and omental tissue is in process to assess temporal as well as spatial genomic heterogeneity, including of metastasis-mediated pathways. In summary, abdominal dissemination of DMG, H3K27-altered is uncommon, but possible, especially in the presence of a VPS. Further research is necessary to determine biologic mechanisms and predictors of extraneural metastases in this disease.
EPEN-04. DEVELOPMENT OF B7-H3-TARGETED CAR T CELL THERAPY FOR EPENDYMOMA
Authors: Mehta Sanya , Houke Haley , Ibanez Jorge , Emanus Erik , Mack Stephen , Krenciute Giedre 
Published: June 2023

Abstract
Targeted treatment options are desperately needed for ependymomas (EPN). Chimeric antigen receptor (CAR) T cells have immense potential to positively transform treatment outcomes; however, little is known regarding antitumor efficacy of CAR T cells against EPNs. We found that B7-H3 is highly expressed on pediatric brain tumor patient-derived orthotopic xenografts from EPN patients, making it a promising target for CAR-T cells. The project’s goal is to develop a safe, effective B7-H3 targeted CAR T cell immunotherapy for EPN. We generated second generation human B7-H3-CAR T cells and examined their anti-EPN cytolytic activity, expansion, and cytokine production. We found that B7-H3-CAR T cells had potent in vitro antitumor activity against all EPN cell lines tested (1425, ST1, CPITT, ST2, EPI, L6SJ). In repeated-stimulation assays, CAR T cells expanded 7-10 times, and up to one million-fold when co-cultured with EPN cells. Interestingly, while the EPN cells were found to secrete high levels of C-C motif chemokine ligand 2 (CCL2) which is consistent with published data, our Multiplex analysis of co-culture supernatants shows that CCL2 production increased up to 10-fold when EPN cells were co-cultured with CAR T cells. Finally, we demonstrate that B7-H3-CAR T cells had antitumor activity in a supratentorial-EPN xenograft mouse model (1425 cell line) without apparent toxicities, although 4 out of 12 tumors re-lapsed. Recurrent tumor analysis is underway. Together, our data shows establishment of a clinically relevant model to study resistance to CAR T cell therapy in EPN, and preliminary mechanistic insight into CAR T cell persistence and modulation through CCL2 signaling. Given that little is known about the function of CCL2 and its receptor, C-C chemokine receptor type 2 (CCR2) in EPNs or CAR T cells, future work will focus on defining these biological mechanisms, enabling us to develop improvements to B7-H3-CAR T cells for EPN.
METB-07. MINIMALLY-INVASIVE DIAGNOSIS OF DIFFUSE LEPTOMENINGEAL GLIONEURONAL TUMOR (DLGNT) USING CEREBROSPINAL FLUID CELL-FREE DNA SEQUENCING IN PEDIATRIC AND YOUNG ADULT PATIENTS
Authors: Larsen Alexandra Giantini , Hill Katherine , Hickman Richard , Bale Tejus , Holle Bridget , Mellinghoff Ingo , Dunkel Ira , Souweidane Mark , Greenfield Jeffrey , Sait Sameer Farouk , Miller Alexandra , Karajannis Matthias 
Published: June 2023

