bluuebunny/crossref_metadata_2025_split huggingface dataset:
TMIC-22. MYELOID CELLS INDUCE IMMUNOSUPPRESSION AND HYPORESPONSIVENESS OF CHIMERIC ANTIGEN RECEPTOR T CELLS IN THE NEURONAL MICROENVIRONMENT OF GLIOBLASTOMA
Authors: Zhang Junyi , von Ehr Jasmin , Look Thomas , Benotmane Jasim Kada , Neidert Nicolas , Kueckelhaus Jan , Weiss Tobias , Heiland Dieter Henrik , Abubakar Yabo Yahaya 
Published: November 2024

Abstract
Chimeric antigen receptor T (CART)-cell therapy has shown little success in glioblastoma and other solid tumors, unlike in hematologic malignancies. The immunosuppressive milieu and intratumoral heterogeneity are some obstacles limiting the effectiveness of CART-cell therapy in glioblastoma. Our goal in this study is to investigate how CART-cell transcriptional adaptations and cellular interactions in the glioblastoma microenvironment drive T-cell hyporesponsiveness. Using a human neocortical slice model, we performed physically interacting cell sequencing (PIC-seq) to investigate the transcriptional diversity of NKG2D CAR-T cells and control T-cells in the glioblastoma ecosystem. Through the integration of spatially resolved T-cell receptors sequencing and PIC-seq, we investigated the cellular and receptor-ligand interactions in CART-treated glioblastoma cells. Gene regulatory networks were reconstructed to identify key transcriptional regulators of CD8 CART-cell dysfunction. We used in-silico perturbation to understand transcriptional drivers that could potentially enhance the cytotoxic response of CAR-Ts. Primary patient-derived glioma cells were inoculated into human neocortical slices from surgical access tissue specimens. After 3 days of tumor growth, slices were treated with NKG2D CARTs or control T-cells. We observed that after 2 days of anti-tumor T-cell response in the CART group, tumor growth did not differ between conditions. After 7 days of treatment, PIC-seq was used to profile CARTs (tomato+) and tumor cells (GFP+) as well as physically connected cells (+/+). We identified a transcriptional shift towards T-cell exhaustion in CART cells compared to T-cells lacking the NKG2D construct. This transcriptional phenotype was driven by myeloid-CART and myeloid-tumor interaction. We identified that tumor-associated macrophages with enhanced phagocytic function in proximity to mesenchymal-like tumor cells located within hypoxic niches are the main drivers of T-cell dysfunction. Gene regulatory network analysis demonstrated that MAF and BACH2 are key transcriptional regulators of CART-cell function. Myeloid-tumor and myeloid-T-cell interactions promote an immunosuppressive microenvironment and hyporesponsive T-cells in glioblastoma. Our data has the potential to catalyze the re-engineering and optimization of CART-cell therapies with sustained cytotoxic effect against glioblastoma.
TMIC-23. INVESTIGATING THE FUNCTIONAL ROLE OF SECRETED FERRITIN FROM GLIOBLASTOMA CELL TYPES
Authors: Tufano Emily , Palsa Kondaiah , Serpa Rebecka , Connor James 
Published: November 2024

Abstract
Glioblastoma (GBM) is the most common primary malignant brain tumor in adults with a survival rate ~5%. Our laboratory previously showed the inhibitory effect of iron and ferritin-bound iron on migration and spheroid formation in GBM cells. Furthermore, studies have shown the release of extracellular vesicles (EVs) and their iron-related cargoes are regulated by cellular iron concentrations. Building on these findings, this study aims to investigate the impact of EV-associated ferritin on migration in GBM. To achieve this, we examined the influence of iron on EV release from four GBM cell lines, comprising two cancer stem cells (CSCs) and two non-CSCs: T387, T3691, LN229, and T98G. Using Nanoparticle tracking analysis (NTA), we demonstrated that 100 μM of ferric ammonium citrate (FAC) significantly increases EV release in the CSC line after 24 hours (p-value &lt;0.01). Conversely, there was no significant impact on the non-CSC line under the same conditions. Based on these results, the ability of iron to regulate EV release is dependent on CSC versus non-CSC status. Given the functional role of ferritin on migration potential in GBM, we used immunoblotting to examine ferritin release from cells following iron treatment. We observed a reduction in ferritin release from CSCs following FAC treatment compared to the control, while non-CSCs, in contrast, released more ferritin after treatment. Interestingly, ferritin released from CSCs was predominantly within EVs, whereas ferritin released from non-CSCs was not associated with EVs. To assess the functional impact of ferritin release on migration, we utilized a Transwell migration system, and observed that EVs from iron-treated cells reduced migration in the recipients. Together, these findings expand our understanding of differential responses of ferritin in CSC and non-CSC EVs to iron treatment and will allow us to identify new targets and strategies for managing GBM progression.
TMIC-32. INFIGRATINIB INHIBITION OF THE THBS1 PATHWAY IN MELANOMA BRAIN METASTASIS
Authors: Tran Andrew , Choi John , Bergsneider Brandon , Kim Lily , Schonfeld Ethan , Abikenari Matthew , Verma Rohit , Cho Kwang Bog , Lim Michael 
Published: November 2024

