bluuebunny/crossref_metadata_2025 huggingface dataset:
BIOM-31. TARGETED DNA SEQUENCING OF RECURRENT GLIOBLASTOMA IDENTIFIES MOLECULAR FACTORS ASSOCIATED WITH CLINICAL OUTCOMES
Authors: Al-Adli Nadeem , Chen William , Young Jacob , Mirchia Kanish , Peeran Zain , Braunstein Steve , Perry Arie , de Groot John , Raleigh David , Costello Joseph , Berger Mitchel , Vasudevan Harish
Published: November 2023

PURPOSE

The goal of our project was to define the molecular features of recurrent glioblastoma (rGBM) and their prognostic significance.

METHODS

We retrospectively reviewed 121 consecutive glioblastoma patients who underwent re-resection at a single institution from 2016-2021. Primary treatment in all patients included maximal safe resection followed by radiotherapy with concurrent and adjuvant temozolomide (TMZ). Capture-based targeted DNA sequencing for point mutations and copy number analysis was performed on all re-resection samples. Overall (OS) and progression-free survival (PFS) from re-resection were analyzed with the Kaplan-Meier method and Cox regression analyses.

RESULTS

Median age at first recurrence was 57 years (range, 27-78) and follow up from re-resection was 11 months. Salvage therapy comprised re-irradiation (n = 52, 43%), temozolomide (TMZ) (n = 39, 32%), lomustine (n = 47, 39%), and/or bevacizumab (n = 22, 18%). Median OS and PFS were 12.1 and 4.4 months, respectively. Median tumor mutation burden (TMB) was 4 mut/Mb (range, 1-650), average unstable microsatellite sites (MSI) were 1.16% (range, 0-20), and MGMT was methylated in 71 (59%) patients. Longer OS was found after postoperative TMZ (19.1 vs. 9.1), KPS > 70 (13.6 vs. 11.1), and/or re-irradiation (12.6 vs. 8.4) (p< .05). On multivariate analysis, TMZ (HR 0.2, 0.079-0.48), re-irradiation (HR 0.27, 0.12-0.58), and mutations in PDGFRA (HR 0.24, 0.067-0.83) or PTEN (HR 2.3, 1.1-4.8) genes were associated with longer OS after re-resection (p< .05). Multivariate predictors of re-irradiation response included MSI >1.16 (HR 0.25, 0.084-0.75), and mutations in PDGFRA (HR 0.04, 0.005-0.33), TERT (HR 0.16, 0.033-0.78), or PIK3R1 (HR 0.18, 0.041-0.8) genes. In the 46 matched primary/recurrent tumors, 40 (87%%) had a change in a mutation and 9 (20%) demonstrated hypermutation, of which 4 (44%) had mismatch repair deficiencies.

CONCLUSIONS

Our study outlines the molecular factors associated with survival and response to re-irradiation in rGBM as well as, the molecular differences between primary/recurrent samples.
BIOM-42. LONGITUDINAL IMMUNE PROFILES OF LOWER GRADE ADULT GLIOMA PATIENTS
Authors: Guerra Geno , Francis Stephen , Molinaro Annette , Taylor Jennie , Warrier Gayathri , Bracci Paige , Rice Terri , Hansen Helen , McCoy Lucie , Clarke Jennifer , Wiencke John , Wrensch Margaret
Published: November 2023

Immune dysregulation is a known characteristic of glioma, however there is little known about the peripheral immune status of lower-grade (LrGG) glioma patients across the course of disease. Using immunomethylomic (deconvoluted blood DNA methylation) data from the UCSF Immune Profiles Study, we explore changes in patient peripheral blood immune cell profiles in newly diagnosed and recurrent LrGG patients. For patients receiving radiation (RT) and temozolomide (TMZ) (n = 75), blood is collected pre-RT/TMZ, post-RT/TMZ, before, during, and after adjuvant TMZ (if applicable). For patients followed only by MRI (n = 53) and those with other treatment plans (n = 67), blood is collected at MRIs, and if applicable, before radiation. For all patients, blood is also collected before new treatment intervention. For patients (LrGG IDH mutated) with paired pre- and post-surgery samples (n = 139), we observed a significant post-surgery increase in the count of memory B-cells (p = 1.7e-5), neutrophils (p = 0.0001), basophils (p = 8.7e-5), monocytes (p = 0.0001), and T-regulatory cells (p = 0.0001) along with a significant decrease in natural killer cells (p = 0.0003). Treatment was also associated with altered patient immune profiles. The use of dexamethasone was associated with an increased count of B-memory (p = 0.0001) and B-naïve cells (p = 0.0001), monocytes (p = 2.1e-5), and neutrophils (p = 3.7e-6) post-surgery. Patients receiving chemoradiation (n = 12) exhibited significant and temporally sustained decreases in B-memory (p < 0.0001), B-naïve (p = 0.005), CD4 naïve cells (p = 0.002), and in neutrophils (p = 0.001). Patient follow-up, data collection, and further analyses are ongoing. These results shed light on the dynamic and variable immune changes in LrGG patients and provide a guide for future prognostic and therapeutic studies.
BIOM-43. LOSS OF NF1 PROTEIN EXPRESSION IS A POOR PROGNOSTIC BIOMARKER IN A SUBSET OF IDH-WILDTYPE GLIOBLASTOMA
Authors: Chang Michael , Chinnasamy Viveka , Ioannou Maria , Rodriguez Fausto , Lucas Calixto-Hope , Schreck Karisa
Published: November 2023

BACKGROUND

Loss of NF1, a tumor suppressor and negative regulator of RAS signaling, is common in IDH-wildtype glioblastoma and occurs through two primary mechanisms – genomic inactivation or proteasomal degradation. NF1 loss is associated with poor clinical outcomes but may also confer sensitivity to MEK inhibitors in some tumors. No clinically validated biomarker for NF1 loss exists, limiting clinical development of therapeutics for these patients. Here, we describe an immunohistochemical biomarker for NF1 expression in IDH-wildtype glioblastoma.

