bluuebunny/crossref_metadata_2025_split huggingface dataset:
STEM-05. SHARED ONCOGENIC MUTATIONS IN THE SUBVENTRICULAR ZONE AND GLIOBLASTOMA: GENETIC AND CLINICAL IMPLICATIONS
Authors: Lee Joo Ho , Kim Hyun Jung , Yoo Jihwan , Shim Jin Kyoung , Kim Eui-Hyun , Chang Jong Hee , Park Jung Won , Li Xue , Lee Jeong Ho , Kang Seok-Gu 
Published: November 2024

Abstract
The subventricular zone (SVZ), a neurogenic niche in the adult brain, is suspected to harbor neural stem cells (NSCs) that can acquire cancer-driving mutations and give rise to glioblastoma (GBM). Nevertheless, the corroborating genetic data are restricted to a small patient cohort. This study aimed to validate the presence of low-level oncogenic mutations in the SVZ of GBM patients and assess their clinical implications.. We performed deep hybrid capture sequencing of 15 GBM-associated genes and TERT promoter amplicon sequencing on triple-matched samples from the SVZ, primary tumor, and blood of 60 GBM patients. Approximately 62% of patients (40 of 61 patients) showed shared oncogenic mutations between the tumor and tumor-free SVZ, including TERT promoter, PTEN, and TP53. While shared mutations did not affect overall clinical profiles, TERT promoter mutations were significantly enriched in the shared group (94.9% of patients, p ≤ 0.001). RNA sequencing revealed upregulation of proliferation-related genes in tumors with shared SVZ mutations, and these patients had worse progression-free and overall survival (p = 0.038 and 0.031, respectively). This study provides the largest-scale genetic validation of the SVZ as a potential cellular origin of GBM, highlighting the distinct genetic and clinical features associated with shared oncogenic mutations. These findings warrant further investigation into the role of SVZ mutations in GBM genesis and their potential as therapeutic targets.
STEM-08. LIPID SATURATION HOMEOSTASIS PRESENTS A TARGETABLE VULNERABILITY IN MYC-AMPLIFIED GROUP 3 MEDULLOBLASTOMA
Authors: Custers Stefan 
Published: November 2024

Abstract
Medulloblastoma (MB) is the most common malignant pediatric brain cancer, originating from the cerebellum. Despite advancements in standard of care (SoC), MB remains fatal for 30% of patients. Tumor relapse, spinal metastasis and treatment resistance are the most prevalent in MYC-driven Group 3 MB (G3-MB). Patients surviving SoC are faced with life-long neurocognitive and neurodevelopmental deficits. These issues highlight the urgent need for improved treatment modalities. Reprogramming of cellular lipid metabolism is an emerging hallmark of cancer and may yield novel cancer-specific therapeutic options. Here we explore the lipidome of both G3-MB and its proposed cell of origin, human neural stem cells (hNSCs) by comparing untargeted lipidomics using Liquid-Chromatography-Mass Spectrometry (LC-MS). Comparative analyses revealed a differential abundance of distinct lipid species in G3-MB, with an overall reduced saturation level of fatty acids (FAs). These findings implicate the de novo lipid synthesis (DNL) pathway. We identified the enzymes involved in DNL to be essential for MB survival in our genome-wide CRISPR KO screen. Additionally, mRNA expression of the DNL enzymes increases at relapse in our SoC-adapted murine patient-derived xenograft (PDX) model. Pharmacological and genetic targeting of the DNL enzyme Stearoyl-CoA Desaturase 1 (SCD1) selectively targets G3-MB. Furthermore, small molecule treatment of SCD1 demonstrates efficacy against G3-MB PDX models in vivo. We identified SCD expression as a prognostic marker in Group 3 and 4 MB patients and identified possible pathways for MB treatment. Overall, these findings indicate that SCD1 is a potent target for the treatment of G3-MB.
STEM-09. A GROUP 3 MEDULLOBLASTOMA STEM CELL PROGRAM IS MAINTAINED BY OTX2-MEDIATED ALTERNATIVE SPLICING
Authors: Saulnier Olivier , Zagozewski Jamie , Liang Lisa , Hendrikse Liam , Layug Paul , Gordon Victor , Aldinger Kimberly , Haldipur Parthiv , Borlase Stephanie , Coudière-Morrison Ludivine , Cai Ting , Martell Emma , Gonzales Naomi , Palidwor Gareth , Porter Christopher , Richard Stephane , Sharif Tanveer , Millen Kathleen , Doble Brad , Taylor Michael , Werbowetski-Ogilvie Tamra 
Published: November 2024