Abstract
Diffuse leptomeningeal glioneuronal tumor (DLGNT) presents with nodular leptomeningeal disease throughout the neuroaxis. Histology shows variable presence of a low density intraparenchymal lesion containing OLIG2-expressing neoplastic cells. Molecularly, DLGNT is characterized by the presence of KIAA1549::BRAF fusion and chromosome arm 1p deletion. Concomitant gain of chromosome arm 1q is associated with poor prognosis. Diagnosis by meningeal biopsy is often non-diagnostic and we hypothesized analysis of cell free DNA (cfDNA) from cerebrospinal fluid (CSF) could represent a superior diagnostic modality. We analyzed CSF samples from 7 patients, median age of 18 years, with DLGNT including matched germline DNA. MSK-IMPACT, a hybridization capture-based next-generation DNA sequencing panel, was utilized to analyze the DNA. Prior tissue biopsy was done in 5/7 patients. Of these, biopsy was non-diagnostic in 3 patients and positive for the KIAA1549:BRAF fusion in 2. Analysis of CSF cfDNA detected KIAA1549::BRAF fusion in 5/7 (71%) of patients, including all patients with prior non-diagnostic biopsy or tissue quantity insufficient for molecular testing. Analysis of CSF cfDNA copy number profiles revealed 1p-/1q+ in 6 patients and 1p- only in the remaining 1 patient, fully concordant with available prior tissue testing in all instances. In 2 patients with prior tissue-confirmed KIAA1549::BRAF fusion, the fusion was not detected in CSF cfDNA during/after treatment with a MEK inhibitor, but additional somatic alterations and DNA copy number profiles were similar to the prior biopsies. Additional somatic alterations were identified in ATRX, BCOR, RARA, STAT5A, SPOP, PPM1D, MED12, KMT2C, RECQL and SETD2, including likely oncogenic secondary driver alterations that have not been previously reported in DLGNT. Given the high molecular diagnostic yield and concordance with open biopsy, cfDNA CSF sequencing should be considered as a first line diagnostic maneuver for patients with MRI findings suspicious for DLGNT, followed by open biopsy only if CSF is non-diagnostic.
HGG-07. OVEREXPRESSION OF A PLAG-FAMILY TRANSCRIPTION FACTOR DURING MURINE BRAIN DEVELOPMENT CAUSES BRAIN TUMOR FORMATION
Authors: Vaillant Jan , Wittmann Andrea , Keck Michaela-Kristina , Rozowsky Jacob S , Filbin Mariella G , Pfister Stefan M , Jones David T W , Kutscher Lena M 
Published: June 2023

Abstract
Pediatric-type gliomas are the most common type of central nervous system tumors in children. They are highly diverse regarding their molecular entities and clinical associations, and they differ from their adult counterparts. A subset of pediatric-type high grade gliomas, including H3K27M mutated diffuse midline glioma, overexpresses the developmentally regulated transcription factor gene PLAG1. Two novel tumor types displaying amplifications or fusions with the related PLAGL1 and PLAGL2 genes have also recently been described (PMIDs 36437415, 34355256). How PLAG-family dysregulation drives tumor formation is currently unknown, however. We generated a novel mouse model that overexpresses hPLAG1 during brain development using a Cre-LoxP approach. Mice that overexpress hPLAG1 together with loss of trp53 in the Nestin-Cre lineage develop brain tumors by 3 months of age with 100 % penetrance. Intriguingly, tumors develop in different brain regions, with hotspots in the thalamus or inferior hypothalamus, the latter likely emerging from the brainstem. All mouse brains contain regions in the brainstem, thalamus and the prefrontal cortex with diffuse patterns of PLAG1+; KI67+ proliferating single cells. Using bulk-RNA sequencing, we identified several genes involved in embryonic brain development, which are still active in the tumors of adult mice. We hypothesize that continued overexpression of PLAG1 locks cells in an aberrant developmental state, making them susceptible for further oncogenic hits and ultimately leading to pediatric glioma formation. We are currently using single-nucleus RNA-seq to determine the developmental trajectories of single tumor cells to identify putative cells-of-origin and primary developmental pathways. Our work will elucidate the consequences of PLAG1 overexpression in pediatric brain tumors and its contribution to tumorigenesis. In the future, we hope to identify genetic dependencies of PLAG1 to generate novel treatment options.
HGG-14. RAS PATHWAY MEDIATED REGULATION OF HISTONE METHYLTRANSFERASE AND TUMOR SUPPRESSOR SETD2 IN PEDIATRIC HIGH GRADE GLIOMA
Authors: Liou Angela , Moran Deborah , Dougherty Jacquelyn , Harvey Kyra , Brosius Stephanie , Labella Kay , De Raedt Thomas 
Published: June 2023

Abstract

BACKGROUND
Pediatric High-Grade Gliomas (pHGG) are a significant cause of cancer-associated deaths in children. Hemispheric pHGG have frequent loss of the NF1 tumor suppressor (15%) causing aberrant RAS pathway activation. Intriguingly, loss of SETD2, a H3K36 methyltransferase, significantly co-occurs with loss of NF1. We found that nearly 80% SETD2-mutant pHGG have inactive NF1 (Pedcbio, 18/23 cases, 78%; odds-ratio 25; FDR adjusted P-value &lt;0.001), suggesting a functional link between SETD2 and RAS signaling. NF1 is a GTPase-activating protein that negatively regulates the RAS oncogenic pathway. SETD2 is an epigenetic regulator and tumor suppressor that tri-methylates H3K36 on active gene bodies during transcription. We found that RAS pathway activation regulates SETD2 function.