METHODS

We constructed a tissue microarray of 73 genomically-characterized glioblastoma treated at Johns Hopkins - 45 NF1-wildtype and 28 NF1-mutant. Immunohistochemistry was performed to assess NF1 expression, then correlated with clinical and genomic characteristics.

RESULTS

Among NF1-mutant tumors, NF1 expression was lost in 17 specimens (77%) and retained in 5 (23%). Among NF1-wildtype tumors, NF1 expression was lost in 11 (25%), equivocal in 3 (7%), and retained in 30 (68%). Loss of NF1 expression was mutually exclusive with EGFR alterations (p = 0.034) but co-occurred with alterations in TSC2 (p = 0.02) and PIK3R1 (p = 0.049), consistent with our larger genomic patient dataset (n = 277). Loss of NF1 expression was associated with decreased median overall survival (7.3 vs 14.4 months, p = 0.027), even after correcting for prognostic variables (age, MGMT promoter methylation, extent of resection, treatment). This survival difference was driven by the subset of patients with intact genomic NF1 but loss of NF1 expression. Median overall survival in this subset (n = 11) compared to the rest of the cohort (n = 49) was 4.1 vs 13.2 months (p< 0.0001). Cox proportional hazards model correcting for prognostic variables revealed HR 5.4 (95%CI 1.94-15, p = 0.001) associated with loss of NF1 expression.

CONCLUSION

Loss of NF1 expression assessed by immunohistochemistry, particularly when paired with genomically-intact NF1, correlates with inferior outcomes in patients with glioblastoma. These results will need to be validated in a larger cohort.
BIOM-52. PROTEOMIC PROFILE OF CEREBROSPINAL FLUID IN MEDULLOBLASTOMA AND THEIR RELATED EXTRACELLULAR VESICLE PROTEIN
Authors: Choi Seung Ah , Kim Joo Whan , Dan Kisoon , Ha Saehim , Phi Ji Hoon , Han Dohyun , Kim Seung-Ki
Published: November 2023

The development of a sensitive diagnostic method for medulloblastoma (MBL), a common malignant brain tumor in children, is still a challenge. Cerebral spinal fluid (CSF) contains various substances such as soluble proteins and extracellular vesicles (EVs) and may reflect pathological changes in the central nervous system. Especially, EVs within CSF have great potential in the dynamic monitoring and intervention strategies of MBL. This study attempted to determine whether the total CSF protein selected based on proteome information distinguished from the CSF in MBL is also considered in EVs cargo proteins. CSF samples from patients with MBL (N = 44) and hydrocephalus (HC, control, N = 18) were analyzed with LC-MS. A machine-learning approach was used to identify protein biomarkers discriminating MBL CSF. The biomarker potential of the selected proteins was verified through ELISA and ROC analysis. TEM, Nanosight, and Exoview systems were used to analyze the physiological and biological characteristics of proteins in EVs from MBL CSF. Proteomic analysis revealed four elevated soluble proteins (Transketolase (TKT), SPTBN1, HSP90AA1 and NME1-NME2) in MBL CSF. Verification of the selected proteins in whole CSF confirms that TKT was significantly elevated in MBL than in HC. Importantly, the AUC value of TKT was 0.94, which was recognized as an excellent diagnostic marker. In additional TKT analysis, when MBL was divided into subtypes, Group 4 was the most significant compared to HC, and when divided into metastasis group, metastasis was more significant than non-metastatic compared to HC. Consistently, TKT was more abundant in MBL CSF EVs compared to HC CSF EVs. Our results provide insights into the relevance of the proteins in CSF to the cargo proteins reflected in EVs, while also proposing TKT in CSF EVs as an MBL diagnostic marker.
BIOM-53. MENINGIOMA COPY NUMBER VARIANT THRESHOLDS REVEAL OPTIMAL SIZES FOR GAINS OR LOSSES THAT PREDICT LOCAL FREEDOM FROM RECURRENCE OR OVERALL SURVIVA
Authors: Nguyen Minh , Chen William , Choudhury Abrar , Magill Stephen , Raleigh David
Published: November 2023

INTRODUCTION

Meningiomas demonstrate heterogeneous rates of postoperative local freedom from recurrence (LFFR) and overall survival (OS). Risk-stratification models incorporating copy number variants (CNVs) improve outcome prediction, but optimal size thresholds for defining CNVs in meningiomas are unknown, and previous models have used thresholds ranging from 5-80% that are applied uniformly across all chromosome arms. Here we show that optimizing chromosome-specific CNV size thresholds improves meningioma risk-stratification.

METHODS

565 retrospective meningiomas with clinical data and DNA methylation profiles from 2 international institutions were included. CNVs were defined from DNA methylation profiles using iterative thresholds ranging from 5-95% for each chromosome arm. The area under the curve (AUC) for 5-year LFFR or OS was calculated using CNVs across iterative thresholds for each chromosome arm. LASSO and Elastic Net Cox regressions were used to generate multi-CNV models for LFFR or OS.

RESULTS

AUC for LFFR or OS peaked with CNV thresholds that were unique to each chromosome arm. Chromosome 1p deletion had optimal AUCs for LFFR and OS of 0.72 and 0.70 at thresholds of 40% and 30%, respectively. Chromosome 1q amplification had optimal AUCs for LFFR and OS of 0.58 and 0.60 at thresholds of 25% and 20%, respectively. Two previously published CNV-based risk-stratification models (integrated score and integrated grade, optimal LFFR thresholds of 5% and 20%, AUCs of 0.77 and 0.78, respectively) demonstrated heterogeneous sensitivity to CNV thresholds. LASSO and Elastic Net Cox regression identified size-dependent risk-stratification models incorporating loss of 1p, 3q, 4p, 5p/q, 6p/q, 9p, 10q, 11p, 12q, 13q, 14q, 17q, and 18p/q, and gain of 1q, 9q, 10p, and 17q. The optimal AUCs for LFFR and OS from multi-CNV models were 0.77 and 0.77 at thresholds of 25% and 35%, respectively.

