bluuebunny/crossref_metadata_2025_split huggingface dataset:
TMIC-15. AGE-RELATED MARKERS FOR SENESCENCE INCREASE IN THE OLDER ADULT EXTRATUMORAL MOUSE BRAIN DUE TO GLIOBLASTOMA
Authors: Bell April , Johnson Margaret , Ladomersky Erik , Kim Miri , Zhao Junfei , Miska Jason , Zhai Lijie , Lauing Kristen , Asimakidou Evridiki , Bommi Prashant , Chandler James , Mi Xinlei , Kocherginski Masha , Lukas Rimas , Wainwright Derek 
Published: November 2022

Abstract

OBJECTIVE
The median age of onset for glioblastoma (GBM; IDHwt) is 68-70 years. Age is a strong prognostic factor for GBM patient outcomes such that overall survival in older adults is less than their younger counterparts – even after adjustment for MGMT promoter methylation status. Aging is associated with increased levels of senescence in the brain. Several age-related neurological disorders have been shown to improve with senolytic treatments. Here, we explored the effects and therapeutic neutralization of syngeneic brain tumors on increasing senescence levels in the extratumoral brain (ie. outside of the brain tumor) of young and old mice.

METHODS
General RNA-sequencing, as well as single-cell (sc) RNA-sequencing was performed on extratumoral tissue from young (8-12 weeks) and older adult (80-90 weeks) C57BL/6 mice with or without GL261 and key markers were validated with RT-PCR. The combined effects of the senolytics, dasatinib and quercetin, with radiation, anti-PD-1 mAb, and IDO enzyme inhibitor treatment, was also investigated.

RESULTS
General- and sc-RNA sequencing revealed a distinct gene expression profile in the extratumoral brain of older mice with syngeneic GL261 as compared to all other groups. RT-PCR results confirmed that the brain tumor increased gene expression for senescence levels, p53, and NFkB signaling in the older adult extratumoral brain. Expression of the senescence marker p16INK4A was primarily localized to oligodendrocyte progenitor cells in the older adult brain. The combinatorial treatment of senolytics with RT, anti-PD-1 mAb, and IDO enzyme inhibitor led to a synergistic survival benefit in older adult mice with GL261 as compared to the treatment with senolytics, or immunotherapy, alone.

CONCLUSIONS
The data suggest that the extratumoral brain may be responsible in-part for the poorer outcomes of older adults with GBM and that treatment approaches that target senescent cells may provide clinical benefit.
TMIC-21. THE ROLE OF CLEC5A ON M2-LIKE TUMOR-ASSOCIATED MACROPHAGES POLARIZATION AND DISEASE PROGRESSION IN GLIOBLASTOMA
Authors: Li Jiabo , Wang Xuya , Markwell Steven , Carneiro-Lobo Tatiana , Olson Cheryl , Shih Ling-Kai , Tong Luqing , Yang Xuejun , Brat Daniel 
Published: November 2022