Abstract
Group 3 medulloblastomas (MBs) are highly metastatic pediatric brain tumors with limited therapeutic options. The transcription factor orthodenticle homeobox 2 (OTX2) is a known driver and molecular hallmark of Group 3 MB. We previously demonstrated that OTX2 regulates Group 3 MB cell fate decisions that favor self-renewal or stemness and the retention of a primitive cerebellar rhombic lip signature. Here, we show that OTX2-regulated alternative splicing (AS) is a major driver of Group 3 MB progression. Surprisingly, the functional role of post-transcriptional events, such as AS, in Group 3 MB development has never been explored. This gene regulatory layer represents an untapped source of therapeutic targets. Using a combination of TurboID, RNA-sequencing and single cell RNA-seq, we characterized the Group 3 primary MB AS landscape with a focus on genes that control MB stem cells. OTX2 associates with the multimeric Large Assembly of Splicing Regulators (LASR) complex through protein-protein interactions and can directly or indirectly bind RNA. Together, OTX2 and LASR complex proteins oversee a pro-tumorigenic splicing program that is mirrored in the human cerebellar rhombic lip origins and is associated with Group 3γ MBs that exhibit the worst prognosis. Among our core set of OTX2-regulated spliced genes, periphilin 1 (PPHLN1) is expressed in the most primitive cerebellar rhombic lip stem cells, and mimicking PPHLN1 AS by pre-treating cells with a splice blocking antisense morpholino reduces tumor growth while increasing survival in vivo. We propose that OTX2-mediated AS is a key determinant of cell fate decisions favoring the maintenance of the stem cell state. Targeting transcription factors like OTX2 is notoriously difficult; thus, our work offers fresh opportunities to exploit MB-specific AS events therapeutically to abrogate tumor progression.
STEM-17. THE UROKINASE RECEPTOR AS A NOVEL IMMUNOTHERAPEUTIC TARGET IN BRAIN CANCERS
Authors: Maich William , Shaikh Muhammad Vaseem , Puri Anish , Mobilio Daniel , Ananda Alisha , Venugopal Chitra , Rossotti Martin , Mikolajewicz Nick , Chokshi Chirayu , Salim Sabra , Savage Neil , Kislinger Thomas , Henry Kevin , Singh Sheila 
Published: November 2024

Abstract
Glioblastoma (GBM) is the common malignant brain tumor in adults, accounting for approximately 15% of all CNS tumors, and 48.6% of malignant brain tumors, with a median survival of approximately 15 months. GBM is characterized by extensive inter- and intra-tumoral heterogeneity and an extremely immunosuppressive tumor microenvironment. Following Standard-of-Care surgical resection and chemoradiotherapy, patients inevitably relapse, at which point few therapeutic avenues exist, owing in part due to a lack of clinically relevant targets. Data from our target identification pipeline shows the urokinase plasminogen activator receptor (uPAR) is significantly upregulated at recurrence on putative GBM brain tumor initiating cells (BTICs), which are believed to drive de novo tumor formation, recurrence, and therapeutic resistance. uPAR plays an important role in the plasminogen activation system, and in the context of cancer, has been implicated in numerous pro-tumorigenic processes such as invasion, proliferation, and therapy resistance. We used CRISPR-Cas9 to genetically delete uPAR from our in-house patient-derived GBM cell lines, and found that knockout of uPAR in recurrent GBM cells significantly reduces various functional characteristics of BTICs in vitro. In our patient-derived mouse model of GBM, we found that knockout of uPAR significantly increases survival, and decreases tumor burden. Further, we generated novel anti-uPAR single domain antibodies (sdAbs) which specifically and efficiently bind uPAR at low nanomolar concentrations in vitro. We developed anti-uPAR CAR Ts using the binder sequence of the sdAbs, which showed potent cytotoxicity in vitro, and drastically reduced tumor burden and extended survival in vivo. Further, we identified uPAR as being highly expressed on brain metastases from multiple origins (lung-to-brain, melanoma-to-brain, etc.), and demonstrate that anti-uPAR CAR Ts effectively kill brain metastases in vitro, and significantly extend survival using in vivo brain metastasis models. From this work we believe uPAR to be a clinically relevant target in both recurrent GBM and brain metastases, and investigation into therapeutic strategies targeting uPAR-positive brain cancers should be explored further.
CSIG-06. BRAF GERMLINE MUTATIONS AND PREDISPOSITION TO BRAIN TUMORS
Authors: Abdelmoula Nouha Bouayed , Abdelmoula Balkiss , Smoui Walid , Trigui Khaled , Louati Rim 
Published: November 2024