CONCLUSIONS

Interrogating CNV thresholds reveals optimal chromosome-specific CNV sizes for predicting LFFR and OS.
BIOS-06. MACHINE LEARNING MADE EASY: DESIGN OF A PROPOSAL FOR A MEDICAL STUDENT EDUCATIONAL SESSION ON HIGH YIELD STRATEGIES FOR LEARNING AND APPLYING DATA SCIENCE
Authors: Brown Nolan , Kuo Cathleen , Neil Zachery , Gendreau Julian
Published: November 2023

Machine learning is a burgeoning field in data science that has many applications in medicine, and it is especially useful in neurosurgery due to the often-complex and potentially emergent nature of day-to-day cases. Machine learning models have the potential to deliver accurate patient prognostication while offering the benefit of providing real-time AI supported clinical decision-making with high data granularity. These models can utilize numerical inputs for more straightforward applications and, with practice, can enable clinicians and trainees to incorporate advanced imaging sequences as inputs to optimize the accuracy of patient prognostication. In the future, these models will become much more commonplace in the clinical setting for neurosurgeons. As medical school curricula do not typically incorporate these statistical techniques into biomedical statistics courses, symposia centered on machine learning could prove essential to medical students seeking to learn these skills. This session would include a brief overview of machine learning models and the procedures for using the programming language of RStudio for the development of novel internet-based prognostication calculators. The benefit of RStudio is that it is user friendly and pre-designed templates are available that can enable students to perform facile customization in tailoring the code to the needs of their clinical investigation. There will be a brief overview of accuracy verification involving discussion of receiver operating curves with the use of training and testing datasets - the heart of machine learning. We have designed and outlined a curriculum that will enable students to identify and classify different techniques (support vector machines, logistic regression, clustering, nearest neighbor classifiers), and recognize the utility of machine learning approaches to neurosurgical outcomes prognostication. Our hope is that students will leave this symposium well-equipped with a newfound confidence and the baseline conceptual framework needed to begin exploring the awesome world of machine learning throughout their training.
BIOS-08. A COMPREHENSIVE ANALYSIS OF METRICS AFFECTING GLIOBLASTOMA RANDOMIZED CONTROLLED TRIALS ROBUSTNESS
Authors: Hameed Farrukh , Sathyamurthi Sam , Sajjad Omar , Hassan Maadeha , Arikawa Kinji , Priyadarshini Mamindla , Wang Yan , Zinn Pascal
Published: November 2023

INTRODUCTION

Randomized controlled trials (RCTs) are the gold standard for evaluating the effectiveness of oncological interventions. RCTs are influenced by various factors, including trial design, outcome measures, sample size, recruitment, and retention. These factors can impact the robustness of a trial.

OBJECTIVE

We comprehensively evaluate the characteristics and robustness of RCTs supporting current GBM management by evaluating trial design, treatment effects, reporting strategies, primary outcome description, patients’ loss to follow-up as well as author and patient demographics.

METHODS

We conducted database searches and all GBM RCTs with significant outcomes published between 2010-2022 were included. Trial characteristics, survival times, the fragility index (FI-minimum number of patients needed to switch to nullify trial outcome) were assessed. Multivariable regression was performed to assess association of trial characteristics with robustness.

RESULTS

Of the 310 RCTs identified, 95 (30.6%) trials reported overall survival as primary end point, 92 (29.7%) progression free survival and 123 (39.7%) examined safety profiles of drugs. Of these, 178 (57.4%) were published from 2015-2022. The trials had a median power of 80.0% (interquartile range [IQR] 80.0-90.0). The median number of enrolled patients was 46 (IQR 24-108) with a median of 2 (IQR 1-5) loss to follow up. Of these trials, 68 were eligible for FI calculation. The median FI was 4.0 (IQR 2.0-11.50), with 38 trials (55.9%) having an FI < 4 and a third of the trials had FI less than number of patients loss to follow up. After adjusting for co-variates, recently published trials, multiple centers trials and commercial sponsorship was associated with significantly higher FI (p< 0.05).

CONCLUSION

Trials supporting current GBM management guidelines have alarmingly low robustness. While the robustness of trials has improved over time, future trials must consider additional metrics in their planning.
CNSC-05. LOCAL AND GLOBAL FUNCTIONAL CONNECTOME DISRUPTION IN PATIENTS WITH GLIOMAS
Authors: Sprugnoli Giulia , Rigolo Laura , Pandit Siddharta , Juvekar Parikshit , Faria Meghan , Tie Yanmei , Sverzellati Nicola , Rossi Simone , Golby Alexandra , Santarnecchi Emiliano
Published: November 2023

The aim of the study was to map the disruption of functional connectome in patients with gliomas using resting-state functional MRI (rs-fMRI) acquired before surgery (3T scanner). 54 patients with gliomas were included (M = 34, mean age = 50.8 y). Manual segmentation of lesions was performed on T1w and T2w MRI scans. Whole-brain measures of network centrality, modularity, integration and segregation were computed for a set of Regions of Interest (ROIs) including the Harvard Oxford anatomical atlas and tumor-related ROIs (i.e., edema, solid tumor, necrotic core). The analysis was performed on adjacency matrices obtained with a cost of 0.15, i.e. considering the 15% of the edges with the largest correlation coefficient values for each subject. A p< 0.05, two-sided, FDR corrected threshold was used for statistical analysis. In newly diagnosed patients (n = 18), we found a decrease of Integration (Degree, Global Efficiency) and Centrality (Betweenness Centrality) and an increase of Segregation (Local Efficiency) in the tumor ROIs compared to healthy brain regions. A similar integration/segregation imbalance pattern was present across the edema, solid tumor and necrotic core, with the edema being the most similar to healthy grey matter and the necrosis being the more altered, possibly reflecting tumor cells’ density and neuronal disruption. In patients at recurrence (n = 36), the same pattern was found, though with a less clear separation among tissue classes, possibly due to the altered anatomical/functional environment caused by functional plastic rearrangement and the effects the treatment(s). Finally, a significant correlation with overall survival was found for local efficiency values of the edema (p< 0.05, r 0,53=, R2 = 28%). We found significant alterations of functional connectome measures in brain regions affected by the tumor, with a decrease of network integration and an increase of network segregation measures characterizing different levels of tumor infiltration and potentially correlating with clinical outcomes.
CNSC-07. USING CRISPR-CAS9 SCREENING TO IDENTIFY BIOMARKERS FOR MMF SENSITIVITY IN GLIOBLASTOMA MULTIFORME
Authors: Buechler Killian , Shireman Jack , Ventarapragada Divya , McManus Graysen , Fares Jawad , Park Cheol , Kumthekar Priya , Ahmed Atique
Published: November 2023