METHODS
First, we used mass spectrometry to identify phosphorylation sites on SETD2 and generated phospho-mimetics/mutants to determine if these sites alter H3K36me3 levels. Second, we assessed if modulating downstream RAS effector pathways with MEK and PI3K inhibitors altered the phosphorylation of identified sites using the PhosTag gel electrophoresis assay.

RESULTS
We found that T2441 phosphorylation was upregulated upon MEK inhibition and that expressing T2441D phospho-mimetic and T2441A phospho-mutants had a differential effect on global H3K36me3 levels, implicating T2441 as an important regulatory node. Additionally, we identified that Cyclin Dependent Kinases are likely responsible for T2441 phosphorylation and are now investigating how RAS regulates these kinases.

CONCLUSION
To our knowledge it is the first time a functional connection between RAS pathway activation and SETD2 function has been found. It remains to be seen how of if RAS pathway activation alters the tumor suppressor function of SETD2 in pHGG.
HGG-25. TARGETED DRUGS SYNERGIZE WITH DOUBLE ALKYLATOR THERAPY IN H3.3 G34R/V PEDIATRIC HIGH-GRADE GLIOMA
Authors: Haase Rebecca , Green Adam , Kilburn Lindsay , Filbin Mariella 
Published: June 2023

Abstract
Diffuse hemispheric H3 G34R/V mutant gliomas (G34R/V DHG) are lethal brain malignancies occurring in the cerebral hemisphere of adolescent patients with no effective treatment options. To date, standard care is adapted from adult treatment protocols which include maximal tumor resection, radiotherapy, and chemotherapy with temozolomide (TMZ). In a recent study led by the Children’s Oncology Group (COG, ACNS0423), the combination of the alkylators TMZ and lomustine (CCNU) showed improved event-free survival for pediatric high-grade glioma patients. Still, patient benefits remain limited and thus demand further improvement of treatment protocols. Therefore, the aim of this study is to identify FDA-approved blood-brain-barrier crossing compounds that specifically targeting known vulnerabilities of pediatric high-grade glioma that achieve synergistic treatment results when combined with the double alkylator approach. We conducted a compound screen in combination with TMZ and CCNU on G34R/V-tumor derived gliomaspheres to identify synergies based on cell viability assays in vitro. We identified three candidate agents - Fimepinostat, trametinib, and avapritinib, which inhibit different key oncogenic targets in pediatric high-grade glioma - Pi3K and histone-deacetylases, MAPKs, and PDGFRA, respectively. Our ongoing in vivo study elaborates the translatability of the identified combinatorial therapies in patient derived G34R/V DHG orthotopic xenograft models. We are comparing different study arms of single agent treatment and double as well as triple combinations of alkylators and the candidate agents. This study will serve to determine (i) treatment efficacy, as measured by tumor growth and survival, (ii) occurring side effects, especially alkylator-induced suppression of platelet counts, and (iii) molecular changes upon different treatments using single cell transcriptomics and immunohistochemistry. In summary, we present valuable pre-clinical data supporting the combination of alkylators with targeted agents as a novel treatment strategy in G34R/V DHG, which might be translated into a promising strategy of a new comparative study based on COG trial ACNS0423.
HGG-26. GENOMIC AND IMMUNE ANALYSIS OF PRIMARY REPLICATION-REPAIR DEFICIENT (RRD) GLIOMAS REVEALS THREE SUBGROUPS WITH DISTINCT DRIVERS AND RESPONSE TO IMMUNOTHERAPY: AN IRRDC REPORT
Authors: Fernandez Nicholas R , Das Anirban , Levine Adrian , Negm Logine , Nobre Liana , Bianchi Vanessa , Stengs Lucie , Chung Jiil , Nunes Nuno M , Edwards Melissa , Bouffet Eric , Hawkins Cynthia , Tabori Uri 
Published: June 2023

Abstract

BACKGROUND
Replication-repair deficiency (RRD) caused by germline/somatic defects in mismatch repair (MMRD) and/or polymerase-proofreading genes (PPD) drives 5-10% of gliomas in children, adolescents, and young adults (CAYA). Although RRD-gliomas harbour high mutation-burden (TMB), the basis of their heterogenous biology, clinical behavior, and response to immune-checkpoint inhibitors (ICI) is unknown.

METHODS
We analyzed the genome (whole exome, low-coverage genome), methylome, transcriptome (bulk, single-nuclei), and the immune-microenvironment of RRD-gliomas in a large cohort of IRRDC patients and correlated these with clinical outcomes and response to ICI.

