bluuebunny/crossref_metadata_2025 huggingface dataset:
BIOM-37. EXPLORING L-TYPE AMINO ACID TRANSPORTER 1 (LAT1) AS A DIAGNOSTIC MARKER AND THERAPEUTIC TARGET IN MALIGNANT PERIPHERAL NERVE SHEATH TUMORS (MPNST)
Authors: Wang Guangfeng , Borcherding Dana , Zhang Xiaochun , He Kevin , Fettig Nikki , Shoghi Kooresh , Hirbe Angela
Published: November 2024

OBJECTIVES

Malignant peripheral nerve sheath tumors (MPNST) are aggressive soft tissue sarcomas that frequently arise from benign plexiform neurofibromas (PN). Approximately half of MPNST are associated with Neurofibromatosis type 1 (NF1). Despite partial elucidation of the molecular mechanisms underlying MPNST, diagnosis remains challenging, and prognosis is poor. Previously, we found that MPNST cells have elevated βIII-spectrin, a cytoskeletal protein involved in cell morphology and regulation of transporters, including L-type Amino Acid Transporter 1 (LAT1). LAT1 facilitates transport of large neutral amino acids and is upregulated in other cancer types. Our objective is to investigate LAT1 as a diagnostic marker and therapeutic target for MPNST.

METHODS

Gene and protein expression were determined by qPCR and WES protein analysis, respectively. The effect of shRNA knockdown of LAT1 in MPNST cells was assessed using IncuCyte proliferation assays as well as multiple mouse models. Tumor uptake of [18F] FDOPA, primarily transported by LAT1, in MPNST PDX tumors was assessed using PET/CT imaging.

RESULTS

LAT1 was expressed in MPNST (n=14) vs. benign PN tissues (n=6). Additionally, in human and murine MPNST cells, shRNA-mediated knockdown of LAT1 suppressed proliferation of MPNST cells in vitro and tumor growth in vivo. In a subcutaneous tumor growth model, intratumor injection of shLAT1 lentivirus reduced tumor volume to approximately 50% of control. Furthermore, uptake of [18F] FDOPA by LAT1 in MPNST PDX tumors was detected by PET/CT imaging. WU-356 PDX tissues exhibited higher LAT1 expression as well as increased uptake of [18F] FDOPA in tumors. Conversely, PET imaging showed significantly lower uptake of [18F] FDOPA in LAT1 knockdown tumors (p<0.05).

CONCLUSIONS

These results suggest that LAT1 promotes the growth and survival of MPNST cells. In addition, these findings demonstrate the potential use of LAT1 and the radiotracer [18F] FDOPA as a diagnostic biomarker in the management of MPNST.
BIOM-41. HMGB2 IS A CANDIDATE PROGNOSTIC BIOMARKER IN BOTH ADULT AND PEDIATRIC DIFFUSE GLIOMAS
Authors: Becker Aline , Becker Valesio , Sirhan Zaid , McElroy Joseph , Haque S Jaharul , Chakravarti Arnab
Published: November 2024

Few prognostic molecular markers are common to pediatric- and adult diffuse gliomas (PDG and ADG). HMGB2 is a protein expressed in human brain development and in de-differentiated high-grade gliomas (HGG), which we have recently described as a putative prognostic and predictive biomarker in ADG, associated with proliferative activity and EIF2 signaling pathway upregulation. Additionally, we showed that HMGB2 shRNA knockdown (HMGB2-KD) reduced cell proliferation and colony formation, and radio-sensitized ADG cell lines. Aims: To assess the prognostic potential of HMGB2 in PDG and its association with stemness.

METHODS

1) We singled out HMGB2 from proteomic profiles of an independent PDG cohort previously associated with EIF2 and MYCN signaling upregulation for survival analysis. 2) PDG cells SF188 were de-differentiated with substitution of DMEM by a neurobasal (NB) medium devoid of serum for stemness association studies. 3) We generated HMGB2-KD clones of PDG cell lines SF188 (grown in DMEM and NB) and KNS42 with 3 different shRNAs.

RESULTS

HMGB2 expression was significantly related to histological grade in PDG (grade 3 vs. grade 2 and grade 4 vs. grade 2: fold change= 8.16 and 9.39; p=0.0045 and <0.001; FDR= 0.04 and 0.0006, respectively) and survival (p=0.004, FDR=0.142 for PFS and p=0.03, FDR=0.291 for OS), in a Cox model including age and grade as covariables. Compared with cells growing in DMEM, SF188 cells growing in NB medium had a morphological change from stellate cells to round cells growing in spheres, with 2-fold increase in C-MYC and HMGB2 expression by Western analysis. HMGB2-KD resulted in >90% reduction of HMGB2 expression in SF188 and >70% in KNS42.

CONCLUSION

Collectively, our preliminary results suggest that HMGB2 is associated with stemness, has similar roles in PDG and ADG, and may be validated as prognostic/predictive biomarker in both adult and pediatric populations. Mechanistic studies of radiation response are underway.
BIOM-62. CIRCULATING TUMOR CELLS IN MEDULLOBLASTOMA: A NOVEL PERIPHERAL BIOMARKER FOR CNS DISEASE
Authors: Ozkaya Tevhide , Chien Franklin , Boya Mert , Gray Rachel , Arthur Robert , Claussen Henry , Velazquez Vega Jose Velazquez-Vega , Schneiderjan Matthew , Bhasin Manoj , Sarioglu Fatih , MacDonald Tobey
Published: November 2024

BACKGROUND

Medulloblastoma is an embryonal tumor of the cerebellum and the most common pediatric malignancy of the central nervous system (CNS). Diagnosis, staging, and disease monitoring require invasive neurosurgical biopsy or MRI. No blood or cerebrospinal fluid (CSF) biomarkers exist. We demonstrate the presence of circulating tumor cell clusters (CTCCs) in patients with medulloblastoma and use a transcriptomic approach to describe their gene expression. Design/Method: We enrolled 34 total patients in a longitudinal study: 24 with medulloblastoma and 10 without malignancy. CTCCs are captured from unprocessed whole blood using a physics-based label-free approach to CTCC isolation. RNA sequencing was performed on 21 samples at time of diagnostic resection from 6 patients with medulloblastoma. Sequencing reads were assessed for quality control, trimmed, and aligned to the human genome. Differential gene expression (DGE) was performed against whole blood from healthy controls. Reactome pathway database of curated, peer-reviewed gene sets are used for gene set enrichment analysis (GSEA).