Abstract

INTRODUCTION
Despite promising preclinical studies, BRAF/MEK inhibition with vemurafenib in melanoma brain metastases (MBM) has suboptimal durable response in humans. Thrombospondin 1 (THBS1), a matricellular protein, is upregulated in vemurafenib-resistant metastatic melanoma, and its inhibition reverses BRAF inhibitor chemoresistance. However, the characteristics of THBS1-mediated signaling pathways as well as methods of inhibition require further exploration in the context of MBM.

OBJECTIVE
To characterize THBS1-mediated signaling pathways in MBM and evaluate the therapeutic potential of targeting THBS1 pathways.

METHODS
We analyzed human parenchymal MBM samples using publicly available single-cell RNA sequencing (scRNA-seq) data. Protein-protein interaction networks were assessed using the STRING database. Syngeneic B16 melanoma cells were intracranially implanted in C57BL/6J mice and treated with intraperitoneal injection of FGF7/FGFR2 small-molecule inhibitor Infigratinib or PBS (sham). Mice were immunophenotyped using flow cytometry and monitored for survival.

RESULTS
THBS1 is highly upregulated in immunosuppressive MBM-associated cell populations such as S100A8+ Metastasis-Associated Macrophages and Mesenchymal Stromal Cell-like cells. Gene set enrichment analysis of THBS1-positive cells paradoxically revealed upregulation of inflammation-associated gene sets. Differential expression analysis identified Fibroblast Growth Factor 7 (FGF7) as strongly co-expressed in THBS1-positive MBM cells, and STRING-DB confirmed close protein-protein interactions between FGF7 and THBS1. In vivo inhibition using Infigratinib for FGF7’s receptor, FGFR2, decreased the proportion of monocytic myeloid-derived suppressor cells (M-MDSCs) (P=0.030), increased T-cell activation marker IFN-y (P=0.027) and proliferation marker CD69 (P=0.047), and resulted in improved overall survival (P=0.0001).

CONCLUSION
MBM scRNA-seq data suggests that THBS1 is associated with inflammation-induced immunosuppression. Inhibiting the FGF7/FGFR2-THBS1 axis with Infigratinib modulates the tumor microenvironment by reducing immunosuppressive M-MDSCs and increasing T cell activation and proliferation. This data supports THBS1 inhibition as a viable therapeutic option that in the future can also be combined with other current immunotherapeutic strategies.
TMIC-34. INFILTRATIVE BEHAVIOR OF PATIENT-DERIVED GLIOBLASTOMA CELL LINES IN REGION-SPECIFIC HUMAN BRAIN ORGANOIDS
Authors: Ganta Saisrinidhi , Bhatia Tarun , King Alexia , Sing Anson , Read Renee , Sloan Steven 
Published: November 2024

Abstract
Glioblastoma (GBM) is the most common malignant brain tumor in adults. GBMs are known for their highly invasive and proliferative properties and are associated with a dismal prognosis (~12-18 months post-diagnosis). Neuroanatomical disparities in the occurrence of GBMs suggest that in addition to microenvironmental impacts, tumor cells are impacted by the cellular and molecular diversity across varying regions of this nervous system. This idea has been difficult to study due to the lack of human-specific models that effectively recapitulate the biological behaviors of GBMs in region-specific microenvironments. To address these challenges, we engrafted GFP-tagged patient-derived glioma cell lines into human induced pluripotent stem cell (hiPSC)-derived organoids patterned to mimic the most forebrain (dorsal cortex) and hindbrain (spinal cord) regions of the nervous system. Immunostaining assays at 2, 7, 14, and 21 days post-engraftment compared the extent of infiltration, as measured by the distance traveled by glioma cells, in both regions. We found that glioma cells infiltrated a significantly greater distance in the dorsal cortical organoids compared to the spinal cord organoids. This suggests a more conducive environment for glioma infiltration in the forebrain vs. hindbrain at baseline. We are currently determining if these regional biases in infiltration capacity are modified with either optogenetic stimulation of the host brain organoids or in the presence of interregional interactions in an assembloid system. Thus, using this fully human-specific platform, we aim to gain insights into the infiltrative behaviors of GBM cells, which will advance our understanding of this devastating disease.
TMIC-35. MITOCHONDRIAL TRANSFER PROMOTES ASTROCYTE REACTIVITY IN GLIOBLASTOMA
Authors: Mitchell Jonathan , Bayik Defne , Watson Dionysios 
Published: November 2024

