bluuebunny/crossref_metadata_2025_split huggingface dataset:
MODL-25. ANALYSIS OF TUMOR CELL HETEROGENEITY IN A HUMAN NEURAL PROGENITOR-BASED MODEL OF GLIOBLASTOMA REVEALS NEURODEVELOPMENTAL PROGRAMS AND CYTOSKELETAL DYNAMICS
Authors: DeSouza Patrick , Qu Xuan , Ishahak Matthew , McCornack Colin , Annamalai Devi , Batista Luis , Wang Ting , Maher Christopher , Millman Jeffrey , Kim Albert 
Published: November 2022

Abstract
Glioblastoma (GBM) is the most common primary malignant neoplasm with poor survival despite treatment. Developing effective therapies remains challenging due to intratumoral heterogeneity, which drives therapeutic resistance and recurrence. To understand how genetic events alter the epigenome to enhance clonal fitness, we developed an isogenic human neural progenitor cell (NPC)-based model of the proneural (PRO) GBM subtype. We introduced TERT promoter (TERTp) C228T and gain-of-function TP53 R248Q mutations in H1 human embryonic stem cells. Wildtype (WT) cells, single TERTp, and double TERTp/TP53 mutants underwent differentiation to NPCs. Lentiviral transduction of double mutants with PDGFRA D842V resulted in triple mutant PRO NPCs. Bulk and single cell transcriptomics of our model system revealed hundreds of gene expression changes with increased mesoderm and human GBM mesenchymal (MES) subtype signatures in single TERTp and double TERTp/TP53 mutants, versus WT cells. TERTp mutation increased telomerase expression and activity, conferring proliferative advantage and immortalization in NPCs and astrocytes. Additionally, TERT expression was further increased in TERTp/TP53 mutants. Surprisingly, triple mutant PRO NPCs, versus WT, displayed &lt; 100 differentially expressed genes, associated with neurodevelopmental and dynamic cytoskeletal processes. Evolution analyses using gene counts signature and splicing dynamics revealed a developmental trajectory model from WT to additive mutants to PRO NPCs. Only triple mutant PRO NPCs formed tumors after intracranial injection in athymic nude mice, with mean survival of 100 days. Tumors presented histopathological features of GBM, and single cell transcriptomic analyses revealed evolution from immune-interacting to both neural progenitor- and neuronal-like subpopulations, with similar cell cycling signatures. Transcription factor genes related to WNT signaling and lineage commitment as well as glial and neuronal cytoskeletal genes exhibited epigenetic selection in vivo, signatures also observed in PRO NPCs in vitro. Our model thus provides opportunity for dissection of epigenetic and functional mechanisms underlying serial mutations during PRO tumor evolution.
MODL-32. IMPACT OF PRDM6 ON CHROMATIN ACCESSIBILITY, GENE EXPRESSION, AND MEDULLOBLASTOMA FORMATION
Authors: Schmidt Christin , Husain Sarah , Carlson Annika , Cohen Sarah , Westelman Samantha , Wang Linyu , Weiss William , Schwer Bjoern 
Published: November 2022

Abstract
Group 4 medulloblastoma (MB) is the most common medulloblastoma subgroup and shows high incidence of metastasis and late-onset relapse. Group 4 MBs lack a unifying oncogenic driver and treatment targets, despite extensive genomic characterization. Group 4 MBs are characterized by recurrent genetic alterations in chromatin modifiers, amplification of stemness genes, and putative enhancer "hijacking" events. A substantial fraction of Group 4 MBs are characterized by enhancer hijacking through tandem duplication of SNCAIP, resulting in high expression of PRDM6, a putative transcriptional repressor and histone methyltransferase. Some PRDM6-overexpressing MBs show additional mutations in chromatin regulators and high MYCN expression. We set out to elucidate the impact and oncogenic potential of sustained PRDM6 expression in early neural stem cell populations and patient-derived medulloblastoma cells. We find that PRDM6 expression in human iPSC-derived neuroepithelial stem cells (NESCs) results in tumor growth in mice, albeit at low penetrance. Moreover, we find that PRDM6 expression in MYCN-overexpressing NESCs does not further alter tumor aggressiveness or survival in vivo. Notably, PRDM6 overexpression in the patient-derived Group 3 MYC-amplified D283-Med cell line causes significantly increased aggressiveness of tumor growth and shorter survival in mice. At the cellular level, PRDM6 localizes to the nucleus, suggesting a role in gene expression regulation. Consistent with this notion, ATAC-seq and RNA-seq analysis of PRDM6-NESCs and PRDM6-D283-Med cells reveals major changes in chromatin accessibility and gene expression, including upregulation of integrin family members and genes related to focal adhesion and extracellular matrix-receptor interaction pathways. We conclude that PRDM6 promotes tumor growth in NESCs and may play a role in tumor maintenance through interactions with the extracellular matrix and tumor microenvironment.
MODL-34. DECELLULARIZATION OF HUMAN AUTOPSY BRAIN TISSUE TO GENERATE A 3D EXTRACELLULAR MATRIX FOR MEDULLOBLASTOMA AND ATYPICAL TERATOID/RHABDOID TUMOUR MODELLING
Authors: McCrorie Phoebe , Porter Harry , Vinohar Jeffy , Diksin Mohammed , Scurr David , Rahman Ruman 
Published: November 2022