RESULTS

The presence of peripherally CTCC patients with medulloblastoma is not previously known and was detected in all patients with medulloblastoma whereas none were observed from patients with non-malignant hematological conditions. RNA-sequencing demonstrate separation on principal component analysis and gene-expression heatmap against whole blood healthy control. DGE shows upregulation of genes indicating stem-like state. GSEA show enrichment of medulloblastoma molecular pathways and gene sets defining integrin, extracellular matrix remodeling, and cell adhesion pathways. Transient elevation of peripheral CTCC followed by return to baseline levels was associated with durable disease response to therapy, whereas persistence or elevation of CTCC in CSF correlated with disease recurrence.

CONCLUSION

The universal and specific finding of peripherally captured CTCC arising from parent tumor in the CNS within our patient cohort suggests a utility as a tumor biomarker. Future sequencing is planned comparing CTCC transcriptome against parent tumor.
BIOM-63. LONG-TERM SURVIVAL AND MOLECULAR CHARACTERIZATION OF IDH-WILDTYPE GLIOBLASTOMA IN AN ASIAN COHORT
Authors: Lee Wing Lun , Ko Elaine Yee-Ling , Lecoultre Marc , Ho Isaac , Chiu Matthew Kin-liang , Chiang Chi-Leung , Leung Dennis , Leung Gilberto Ka-Kit , Chow Joyce Shuk-wan , Woo Peter Yat Ming , Lam Tai Chung , Aya El Helali
Published: November 2024

BACKGROUND

Glioblastoma (GB), the most aggressive form of brain tumor, presents a significant clinical challenge with a devastating prognosis. Globally, the median overall survival (OS) for GB is typically 12-15 months, largely due to its inherent resistance to chemotherapy and radiotherapy. However, some patients exhibit prolonged survival, and their specific demographic and genomic characteristics remain poorly defined. Understanding the clinical profile of these exceptional responders holds great promise for informing improved management strategies for GB.

METHODS

In a retrospective two-center study conducted in Hong Kong (IRB: UW 21-189), we recruited 80 patients diagnosed with IDH wildtype glioblastoma. All patients underwent next-generation sequencing as part of their treatment. Long-term survivor (LTS) was defined as >=36 months from diagnosis, while medium-term survivor (MTS) was defined as >=18 months, and <18 months as short-term survivors (STS). When appropriate, descriptive statistics were used to report frequency with percentage or median with range. Fisher’s exact test was employed to analyze categorical variables, while continuous variables were assessed using the Kruskal-Wallis test. All statistical analyses were conducted using R (version 4.2.3). Result: Among the cohort of 80 recruited patients, the median survival was 18.8 months (95% CI 17.2-24.8). Notably, 21% (n=17) were LTS, 31% (n=25) were MTS, and 48% (n=38) were STS. A statistically significant variation (p < 0.05) was evident across three groups: gender distribution, the number of resections performed, the proportion of patients receiving genomic-guided treatment, and their participation in clinical trials. However, no significant difference was observed in MGMT promoter status and ECOG score.

CONCLUSION

Identifying these clinical factors enhances the clinical management of GB. The implications of these findings underscore the potential importance of incorporating genomic-guided treatment in both local and global clinical practices in GB.
BIOM-64. EXPLOITING CELL-FREE DNA TO DISSECT MECHANISMS OF RESISTANCE TO TEMOZOLOMIDE IN GLIOBLASTOMA CELLS
Authors: Mankame Sharvari , Kyriakidou Maria , Tang Nanyun , Reid George , Berens Micheal , Barthel Floris P
Published: November 2024

Temozolomide (TMZ) is a cytotoxic alkylating agent that serves as a cornerstone in glioblastoma (GBM) treatment, yet is notorious for eliciting hypermutation in recurrent tumors that have developed drug resistance by downregulating mismatch repair (MMR). We reasoned that cell-free DNA (cfDNA), a non-invasive measure for detecting tumor-derived biomarkers, could be used to temporally examine the mechanism of resistance to TMZ. We therefore designed an in-vitro experiment using the patient-derived GBM43 cell line. Over five days, batches of cells were treated with either TMZ or vehicle. Cell counts and cfDNA were collected every 24 hours. We observed an associated increase in both dead cell counts and cfDNA yield in treated cells compared to untreated cells. In TMZ-treated samples, cfDNA showed a periodic fragmentation pattern consistent with internucleosomal cleavage and TMZ-induced apoptosis. Intriguingly, higher numbers of variants were found in the cfDNA of untreated samples when compared to treated cfDNA, possibly reflecting TMZ-induced selection of resistant subclones in treated samples. This was further supported by the finding that the average variant allele frequency (VAF) increased over time in TMZ-treated samples, consistent with purifying selection. Conversely, cfDNA from vehicle-treated samples demonstrated a decrease in the average VAF over time, consistent with clonal diversification. Although we did not observe an enrichment of the TMZ-associated mutational signature SBS11, we did find higher levels of defective MMR mutation signature SBS44 in treated samples, potentially indicating the selection of subclones with defective MMR mechanisms. Overall, our preliminary findings show that cfDNA can be used to resolve mechanisms of TMZ resistance over time. We are addressing open questions by performing longer treatment times, testing other patient-derived cell lines, and performing murine xenotransplantation experiments to test TMZ-associated hypermutation in vivo. Our work will contribute to our understanding of TMZ resistance, and importantly push the development of cfDNA-based biomarkers for measuring TMZ-sensitivity in patients.
BIOM-65. ELEVATED MMP9 EXPRESSION IN ADHERENT AND LARGE VESTIBULAR SCHWANNOMAS: A PROMISING THERAPEUTIC TARGET
Authors: Nguyen Han T N , Duhon Bailey H , Kuo Hsuan-Chih , Fisher Melanie , Brickey Olivia M , Zhang Lisa , Otero Jose J , Prevedello Daniel M , Adunka Oliver F , Ren Yin
Published: November 2024