Glioblastoma (GBM) the most common malignant brain tumor with a median survival of 21 months. Recurrent GBM remains resistant to available therapies. Our phase I clinical trial investigates combining mycophenolate mofetil (MMF) and temozolomide (TMZ) to target de novo purine biosynthesis in GBM. Previously, our lab revealed that GBM cells upregulate purine biosynthesis and inhibit purine salvage through the ARL13B-IMPDH2 axis during TMZ therapy. Targeting IMPDH2 with MMF/TMZ extended survival in GBM patient-derived xenograft models. In preparation for Phase II of our clinical trial, we aim to identify a predictive biomarker for MMF+ TMZ therapy. In a CRISPR-Cas9-mediated knockout screen, we identified ten genes, including SLC1A5, showing enrichment (differential β score mean values > 60), indicating sensitivity to MMF/TMZ. SLC1A5 displays a wide distribution of expression in Glioviz patient datasets and encodes the glutamine transporter ASCT2, which is crucial for de novo purine biosynthesis. RNA-seq and protein expression analysis in six glioma cell lines confirmed that high SLC1A5 expression appears to correlate with sensitivity to MMF/TMZ (IC50 0.58-21.30 μM), while low expression led to resistance (IC50 73.43-1148 μM). Knockdown of SLC1A5 in a high-expression cell line demonstrated reduced sensitivity to MMF/TMZ (IC50 2.099 vs IC50 1007 μM). To further validate our findings, we plan to perform SLC1A5 overexpression in low-expression glioma cell lines. We hypothesize that GBM patients with higher SLC1A5 expression may respond favorably to MMF/TMZ therapy due to the reliance of their tumors on de novo purine biosynthesis. We aim to combine this data with analysis of our Phase I clinical samples to retrospectively establish SLC1A5 as a biomarker for MMF/TMZ sensitivity and investigate its role in tumorigenic purine metabolism. Identifying predictive markers will enhance personalized treatment strategies for GBM, offering hope for improved outcomes.
CNSC-11. SEIZURE PRESENTATION IN IDH WT GLIOMA IS RELATED TO UPREGULATED SYNAPTIC SIGNALING AND IONIC TRANSPORT GENES
Authors: McDonald Malcolm , Khan Abdul , Athukuri Prazwal , Lozzi Brittney , Deneen Benjamin , Mandel Jacob , Rao Ganesh
Published: November 2023

Isocitrate dehydrogenase wildtype (IDH WT) glioblastoma (GBM) is the most common malignant brain tumor in adults and is universally lethal. Seizures often occur as a presenting sign of tumor. Recent studies have implicated tumor-neuron communication facilitating synaptic remodeling and peritumoral circuit hyperactivity influencing tumor growth. We assessed the clinical and transcriptional effect of seizure presentation on GBM biology. 63 tumor samples were analyzed for clinical characteristics. Bulk RNA sequencing data was used to identify differentially expressed genes (DEGs), gene sets enrichment analysis (GSEA), and gene ontologies (GO). 24 IDH WT GBM patients presented with seizures compared to 39 patients presenting with other symptoms. There were no significant differences between age at presentation, sex, tumor lobe, tumor location, or grade of tumor. GSEA of gene ontologies revealed upregulated genes in the seizure cohort were largely related to synaptic signaling, ionic transport, and receptor signaling (p-val< 0.05). Alternatively downregulated gene sets were related to extracellular matrix organization, and inflammatory and immune response (p-val< 0.05). Focusing on significantly upregulated DEGs (padj< 0.05,) biological processes related to glutamatergic synaptic transmission and cellular components related to synaptic structures were upregulated. Alternatively, significantly downregulated DEGs (padj< 0.05) were related to biological processes of immunity and extracellular matrix organization and cellular components related to the extracellular matrix. Upregulated genes include IGFN1 and THBS4, both involved in synapse assembly, and SLC17A3, involved in presynaptic glutamate sorting (Log2FC >1,padj <0.05). The downregulated genes include numerous collagen related genes and immune related genes and KCNJ15, an inward rectifying potassium channel and GABRR1, a GABA-rho subunit. Using bulk RNA sequencing in a clinically similar set of IDH WT tumors, we found gene sets and gene ontologies related to synaptic signaling and glutamatergic signaling upregulated in patients with seizures while genes related to both extracellular matrix and immune inflammation were downregulated.
CNSC-12. PATHOLOGY SPECIFIC, PHARMACOLOGICALLY REVERSIBLE LOSS OF NEURONAL FUNCTION IN HUMAN GLIOMAS
Authors: Umbach Gray , Hervey-Jumper Shawn
Published: November 2023