Abstract
We report on two BRAF-mutated children and discuss genetic counseling regarding predisposition to brain tumors. Molecular analysis of the PTPN11 gene and twelve other genes in the RAS-MAPK pathway was carried out in a series of 54 dysmorphic patients with suspected Rasopathies. Two heterozygous BRAF germline mutations were detected in two 7-year-old girls, at exon 6 and 12 respectively. Both patients had the distinctive facial appearance of cardio-facio-cutaneous syndrome (CFC) type 1. Patients with CFC are rare and characterized by phenotypic and genetic heterogeneity, with overlaps with Noonan syndrome. While nearly 75% of CFC cases have germline BRAF mutations, the frequency of BRAF mutations is inferior in Noonan and LEOPARD syndromes. BRAF mutations in CFC are often found in the cysteine-rich domain of the BRAF protein (exons 6, 11, 12, 13, 14, 15 and 16). Conversely, somatic BRAF mutations are more common, grouped into classes according to their kinase activity level, and have been associated with a variety of malignancies, including pediatric and adult brain tumors (low- and high-grade gliomas, glioblastomas and low-grade astrocytomas). BRAF V600E is the most common somatic mutation, but other mutations are currently being identified. Many of these mutations are within the kinase domain, while others are located across the gene and have less well-understood functional consequences. Germinal V600E mutation has been described once, in a CFC patient having a severe phenotype. Nevertheless, it seems that patients with germline BRAF mutations and gliomas have not yet been reported. Given the rarity of CFC, it is unclear whether there is a high risk of brain tumors. Further molecular studies will enable us to refine recommendations for the prevention, monitoring and care of people with germline BRAF mutations, given the success of BRAF inhibitors in certain cancers, with encouraging results in gliomas.
CSIG-11. TUMOR-CELL INTRINSIC CALCIUM OSCILLATIONS DRIVE BRAIN METASTASIS
Authors: Hebach Nils R , Olshausen Niels , Schlag Joana S , Bang Severin , Kraft Theresa , Mayer Chanté , Linke Zoe , Westphal Dana , Seifert Michael , Timmer Jens , Wick Wolfgang , Winkler Frank , Karreman Matthia A 
Published: November 2024