Vestibular schwannoma (VS) is the most common tumor of the cerebellopontine angle affecting up to 1 in 2000 adults and presents significant treatment challenges, especially when the tumor is adherent to the brainstem and cranial nerves. These tumors often require complex surgeries with high risks of complications such as hearing loss, facial paralysis, and incomplete tumor resection. Previous studies have linked immune cell infiltration and protease activation to aggressive VS. MMP9, a protease involved in extracellular matrix (ECM) remodeling, is associated with cancer invasion and progression, suggesting its potential as a biomarker and therapeutic target. This study explores whether MMP9 can classify VS and improve surgical outcomes.

METHODS

Fresh tumors were collected from patients undergoing surgery to establish primary VS cultures. Transcriptomic profiling was performed to examine expression of genes related to ECM remodeling, protease activation, and immune cell infiltration. MMP9 expression during schwannoma progression was validated using immunofluorescence staining and ELISA. Transwell invasion assays were performed to evaluate the role of MMP9 in schwannoma invasion. The efficacy of small molecule inhibitors against eIF4F (a transcription factor that binds to 5’ UTR of MMP9) and against MMP9 was tested in a mouse model of schwannoma.

RESULTS

MMP9 was the most abundantly expressed protease in VS. MMP9 expression increased with tumor size, and was higher in adherent than non-adherent VS (49.7 vs. 13.8 ng/mL, p=0.014). MMP9 expression was abundant in perivascular regions in adherent VS. Inhibition of eIF4F reduced schwannoma cell invasion mediated by MMP9.MMP9 inhibition in vivo- reduced tumor burden by 50% in mice.

CONCLUSIONS

MMP9 is a highly active protease in VS growth and adherent VS. Developing MMP9-targeted therapies could be a breakthrough in managing these challenging tumors.
BIOM-72. INTEGRATION OF ULTRASENSITIVE ELECTROLUMINESCENT IMMUNOASSAY AND CELL-FREE DNAMETHYLATION ANALYSIS FOR NON-INVASIVE MONITORING OF ADULT DIFFUSE GLIOMAS
Authors: Ajisebutu Andrew , Chatanaka Miyo , Patil Vikas , Prassas Ioannis , Eleftherios Diamandis , Zadeh Gelareh
Published: November 2024

Gliomas are highly aggressive brain tumors with nearly universal recurrence rate. Despite this, the ability to accurately predict and identify tumor recurrence relies solely on serial MRI imaging, which is marred by treatment effects such as radiation necrosis, and necessarily identifies progression retrospectively. It is believe that tumor undergo molecular changes upon tumor progression, however the resampling of tumors upon recurrence imposes significant risks. This highlights the need for novel non-invasive biomarkers capable of identifying tumor recurrence. Due to the low accuracies of individual biomarkers, we have proposed the use of an integrated, multi-platform approach to biomarker discovery. A cohort of 107 glioma plasma samples, including 30 pairs, underwent plasma proteomic analysis, consisting of a panel of serum proteins (FABP4, GFAP, NFL, Tau and MMP3,4 &7) quantified through ultrasensitive electrochemiluminescence multiplexed immunoassays, and plasma DNA methylation analysis, captured through cell-free methylated DNA immunoprecipitation and high-throughput sequencing. Unsupervised hierarchal clustering revealed robust separation of primary and recurrent tumors through plasma proteomics, associated with a distinct plasma methylation signature. NFL, Tau and MMP3 levels differed between primary and recurrent samples; pair-wise analysis revealed increased in NFL and Tau concentrations upon recurrence. Tau levels predicted outcome independent of WHO Grade and IDH status. A predictive generalized linear regression model created through the integration of the proteomic and methylation signatures allowed for the discrimination of primary and recurrent samples in 83% of cases. This work suggests that the combination of DNA methylation and plasma proteomics may improve the ability of these techniques for the serial monitoring of gliomas patients.
BIOM-74. IDENTIFICATION OF EMP2 AS A TARGETABLE BIOMARKER IN GLIOBLASTOMA TUMORS RESISTANT TO ANTI-ANGIOGENIC THERAPIES
Authors: Mekonnen Mahlet , Aguirre Brian , Mehta Riddhi , Chandla Anubhav , Wadehra Madhuri , Yang Isaac
Published: November 2024

BACKGROUND

The role of Epithelial Membrane Protein-2 (EMP2) in mediating resistance to current anti-angiogenic drugs in the treatment of glioblastoma (GBM) is an intriguing hypothesis that may provide insights into improving the limited survival benefits observed in current therapies. Investigating the interaction between EMP2 and anti-angiogenic drugs such as bevacizumab or Aflibercept could lead to a better understanding of the mechanisms underlying resistance, potentially opening up new avenues for more effective treatment strategies.

OBJECTIVE

To use EMP2 as a targetable biomarker for anti-VEGF resistance and test combination therapy with anti-VEGF (Aflibercept or bevacizumab), and a novel anti-EMP2 mAb in SB28 and GL261 GBM mouse models.