Abstract
Astrocytes are the most abundant glial cell in the brain and have been shown to adopt reactive phenotypes upon interaction with glioma cells. While tumor-associated astrocyte (TAA) reactivity is linked to increased inflammation, proliferation, invasion, and treatment resistance of glioblastoma (GBM), the mechanisms by which cancer cells reprogram TAA are poorly defined. We have previously shown that mitochondrial transfer from astrocytes to GBM cells via GAP43-dependent microtubes alters GBM cellular metabolism and increases tumorigenicity. However, the potential role of this communication mechanism in TAA function remains unknown. Patient-derived GBM cells (PDCs) were transduced with a mito-GFP lentiviral vector and were co-cultured with hTERT-immortalized, mito-mCherry human astrocytes. Using flow cytometry, we observed that the frequency of mitochondrial transfer to hTERT astrocytes was dependent on GBM cell type and noted transfer rates as high as ~40% in co-cultures with L1 PDCs. Utilizing high-resolution confocal microscopy, we subsequently confirmed co-localization of fluorescent protein signal with MitoTracker Deep Red and observed increases in mitochondrial biomass in transfer-positive astrocytes. To evaluate whether mitochondrial transfer occurs in the tumor microenvironment, we implanted immunocompromised mice with L1 mito-mCherry PDCs. Approximately 17.7% of mouse TAAs in the tumor core were positive for GBM mitochondria, whereas long-distance transfer was not observed in the contralateral hemisphere. Functionally, acquisition of tumor mitochondria promoted astrocyte proliferation as measured by cells in G2/M phase and significantly increased total cellular reactive oxygen species (ROS) levels. These findings highlight a process by which astrocytes may become reactive in response to increased ROS from GBM mitochondrial transfer. Future studies will investigate the cellular response to increased ROS in astrocytes and investigate potential links to inflammatory and immunosuppressive signaling. Elucidating how GBM-to-astrocyte mitochondrial transfer reprograms TAA would help identify therapeutic vulnerabilities that can be exploited by blocking mitochondrial transfer.
TMIC-39. SPATIAL MULTIOMICS PROFILING REVEALS HETEROGENEITY OF B:T CELL INTERACTIONS AND PLASMA CELL FORMATION IN TERTIARY LYMPHOID STRUCTURES IN HUMAN GLIOMAS
Authors: Cakmak Pinar , Lun Jennifer H , Singh Aakanksha , Macas Jadranka , Burger Michael , Hasse Lucie Marie , Hattingen Elke , Köhler Miriam , Schupp Jonathan , Starzetz Tatjana , Steidl Eike , Plate Karl H , Reiss Yvonne , Imkeller Katharina 
Published: November 2024

Abstract
Adult-type diffuse gliomas, the most common primary brain tumours, remain a clinical challenge in oncology with limited treatment options, restricted anti-tumour immune response and a dismal patient prognosis. Despite their immunosuppressive microenvironment, the formation of lymphoid aggregates containing adaptive immune cells has been reported in gliomas. However, the cellular compositions, immunological function, and relevance of lymphoid aggregates for adaptive anti-tumour immunity is not well understood. Therefore, we performed a comprehensive, unbiased analysis of lymphoid aggregation in 642 adult-type diffuse gliomas using a multi-modal approach, combining DNA methylation and RNA sequencing with spatial transcriptome and proteome profiling. Overall, B cell aggregates and tertiary lymphoid structures (TLS) were observed in 15% of tumours, with the highest prevalence in primary glioma localised to the temporoparietal lobe. The presence of TLS was associated with an improved overall survival. Gliomas containing TLS displayed a remodelled perivascular space, characterised by transcriptional upregulation and spatial redistribution of collagens associated with barrier functions. Spatial transcriptome and proteome profiling revealed heterogenous B:T cell interactions that were associated with elevated CD8 T-cell numbers and differences in IgA and IgG plasma cell forming capacity, suggestive of dynamic adaptive immune response.
TMIC-40. TARGETING GLIOMA TUMOR MICROENVIRONMENT BY SPECIFIC NEUTRALIZATION OF EXTRACELLULAR NICOTINAMIDE PHOSPHORIBOSYLTRANSFERASE (ENAMPT) USING ALT-100, A NOVEL HUMANIZED MONOCLONAL ANTIBODY
Authors: Sharma Pratibha , Saindane Madhuri , Zhang Katherine , Xu Jihong , Huo Longfei , Ju Zhenlin , Miele Stan , Garcia Skip , Puduvalli Vinay 
Published: November 2024