Abstract
Glioblastoma, IDH-wildtype (GBM, WHO grade 4) is the most common primary intracranial malignant tumor of adults and is characterized by an immunosuppressive tumor micro-environment (TME) and poor clinical outcomes. CLEC5A is a protein-coding gene involved in inflammatory and infectious diseases, where its activation enhances immune response. However, its role in GBM immune regulation is unclear. Seurat analysis of scRNA-seq data and CIBERSORTx analysis of GBM RNA-seq data showed that M2-like TAMs accounted for the largest proportion of immune cells in the GBM TME. Among CD163+ cells (M2-like TAM marker) in GBM, CLEC5A expression was most highly associated with overall patient survival. IHC on tumor microarrays (TMA) and analysis of Ivy GAP data indicated that CLEC5A and M2-like TAMs were preferentially expressed in tumor peri-necrotic zone. THP-1 cells exposed to GBM conditioned media showed increased CLEC5A and CD163 expression by RT-qPCR and Western blot compared to other immune-activating factors (IL-4, IL-13). CLEC5A overexpression in THP-1 cells enhanced their migration toward glioma cells in vitro, and also led to increased M2-like TAM biomarker expression. Cytokine antibody microarray results revealed that overexpressing CLEC5A in THP-1 cells also significantly increased the expression of IL-1β, IL-4, IL-6, IL-8, IL-10, IL-13, GM-CSF, and other cytokines, which all contribute to the immunosuppressive TME. In TCGA GBM data, tumors with higher CLEC5A expression were enriched in IL6-JAK-STAT3 signaling. In vivo, silencing CLEC5A delayed glioma growth and prolonged survival of tumor-bearing mice, and analysis of tumors showed reduced expression of CLEC5A, CD163 and Ki-67. Collectively, CLEC5A is expressed by M2-like TAMs and enhances M2-like TAM polarization, likely through IL6-JAK-STAT3 signaling.
TMIC-23. EXPLORING IBRUTINIB'S EFFECTS ON BLOOD-BRAIN BARRIER INTEGRITY AND GLIOMA PROGRESSION
Authors: Dalmage Mahalia , Kundu Nabanita , Kwak Minhye , Lim Sanghee , Jackson Sadhana 
Published: November 2022

Abstract

BACKGROUND
The blood-brain barrier (BBB), among malignant gliomas limits certain agents from impairing tumor growth. Thus, identifying agents that can transiently increase BBB permeability may prove useful in enhancing glioma treatment response. Previous ibrutinib studies showed it independently disrupted gut epithelial integrity and hampered glioma growth through BMX (bone marrow x-linked tyrosine kinase) inhibition. We propose BMX inhibition via ibrutinib disrupts BBB integrity while impairing glioma progression.

METHODS
To evaluate ibrutinib’s effect on brain endothelium, we evaluated electrical cell-cell impedance, junctional/cytoskeletal expression, downstream protein expression and functional ABC transporter activity; monitoring changes over time (0 to 8h) and at varied drug concentrations (1-10μM ibrutinib). Using rat glioma cells (S635) in vitro and in models, we examined cytotoxicity, apoptosis, model survival, and drug concentrations with ibrutinib alone or combined with poorly permeable Abcb1 substrate doxorubicin.

RESULTS
Ibrutinib dose-dependently decreased brain endothelial cell-cell impedance by 60% at 2h, without affecting cell viability. We observed decreased ZO-1 junctions, actin cytoskeletal rearrangement, and downstream pErk and pMek decreased expression optimally 2h (10μM ibrutinib). Dose-dependently ibrutinib inhibited Abcb1 efflux activity, further favoring a more permeable endothelium. Synergy with doxorubicin was seen in rat glioma cells treated with ibrutinib via increased apoptosis and cytotoxicity. However, while combined treatment resulted in decreased systemic doxorubicin concentrations, no differences in brain:plasma or tumor:plasma concentrations were evident. Combination therapy also did not increase rodent glioma model survival nor decrease tumor size.

CONCLUSION
Our results suggest that while ibrutinib induces brain endothelial permeability by junctional/cytoskeletal disruption and inhibition of Abcb1 efflux, no sustained effect can be seen in rodent glioma models in combination with a cytotoxic agent. Additional studies exploring similar agents that can enhance BBB permeability while slowing glioma growth are warranted, so as to improve upon current bleak malignant glioma treatment options.
TMIC-26. USING INTEGRATED MULTIDIMENSIONAL MASS CYTOMETRY AND MULTIPLEX IMMUNOHISTOCHEMISTRY TO INFER SPATIAL RELATIONSHIPS BETWEEN PHENOTYPICALLY DISTINCT GLIOBLASTOMA INFILTRATING IMMUNE CELLS
Authors: Bartkowiak Todd , Brockman Asa , Lima Sierra , Hayes Madeline , Roe Caroline , Sinnaeve Justine , Mistry Akshitkumar , Leelatian Nalin , Mobley Bret , Chambless Lola , Thompson Reid , Weaver Kyle , Irish Jonathan , Ihrie Rebecca 
Published: November 2022

