bluuebunny/crossref_metadata_2025_split huggingface dataset:
IMMU-01. DE NOVO-DESIGNED BINDER-BASED CAR T CELLS ENABLE EFFECTIVE TARGETING OF GLIOBLASTOMA
Authors: Xie Qi 
Published: November 2024

Abstract
Glioblastoma is a devastating disease with poor responses to all current therapies. Chimeric antigen receptor (CAR) T cell therapy has achieved impressive success against hematological malignancies but produces responses in only a limited number of glioblastoma patients. Here, we developed a novel platform for CAR constructs using de novo designed protein binders to target tumor antigens. We firstly designed a binder CAR targeting epidermal growth factor receptor (EGFR), which is highly expressed in glioblastoma, to validate the feasibility of our platform. Our EGFR binder CAR design promoted CAR T cell proliferation, cytotoxic cytokine secretion and exhaustion resistance, leading to better antitumor performance both in vitro and in vivo. Bulk and single-cell transcriptional profiling of binder CAR T cells revealed an underlying mechanism involving enhanced effector activity and reduced exhaustion responses. Mechanismly, the binder CARs exhibited higher surface expression and greater resistance to degradation triggered by tumor antigens. Moreover, we designed and engineered a novel CD276 binder CAR T cell product and verified its antitumor function, further demonstrating the general applicability of our binder-based CAR platform. Overall, our study provides an alternative avenue for the design of a CAR antigen-binding domain to potentiate CAR T cell antitumor efficacy.
IMMU-03. EXPLORING CANCER-SPECIFIC T CELLS AND ANTIGENS IN GLIOBLASTOMA BASED ON SINGLE-CELL SEQUENCING OF CD8+ TILS
Authors: Okamoto Takanari , Mizuta Ryo , Demachi-Okamura Ayako , Muraoka Daisuke , Sasaki Eiichi , Masago Katsuhiro , Onoguchi Kazuhide , Yamashita Yoshiko , Muto Osamu , Yamaguchi Rui , Takahashi Yoshinobu , Hashimoto Naoya , Matsushita Hirokazu 
Published: November 2024

Abstract

BACKGROUND
The effectiveness of cancer immunotherapy against glioblastoma (GBM) remains limited. This study aims to identify cancer-specific antigens to develop antigen-based cancer immunotherapies for GBM. We investigated candidate tumor antigen-specific T cells in GBM by single-cell RNA sequencing (scRNA-seq) and single-cell TCR sequencing (scTCR-seq), and we explored candidate antigens through genetic analysis of tumor tissues.

METHODS
Flow cytometry analysis was conducted on fresh tumor digest samples to evaluate tumor-infiltrating lymphocytes (TILs) of GBM. Single-cell RNA and TCR sequencing were performed on CD8+ T cells in TILs. Both data generated by Cell Ranger software were loaded into Seurat at R studio. The dimensional reduction for clustering was performed with uniform manifold approximation and projection (UMAP). Additionally, we performed bulk RNA sequencing (RNA-seq) and whole exome sequencing (WES) on tumor tissues.

RESULTS
Two out of 20 patients (10%) exhibited abundant TILs in GBM. From these two patients, approximately 20,000 CD8+ T cells in total were analyzed in single-cell analysis. Clustering based on UMAP revealed a predominance of clusters expressing exhaustion markers, with high TCR clonality centered on exhausted T cell (TEX) clusters, like our previous date of lung cancer TILs recognizing tumor antigens. Moreover, within the TEX clusters, the expression of CXCL13, often reported as an antigen-specific marker in recent years, was significantly higher in our data compared to two publicly available datasets. Subsequently, based on RNA-seq and WES, 61 candidate neoantigens were estimated using machine-learning prediction algorithms from NEC Corporation. Furthermore, 5 overexpressed cancer/testis antigens, and 3 bacterial species-derived microbial peptides using Kraken2 were selected as antigen candidates.