Abstract

BACKGROUND
Intracellular nicotinamide phosphoribosyltransferase (iNAMPT is the rate-limiting enzyme of NAD salvage pathway and is preferentially used by glioma cells for regulating energy metabolism and homeostasis. When secreted, extracellular NAMPT (eNAMPT) exerts cytokine-like functions and is associated with poor prognosis in cancer patients. We have previously reported an essential role of iNAMPT for glioma cell proliferation and survival. Here, we examined whether eNAMPT influences glioma biology and tumor microenvironment by neutralizing eNAMPT using ALT-100, a novel humanized monoclonal antibody.

METHODS
Native levels of eNAMPT in blood samples of patients with primary/recurrent gliomas were determined using ELISA under an IRB approved protocol. Next, the effects of ALT-100 on glioma microenvironment were determined by treating viable human glioma tissue slices with ALT-100 and assessing downstream effects on RNA and protein using RNASeq, reverse phase protein array and spatial transcriptomics. Lastly, the in vivo effect of ALT-100-mediated eNAMPT neutralization on survival was determined in a mouse intracranial glioma model.

RESULTS
Measurable levels of eNAMPT were noted in all patient samples with a range of (3.7-71.2 ng/ml). Preliminary results showed a correlation of eNAMPT levels with grade of tumor. Human organotypic glioma slice cultures treated with ALT-100, showed a profound alteration of gene and protein expression particularly related to key energy metabolism and proliferation pathways and the PD-1/PD-L1 axis. Additionally, results of spatial profiling of ALT-100 treated human organotypic glioma slices will be presented. Survival and response data from ALT-100 treatment in ongoing in vivo experiment will also be reported.

CONCLUSIONS
This is the first report of ALT-100-mediated eNAMPT neutralization in gliomas and demonstrates profound effects on the tumor and its microenvironment. Given that ALT-100 is in phase 2 trials (NCT05938036) against ARDS, data from our studies can potentially accelerate translating ALT-100-mediated eNAMPT neutralization to treat gliomas.
TMIC-44. ANDROGEN LOSS WEAKENS ANTI-TUMOR IMMUNITY AND ACCELERATES BRAIN TUMOR GROWTH
Authors: Lee Juyeun , Lathia Justin , Chung Yoon-Mi , Sharifi Nima , Curtin Lee , Swanson Kristin 
Published: November 2024

Abstract
Many cancers, including glioblastoma (GBM), exhibit a male-biased incidence and outcome disparity. The underlying reasons for this sex bias are unclear but likely involve differences in tumor cell state and immune response. This effect is further influenced by sex hormones, including androgens, which have been shown to inhibit anti-tumor T cell immunity in other solid tumors. Using a series of pre-clinical models, we observed that androgens drive anti-tumor immunity in brain tumors, contrasting with their effect in other tumor types. Upon castration, tumor growth was accelerated in GBM and brain tumor models, while the opposite effect was observed when tumors were located outside the brain. This castration effect could be reversed by testosterone administration. Flow cytometry revealed decreased T cell function, indicated by reduced anti-tumor cytokines (IFNγ and TNFα), and increased T cell exhaustion in both tumors and peripheral tissues in castrated mice, suggesting systemic immunosuppression. Indeed, the survival difference was completely abrogated in T cell deficient mice (RAG1 knockout). Mechanistically, increased hypothalamus-pituitary-adrenal (HPA) axis activity in castrated mice was responsible for the observed systemic immunosuppression, particularly in those with brain tumors. Blocking glucocorticoid receptors reversed the accelerated tumor growth in castrated mice, indicating that elevated glucocorticoid signaling mediated the castration effect. This mechanism was brain-specific rather than GBM-specific, as hyperactivation of the HPA axis was also observed with intracranial implantation of non-GBM tumors. Neuroinflammation caused by brain tumors was stronger in castrated mice, and proinflammatory cytokines like IL-1 and TNF induced HPA axis hyperactivation. Together, these findings highlight that brain tumors drive distinct endocrine-mediated mechanisms in the androgen-deprived setting and highlight the importance of organ-specific effects on anti-tumor immunity.
TMIC-47. HUMAN BLOOD-BRAIN TUMOR BARRIER ON A CHIP TO INVESTIGATE PERSONALIZED TREATMENT FOR GLIOBLASTOMA PATIENTS
Authors: Lim Jaejoon , Cho Sujin , Lee Gaeun , Ryoo Minsu , Kim Yu Jin , Sim JeongMin , Kim Dongkil , Kang Youn-Jung , Ahn Song Ih , Ahn Jungho 
Published: November 2024