Abstract
Calcium is a ubiquitous secondary messenger which regulates many cellular processes. Recently, we have shown that a subpopulation of glioblastoma cells can intrinsically generate calcium oscillations which drive proliferative signalling. Here, we set out to investigate whether such cancer cell autonomous oscillations are also present in secondary brain tumours. Employing longitudinal intravital two-photon microscopy in awake mice, we demonstrate that brain metastases of melanoma and lung cancer exhibit spontaneous calcium oscillations in vivo along the metastatic cascade. These oscillations are already observed during intravascular arrest and in in vitro monocultures, suggesting a tumour-cell intrinsic origin. We developed a fully automatic image analysis pipeline to accurately quantify calcium oscillations both in vivo and in vitro. Mathematical modelling of cytosolic and ER calcium stores can predict stable calcium oscillations with similar frequency and peak characteristics as those experimentally observed. We discovered that these oscillations are tightly regulated by an interplay of calcium-induced calcium release and store-operated calcium entry (SOCE). Similarly, SOCE-related ion channels are enriched in tissue from human melanoma brain metastases compared to healthy brain controls and calcium activity can also be observed ex vivo in patient-derived brain metastasis resections. Finally, by correlating calcium activity and EdU incorporation we discovered that the presence of calcium oscillations predicts their proliferative potential in vitro. Importantly, by intravital two-photon microscopy we show that calcium activity correlates with metastasis size in preclinical models, suggesting a potential new therapeutic angle from which to target brain metastases.
CSIG-20. LIPOCALIN-2 DRIVES CELL-INTRINSIC FUNCTIONS IN GLIOBLASTOMA
Authors: Wang Sabrina , Timken J , Bishop Christopher , Deleyrolle Loic , Lathia Justin 
Published: November 2024

Abstract
Glioblastoma (GBM) continues to persist despite aggressive therapies, many of which are cytotoxic, and therapeutic failure is driven, in part, by the tumor cell’s ability to evade multiple forms of cell death, including ferroptosis. Lipocalin-2 (LCN2) functions to sequester iron and is considered an inflammatory marker through its function of limiting iron for bacterial usage; it has also been described to have pro and anti-tumorigenic effects depending on environmental iron status. In previously published studies in brain metastases, LCN2 promotes tumor cell proliferation by sequestering iron for cancer cells growth and survival, but the mechanisms of how LCN2 contributes to GBM progression is unknown. LCN2 expression in patient data revealed that high LCN2 expression is associated with reduced survival in GBM patients. A survey of mouse glioma models demonstrated a range of LCN2 secretion across models with KR158 and CPA being high and medium LCN2 secretors. LCN2 short hairpin knockdown of KR158 and CPA and decreased tumor cell viability and proliferation. Furthermore, when both KR158 and CPA control versus LCN2 knockdown cells were orthotopically implanted into mice, survival was increased. To understand if LCN2 impacted cell death dynamics, we investigated various cell death mechanisms and found that LCN2 knockdown increases tumor susceptibility to ferroptotic death by sensitizing cells to lipid peroxidation as well as reducing key anti-ferroptotic proteins GPX4 and xCT. To elucidate downstream mechanisms, we performed a phosphorylation array and discovered that phosphorylation of AXL, which has recently been linked to ferroptosis, is decreased with LCN2 knockdown. Moreover, AXL phosphorylation was localized to the nucleus, indicating that LCN2 knockdown attenuates AXL processing and downstream signaling. Overall, these findings indicate that LCN2 in GBM promotes tumor cell growth through AXL signaling and evasion of ferroptotic death, ultimately revealing a novel signaling network that can be leveraged for future therapeutic development.
CSIG-28. DISSECTING THE FUNCTIONAL IMPAIRMENT OF WILD-TYPE P53 IN HUMAN GLIOBLASTOMA
Authors: García-Montaño Leopoldo A , Muñoz Yamhilette Licón , Martinez Frank , Gallegos Kandee , Bowers Christian A , Kang Huining , Piccirillo Sara G M 
Published: November 2024