CONCLUSIONS
We identified prospective tumor antigen-specific T cell subsets with high CXCL13 expression from two GBM patients. We plan to identify cancer-specific T cells and antigens from these candidates by T cell activation assay.
IMMU-06. H-FERRITIN AFFECTS IMMUNE CELL INFILTRATION INTO THE GLIOBLASTOMA TUMOR MICROENVIRONMENT IN A SEX-BIASED MANNER
Authors: Pappus Sahu Aurosman , Palsa Kondaiah , Shenoy Ganesh , Helmuth Timothy B , Slagle-Webb Becky , Neely Elizabeth , Connor James R 
Published: November 2024

Abstract
Glioblastoma (GBM) is an aggressive brain cancer that is sexually dimorphic in nature, with females less susceptible to developing it, responding better to therapy, and gaining a survival advantage. Iron metabolism is also sexually dimorphic and has been shown previously to contribute to sex bias in GBM. Ferritin is an iron-storing protein that has elevated expression in tumor cells and is essential to GBM growth. Here, we used an in vivo GBM model consisting of heterozygous FTH1 knockdown (FTH1-KD) mice and GL261 cells to study the role of host ferritin in GBM progression. We have previously published that FTH1 knockdown in female mice decreased overall survival compared to WT (P = 0.03). This work delves into potential mechanisms that contribute to this survival difference. GBM was induced in 6-8 weeks-old mice by injecting 2*10^4 GL261 cells intracranially by a stereotaxic apparatus. The tumor volume was measured 3 weeks post-injection by magnetic resonance imaging, followed by image segmentation. We observed that FTH1 knockdown did not affect tumor volume. Flow cytometry was used to examine the immune infiltrating cells (CD45high) population in the tumor-bearing hemisphere. In females, there was an increase in CD11b+ cells in FTH1KD mice compared to WT mice (P = 0.0607). Similarly, Female FTH1-KD mice had a lower T cell (CD3+) population than Female WT mice (P = 0.0033). This decrease in infiltrating CD3 cells was due to a reduction of CD4 T cells (CD3+ CD4+) in female FTH1-KD mice compared to female WT mice (P = 0.0375). We observed no difference in CD8 T cells (CD3+ CD8+). These data indicate that host ferritin affects immune infiltration into GBM cells in a sex-biased manner.
IMMU-08. PRECLINICAL EVALUATION OF GAL-1-SPECIFIC CAR-T CELL FUNCTION IN AN IN VITRO AND EX VIVO MODEL OF GLIOBLASTOMA
Authors: Das Arabinda , Zukas Alicia M , Santos Jaime Martinez , Stephens Heather R , Mark Joseph , Sekerak Penny , Infinger Libby Kosnik , III William A Vandergrift , Varma Abhay K , Patel Sunil J , Cachia David , Lindhorst Scott M 
Published: November 2024

Abstract

BACKGROUND
Human glioblastoma (GBM), especially those exhibiting anti-VEGF resistance, overexpress the carbohydrate-binding protein galectin-1 (Gal-1). To develop novel anti-GBM approaches, we developed Gal-1 targeted CAR-T cells to investigate potential antitumor effect with in vitro and ex vivo slice culture models.

METHODS
Uniform GBM brain slice cultures were collected using NICO’s Myriad resection tool and their Automated Preservation System (APS), designed to obtain fresh brain tumor samples maintaining tumor cytoarchitecture and microenvironment. To prepare Gal-1 CAR-T cells, we used creative laboratory to utilize a third-generation Gal-1-CAR construct with high binding and codon optimized scFv-Gal-1 fragment fused with CD3ζ and co-stimulatory cytoplasmic domains of CD28 and 4-1BB (Gal-1 scFv-CD28-4-1BB-CD3ζ). We then transfected into the T cells, from peripheral blood, mononuclear cells using a lentiviral system. The anti-tumor capabilities of CAR-T cells were evaluated by co-culturing with GBM cells and ex vivo APS-GBM slices.