Abstract

INTRODUCTION
Childhood medulloblastoma (MB) and atypical teratoid/rhabdoid tumours (AT/RT) are malignant brain tumours occurring in the posterior fossa, for which prognoses remains particularly poor for the MB Group 3 subtype characterised by amplification of the Myc oncogene and for AT/RT in general. Current in vitro models for these neoplasms rely on non-coated plastic, various hydrogels, or animal-derived extracellular matrix (ECM), which fail to recapitulate the physiological environment from which the cells are derived from.

METHODS
We have developed a method to decellularize ex vivo human brain tissue from different anatomical locations for the use in 3D in vitro models. Human cerebellar brain tissue was harvested from autopsy brain and sectioned into small cubes before bathing in a sodium dodecyl sulfate/phosphate-buffered saline mixture for several days, before washing and lyophilising.

RESULTS
The optimised method for generation of decellularized human brain ECM successfully removes nuclei as confirmed by histological staining and DNA quantification (DNA reduction of ≥ 60%). Orbitrap-Secondary Ion Mass Spectrometry analysis confirmed the retention of the ECM components laminin (C9H11N3O2Na+), fibronectin (C9H14N4O2Na+ and C20H33N7O5Na+) and collagen (C4H5N2O2+), with a reduction in cell membrane lipid components (glycerophosphocholine, C9H19NPO4+; phosphocholine, C5H15NPO4+; and choline, C5H14NO+) relative to control tissue, with a &lt; 2 ppm accuracy, which was further corroborated by glycosaminoglycan and collagen assays. Multiple molecular subtype-specific AT/RT and MB cell lines have been successfully grown on decellularized cerebellar-ECM/PEGDA hydrogel, showing no reduction in metabolic viability using PrestoBlue and Cell Titer Glo assays.

CONCLUSIONS
This methodology offers an innovative human-only high-throughput 3D drug screening model, whereby patient-derived MB or AT/RT cells are co-cultured with healthy human cerebellar astrocytes upon decellularized cerebellar ECM, which we term ‘Tumoursphere Matrices’.
MODL-35. A NOVEL PATIENT-DERIVED TUMOR ORGANOID MODEL IDENTIFIES HDACI PANOBINOSTAT AS HIGHLY EFFECTIVE DRUG IN MENINGIOMAS
Authors: Jungwirth Gerhard , Cao Junguo , Warta Rolf , Moustafa Mahmoud , Abdollahi Amir , Andreas Unterberg , Herold-Mende Christel 
Published: November 2022

Abstract
Tumor-organoids (TOs) are novel, complex three-dimensional ex vivo tissue cultures that under optimal conditions accurately reflect the genotype and phenotype of the original tissue with preserved cellular heterogeneity and morphology. They offer a new and exciting platform for studying cancer biology and directing personalized therapies. Our study aimed to establish TOs from meningioma (MGM) and explore drug responses of selected drugs.TOs were established by controlled reaggregation of freshly prepared single-cell suspension of MGM tissue samples in 384-well plates. We were capable of forming several hundred TO equal in size with a success rate of 98%. TOs consisted of largely viable cells, whereas dead cells were predominantly found outside of the organoid. H&E and IF stainings confirmed the successful establishment of dense tissue-like structures. On the molecular level, transcriptional and mutational profiles were conserved in TOs. To query the suitability and reliability of TOs for robust large-scale drug testing by employing nine highly potent compounds, derived from a drug screening performed on MGM cell lines. In total, standardized TOs from 60 patients were formed, including eight grade II and three grade III MGMs. The FDA-approved epigenetic drug panobinostat demonstrated high anti-meningioma effects in the majority of patients. The efficacy was mediated through inhibition of HDAC1/2 validated by siRNA-mediated knockdown experiments. In an orthotopic in vivo model, treatment with panobinostat extended the median survival of mice, however, the therapy was not curative. To further investigate HDACi resistance, transcriptomes of sensitive and resistant TOs were compared. In resistant tumors, a significant induction of the TGFβ pathway was observed.Taken together, we developed a protocol to generate standardized TO from MGM and preserved histological and molecular features of parental tumor. Drug screening of selected drugs in TOs from 60 patients identified HDACi panobinostat as effective drug through inhibition of HDAC1/2.
MODL-37. DISCERNIBLE INTERINDIVIDUAL PATTERNS OF GLOBAL EFFICIENCY DECLINE DURING THEORETICAL BRAIN SURGERY
Authors: Yueh-Hsin Lin , Dadario Nicholas , Crawford Lewis , Tanglay Onur , Dow Hsu-Kang , Young Isabella , Ahsan Syed Ali , Doyen Stephane , Sughrue Michael 
Published: November 2022