Abstract
Glioblastoma (GBM) is the most common malignant brain tumor in adults and is characterized by heterogeneous nature, invasive potential, and poor treatment response. Work from The Cancer Genome Atlas (TCGA) identified the tumor suppressor gene TP53 as a GBM driver. In contrast to several cancers, only ~35% of GBM cases carry mutations in TP53. Work from our lab showed that TP53 mutations emerge as ‘late’ events during GBM growth. Therefore, we hypothesized that wild-type (wt) p53 function is impaired in GBM. We used the TCGA data of 591 GBM patients to compare the clinical and molecular characteristics of wt and mutant TP53 patients. Gene expression data were analyzed to compare the correlation of TP53 with the expression of key p53 regulators and identify differentially expressed regulators between our groups. Furthermore, we evaluated the expression of p53 and selected regulators in patient-derived cells (PDCs) and control lines with known TP53 status. Our results show that wt TP53 patients live less than mutant patients and both groups respond similarly to treatment. No differences in age, sex, race, TP53 levels, and MGMT methylation status were observed between the groups. Gene expression analyses revealed that MAPK8, a p53 activator, negatively correlates with TP53 in the wt group. Moreover, ~21% of p53 regulators are differentially expressed between wt and mutant patients. Among these, we selected SIRT3, a p53 repressor, for experimental validation. We verified the p53 genetic status of PDCs through western blot and confirmed Sirt3 expression in wt and mutant TP53 PDCs. By CRISPR-Cas9-based genomic editing, we are currently evaluating the impact of SIRT3 knockout in GBM-PDC and patient-derived xenografts. We expect that SIRT3 knockout will restore p53 function in the wt group. In conclusion, our results suggest that p53 function is impaired in wt patients and provide insights into mechanisms affecting its activity.
CSIG-31. ANNEXIN A7 ALTERNATIVE SPLICING IN GLIOMA STEM CELLS FACILITATES THE DEGRADATION OF EGFR BY PHOSPHORYLATING TYROSINE RESIDUES EXCLUSIVE TO PROTEIN ISOFORM 1
Authors: Merati Aran , Nair Sindhu , Rajbhandari Rajani , Johnson Nicholas , Bredel Markus 
Published: November 2024

Abstract
The majority of GBMs express aberrant receptor tyrosine kinase (RTK) activity, which are canonically endocytosed and degraded. This process is disrupted in GBM to promote receptor recycling and resulting in hyperactive RTK signaling. Exon six of Annexin A7 (ANXA7) can be alternatively spliced to yield two distinct protein isoforms through its inclusion (I1) or exclusion (I2). We investigated ANXA7 isoform differences for epidermal growth factor receptor (EGFR) trafficking fates in PDX glioma stem cells (GSCs). We demonstrated that ANXA7 exon skipping supports the impaired endosomal trafficking of RTKs to favor recycling in GBM. JX14 GSCs were made to overexpress ANXA7-I1 with lentivirus to conduct immunofluorescence experiments measuring EGFR-ANXA7 colocalization (Pearson) with markers of the endocytic pathway for the early endosomes (EEA1), recycling endosomes (Rab4/Rab11), and lysosomes (LAMP1). EV and I1 cells were also incubated with siRNA targeting both ANXA7 isoforms to determine the impact of ANXA7 loss on endosomal EGFR trafficking. Interestingly, we found drastic differences in endosomal EGFR trafficking in I1 expressing cells. EGFR colocalized with EEA1 in both EV and I1 cells (R2=0.01336, P&gt;0.6275). In EV cells, EGFR was recycled via fast/slow recycling endosomes demonstrated by Rab4/Rab11 colocalization while I1 did not (R2=0.9267, P&lt;0.0001). Furthermore, I1 cells and not EV preferentially degraded EGFR as seen by LAMP1 colocalization (R2=0.9398, P&lt;0.0001). On the other hand, ANXA7 siRNA KD in I1 cells impaired EGFR sorting via a 40% increase (P&lt;0.0010) in total EGFR protein and a 40% reduction (P&lt;0.0001) in colocalization with EEA1 to prefer recycling again while EV cells were unaffected. I1 overexpression significantly reduced EGFR signaling through degradation, which is driven by the phosphorylation of I1-exclusive tyrosine residues in phosphomimetic (p&lt;0.01) and non-phosphorylatable protein mutants. I1-induced reduction in EGFR signaling suggests a new therapeutic vulnerability for GSCs by potentially reducing tumorigenicity or improving the efficacy of EGFR inhibition.
CSIG-34. TYROSINE PHOSPHORYLATION OF NON-CANONICAL NF-ΚB P52 PROMOTES GLIOMA GROWTH AND CHEMORESISTANCE IN GBM
Authors: Driscoll Riley , Wu Longtao , Bernal Giovanna , Dalmage Mahalia , Spurr Liam , Yamini Bakhtiar 
Published: November 2024