RESULTS
The collected APS- GBM slices samples were clear of debris and blood product, not requiring post-surgical mechanical slices, and exhibited viability up to 12 days over traditionally prepared ex vivo cultures. With this, we found Gal-1-CAR-T cells grew exponentially in the presence of cytokines and maintained phenotypic and biological attributes such as cell viability, potency, migration, and T cell activation. Gal-1 scFv-CD28-4-1BB-CD3ζ -CAR-T cells killed more than 80% of GBM tumor cells within 24 hours in Gal-1+ve, but not Gal-1-ve GBM cells in vitro. Additionally, these CAR-T cells selectively caused significant release of IL-2, IFN-γ and TNF-α only from Gal-1+ve co-cultures and induced significant Gal-1+ve GBM cell death in an ex vivo model without any general toxicity.

CONCLUSION
Based on in vitro cell culture and ex vivo APS-brain slice culture results, we conclude that the novel Gal-1 scFv-CD28-4-1BB-CD3ζ CAR-T cell therapy may offer an effective therapeutic option. Further pre-clinical and clinical studies need to be performed to validate this novel approach
IMMU-09. REVERSING SYSTEMIC T CELL DYSFUNCTION TO LICENSE IMMUNOTHERAPY IN INTRACRANIAL TUMORS
Authors: Wachsmuth Lucas , Lorrey Selena , Finlay John , Neff Corey , Ayasoufi Katayoun , Price Mackenzie , Reesman Rachael , Ostrom Quinn , Fecci Peter 
Published: November 2024

Abstract
Intracranial tumors present unique challenges for immunotherapy. These can include both local and systemic modes of immune suppression, the mechanistic underpinnings of which are incompletely understood. Our work reveals that intracranial tumors elicit systemic increases in circulating catecholamine levels, and that this chronic sympathetic hyperactivity results in T cell dysfunction that impedes immunotherapeutic efficacy. We show that treatment with b-adrenergic blockade can partially overcome the negative impacts of chronic sympathetic hyperactivity by increasing NF-kB activity in immune cells, restoring T cell polyfunctionality, favorably modifying the tumor microenvironment, and extending survival in murine models of glioblastoma treated with immune-based therapies. Furthermore, we demonstrate that extended survival is also observed in glioblastoma patients receiving b-adrenergic blockade, as well as in patients with melanoma and lung cancer brain metastases who received b-blockade alongside concomitant immune checkpoint inhibition. Importantly, while local sympathetic hyperactivity in the tumor microenvironment and b-blockade also impact the anti-tumor immune response in the context of extracranial disease, these effects are markedly more pronounced in intracranial disease. Taken together, these data reveal that sympathetic hyperactivity facilitates systemic immune dysfunction in the setting of intracranial tumors and highlight a novel role for the application of b-adrenergic blockade to license immunotherapy in intracranial malignancies.
IMMU-24. GLIOBLASTOMA ENRICHED GLYCOSPHINGOLIPIDS MODULATE THE FUNCTION OF HUMAN INKT CELLS
Authors: Coombs Morgan , Dowdy Tyrone , Alam Md Masud , Khayrullina Guzal , Joshi Vibhuti , Keretsu Seketoulie , Jung Jinkyu , Hara Ayaka , Steffke Emily , Latifi Laila , Kim Hye , Spurgeon Jo , Malik Nargis , Hancock John , Smith Kelsey , Gumperz Jenny , Larion Mioara , Terabe Masaki 
Published: November 2024