Prior evidence has demonstrated that neuronal activity is preserved within glioma-infiltrated cortex and maintains the ability to engage in task specific computations albeit with information loss. These findings support a potential therapeutic target to rescue cognitive impairments seen in nearly all afflicted patients if neuronal activity can be recovered. We developed an in vivo human behavioral and electrophysiological model to assess neuronal population tuning responses within primary sensory cortex including the decodability of stimuli and task-related changes in population spiking in a cohort of 14 human patients undergoing awake resection. We placed high-density electrode arrays to record human local field potentials from gliomas projecting to the cortical surface. We next designed a static sensory detection threshold task, stimulating with a tachometer at two different face and hand sites totaling 11,200 trials while recording from macroelectrode cortical contacts over both tumor-infiltrated (50 electrodes) and normal appearing cortex (52 electrodes). In tumor-infiltrated cortex, the ability to decode the site of stimulation using the oscillatory power in both theta (4-8 Hz) and gamma (32-70 Hz) bands was reduced compared to normal appearing cortex. However, tumor-infiltrated cortex-maintained selectivity between hand and face stimulation. Targeted next-generation tumor sequencing uncovered novel molecular targets of glioma-neuronal integration. The degree of loss of selectivity correlated with WHO grade and pathology. WHO grade 4 tumors and oligodendrogliomas compared to astrocytomas demonstrating greater loss of selectivity. Gabaergic agonist has been postulated to restore neuronal selectivity in aging and neurodegenerative disease. Therefore, we next computed GABA agonist dose (estimated by time off Propofol and beginning of behavioral testing). Higher concentrations of GABA agonist correlated with improved theta band selectivity and decodability. These results suggest that loss of neuronal computations in glioma-infiltrated cortex is caused by loss of neuronal selectivity and is both tumor selective and pharmacologically reversible.
CNSC-14. T-TYPE CALCIUM CHANNELS REGULATE NEURON-GLIOBLASTOMA INTERACTIONS AND PROMOTE TUMOR GROWTH
Authors: Dube Collin , Zhang Ying , Saha Shekhar , Lai Michelle , Gibert Myron , Escalante Miguel , Hudson Kadie , Yener Ulas , Hanif Farina , Kefas Benjamin , Xu Esther , Sorot Aditya , Sontheimer Harald , Abounader Roger
Published: November 2023

Glioblastoma (GBM) is the most common primary malignant brain tumor with a dismal median survival of 15 months. Glioblastoma utilizes calcium signaling to regulate communication between tumor cells and the neuronal microenvironment. T-type calcium channels (TTCCs) are voltage gated calcium channels expressed in various types of neurons throughout the brain regulating glutamatergic synaptic transmission. We analyzed spatial transcriptomics of human GBM tumors and found that TTCCs are expressed in infiltrating tumor regions and enriched for neuronal development spatial patterns and exhibit neurodevelopmental signatures. Analysis of single cells transcriptomic of human GBM tumors demonstrated that TTCCs are enriched in neuronal subtypes. To complement publicly available data,we treated Glioblastoma Stem cells (GSCs) with an FDA-approved repurposed TTCC blocker, mibefradil, and found downregulation of genes associated with neuron processes such as synapse organization, postsynaptic density and glutamatergic synapses. Treatment of GSCs with mibefradil abrogated cell proliferation in the presence of glutamate. We examined the effects of neuronal Cav3.2 on GBM growth. Genetic Cav3.2 knockout (Cav3.2 KO) in the microenvironment of mice xenografted with syngeneic cell lines (GL261 and CT2A) exhibited significant tumor volume reduction as well as increased survival compared to WT mice in both cell lines. Cav3.2 KO tumors exhibited a decrease in proliferation (Ki67+) and neuronal connections (VGlut1) measured by immunofluorescence compared to WT mice. Cav3.2KO neuron-GBM co-cultures exhibited a significant decrease in proliferation compared to WT neuron-GBM co-cultures. Cav3.2KO neuron-GBM co-cultures exhibited a decrease in synaptic connections and a decrease in neuronal projections to GBM cells compared to WT neuron-GBM co-cultures. Transcriptomic analysis of tumors from Cav3.2KO tumors revealed deregulation of genes associated synaptic transmission, glutamatergic synapses and postsynaptic specialization. Altogether, our findings indicate a role of Cav3.2 in regulating tumors and neurons and promoting GBM progression.
CNSC-31. BESPOKE THREE- PHOTON MICROSCOPY AND ANALYSIS ENABLE DEEP INTRAVITAL BRAIN TUMOR IMAGING
Authors: Soyka Stella , Schubert Marc , Tamimi Amr , Maus Emanuel , Denninger Robert , Wißmann Niklas , Reyhan Ekin , Tetzlaff Svenja , Beretta Carlo , Drumm Michael , Schroers Julian , Steffens Alicia , Walshon Jordain , McCortney Katy , Heiland Sabine , Golebiewska Anna , Kurz Felix , Wick Wolfgang , Winkler Frank , Kreshuk Anna , Kuner Thomas , Horbinski Craig , Prevedel Robert , Venkataramani Varun
Published: November 2023

Glioblastoma is the most common primary brain tumor with a poor prognosis due to its aggressive invasiveness, inevitable recurrence, and therapy resistance. One hallmark of the disease is its whole brain dissemination via preexisting brain structures like blood vessels or white matter tracts. While intravital glioblastoma imaging has been limited to the superficial cortical layers, we developed a method of three photon microscopy allowing the investigation of glioblastoma invasion and colonization in the white matter up to a depth of 1.2 mm over extended time periods. Our method, combining adaptive optics 3P imaging, customized deep learning- based denoising and machine learning segmentation, enables longitudinal volumetric multicolor imaging, being able to distinguish GBM cell signal from myelin structures as well as blood vessel signal. Thus, GBM cells and their interactions with and influence on the microenvironment within the white matter can be investigated longitudinally. We found that compared to cortical invasion, a majority of GBM cells in the corpus callosum are aligned with myelinated axonal tracts, showing the influence of the tumor microenvironment on GBM cell directionality. Also, we found that most GBM cells enter the corpus callosum via a perivascular route. We identified two blood vessel-related mechanisms that allow invasion orthogonal to the myelinated axonal tract direction. We discovered that white matter tract directionality did not change significantly in the early stages of GBM colonization. This might explain limitations of modern clinical imaging to depict white matter glioma infiltration. However, we found that the shape of THG discontinuities differs between GBM cells and other resident cells, which could potentially serve as an imaging biomarker. To summarize, this tailored three-photon microscopy methodology allows unprecedented intravital investigation of GBM invasion in the gray and white matter, enabling the discovery of novel (sub-)cellular mechanisms in cancer and its microenvironment deep in the brain.
CNSC-37. THE ASSOCIATION BETWEEN THE NUMBER OF HIGH-DOSE METHOTREXATE CYCLES AND LONGER REMISSION PERIODS IN PATIENTS WITH PRIMARY CENTRAL NERVOUS SYSTEM LYMPHOMA: A RETROSPECTIVE REVIEW
Authors: Nahm Joshua , Sinha Mousumi , Schumann Eva Highberg , Vu Mia , Ware Cornelius , Hsu Sigmund , Zhu Jay-Jiguang
Published: November 2023