Abstract
Resection of infiltrating brain tumors, such as diffuse gliomas, generally involves resecting a portion of a lobe of the brain. While the concept of localized functions, and the risk to removing these areas, is well established in neurosurgical thinking, the potential that the overall global efficiency of the connectome could be disproportionately disturbed by an intervention in ways which are not immediately obvious have not been formally studied. The current article provides evidence that structural patterns exist in the impact resection of various lobes of the brain has, which also differs between subjects. We utilized diffusion tractography to create structural connectivity graphs from the brains of 80 healthy adults, and then performed every plausible brain surgery in every gross anatomic region of the cerebrum by deleting every possible combination of nodes in the graph which were adjacent to each other, and measured the drop in global efficiency (GE) at each nodal deletion. Not surprisingly, the deletion of some nodes was worse than others, such that in every lobe we studied in every subject, there were combinations of deletions which were worse for GE than removing a greater number of nodes in a different part of the brain. Interestingly, while the worst nodes differed between subjects, there were specific nodes which typically showed up as particularly detrimental regardless of which node was the worst in that person, but that there were patterns of so-called ―connectotype, which could determine which nodes were the worst. Progressive removal of a lobe of the brain leads to patterns of global efficiency decline which are reasonably predictable, but which are not the same between subjects. Given evidence that global efficiency relates to specific neuro-cognitive abilities, this provides a path towards reducing the cognitive footprint of brain surgery.
MODL-38. DEVELOPMENTAL EFFECTS OF MYBL1 ACTIVATION ON MURINE BRAIN AND GLIAL DEVELOPMENT
Authors: Azazmeh Narmen , Rouleau Cecile , Schoolcraft Kate , Jacob Etai , Malinowski Seth , Busanovich John P , Ramkissoon Lori , Kang Yun Jee , Ramkissoon Shakti , Pelton Kristine , Meng Alice , Jones Victor , Bronson Rodderick , Piccioni Federica , Kleinman Claudia , Bandopadhayay Pratiti , Ligon Keith , Beroukhim Rameen 
Published: November 2022

Abstract
Low-grade gliomas (LGG) represent 30% of pediatric brain tumors. Their cellular origins are unknown but are presumed to arise from subtle alterations of progenitor cell cycle regulation during brain development. Rearrangements activating MYB and MYBL1 have been identified as drivers of LGG in angiocentric glioma and diffuse astrocytomas, respectively, but the roles of these genes in the normal brain and the development of LGG are poorly understood. We first performed a developmental analysis of human and mouse Mybl1 expression from bulk and single-cell RNA-sequencing and identified exclusive expression of MYBL1 in neural stem and progenitor cells in the ganglionic eminence. We also found that MYBL1high cell transcriptomes are enriched in genes functionally involved in centromere and mitotic processes, strongly suggesting an association between MYBL1 expression and cellular proliferation states. We next hypothesized that C-terminal truncation may drive tumorigenesis through a direct increase in MYBL1 expression and cell proliferation. We developed a novel Cre-dependent knock-in mouse-model for human truncated MYBL1 expression and tested effects in oligodendroglial(Olig2-cre), astrocytic hGFAP-cre), and somatic(Ubiquitin-cre) cell types. In Ubq-cre:R26-MYBL1-tr mice there was expression and dysplasia in the salivary gland but no significant effects on brain development. In Olig2-Cre+/tg:R26-MYBL1-tr+/fl mice we observed higher susceptibility to motor seizures, early postnatal death without gross or microscopic abnormalities of brain morphology. In contrast, expression in stem cells and astrocytes of hGFAP-Cre+/tg:R26-MYBL1-tr+/flmice drove dramatic abnormalities in brain development and altered proliferation of progenitor and stem cells, but not glioma formation. Single-cell RNA sequencing of MYBL1-tr cells from mNSCs and brains of mice were also used to determine patterns of altered expression driven by MYBL1-tr. These results indicate that aberrant MYBL1 activation affects neural stem/progenitor cell and brain development through altered cell proliferation. Future targeting of the pathways identified may be therapeutically beneficial for patients with LGG driven by MYB-family oncogenes.
MODL-42. ESTABLISHMENT AND CHARACTERIZATION OF A NOVEL MOUSE SYNGENEIC GLIOBLASTOMA CELL LINE
Authors: Ye Juan , Wang Herui , Yang Chunzhang , Zhuang Zhengping 
Published: November 2022