RESULTS
Gliomas in 202 CAYA-patients uniformly harbored hypermutation and genomic microsatellite-instability. Median TMB was 297-mutations/megabase, with frequent mutations in TP53 (90%), ATRX (85%), RAS/MAPK (80%) and IDH1/2 (15%). MMRD (but not PPD) mutational signatures contributed to the enrichment of driver mutations in POLE, IDH1, and TP53 while common pediatric mutations (K27M, G34R/V and BRAF;p.V600E) not driven by MMRD signatures were absent. Paired analyses suggested acquisition of novel variants driven by the mutational signatures that also impacted the immune microenvironment. Multi-omic analyses classified RRD-gliomas into three subgroups: RRD1 (MMRD+PPD; 60%), RRD2 (MMRD-only; 23%), and RRD3 (MMRD+IDH1/2; 16%). All RRD1-gliomas were glioblastomas with earlier age of onset, enrichment in CMMRD, arose at diverse locations including the posterior-fossa, classified in proximity to methylation-RTK1-subclass, exhibited balanced copy number profiles, harbored the highest TMB (median: 409-mutations/megabase), and immunogenic PPD signatures. Conversely, RRD3-gliomas were frequent in Lynch syndrome, presented at an older age, with predominant localization in the forebrain, more complex genomic instability, clustered in proximity to IDH1-gliomas, and harbored lower TMB (median: 33-mutations/megabase). RRD1-gliomas revealed the highest immune infiltrates with significantly improved median post-ICI survival of 52-months versus &lt;12-months for RRD2/3 (p&lt;0.0001).

CONCLUSIONS
The distinct genomic subgroups of RRD-gliomas can explain their diverse clinical outcomes, highlighting the need for developing subgroup-specific, immune-directed treatment approaches for these patients.
IMG-06. RADIOMIC-BASED RISK STRATIFICATION OF PEDIATRIC LOW-GRADE GLIOMA REVEALS DIFFERENTIALLY EXPRESSED MOLECULAR PROCESSES CONTRIBUTING TO VARIABLE PATIENT PROGNOSIS
Authors: Kazerooni Anahita Fathi , Kim Meen Chul , Kraya Adam , Mahtabfar Aria , Haldar Debanjan , Rathi Komal , Khalili Nastaran , Familiar Ariana , Ware Jeffrey B , Bagheri Sina , Anderson Hannah , Storm Phillip B , Resnick Adam C , Vossough Arastoo , Davatzikos Christos , Nabavizadeh Ali 
Published: June 2023

Abstract
In pediatric low-grade gliomas (pLGG), prognosis and responses to treatments are heterogeneous. This heterogeneity may be explained by the differences in molecular composition of the tumors of the same histology and the likely upstream alterations following administered radiation or systemic therapy. Integration of radiomics and clinical variables could generate a non-invasive biomarker that provides upfront prediction about patient’s risk of progression. We show that our proposed radiomic-based risk-stratification signature for pLGGs is associated with alterations in transcriptomic pathways. Standard multiparametric MRI sequences of 134 pLGG patients from Children’s Brain Tumor Network (CBTN) were retrospectively collected and 881 quantitative radiomic features were extracted. A multivariate Cox proportional hazard’s (Cox-PH) regression model was fitted on clinical (age, sex, tumor location, and extent of tumor resection) along with radiomic variables using 5-fold cross-validation, to predict patient’s risk of progression. The Cox-PH model showed excellent performance in prediction of PFS and patient’s risk scores, supported by the concordance index of 0.78. Radiogenomic analysis was performed to determine the transcriptomic pathways (1594 pathways, c2 MsigDb Reactome v2022.1) that contribute to the pLGG risk, predicted by the radiomic signature. ElasticNet regression was applied on the scores obtained by gene set enrichment analysis (GSEA) (in 70/134 subjects) to predict radiomic-based risk scores. Increased risk, corresponded to upregulation of DNA repair pathways, dysregulation of lipophagy, fatty acid beta oxidation, and vitamin D pathways, which are tumor-promoting. BRAFfusion signaling inversely correlated with risk, consistent with known favorable prognosis of KIAA1549-BRAF fused pLGGs. Upregulation in immune related pathways and Toll-Like Receptor (TLR) signaling was associated with lower risk. This study elucidates the synergistic dynamics between the biological processes that promote the risk of progression and radiomic-based risk-stratification signature. The proposed biomarker may be used to encourage targeted therapies in patients with increased predicted risk.