Abstract
Glioblastomas (GBM) account for ~60% of adult primary brain tumors. With few advances in therapeutics, median overall survival remains 15-months post-diagnosis. Immunotherapies may provide therapeutic benefit; however, no predictive immune features have informed therapeutic stratification. Radiographic tumor contact with the lateral ventricle (C-GBM) correlates with 7-months worse prognosis compared to patients with ventricle non-contacting GBM (NC-GBM), yet the influence of ventricle contact on anti-tumor immunity is unknown. This study characterized the GBM immune microenvironment and identified targetable mechanisms of immunosuppression correlating with worse outcomes in C-GBM patients.Primary glioblastoma tissue was provided with written informed consent in accordance with the Declaration of Helsinki and with approval of the Vanderbilt Institutional Review Board (IRB #131870). Seventeen patients presented with primary, IDH-wildtype C-GBM and 15 with NC-GBM. Machine learning integrated mass cytometry and matched multiplex immunohistochemistry on FFPE embedded tissue to identify phenotypic, functional, and spatial biomarkers correlating with patient outcome. C-GBM tumors were enriched in STAT3-driven CD32+CD44+HLA-DR+ monocyte-derived macrophages (MDM) compared to NC-GBM (19 ± 8% vs. 6 ± 2%; p&lt; 0.001) and depleted in lymphocytes (2.9 ± 1% vs. 7.6 ± 2%; p&lt; 0.001) and tissue-resident microglia (1.8 ± 0.3% vs. 7 ± 3%; p&lt; 0.001). Exhausted T cells in C-GBM co-expressed checkpoint receptors PD-1 and TIGIT. K-means clustering identified 10 immunological niches in GBM. Macrophage-tumor niches were most common in C-GBM (17.93% of niches), followed by T cell-microglia-tumor niches (17.72%). Within NC-GBM niches, T cell-T cell interactions were more prevalent (log odds ratio = 0.90) and correlated with improved outcome.These findings suggest that factors within the periventricular space negatively influence the immune microenvironment within GBM tumors. Clinically targetable immune biomarkers were identified in C-GBM. Notably, radiologic assessment of lateral ventricle contact may guide clinical trial design for immunotherapies in neuro-oncology based on tumor proximity to the ventricle wall.
TMIC-28. COMBINING INDUCED NEURAL STEM CELL THERAPY AND IMMUNOMODULATION IN GLIOBLASTOMA
Authors: Thang Morrent , Valdivia Alain , Hingtgen Shawn 
Published: November 2022

Abstract
Glioblastoma (GBM) is an aggressive malignant adult brain tumor and is characterized by poor prognosis and survival outcome. Insufficient drug accumulation to the tumor site and development of monotherapy drug resistance are critical challenges to overcome in combating GBM. Human induced neural stem cells (hiNSC) therapy is promising due to the stem cells’ innate capability to migrate to the tumors and deliver therapeutics. We previously have developed spheroidal iNSCs and engineered them to constitutively secrete pro-apoptotic protein TRAIL—hiNeuroS-TRAIL. However, investigating a combination therapy along with hiNeuroS-TRAIL therapy is needed to potentially avoid monotherapy resistance and enhance overall treatment durability. One of the GBM hallmarks is its immunosuppressed tumor microenvironment (TME). GBM-associated macrophages and microglia (GAMs) display pro-tumoral phenotypes in TME—inhibiting their ability to target the tumor cells or recruit other immune cells. In humans, higher glioma grade is associated with higher expression of Mer Tyrosine Kinase Receptor (MerTK) in both GBM cells and GAMs. Activation of MerTK in GAMs and GBM cells exacerbates immunosuppression and tumor progression respectively. Inhibiting MerTK activation via a small molecule drug, MRX-2843 and combining with hiNeuroS-TRAIL demonstrated robust synergistic tumor killing effects in vitro (CDI=0.22) and ex vivo (CDI=0.18). Additionally, the preliminary data for in vivo pilot study revealed that the median survival for the combination therapy treated group of mice extended over 24 days whereas the median survival for control, MRX-2843, and hiNeuroS-TRAIL treated groups of mice were 16, 18, and 21 days respectively. We think that combining these two therapies could significantly extend the survival in our murine model of GBM. Therefore, we will be optimizing dose concentration and cell counts for single agent therapies, and we will then repeat in vivo y synergy study with optimized therapies and a larger sample size to fully uncover the potential of this combination therapy.
TMIC-32. TUMOR CELL CILIA ASSOCIATE WITH SPECIFIC IMMUNE CELL POPULATIONS IN HUMAN AND MOUSE MODELS OF GBM
Authors: Deleyrolle Loic , Harrison Jeffrey , Takacs Gregory , Sarkisian Matthew 
Published: November 2022