Representing 3-4% of newly diagnosed central nervous system tumors, primary central nervous system lymphomas (PCNSL) are rare, extra-nodal non-Hodgkin lymphomas with an overall survival (OS) of 91.9 months. Though patients can achieve a complete response (CR) after first-line therapy, PCNSL typically recurs within two years. Standard treatment involves high-dose methotrexate (HD-MTX)-based polychemotherapy, but the ideal number of HD-MTX cycles that achieves and maintains a CR is unclear. To address this question, we conducted a retrospective review of PCNSL patients. The primary outcome was median OS. Secondary outcomes included the OS of recurrent PCNSL patients (rOS), number of HD-MTX cycles before the first recurrence or CR, and adverse events (AE). Survival analysis was performed using the Kaplan-Meier estimate. Forty-one patients (23 males; median age: 66) were identified. The median OS is 114 months (95% CI 44.9, NA). Twenty-nine patients achieved CR after first-line therapy with a median of five cycles. Eleven patients experienced PCNSL recurrence (rPCNSL), with five patients previously achieving a CR. The median rOS before the first recurrence was 60.4 months (95% CI 6.3, NA). The median number of cycles of HD-MTX before recurrence was three cycles, and the cumulative total of HD-MTX was five cycles. Among the 24 patients without recurrence and with CR, the median OS was not reached, but the median number of HD-MTX cycles before CR was six cycles and cumulatively was six cycles. Complete results of AEs will be updated at the meeting, but they currently include grade III hypokalemia (45.8%), grade III anemia (45.8%), and grade III acute kidney injury (25.0%). Despite the limitations of a retrospective review, our findings suggest that a higher number of HD-MTX cycles is associated with prolonged duration of remission. Future studies are warranted to determine an ideal number of HD-MTX to maintain remission state in PCNSL patients.
CNSC-44. BIOLOGICAL PATTERN DISCOVERY IN GLIOBLASTOMA
Authors: Ayyadhury Shamini , Sachamitr Patty , Kushida Michelle , Park Nicole , Coutinho Fiona , Whitley Owen , Prinos Panagiotis , Arrowsmith Cheryl , Dirks Peter , Pugh Trevor , Bader Gary
Published: November 2023

Glioblastoma(GBM - IDH wildtype) is an adult glioma, showing abysmal prognosis. Sequencing technologies show us the existence of a neurodevelopmental frame-work, upon which heterogeneous molecular patterns are overlaid. The tumor's functional capacity is the net result of extrinsic+intrinsic molecular factors interacting with each other. These factors affect how cells communicate and organize themselves into complex functional architectures that support their evolution, survival and resistance. Understanding the phenotypic significance of a tumor's architecture, will allow us to understand these processes and compute complex patterns in GBM that are "biologically-relevant". First, we showed, as a proof-of-concept, that "biologically-relevant" signatures are imprinted within the spatial organization of cells using spatial pixel analysis. We used a phase-contrast image dataset of glioblastoma stem cells grown in culture, imaged 4-12hrly, over 12-16 days. We applied 29 hand-engineered pixel features per image, deriving spatial pixel signatures for each of our 17’601 phase-contrast images. Using different computational analytical methods and gene expression from matched bulk RNA datasets, we showed that spatial pixel patterning follows biologically relevant phenotypes. We found samples of images with high PC2 scores had higher mesenchymal/microglia scores, as compared to samples of images with low PC2 scores, which showed higher neurodevelopmental signatures. In addition, we found that mathematical algorithms describing entropy, homogeneity, contrast and complexity were clearly enriched in specific biological groups. Hence, our study showed that the organizational/architectural patterns of biological entities show preservation of fundamental organizational principles. Building upon the above data (which is currently a manuscript under preparation), we will apply these principles towards pattern discovery of GBM tissues at multiple architectural hierarchies (i.e subcellular, cellular, stroma) using different GBM models, imaging and spatial transcriptomics. If structure equates to function, understanding GBM architecture will have important applications in surgical tool developments and understanding relevant mechanistic patterns in GBM.
STEM-03. TARGETING ANDROGEN SIGNALING IN GLIOBLASTOMA
Authors: Mason Sofia , Chung Sylvia , Juan Beatriz Perez San , Sim Hao-Wen , Dear Rachel , Day Bryan , Holst Jeff , Chaffer Christine
Published: November 2023

BACKGROUND

Glioblastoma is a lethal brain cancer. Androgen receptor (AR) transcript levels and protein expression are upregulated in glioblastoma compared to normal brain. AR signaling has a role in cancer stem cell function in various cancers. Stem cell activity and cancer cell plasticity represent a key resistance mechanism in glioblastoma. Thus, potent brain-penetrant anti-androgen therapies have the potential to improve treatment outcomes in glioblastoma.

METHODS

Clinical samples and patient derived xenografts were assessed for AR expression by immunohistochemistry. Cytoplasmic AR+ and AR- glioblastoma patient-derived xenografts maintained under stem cell conditions were tested in vitro for their response to anti-androgen therapies (abiraterone, enzalutamide and seviteronel). Stem cell function was assessed using a tumoursphere assay. Changes in plasticity markers, including ZEB1, were measured with immunofluorescence.