METHODS

Tumor tissue was collected prospectively from patients undergoing surgery for suspected glioblastoma and stained for EMP2 and HIF-2a using standard immunohistochemistry. Patients had banked tumor tissue both before and after bevacizumab treatment with 3 months of clinical and radiologic follow-up available. To establish syngeneic models, 10-week-old mice were subcutaneously implanted with murine SB28 or GL261 GBM cells.

RESULTS

Immunohistochemistry conducted on pre- and post-bevacizumab-treated patient samples revealed an increase in EMP2 and HIF-2a protein expression post-treatment. This increase in EMP2 expression correlated with reduced survival time. Similarly, the in vivo models showed a significant upregulation in both markers in the Aflibercept-treated cohort compared to the control. Treatment with either polyclonal sera to VEGF or Aflibercept failed to reduce tumor load or provide a therapeutic benefit. However, combination therapy with Aflibercept and anti-EMP2 mAb was found to significantly reduce tumor volume and weight in both SB28 and GL261 tumors. These findings highlight the potential effectiveness of combination therapies targeting EMP2 and VEGF in reducing tumor load in GBM.

CONCLUSION

Our results suggest that EMP2 can be a promising new therapeutic target that can be combined with anti-VEGF treatment to potentially increase overall survival in GBM patients.
BIOS-02. DIFFERENT GROWTH PATTERN OF VESTIBULAR SCHWANNOMAS IN NEUROFIBROMATOSIS TYPE 2 PATIENTS ACCORDING TO AGE
Authors: Kang Ho , Ji So Young , Hwang Kihwan , Kim Chae-Yong , Han Jung Ho
Published: November 2024

The natural growth pattern of vestibular schwannoma (VS) associated with the rare disease neurofibromatosis type 2 (NF2) has not been sufficiently elucidated, and the rationale for proactive treatment for VS in NF2 is not firmly established. This study aimed to uncover the growth pattern of VS associated with NF2 and identify risk factors for rapid growth. A total of 68 VSs from 41 NF2 patients who were diagnosed in Seoul National University Bundang Hospital from July 2006 to July 2023 were retrospectively reviewed. NF2 was clinically diagnosed according to the Manchester criteria. The mean age was 34±16 years (mean ± standard deviation) at diagnosis. The initial tumor volume was a median of 0.263 cm³ (interquartile range [IQR], 0.053-0.754 cm³), and 51 VSs exhibited serviceable hearing classified as Gardner-Robertson grade 1 or 2. Patients were followed for a median of 45 months (IQR, 30-80 months), during which the median volume increased by 0.145 cm³ (IQR, 0.020-0.783 cm³), representing an annual growth rate of 0.044 cm³/yr (IQR, 0.003-0.202 cm³) or 12.4%/yr (IQR, 2.2-27.5%). Patients younger than 25 years (15 patients with 28 VSs) had significantly smaller initial tumor sizes (0.170 cm³ [IQR, 0.031-0.339 cm³] vs. 0.510 cm³ [0.099-1.399 cm³]) but faster tumor growth rates (annual growth rate; 27.3%/yr [IQR, 11.3-81.4%/yr] vs. 4.7%/yr [IQR, 0.0-16.1%/yr]) compared to those 25 years or older (26 patients with 40 VSs). Among the 51 VSs with serviceable hearing at diagnosis, 40 VSs were followed up for more than 12 months. Among them, 13 (32.5%) experienced unserviceable hearing deterioration at a median of 58 months. The risk of hearing loss significantly increased with faster tumor growth rates. The growth rate of NF2-related VS was higher in patients under the age of 25, which may be helpful in determining indications for proactive treatments like radiosurgery for NF2-related VS.
BIOS-04. THERAPIES FOR LEPTOMENINGEAL DISEASE: A NETWORK META-ANALYSIS
Authors: Suresh Vinay , Panchawagh Suhrud , Muneer Muneeb , Mansouri Alireza , Ozair Ahmad
Published: November 2024

INTRODUCTION

Leptomeningeal disease (LMD) has limited head-to-head comparison of available therapies, including systemic chemotherapy (SysCT), intrathecal (IT) or intraventricular therapy (ITV), whole-brain radiotherapy (WBRT), or cranio-spinal irradiation (CSI). No network meta-analysis (NMA) of randomized controlled trials (RCTs) of LMD has been reported so far.

METHODS

NMA was conducted using published LMD RCTs and reported following PRISMA-NMA guidelines. Primary outcome was survival, assessed using hazard ratios (HRs) and 95% confidence intervals (95%CI). Survival times were converted to HRs under exponential distribution assumption. LogHR standard errors were calculated using deaths per RCT arm. P-scores were used to estimate probability of treatment being superior. Analysis was performed in R using frequentist approach, followed by back-transforming logHRs for indirect and direct comparisons. Incremental effects of individual treatment components were estimated using additive model. Heterogeneity and inconsistency were assessed using tau-squared (τ²), I-squared (I²), and Q-statistics. Analyses were performed separately for main subnetwork with disconnected subnetworks accommodated in additive model.

RESULTS

Seven studies were included, incorporating 7 pairwise comparisons and 9 treatments. 3 subnetworks were identified. Primary subnetwork analysis included 5 studies. Compared to SysCT±RT (reference), logHRs (95%CI) under random effects model were: IT/ITV DepoCyt±SysCT±RT -0.094(-0.468,0.281); IT/ITV methotrexate(MTX)±cytarabine(AraC)±SysCT±RT 0.8567(0.066,1.647). IT/ITV MTX±SysCT±RT 0.318(-0.163,0.798); ITV Thiotepa±SysCT±RT 0.438(-0.294,1.169). Heterogeneity and inconsistency were not detected (τ²=0,I²=0%,Q-statistic=0.57,P=0.45). P-scores were highest for IT/ITV DepoCyt±SysCT±RT (0.894), followed by SysCT±RT (0.769). Disconnected-component-NMA demonstrated incremental logHR: AraC 0.539(-0.347,1.425); IT/ITV DepoCyt 0.052(-0.566,0.670); IT/ITV MTX 0.179(-0.484,0.840); ITV thiotepa 0.298(-0.767,1.362); proton CSI -0.250(-0.726,0.225). No component demonstrated statistically significant incremental benefit, with moderate heterogeneity and adequate fit present (τ²=0.1017,I²=57%,Q-statistic=4.65,P=0.098).