Abstract

INTRODUCTION
Glioblastoma (GBM) typically recurs after standard of care therapies. A major obstacle is that GBMs employ various mechanisms to suppress the host immune system, preventing destruction and removal of cancer cells. A better understanding of the crosstalk mechanisms between tumor and immune cell types is needed to advance immunotherapeutic approaches against GBM. GBMs contain primary cilia, organelles likened to both cellular ‘antennae’ and transmitters, and their presence is associated with more aggressive and treatment resistant tumors.

OBJECTIVE
To explore whether ciliated GBM cells associate with infiltrating immune cells and to determine if these interactions are specific for certain immune cell populations.

METHODS
We immunostained GBM patient specimens and sections from syngeneic mouse models of GBM for markers of cilia (ARL13B and gTub) together with various immune cell markers (CD45, CD11b, Ly-6B.1, CD3). Cilia were also examined in brain tumors generated in CCR2+/rfp/CX3CR1+/gfp mice, to evaluate the proximity of glioma cell cilia to CCR2- and CX3CR1-expressing myeloid cell subpopulations (brain resident microglia as well as peripheral-derived macrophages and monocyctic- MDSCs (M-MDSCs). Proximity of tumor cell cilia to different markers or cell types was quantified using nearest neighbor analyses.

RESULTS
In patient samples, CD45+ cells extended processes that contacted the tumor cilia and cilia tip. Similar observations were made in intracranial GBM-bearing mice where we detected juxtaposition of cilia with recruited immune cells (i.e. CD45, CD11b and Ly-6B.1). Notably, cilia predominately associated with immune-suppressive M-MDSCs. Further, CD3+ T cells rarely juxtaposed tumor cilia and maintained greater distances away from ciliated tumor cells compared to M-MDSCs.

CONCLUSION
Our data suggest specific interactions between ciliated tumor cells and select immune-suppressive cell types, raising the possibility immunologically cold tumors may contain more ciliated cells. Ongoing studies are examining how the tumor immune cell landscape develops depending on the presence of tumor cilia.
TMIC-45. DISTRIBUTION AND TUMOR CELL PROXIMITY OF IMMUNE CELLS IN THE TUMOR CORE, INFILTRATION ZONE AND PERIPHERY OF GLIOBLASTOMAS
Authors: Pedersen Vilde , Knudsen Arnon Møldrup , Michaelsen Signe Regner , Kristensen Bjarne Winther 
Published: November 2022