RESULTS

~55% of people with glioblastoma have detectable cytoplasmic AR based on immunohistochemistry. AR-positive patient-derived glioma stem cell line RN1 was inhibited by low concentrations of anti-androgen agents, with seviteronel having the lowest half maximal inhibitory concentration (IC50) (at 96 hours: enzalutamide 52µM, abiraterone 12µM, seviteronel 7µM). AR-negative line WK1 was also inhibited though with higher IC50s (at 96 hours: enzalutamide 63µM, seviteronel 21µM). Conventional cell lines U87 and U251 were also inhibited by anti-androgen monotherapy. Tumoursphere assays demonstrated that anti-androgen agents inhibit the tumour-forming ability of cells, with seviteronel and abiraterone showing more inhibition than enzalutamide. Immunofluorescence analysis showed downregulation of plasticity marker ZEB1 with anti-androgen treatment (normalised mean fluorescent intensity: abiraterone 0.84, enzalutamide 0.91, seviteronel 0.74).

CONCLUSIONS

Glioblastoma is a devastating disease, lacking effective or targeted treatments. Targeting AR with repurposed anti-androgen drugs may be a promising therapeutic strategy, with the potential to abrogate treatment resistance. We are now evaluating these anti-androgen regimens in animal experiments, which will provide a more replicative microenvironment and confirm blood-brain barrier penetrance.
CCRG-02. DEVELOPMENTALLY PARTITIONED PROLIFERATIVE COMPARTMENTS IN GLIOBLASTOMA
Authors: Plaisier Christopher , O'Connor Samantha , Arora Sonali , Petrecca Kevin , Patel Anoop , Paddison Patrick
Published: November 2023

It is now well established from single-cell RNA-seq studies that glioblastoma (GBM) tumors are complex, maligned neuro-developmental ecosystems harboring diverse tumor cell types. Neoplastic cells can resemble astrocytes, neural progenitors, oligodendrocyte progenitor cells, mesenchymal cells, and radial glial cells that contribute to tumor growth and homeostasis in specific ways. However, GBM single-cell data sets have failed to produce general models for transitions in and out of specific developmental and proliferative states in tumors. One reason is that human GBM tumors do not neatly resolve into developmental hierarchies. Here we focused on modeling GBM tumor cellular heterogeneity by defining "proliferative compartments" in single-cell transcriptomic data derived from primary tumors and early passage tumorsphere cultures. Previously, we observed that each tumor cell entering S-phase has a unique developmental signature that can be leveraged to define broader partitioned proliferative compartments (PPCs) with distinct developmental gene expression and genomic alteration patterns. Thus, we extracted the S-phase cells from a tumor, defined the proliferative compartments in that tumor using de novo clustering, determined the marker genes for each compartment, and defined the broader PPCs across tumors by de novo clustering based on similarity in marker genes. From a cohort of six tumors we observed eight broader PPCs and found that tumors can contain as many as five PPCs or as few as two. Because tumor growth and recurrence both require cell proliferation, we propose that patient-specific PPCs represent the engines of GBM progression.
CSIG-13. DIELECTRIC PROPERTIES OF INTRACRANIAL TUMORS – ROLE OF MYELIN CONTENT
Authors: Proescholdt Martin , Haj Amer , Schebesch Karl-Michael , Riemenschneider Markus , Baborie Atik , Schmidt Nils-Ole
Published: November 2023

INTRODUCTION

Recently, tumor treating fields (TTFields) were established for the treatment of newly diagnosed glioblastoma (GBM). One of the most crucial parameters defining the treatment efficacy of TTFields is the electric field intensity, which depends on the dielectric properties of the tumor tissue. In this study we determined the dieclectric properties of brain tumors by analyzing resected tissue. In GBM patients, histological analysis for cellularity, vascularization and myelin content was performed. In addition, sensitivity analyses for specific parameters were conducted.

METHODS

A cohort of 130 patients with tumors of different histology and malignancy grade have been recruited. Tissue samples were placed into a cylindrical cell with a known diameter. The impedance was recorded at frequencies 20Hz-1MHz. The measured impedance was translated into dielectric properties (conductivity and relative permittivity) based on the parallel plate model. Myelin, which is the most powerful electric isolator in the brain, was assessed in GBM samples by luxol fast blue staining, and quantified in a three-tier scale. To assess the impact of tissue conditions on the measurements, probes were warmed to 35 degree Celsius, dehydrated or irrigated with 0.9% saline solution.

RESULTS

We found significant differences between the conductivity of different types of tumors with meningiomas showing the lowest and GBM tissue exhibiting the highest conductivity values. GBM samples with very high median conductivity values displayed a consistently lower myelin content. In addition, GBM patients with particularly high conductivity had a significantly shorter overall survival (log rank analysis, p = 0.024). While tissue temperature had no detectable effects on the dielectric properties in GBM, saline irrigation tissue hydration significantly affected the results.

CONCLUSION

The dielectric properties of intracranial tumors depend on histological class and malignancy grade. GBM patients with high conductivity values showed a significantly poorer prognosis, indicating this parameter as potential marker for TTF efficacy.
CSIG-22. CAMKIIΓ INTERACTS WITH HIF-1Α IN AN OXYGEN-INDEPENDENT MANNER TO DRIVE PLASTICITY IN GLIOBLASTOMA
Authors: Ventarapragada Divya , McManus Graysen , Zhou Weidong , Park Cheol , Buechler Killian , Stepniak Amelia , Miska Jason , Ahmed Atique
Published: November 2023