CONCLUSIONS

IT/ITV DepoCyt±SysCT±RT and SysCT±RT were found most effective. Lack of significant incremental benefit warrants evaluating older therapies in contemporary RCTs.
BIOS-08. CISD: A NOVEL, USER-FRIENDLY MULTIFACETED ALGORITHM FOR DETECTING STITCHING ARTIFACTS IN LARGE-SCALE CODEX IMAGING DATASETS
Authors: Jahedi Afrooz , khan Aliya , Gupta Pravesh , Huse Jason , Kannan Kasthuri , Bhat Krishna
Published: November 2024

BACKGROUND

CODEX (CO-Detection by indEXing) imaging has emerged as a pivotal tool in cellular biology, providing unprecedented insights into cellular distributions and interactions within the tissue microenvironment, crucial for advancing computational pathology. Despite its advantages, CODEX is susceptible to stitching errors during the merging of image tiles, which can significantly distort spatial analysis by introducing artifacts that obscure true cellular architecture.

METHODS

Addressing this challenge, we introduce CISD (CODEX Image Stitch Detection), a novel, user-friendly algorithm optimized for the detection of stitching artifacts in large-scale CODEX datasets. Developed and validated using a comprehensive dataset from 7 matched glioblastoma patients—encompassing over 2800 images across 29 immune protein markers—CISD utilizes a combination of image preprocessing, statistical measures, and thresholding to detect stitching errors.

RESULTS

CISD demonstrates robust performance, achieving a median Area Under Curve (AUC) value in the gold standard dataset where 73% of markers exceed 70% AUC, and 17% surpass 90%. Further validation shows that in a secondary dataset, 45% of markers maintain AUC values over 70% with 38% exceeding 90%, validating the algorithm’s consistent efficacy across varied datasets.

CONCLUSION

CISD’s significance lies not only in its computational achievements but also in its accessibility, facilitating high-quality analysis of extensive datasets without complex pattern recognition requirements. This democratizes high-precision computational pathology, enabling more researchers to accurately analyze spatial data. By reducing technical barriers, CISD sets a new benchmark in CODEX imaging, enhancing data reliability and accelerating research discoveries in cellular biology. This innovation stands to substantially refine spatial analysis in digital pathology, ensuring dependable interpretation of complex biological data.
CNSC-01. PEDIATRIC GLIOMAS EXPRESS CIRCADIAN GENES AND DEMONSTRATE CIRCADIAN REGULATION OF TEMOZOLOMIDE SENSITIVITY
Authors: Nettnin Ella A , Garcia Cesar A , Schonfeld Ethan , Mehl Lindsey C , Guinle Maria Isabel Barros , Petritsch Claudia K , Gibson Erin M , Prolo Laura M
Published: November 2024

INTRODUCTION

Pediatric high-grade gliomas (pHGG) are highly invasive brain tumors with dismal survival rates, prompting the need for innovative therapeutic strategies. Previous research in adult glioblastoma patients revealed a correlation between the timing of Temozolomide administration and patient survival. There has, however, been limited exploration into circadian gene expression in pediatric gliomas. In this study, we investigate whether pHGGs exhibit rhythmic expression of circadian genes and whether the timing of drug administration influences Temozolomide sensitivity.

METHODS

A bioinformatics approach, immunocytochemistry and immunoblotting were used to assess key circadian genes in low- and high-grade gliomas. We then used timed qPCR to assess rhythmic expression in BMAL1 and REV-ERBα, two key circadian genes, across 48 hours in our pHGG cells following dexamethasone cell synchronization. Lastly, we used CyQuant Proliferation Assays to examine the effects of Temozolomide applied at peaks versus troughs in BMAL1 expression.

RESULTS

Using data from PedcBioPortal, we found significantly higher expression of BMAL1 in pHGGs compared to pLGGs (0.349 + 1.05 vs 0.0545 + 0.666, p < 0.001). We identified cellular localization of BMAL1 and REV-ERBα in pHGGs along with rhythmic expression of BMAL1 and REV-ERBα in our three pHGGs primary cell cultures across 48 hours (JTK_CYCLE, p < 0.005). Lastly, we demonstrated a chronotherapeutic response to Temozolomide in two of our pHGG cell lines, with significantly decreased proliferation when treatment occurred at trough versus peak BMAL1 expression (p < 0.01).

CONCLUSION

In this study, we found that circadian genes BMAL1 and REV-ERBα are expressed in pHGGs and expression patterns differ between pHGGs versus pLGGs. Circadian gene expression is rhythmic in pHGGs, and pHGGs can have differential Temozolomide-sensitivity based on circadian cycles. These findings suggest that circadian timing of chemotherapy may influence the efficacy of treatment, and next steps will be to explore the mechanisms underlying circadian regulation of Temozolomide sensitivity.
CNSC-05. MAPPING LANGUAGE PROCESSING IN SURGICAL GLIOMA PATIENTS
Authors: Muir Matthew , Noll Kyle , Prinsloo Sarah , Traylor Jeffrey , Liu Ho-Ling , Kumar Vinodh , Prabhu Sujit
Published: November 2024