Abstract
Glioblastoma (GBM) is the most common and the deadliest type of brain tumor affecting the adult central nervous system, carrying a median survival of 14 months. In vivo syngeneic animal models of GBM that recapitulate human GBM are urgently needed to decipher glioma biology and develop novel therapeutics, including immunotherapy. Here, we generated a syngeneic mouse glioblastoma stem cell line (RCAS-TVA-GSC01) through repeated in-vitro passages of stem-like tumor cells isolated from the RCAS-TVA glioma mouse model, which overexpresses PDGFA and carries a homozygous deletion of Ink4a-Arf locus. We characterized this new GSC line both in vitro and in vivo. This cell line expresses stemness markers CD133 and Nestin, and is able to differentiate into astrocyte, neuron, and oligodendrocyte-like cells in 2% FBS differentiating medium with markers GFAP, β-III tubulin, and O4, respectively. Transcriptomic and genomic profiling of RCAS-TVA-GSC01 cells revealed molecular heterogeneity comparable to human glioblastoma. When intracranially transplanted into wild-type C57BL/6 mice, this cell line formed high-grade gliomas with a median survival time of 56.5 days. The differentiation potential and molecular heterogeneity make RCAS-TVA GSC01 line a unique and useful tool for studying multiple aspects of glioblastoma pathogenesis and a promising platform for testing new GBM therapeutics study, especially for immunotherapy that requires a syngeneic mouse model.
STEM-04. CONNEXIN 43 DRIVES GLIOBLASTOMA CANCER STEM CELL PHENOTYPES THROUGH A WNK1 SIGNALING AXIS
Authors: Mulkearns-Hubert Erin , Hajdari Nicole , Lathia Justin 
Published: November 2022

Abstract
Tumors represent a highly dynamic system that relies on coordinated signaling to drive growth and adapt to selective pressures, including those induced by anti-cancer therapies. Given the need for tumors to engage in rapid and coordinated cell-cell communication, mechanisms including gap junction intracellular communication would be essential. Surprisingly, the connexin proteins that make up gap junctions have traditionally been considered tumor suppressors. This hypothesis was based on the loss of GJIC in cancer cells and, frequently, lower connexin expression in tumor cells compared to non-neoplastic tissue, including in glioblastoma (GBM), where connexin 43 (Cx43) was shown to be decreased. However, recent data suggest that this tumor-suppressive effect is context dependent and that connexins can function in a tumor-promoting role. We previously found that GBM cancer stem cells (CSCs) rely on gap junctional intercellular communication through connexin 46 (Cx46) for survival. We now identify a subset of GBM patient-derived xenograft (PDX) CSC models that also rely on Cx43 in addition to Cx46. This dependence on Cx43 is independent of Cx43 expression level. Knockdown of Cx43 using shRNA in these PDX models compromises their survival and self-renewal while inducing apoptosis. We leveraged a protein kinase phospho array to identify signaling changes associated with Cx43 knockdown and observed reductions in phosphorylation of WNK lysine-deficient protein kinase 1 (WNK1) in addition to ERK1/2, AKT, and p38MAPK. Immunoblotting validated that WNK1 phosphorylation is lost in the absence of Cx43. Treatment with a WNK1 inhibitor was toxic to CSCs. Current work is focused on interrogating the link among Cx43, WNK1 phosphorylation, and downstream signaling events and the therapeutic window of WNK1 inhibition. This work identifies a potential new signaling axis downstream of Cx43 in glioblastoma CSCs and further strengthens the hypothesis that connexins may act as both tumor suppressors and tumor drivers depending on context.
STEM-09. NEURAL STEM CELLS OF THE SUBVENTRICULAR ZONE EVOLVE INTO TUMORS AROUND THE RESECTION CAVITY AFTER SURGICAL RESECTION OF GLIOBLASTOMA VIA THE CXCL12/CXCR4 AXIS
Authors: Lee Joo Ho , Li Xue , Kim Hyun Jung , Lee Jeong Ho , Kang Seok-Gu , Wu Hong-Gyun 
Published: November 2022