Abstract
p52 is a nuclear factor-κB (NF-κB) transcription factor that comprises the non-canonical NF-κB pathway. The impact and mechanism of non-canonical NF-κB signaling remains understudied in glioblastoma (GBM). Subunit post-translational modification is a known mechanism of NF-κB activation, however phosphorylation of p52 has never been demonstrated. Elevated tyrosine kinase activity is a known-feature of GBM and portends worse survival. Further, p52 has been shown to upregulate factors that likely promote invasion of GBM, thereby reducing survival. Activation of p52 may serve as a link between increased kinase activity and poor survival in GBM. p52-knockout cell lines were generated using the CRISPR-Cas9 system and assessed for growth and survival differences in vitro and in syngeneic murine glioma models. Public glioma datasets and an institutional cohort were surveyed to corroborate p52-dependent survival differences in GBM. To determine whether p52 is post-translationally modified, we performed tandem mass spectrometry on p52 in GBM cells. Interacting factors were also analyzed and tested for site-specific modification of p52 via co-IP studies. We then generated PTM-null, knock-in glioma cell lines using CRISPR-CAS9 editing to demonstrate the functional significance of p52 PTMs, which were assessed using assays of relative cell growth, stemness, chemosensitivity and survival. Finally, we performed unbiased analysis of the p52 PTM-dependent transcriptome via RNA-seq. p52 loss significantly reduced glioma growth rate in vitro and improves murine survival. Retrospective analysis of GBM in patients suggests that p52 levels are negatively prognostic. Mass spectrometry identified multiple p52 PTMs with tyrosine phosphorylation sites as top hits. Identified phospho-tyrosine residues specifically interact with SRC-family kinases. PTM site editing significantly reduces glioma cell growth and stemness while enhancing chemosensitivity, reflecting broad transcriptome level changes. In sum, p52 activity enhances GBM growth and chemoresistance, which is partially modulated by tyrosine-specific phosphorylation. p52 warrants investigation as a therapeutic target in GBM, possibly in combination with kinase inhibitors and standard alkylating chemotherapy.
CSIG-35. RAB3B DICTATES MTORC1/S6 SIGNALING IN CHORDOMA AND PREDICTS RESPONSE TO MTORC1-TARGETED THERAPY
Authors: Meng Tong 
Published: November 2024

Abstract
Chordoma is a rare mesenchymal malignancy, exhibiting a high tendency to postoperative recurrence and poor prognosis. To date, its tumorigenic regulatory mechanisms remain elusive leading to a lack of effective therapeutic targets and drug sensitivity indicators. Here, via transcriptome and proteome analyses, we unveiled RAB3B as the prominent oncogenic regulator in chordoma. Multidimensional tissue samples further indicated its overexpression and enhancer-associated high transcriptional activity in chordoma. Notably, RAB3B ablation attenuated the chordoma cell growth, stemness, migration and tumorigenicity in vivo and in vitro. Through determining the RAB3B-mediated program in chordoma, we identified that RAB3B was highly associated with PI3K-AKT-mTOR signaling and directly bound to S6. Mechanistically, RAB3B increased the S6K-mediated phosphorylation of S6 by physically interacting with dephosphorylase DUSP12, to block its dephosphorylating of p-S6 specifically at S235/236. Pharmacological targeting mTORC1 pathway via dactolisib and rapamycin dramatically impeded the RAB3B-induced chordoma cell oncogenesis, while RAB3B knockout desensitized mTORC1 inhibition. Clinically, RAB3B/p-S6 suggested a good prognostic value and response to mTORC1 inhibitors for chordoma patients. Altogether, this work uncovers RAB3B as a novel activator of mTORC1 pathway, and suggests the oncogenic and predictive roles of RAB3B/p-S6 in chordoma, indicating therapeutic potentials of mTORC1-targeted therapy in advanced chordoma patients with aberrant RAB3B/p-S6 hyperactivation.