Abstract
Glioblastoma is the most frequent and malignant brain tumor. Immune therapies have had limited effect, but not much is known about the frequency and type of immune cells in the infiltration zone and periphery. In these areas, tumor cells migrate into the brain parenchyma and prevent total tumor resection thereby leading to recurrence. This study aims to quantify the type and distribution of immune cells in glioblastomas. A cohort of 67 glioblastomas was established containing tissue from central tumor and/or infiltration zone and/or tumor periphery. Patients were included if most tumor cells were positive in immunohistochemical staining for P53. A multiplex staining was established, where tissue sections were stained with P53 (tumor cells), FOXP3 (FOXP3+ regulatory T cells), CD8 (CD8+ cytotoxic T cells) and IBA1 (microglia/macrophages). Cells were counted in tumor core, infiltration zone and periphery using Visiopharm software based on training of convolutional neural networks. Moreover, the number of tumor cells with immune cells in proximity (30 µm) was quantified. The number of CD8+, FOXP3+ and Iba1+ cells was significantly higher in the core than in the periphery (P &lt; 0.001), and in the infiltration zone than in the periphery (P &lt; 0.001). However, the CD8+, FOXP3+ and Iba1+ cells/tumor cell ratio increased from core to infiltration zone and to periphery (P &lt; 0.001). There was a higher amount of tumor cells with CD8+ cells in proximity in the tumor core (P &lt; 0.001), whereas the highest amount of tumor cells with IBA1+ cells and FOXP3+ cells in proximity was the infiltration zone (P &lt; 0.001). In conclusion, although levels of immune cells are highest in the tumor core - the higher proximity and ratio of immune cells to tumor cells in the infiltration zone/periphery suggest immune-suppressive mechanisms to be active in these areas.
TMIC-48. HYPOXIC AND PERI-NECROTIC ZONES IN GLIOBLASTOMA CAUSE HIPPO-OFF SIGNALING, POTENTIALLY SUPPORTING GLIOMA STEM CELLS
Authors: Shih Ling-Kai , Li Jiabo , Markwell Steven , Carneiro-Lobo Tatiana , Olson Cheryl , Brat Daniel 
Published: November 2022

Abstract
Glioblastoma (GBM) is the most common and malignant primary brain tumor with pre-necrotic and post-necrotic growth phases. The development of necrosis is associated with extensive tumor micro-environmental (TME) rearrangement and rapid biologic advancement. Compared to pre-necrotic GBMs, post-necrotic GBMs are heterogenous and contain multi-focal regions of necrosis surrounded by pseudopalisading cells and characterized by severe hypoxia. Hypoxic, peri-necrotic zones are highly enriched for glioma stem cells (GSCs) and tumor associated macrophages (TAMs), the latter accounting for 30-40% total tumor cellularity. Using Ivy-GAP data, we performed gene set enrichment analysis (GSEA) for stem-cell associated markers, comparing peri-necrotic to non-necrotic regions. We also used TCGA/CGGA data to identify correlations between hypoxia markers and classic stem-cell associated markers (SOX2, SOX9, nestin, STAT3, prominin). None of these showed enrichment in the peri-necrotic zone or association with hypoxia markers. However, we uncovered strong associations of Hippo-off signaling with peri-necrotic regions, signifying activation of YAP/TAZ transcriptional activation. We therefore hypothesized that Hippo-off (YAP/TAZ transcriptional activation) in hypoxic peri-necrotic zones promotes GSC self-renewing division. To support this, we performed GSEA for Hippo pathway readout genes and found peri-necrotic enrichment of Hippo regulated genes CYR61, BIRC2, SERPINE1 and AREG. Using patient-derived GBM neurosphere cultures, we exposed glioma cells to 1% hypoxia and found that p-YAP1 and p-TAZ protein levels were downregulated compared to normoxia (21%) at 24 hrs, correlating with Hippo-on. Hippo pathway gene targets were also upregulated, indicating activation of YAP/TAZ transcription. Using Ivy-GAP, TCGA and CGGA data, we also found peri-necrotic/hypoxic enrichment of two cytokines, IL-1β and IL-6, known to trigger YAP/TAZ activation. Using cytokine multiplex analysis, we observed an upregulation in both IL-1β and IL-6 secretion from hypoxic monocytes and microglia, raising the possibility that these cytokines directly stimulate Hippo-off and promote GSC self-renewing division in the GBM peri-necrotic niche.
TMIC-51. SEX-SPECIFIC T CELL BEHAVIOR DRIVES DIFFERENTIAL IMMUNE RESPONSES IN GLIOBLASTOMA
Authors: Lee Juyeun , Sneiderman Chaim , Kohanbash Gary , Sangwan Naseer , Lathia Justin 
Published: November 2022