Previous work from our group has shown that the resection of white matter tracts localized by transcranial magnetic stimulation (TMS) predicts postoperative neurological deficits. Here we use advanced preoperative brain mapping along with formal language evaluations to localize the neuroanatomical underpinnings of language processing in surgical glioma patients. In patients who had TMS tracts resected, the resected portions (“tract-cavity segmentations”) were classified according to the functional outcome (“true positives”-corresponding deficit; “false positives”-no deficit) and normalized to MNI space. For patients who sustained long-term deficits, TMS tracts were segmented according to spatial overlap with normative tracts to create ROIs. Each normalized resection cavity (n=79) was analyzed for overlap with each ROI associated with postoperative deficits (n=8). The percentage of patients who had positive ROI resections that included TMS tracts versus patients who had positive ROI resections irrespective of TMS tracts were analyzed according to the percent who sustained deficits in each group. ROIs from all patients were summed to create a probabilistic atlas of functional white matter. “True positive” tract-cavity segmentations localized into dual anterior/posterior “hubs” and demonstrated significantly higher spatial association with normative tracts across three different measures (p<.05). 92% of patients who underwent ROI resections that included TMS tracts sustained long-term deficits while 34% of patients who underwent ROI resections irrespective of individualized mapping sustained deficits (p<.05). The probabilistic atlas of functional white matter demonstrated significantly higher voxel intensities within the white matter deep to the temporoparietal junction defined by the same posterior “hub” ROI from Figure 1 (p<.001). This study demonstrates that essential language processing localizes to ROIs within normative white matter tracts that require individualized mapping for surgical preservation. The white matter deep to the temporoparietal junction serves as a crucial hub for language-processing circuitry in glioma patients.
CNSC-07. OLIGODENDROGLIAL IMPAIRMENT IN WHITE MATTER TRACTS FOLLOWING NEUROFIBROMATOSIS TYPE 1 (NF1) GERMLINE MUTATION OR CARBOPLATIN TREATMENT
Authors: Aghoghovwia Benjamin Emoefe , Alnassiry Sama , El-Toukhy Hana , Hamblett Ava , Sharafaldin Samer , Harris Momo , Pan Yuan
Published: November 2024

Neurofibromatosis type 1 (NF1) is a cancer predisposition syndrome due to mutations in the NF1 gene. The disease affects 1 in 2500-3000 people worldwide and patients are predisposed to neoplasms such as optic pathway glioma, plexiform neurofibroma. In addition, NF1 patients often experience neurological issues such as learning difficulties. Unfortunately, there is no cure for NF1 yet. Chemotherapy (e.g., carboplatin) remains the first-line treatment for NF1-associated low-grade glioma; while effective in reducing tumor burden, it does not reliably restore neuronal function. In this study, we sought to determine how chemotherapy and Nf1 germline mutations influence neurological function, using genetically engineered mice of NF1. Nf1-heterozygous mice received carboplatin (60 mg/kg/day) or vehicle every other day for one week. Their optic nerves were collected and processed for immunofluorescence and transmission electron microscopy (EM). We found that carboplatin treatment reduces the density of oligodendrocytes in Nf1-heterozygous mice and impairs myelination. Moreover, a similar effect of carboplatin was observed in wild-type mice, suggesting that carboplatin impairs oligodendroglia independent of the NF1 status and the effect of carboplatin should also be evaluated in the non-NF1 context. In another set of experiments, we sought to understand how germline Nf1 mutations influences oligodendroglia. Whereas we did not observe a difference in optic nerve oligodendrocyte density in mice with an engineered mutation targeting the GTPase-activating protein-related domain (GRD) domain of the Nf1 gene, a reduction of oligodendrocyte density was observed in mice harboring a patient-derived mutation (R1809C), which localizes outside of the GRD. A similar effect was observed in the corpus callosum of Nf1+/R1809C mice together with myelin impairment, suggesting a RAS-independent mechanism of Nf1 regulation of oligodendrocytes. Together, these findings indicate that both Nf1 germline mutations and chemotherapy could contribute to oligodendrocyte and myelin deficits in white matter tracts.
CNSC-08. PROGNOSTIC BIOMARKER DISCOVERY THROUGH PROTEOMIC ANALYSIS OF GLIOBLASTOMA MULTIFORME
Authors: Liu Szu-Yuan
Published: November 2024

BACKGROUND

Glioblastoma multiforme (GBM) is an aggressively malignant brain tumor known for its high recurrence rates and dismal survival outcomes. Despite advancements in surgical and therapeutic strategies, the prognosis for GBM patients remains poor. This study applies proteomic analysis to postoperative GBM tumor specimens to identify molecular markers that could enhance prognosis prediction. Through the analysis of protein expression profiles before and after tumor recurrence, we identified significant alterations in protein pathways via Gene Set Enrichment Analysis (GSEA), highlighting potential biomarkers for disease outcome prediction.

METHODS

We employed Sequential Window Acquisition of All Theoretical Fragment Ion Spectra (SWATH-MS) proteomics to analyze protein samples from both primary and recurrent GBM patients. Gene Set Enrichment Analysis (GSEA) was utilized to compare protein expression profiles and discern significantly altered pathways.

RESULTS

Analysis revealed 778 differentially expressed proteins (DEPs), with notable alterations in pathways such as glycolysis, interleukin signaling, and MAPK signaling. Proteins such as TOLLIP, ENO1, and ITGA2B emerged as potential prognostic markers. Validation through an online tumor prognostic analysis platform (ToPP), integrated with The Cancer Genome Atlas (TCGA) data, facilitated the development of a prognostic risk score model. This model effectively categorized patients into high and low-risk groups based on their protein expression profiles, enhancing the prediction of patient outcomes. The proteins and pathways identified offer new insights into the molecular dynamics of GBM postoperatively. The association between protein expression profiles and patient prognosis underscores the potential of these markers in developing customized therapeutic strategies. Additionally, the prognostic risk score model serves as a valuable tool for improving prognostic accuracy and could guide personalized treatment plans.