Abstract
The inherent characteristics of glioblastoma multiforme (GBM), including tumoral heterogeneity and invasive capacity, combined with the presence of the blood-brain tumor barrier (BBTB), present challenges in developing effective treatment for GBM. Especially, the margins of GBM, where GBM cells infiltrate normal brain tissue, exhibit high resistance to therapies. Despite the difficulties in controlling tumor progression from this region, the GBM margin remains a critical area to be studied. Here we report a microengineered model that mimics the BBTB within the GBM margin, incorporating a 3D network of normal astrocytes and GBM cells isolated from patients newly diagnosed with GBM. The interaction between GBM cells and stromal cells results in increased vascular permeability, reactive gliosis, and alterations in astrocyte behavior to foster tumor invasiveness and progression. We compare patient-specific tumor responses to conventional chemotherapy in our BBTB on a chip model with clinical outcomes, demonstrating the capability of the model to predict personalized drug responses. Our BBTB model may serve as a personalized tool to examine the interactions between tumors and normal brain tissue, ultimately facilitating the screening of personalized medicine for GBM treatment.
TMIC-49. TUMOR-DERIVED SPERMIDINE DRIVES IMMUNE SUPPRESSION IN THE GLIOBLASTOMA MICROENVIRONMENT VIA CD8 T CELL DEPLETION AND FUNCTIONAL ATTENUATION
Authors: Lathia Justin , Kay Kristen , Lee Juyeun , Wang Zeneng , Grabowski Matthew , Brown Mark , Hazen Stanley , Bayik Defne 
Published: November 2024

Abstract
A hallmark of glioblastoma (GBM) is an immunosuppressive microenvironment that is partially driven by tumor cell-derived secreted factors, highlighting the need for a more complete understanding of local immune suppressive mechanism. Altered secreted metabolite levels are observed in the GBM microenvironment; specifically, spermidine (a polyamine), which functions to drive cell proliferation, apoptosis, T cell lineage commitment, and myeloid cell-mediated suppression. However, the direct link between GBM cell-derived spermidine and the tumor microenvironment has yet to be determined. In syngeneic mouse GBM models, spermidine levels were increased in the tumor microenvironment and exogenous administration of spermidine accelerated tumor aggressiveness in an immune dependent-manner. Mechanistically, spermidine administration increased apoptosis of CD8+ T-cells and generated an exhausted phenotype. When ornithine decarboxylase (ODC1, the rate limiting enzyme in spermidine synthesis), was knocked down in tumor cells, no direct effect was observed on tumor cell growth. However, there was an increase in time to tumor succumbence and a concomitant increase in CD8+ T-cells and decrease in CD8+ T cell exhaustion. In GBM patients, there was a negative correlation between ODC1 expression and CD8+ T-cells present near the tumor. Additionally, patients with a favorable prognosis had significantly lower intratumoral levels of spermidine compared to patients with a poor prognosis. Taken together, these results demonstrate that spermidine functions as a tumor cell-derived metabolite that drives tumor progression through reducing CD8+T cell number and altering their phenotype. These findings have clinical implications as clinical trials using a polyamine biosynthesis inhibitor alone have not been very effective in GBM patients, suggesting the need for additional combination immunotherapies. Finally, spermidine is also produced at prominent levels by commensal gut microbes as well as absorbed from consumption of foods such as mushrooms; investigation into how these distal spermidine sources impact GBM growth via the gut-brain axis is actively ongoing.