Abstract
Glioblastoma (GBM) is the most common primary malignant brain tumor and shows poor outcomes as a median survival is 12-18 months with current standard-of-care of therapy. GBM exhibits sex differences in incidence and overall survival, with males experiencing a higher incidence and a worse prognosis compared to females. However, the role of immune cells in GBM sex differences outside of myeloid cells remains poorly understood. Using a syngeneic mouse GBM model, we recapitulated the sex differences observed in patients, with shortened survival in male hosts compared to female hosts. These findings were not recapitulated in immuno-deficient mouse strains such as NSG and RAG1KO mice, suggesting a role for immune system, specifically T cells, in GBM sex differences. Flow cytometry analysis of tumor-infiltrating leukocytes revealed that more T cells were found in female tumors, whereas male tumors were enriched in macrophages. Additionally, more T cells in male tumors exhibited high levels of inhibitory receptors, whereas most female T cells were more functional as measured by expression of anti-tumor cytokines such as TNF, IFN-gamma, and granzyme B. A bone marrow chimera model (BMC) revealed that male T cells retained their phenotype in female hosts, whereas female T cell behavior was affected by the male environment. Yet survival analysis on BMC model and adoptive transfer model suggests strong immune cell-intrinsic effect on controlling tumor progression, further supported by in vitro T cell exhaustion model. Lastly, we found that males have more progenitor exhausted T cells, which led to better response to anti-PD1 treatment. Collectively, these results suggest that both cell-intrinsic and cell-extrinsic factors regulate T cell activity in a sex-specific manner, providing insights to develop sex-specific therapeutic approaches.
TMIC-54. THE ROLE OF TUMOR MICROENVIRONMENT DERIVED GROWTH FACTORS IN PEDIATRIC BRAIN TUMORS
Authors: Cruz Andrea , Locke Abigail , Halligan Katharine , Sanders Lauren , Cheney Allison , Stanton Ann-Catherine Jean , Koncar Robert , Broniscer Alberto , Vaske Olena M , Pearce Thomas , Marker Daniel , Wiley Clayton , Mack Stephen C , Filbin Mariella , Pollack Ian F , Agnihotri Sameer 
Published: November 2022

Abstract

BACKGROUND
High-grade gliomas (HGGs) are the most common fatal intrinsic brain tumors in pediatric patients. H3K27-altered diffuse midline gliomas (H3K27-DMGs), a subgroup of HGGs defined by a histone 3 position 27 alteration, are especially aggressive and result in the poorest patient outcomes. Despite in-depth genomic characterization, the 5-year survival rate has yet to improve beyond 2% following diagnosis. A common feature of H3K27-DMGs is infiltration of microglia, macrophages, other myeloid cells, collectively referred to as GAMs, and a small population of T-cells. The contribution of non-tumor cells in the tumor microenvironment (TME) can both promote and or inhibit tumor growth, thus representing an opportunity in the pursuit of novel therapeutics. Using bioinformatic analysis on a human H3K37-DMG single cell-RNA sequencing dataset, we reveal several cell-to-cell communication signaling networks, mediated by ligand and receptor pairs, between GAMs and tumor cells, respectively.

HYPOTHESIS
Microglial-derived growth factors activate oncogenic signaling pathways via paracrine signaling axes, thus promoting H3K27-DMG tumor cell proliferation and growth.

METHODS
I will validate these findings and test their therapeutic potential using co-culture studies, CRISPR and shRNA gene silencing, and phospho-proteomics technology.

RELEVANCE
This research provides further insights on the contribution of non-tumor cells in the TME towards H3K27-DMG cell proliferation and growth and could potentially inform future therapy paradigms.