CONCLUSION

Proteomic profiling of GBM tumors significantly contributes to uncovering the molecular mechanisms driving this aggressive cancer and facilitates the development of dependable prognostic tools. These findings not only affirm the critical role of proteomics in cancer research but also advance personalized medicine strategies in GBM management.
CNSC-09. OTOSYPHILIS MIMICKING A VESTIBULAR SCHWANNOMA
Authors: Nasany Ruham Alshiekh , Blaquier Juan , de la Fuente Macarena , Benjamin Carolina , Mellon Eric
Published: November 2024

Vestibular schwannomas are common nonmalignant primary brain tumors representing up to 80% of the cerebellopontine angle (CPA) tumors. Their diagnosis is made radiographically in most cases with the presence of an enhancing mass along the 8th nerve complex. The United States is experiencing a surge in the number of recorded syphilis cases with an approximate 80% increase since 2018 according to the CDC, after reaching an all-time low in the year 2000. Syphilis can involve the Central Nervous System (CNS) in its early, late, and latent forms, and in its early stages it typically presents in the form of meningitis which can be accompanied by cranial neuropathies especially of the nerves II, VII, and VIII. We present a case of a 35-year-old man who presented with subacute onset unilateral left-sided hearing loss and dizziness that did not respond to prescribed steroids. Magnetic resonance imaging (MRI) revealed an enhancing mass in the left cerebellopontine angle presumed to be a vestibular schwannoma. The patient was subsequently planned for radiosurgery, however, MRI done during the procedure revealed that the mass was of a smaller size. The patient was referred to neuro-oncology and work up included cerebrospinal fluid (CSF) analysis that showed 3 white blood cells, normal protein and glucose, and negative VDRL. Cytology, flow cytometry, and inflammatory markers were negative. However, the Treponema pallidum antibodies were positive confirming otoshyphilis. The patient was treated accordingly with a course of Penicillin G with near resolution of the enhancement of the 8th nerve complex. His hearing remained impaired on his initial follow up, however, his facial weakness near resolved. This case brings up an important diagnostic question in cases of presumed schwannomas where patients would be otherwise treated solely based on the MRI appearance, especially given the re-emergence of syphilis.
CNSC-17. UPREGULATION OF CCL4 AFTER TREATMENT WITH A NOVEL MRNA VACCINE SHIFTS THE PHENOTYPE OF CCR5-EXPRESSING MICROGLIA WITHIN TUMOR MICROENVIRONMENT
Authors: Chauhan Anjali , Thomas Nagheme , Dastmalchi Farhad , McGrath Kyle , Takacs Gregory Pal , Harrison Jeffrey K , Subramaniam Vignesh , Angelini Thomas , Ebrahim Ghaidaa , Tomdio Macaulay Y , Frain Matthew , Mendez-Gomez Hector , Sayour Elias J , von Roemeling Christine , Mitchell Duane A Mitchell A , Rahman Maryam
Published: November 2024

INTRODUCTION

Intra-tumoral immunosuppression is a major cause of treatment failure for patients with glioblastoma (GBM). Our group has developed a proprietary vaccine that combines mRNA encapsulated in lipid nanoparticles with PEG hydrogel and a chemokine (CXCL9). This hydrogel-chemokine-mRNA (HCM) vaccine is delivered subcutaneously (SQ) and results in significant anti-tumor efficacy in syngeneic murine glioma models. The objective of this study was to evaluate the effects of the vaccine on the tumor microenvironment (TME).

METHODS

C57BL/6 mice underwent intracranial implantation of KR158-luc tumor cells and underwent SQ injection of the HCM vaccine using total tumor mRNA. Tumors were collected for analysis at different time points. Q-PCR and flow cytometry were done to evaluate the differential gene expression and CCR5 expression (receptor for CCL4) in the TME following HCM vaccination.

RESULTS

CCL4 was found to be upregulated in tumor-bearing animals treated with HCM vaccine using PCR. In a separate experiment, CCL4 blockade was given prior to HCM vaccination and this resulted in abrogation of the survival benefit of the vaccine. Next, flow cytometry results revealed that microglia in the TME had the highest expression of the CCR5 receptor. Further, QPCR on the isolated microglia from TME revealed significant up-regulation of inflammatory gene CCL5 (25 fold) and down-regulation of anti-inflammatory genes as compared to untreated control and empty hydrogel control.

CONCLUSION

The HCM vaccine results in significant anti-tumor efficacy in murine syngeneic murine glioma which is dependent on upregulation of CCL4 in the TME. The CCL4 exerts its effect through microglia expressing CCR5 in the TME. HCM vaccination shifted the microglia phenotype from immune-suppression to inflammatory state. Further analysis of the effect of CCR5 knock out on the HCM vaccine survival efficacy are under current investigation.
CNSC-20. MONOSYNAPTIC TRACING DEFINES BRAIN-WIDE CIRCUIT CONNECTIVITY OF HUMAN GLIOBLASTOMA
Authors: Sun Yusha , Wang Xin , Zhang Daniel , Zhang Zhijian , Bhattarai Janardhan , Wang Yingqi , Park Kristen , Dong Weifan , Yang Qian , Zhang Feng , Rajamani Keerthi , Mu Shang , Kennedy Benjamin , Hong Yan , Galanaugh Jamie , Sambangi Abhijeet , Kim Sang Hoon , Wheeler Garrett , Goncalves Tiago , Wang Qing , Geschwind Daniel , Kawaguchi Riki , Viaene Angela , Helbig Ingo , Kessler Sudha , Hoke Ahmet , Wang Huadong , Xu Fuqiang , Binder Zev , Chen Isaac , Pai Emily Ling-Lin , Stone Sara , Nasrallah MacLean , Christian Kimberly , Fuccillo Marc , Toni Nicolas , Wu Zhuhao , O’Rourke Donald , Ma Minghong , Ming Guo-li , Song Hongjun
Published: November 2024