Glioblastoma (GBM) is a highly aggressive brain tumor that demonstrates treatment resistance. Hypoxia-inducible factor 1-alpha (HIF-1α) plays a crucial role in GBM progression and recurrence. While HIF-1α overexpression is correlated with poor GBM survival, clinical inhibitors have shown limited efficacy due to compensatory pathways that are not well understood. In previous studies, we discovered GBM upregulates HIF-1α after temozolomide (TMZ) through an unknown mechanism independent of physiological hypoxia. We demonstrated HIF-1α but not HIF-2α knockdown reduces the stem-like properties of GBM and enhances their sensitivity to TMZ therapy in vitro and in vivo. To elucidate the mechanism of TMZ-induced HIF-1α stabilization, we performed co-immunoprecipitation of HIF-1α followed by mass spectrometry analysis (IP-MS). Our findings revealed 99 binding partners that were significantly and uniquely upregulated after TMZ treatment compared to GBM cells cultured under normal oxygen tension (20% O2), hypoxia (1% O2), or DMSO. Among these interactions, we focused on the HIF-1α-CAMKIIγ axis for further investigation based on pathway enrichment, patient survival data, and co-expression analysis in the TCGA_GBM dataset. Co-immunoprecipitation experiments confirmed enhanced binding between HIF-1α and CAMKIIγ in different GBM patient-derived xenograft (PDX) lines while this interaction was significantly reduced in normal astrocytes. Knockdown of CAMKIIγ inhibited the transcriptional activity of HIF-1α, increased its protein turnover, and improved TMZ sensitivity compared to control cells following TMZ treatment. Additionally, the expression of stem-cell markers and neurosphere formation, which are associated with GBM aggressiveness, were attenuated in hypoxia and TMZ conditions when CAMKIIγ was absent. Our data demonstrated that the HIF-1α-CAMKIIγ axis is critical for TMZ-induction of HIF signaling and cellular plasticity.
CSIG-28. R17 INDUCES A PRO-ONCOGENIC PROTEIN HYPERPHOSPHORYLATION PROGRAM IN CUSHING’S DISEASE
Authors: Asuzu David , Mullaney Dustin , Nwokoye Diana , Stoica Stefan , Bhatt Shyama , Mandal Debjani , Maric Dragan , Elkahloun Abdel , Sisay Bayu , Tatsi Christina , Nieman Lynette , Chittiboina Prashant
Published: November 2023

OBJECTIVES

to investigate molecular drivers of CD.

METHODS

We performed single-nucleus assays for transposase-accessible chromatin (snATACseq) and single-nucleus RNAseq (snRNAseq) in 4 CD adenomas including two adenoma-normal pairs. To assess proteins bound to the R17 promoter, we performed reverse chromatin immunoprecipitation (rChIP) using CRISPR dCas9-3XFLAG-Biotin and R17 promoter gRNAs followed by mass spectrometry (LC-MS). Mechanistic studies were performed in murine corticotroph cells (mCort) stably overexpressing R17 versus GFP. Using LC-MS, we quantified total and phospho-peptides from human CD adenomas versus adjacent normal pituitary glands (n = 3/group) and from mCort-R17 versus mCort-GFP cells (n = 5/group).

RESULTS

We found increase in R17 chromatin accessibility and gene expression in CD versus normal corticotrophs. rChIP identified R17 promoter enrichment of SUB1 and histone variant H1.4 indicating active remodeling. In CD specimens and in mCort-R17 cells, PP2A targets EGFR and ERK were hyperphosphorylated. LC-MS identified 30 peptides differentially phosphorylated in both CD adenomas and mCort-R17 cells. mCort-R17 cells demonstrated accelerated cell cycle progression with increased proliferation (P < 0.001). In mCort-R17 cells, the PP2A agonist Fingolimod partially rescued AKT phosphorylation and induced apoptosis.

CONCLUSIONS

R17 is activated in CD due to upstream chromatin remodeling events. Aberrant R17 signaling leads to protein hyperphosphorylation and cell cycle acceleration, which is partially reversible by the PP2A agonist Fingolimod. Fingolimod may represent a novel therapeutic option for patients with CD.
RADT-02. WHOLE-BRAIN RADIATION THERAPY WITH SIMULTANEOUS INTEGRATED BOOST VERSUS STEREOTACTIC RADIOTHERAPY FOR THE TREATMENT OF BRAIN METASTASIS FROM NON-SMALL CELL LUNG CANCER
Authors: Ziwei Wu , Lixia Lu , Yuanyuan Chen , Ming Chen
Published: November 2023

OBJECTIVE

To compare whole-brain radiotherapy with a simultaneous integrated boost (WBRT-SIB) to stereotactic radiotherapy (SRT) in treating non-small cell lung cancer (NSCLC) patients with brain metastases (BMs).

METHODS

Between April 2013 and June 2021, NSCLC patients receiving WBRT-SIB or SRT for 1-10 BMs were reviewed. Propensity score matching was carried out without replacement using a ratio of 1 WBRT-SIB: 1 SRT patient and a caliper width of 0.03 to yield sufficient power and homogeneity between treatment groups. Intracranial progression-free survival (iPFS), overall survival (OS), the cumulative incidence of local tumor progression, and intracranial distant tumor progression were compared between groups. Toxicities were assessed utilizing the National Cancer Institute Common Terminology Criteria for Adverse Events Version 5.0 (CTCAE 5.0).

RESULTS

A cohort of 181 patients with was included in this study. After propensity score matching, there were 37 patients in the WBRT-SIB and the SRT group, respectively. The WBRT-SIB group had a significantly longer median iPFS than the SRT group (10.8 months vs. 6.5 months, p = 0.018). The median OS (p = 0.861) and the cumulative incidence of local tumor progression (p = 0.437) were similar between the two groups. WBRT-SIB was associated with reduced intracranial distant tumor progression (p = 0.043). No toxicities higher than grade 3 were observed in both groups. KPS score was an independent prognostic factor of OS (HR 0.459, p = 0.032).

CONCLUSIONS

Compared to SRT, the iPFS of NSCLC patients with BMs was prolonged after WBRT-SIB, mainly related to better control of distant intracranial tumors. WBRT-SIB is safe and can be an alternative option for patients with BMs. We also found that the KPS score is an independent prognostic factor of OS.