Abstract
Neural stem cells (NSCs) harboring cancer-driving mutations in the subventricular zone (SVZ) have recently been reported as the cells of origin for human GBM. However, the pathological role of residual tumor-initiating NSCs (tNSCs) in the SVZ is undetermined for recurrent tumors after surgical removal of the primary tumor. After the primary resection of GBM, most recurrences still appear as local tumors around the resection cavity (RC). Accordingly, after surgical removal, we hypothesized that the tNSCs in SVZ might be the source for tumor construction at RC as local recurrence. Here, we examined the role of tNSCs in developing local tumors around RC using a mouse model and human tissues. First, cancer-driving mutations, including Trp53, Pten, and Egfr mutations, were introduced to NSCs in SVZ through in vivo electroporation, and the RC was generated in the remote cortex through surgical resection. We found that the NSCs harboring driver mutations migrated specifically to the RC through the aberrant growth of oligodendrocyte-precursor cells (OPCs). Furthermore, the SVZ-originated cells constituted the GBM around the RC at four weeks post-resection in 64% of mouse models. Second, the CXCR4/CXCL12 axis was upregulated through migration from the SVZ and tumor formation at the RC compared with primary tumors without the RC. Blockage of the CXCR4/CXCL12 axis decreased the number of immigrating OPC lineage and switched the differentiation of tNSC to astrocyte lineage. Furthermore, it improved survival rates in vivo. Third, we validated the independent clonal evolution of recurrent tumor from SVZ and the upregulation of CXCR4 axis in the recurrent tumor in a human patient. Taken together, our results reveal that the residual tNCSs harboring cancer-driving mutations in the SVZ can re-construct tumors after surgical resection of GBM. Blocking CXCL12/CXCR4 might be a potential treatment strategy for preventing recurrences after resection.
STEM-13. GLIOMA STEM CELLS MIMIC REGULATORY T CELL FUNCTION TO SUPPRESS THE IMMUNE RESPONSE AND PROMOTE TUMOR PROPAGATION
Authors: Johnson Amanda , Korleski Jack , Sall Sophie , Lopez-Bertoni Hernando , Laterra John 
Published: November 2022

Abstract
The advent of immunotherapy has transformed our approach to cancer treatment; however, it is virtually ineffective in glioblastoma (GBM), the most common & malignant brain cancer in adults, due in part to the inherently immunosuppressive tumor microenvironment (TME). Recent research has identified GBM cancer stem cells (GSCs) as key modulators of the TME, mainly through activation and recruitment of pro-tumor immune cells (e.g., TAMs, regulatory T-cells (Tregs), MDSCs). We identified a novel immunosuppressive phenotype whereby GSCs mimic Treg function via TGF-b type II receptor (TGFBR2) signaling to suppress the immune response with the potential to promote tumor propagation and resist immunotherapy. Preliminary experiments revealed that TGFBR2 drives GBM cells to a CD44-high, mesenchymal-like GSC state. Transgenic TGFBR2 was found to induce a Treg-like transcriptional profile defined by expression of canonical Treg effector genes (e.g., CD274, ENTPD1, NT5E, LGALS1, TGFB1, PDCD1LG2) in human GBM neurospheres. Single-cell RNA-Seq and bioinformatics analyses confirmed the presence of this TGFBR2-mediated transcriptional signature in both patient-derived GSCs and patient tumors. Notably, in silico deconvolution of bulk GBM sequencing predicts that high expression of a TGFBR2-induced transcriptome associates with an immunosuppressive TME characterized by high infiltration of pro-tumor immune cells (e.g., TAMs, neutrophils). To target this Treg-like, immunosuppressive GSC phenotype, we utilized ITD-1, a small molecule inhibitor of TGFBR2. In vitro ITD-1 treatment reduced GSC proliferation capacity and downregulated expression of CD44 and Treg effectors. In T-cell co-cultures, pre-treating GSCs with ITD-1 increased T-cell viability and T-cell-mediated tumor cell killing. Administration of ITD-1 in tumor-bearing, immunocompetent mice reduced tumor volume and increased anti-tumor immune cell infiltration relative to vehicle controls. In all, ITD-1-mediated TGFBR2 inhibition attenuates a unique, Treg-like immunosuppressive and potentially tumor-protective phenotype in GSCs. Future studies will combine ITD-1 and checkpoint inhibitor therapy in an effort to increase the efficacy of immunotherapy in GBM.