Glioblastoma (GBM), a deadly brain cancer, infiltrates the brain and can be synaptically innervated by neurons. Synaptic inputs onto GBM cells identified so far are largely short-range and glutamatergic in nature. The extent of integration of GBM cells into the brain-wide neuronal circuitry is therefore not well understood. We report the application of transsynaptic viral tracing approaches to study the neuronal connectome of GBM. We applied rabies virus-mediated retrograde tracing and herpes simplex virus (HSV)-mediated anterograde tracing approaches to characterize presynaptic partners in vivo in the adult mouse brain and ex vivo with iPSC-derived neurons, cortical organoid-GBM assembloids, and human surgical specimens. After transplantation into adult mice, GBM cells derived from multiple patients rapidly integrated into brain-wide neuronal circuits. GBM exhibited increased connectivity rates compared to transplanted neural progenitor cells, highlighting functional connectivity as a hallmark of malignant cells. Beyond glutamatergic inputs, we identified neuromodulatory inputs across the brain, including cholinergic inputs from the basal forebrain. We validated these long-range cholinergic neuron-to-glioma synapses signaling through the metabotropic CHRM3 receptor by high-resolution microscopy, anterograde monosynaptic HSV tracing, calcium imaging, and patch-clamp electrophysiology. To interrogate the functional effects of acetylcholine (ACh) on GBM, we performed calcium imaging and RNA sequencing analyses, which showed that ACh stimulation induced sustained calcium oscillations and long-lasting transcriptional reprogramming of GBM cells into a more invasive state via CHRM3. Importantly, CHRM3 activation promoted GBM cell invasion, whereas CHRM3 downregulation suppressed GBM cell invasion in vitro and in vivo, suggesting CHRM3 as a potential therapeutic target for this disease. Together, these results reveal the capacity of human GBM cells to robustly integrate into anatomically and molecularly diverse neuronal circuitry in the adult brain. They also support a model wherein rapid synapse formation onto GBM cells may promote a long-lasting increase in tumor cell fitness.
CNSC-26. INVESTIGATING THE ROLE OF QUAKING IN REGULATING MHC II ANTIGEN PRESENTATION IN TUMOR-ASSOCIATED MACROPHAGES OF GBM
Authors: Hwang Spring , Zeng Ailiang
Published: November 2024

Glioblastoma (GBM) is the most common and malignant primary brain tumor with a median survival rate of 14 months. Despite the success of immunotherapy in treating various cancer types, its efficacy remains poor in GBM. Such limitation can be attributed to the profound immunosuppressive tumor microenvironment of GBM enriched with the tumor-associated macrophages (TAMs). Quaking (Qki) is an RNA-binding protein that is mutated or deleted in more than 35% of GBM patients. Decreased levels of Qki in TAMs associates with worsened survival rates and responsiveness to immunotherapy in GBM mouse and patients. Moreover, Qki is a tumor suppressor gene in GBM and a major regulator in phagocytosis, a fundamental biological process in delivering antigens to major histocompatibility complex class II (MHC II). MHC II-restricted antigen presentation in macrophages of GBM is important in maintaining the fitness of cytotoxic and helper T cells required in anti-tumor response. However, the mechanism through which Qki regulates MHCII expression and presentation has yet to be elucidated. I hypothesize that Qki promotes MHC II expression in TAMs, thereby constraining the development and progression of GBM. In vitro assays like immunofluorescent staining, qRT-PCR, cell co-culture system, and flow cytometry will be performed involving QPP (Qk-/-Trp53-/-; Pten-/-) mouse brains, QPP cells, and mouse macrophage cell lines. A positive correlation between transcriptomic and protein expression levels of Qki, MHC II, and CIITA (a master regulator of MHC II) was confirmed in both mouse QPP and human GBM samples. Moreover, bone-marrow-derived macrophages activated by QPP cell supernatant shows upregulated levels of Qki and thus MHC II genes. A preliminary conclusion is that Qki plays a permissive role in MHCII expression in TAMs of GBM. Further downstream impact of Qki in TAMs will be evaluated in T cell activity and in transcription of MHC II in TAMs.
CNSC-37. ROLE OF PGE2-REGULATED NEUROINFLAMMATION IN CONSTRUCTING NEURON-NK-GLIOMA CIRCUIT FOR GLIOBLASTOMA
Authors: Chen Hsien-Chung , Chang Wen-Chang , Hsu Tsung-Yi
Published: November 2024

Glioblastoma (GBM), known for its aggressive nature and poor prognosis, frequently recurs and resists standard treatments such as temozolomide (TMZ). This resistance is partly due to complex interactions within the tumor microenvironment, particularly involving neuronal excitation. Recent insights suggest that neuronal activity, through mechanisms like neurogliomal synapses, not only promotes glioma cell proliferation but also contributes to the chemoresistance observed in recurrent tumors. Our focus is on Prostaglandin E2 (PGE2), a mediator that regulates neuronal excitation, and its role in the recurrence and resistance of GBM. We discovered that PGE2 upregulates synaptic proteins and alters neurotransmitter profiles, enhancing tumor cell survival and resistance to TMZ. Specifically, increased levels of neurotransmitters such as glutamine and asparagine were observed in PGE2-rich environments, correlating with heightened chemoresistance and tumor recurrence. Co-culture experiments further demonstrated that PGE2-treated neuronal cells could induce resistance in adjacent GBM cells. These findings highlight the potential of targeting PGE2 pathways as a therapeutic strategy. A significant advancement in our study was the development and application of compound #038, a novel celecoxib derivative designed to cross the blood-brain barrier (BBB). This compound demonstrated enhanced efficacy in inhibiting GBM growth and improving survival rates in mouse models, indicating its potential as an effective therapeutic agent against GBM.