Abstract
Glioblastoma is an aggressive primary brain tumor highly resistant to currently available immunotherapies. A deeper understanding of its underlying immunoregulatory mechanisms is paramount to developing future immunotherapeutic treatments. Invariant natural killer T (iNKT) cells are unconventional T cells that recognize lipid antigens presented by an MHC-like molecule called CD1d. Although iNKT cells have been shown to regulate tumor immunity in other cancer types, their role in glioblastoma is not well characterized. Given the lipid-rich nature of the brain and the unique metabolic activity of glioblastoma cells, we hypothesized that interactions between glioblastoma and iNKT cells through glioblastoma-produced lipids could contribute to the immunosuppressive nature of the disease. We report multiple lipid species enriched in aggressive human glioblastoma stem-like cell (GSC) lines that activate human iNKT cells and modulate their functions. Lipidomic LC-MS analysis of GSCs, low-grade glioma stem-like cell lines, and normal human astrocytes revealed various glycosphingolipid species, including sulfatides, highly enriched in the GSCs. Multiple enriched sulfatide species, when presented by CD1d, were recognized by and activated iNKT cells in a dose-dependent manner. Additionally, cytokine analysis of stimulated human PBMC-derived iNKT cells demonstrated that the enriched sulfatides did not induce Th1 cytokine production, but rather many non-Th1 cytokines that potentially counteract Th1-type immune responses. This modulation of iNKT cell function by glioblastoma-enriched glycosphingolipids may contribute to the immunosuppression of glioblastoma and could highlight sulfatide production as a potential therapeutic target for glioblastoma treatment.
IMMU-29. B7-H3-TARGETED CAR-T AND CAR-NK CELLS AGAINST HUMAN GLIOBLASTOMA
Authors: Tachi Tetsuro , Kijima Noriyuki , Kuroda Hideki , Murakami Koki , Hosen Naoki , Kishima Haruhiko 
Published: November 2024

Abstract
Glioblastoma multiforme (GBM) is the most lethal primary brain tumor, necessitating novel therapies. Chimeric antigen receptor (CAR) T-cell therapy has recently shown efficacy against GBM. However, this approach is costly and time-consuming due to its personalized nature. Alternatively, CAR-transduced natural killer (NK) cells offer a potential “off-the-shelf” immunotherapy option, as they do not cause graft-versus-host disease. This study aimed to assess the anti-GBM efficacy of CAR T and NK cells targeting B7-H3, a protein highly expressed in GBM. To achieve this, CAR T cells targeting B7-H3 were developed using known anti-B7-H3 scFv sequences. Additionally, cord blood-derived NK cells were transduced with the B7-H3 CAR. Both types of CAR-modified cells were tested for their anti-GBM activity in vitro. Moreover, their anti-tumor effects were evaluated in an in vivo xenograft model using patient-derived GBM cells. The results demonstrated that both B7-H3 CAR T cells and CAR NK cells exhibited significant cytotoxicity against patient-derived GBM cells in vitro. Furthermore, intracranial injection of these CAR-modified cells into the xenograft model significantly reduced tumor growth. In conclusion, B7-H3 targeted CAR T cells and cord blood-derived CAR NK cells both show potential in eliminating GBM cells, suggesting promising avenues for future GBM treatment strategies.
IMMU-30. IGV-001: A BIOLOGIC-DEVICE COMBINATION PRODUCT ELICITS POTENT ANTI-GBM RESPONSES AS A MONOTHERAPY AND IN COMBINATION WITH CHECK POINT INHIBITION
Authors: Zilberberg Jenny , Exley Mark 
Published: November 2024

Abstract

INTRODUCTION
Imvax is developing GoldspireTM, a personalized immunotherapy that combines whole tumor-derived cells with an antisense oligonucleotide against insulin-like growth factor 1 receptor (IGF-1R; IMV-001) in biodiffusion chambers (BDCs; 0.1 µm pore). BDCs are irradiated and implanted at abdominal sites for ~48h to deliver an antigenic payload and immunostimulatory factors that together train the immune system to attack tumor cells. The lead product, IGV-001, was evaluated in newly diagnosed glioblastoma (ndGBM) patients in a phase 1b study. Median overall survival (OS) of highest exposure IGV-001-treated Stupp-eligible patients (n=10) was 38.2 mos compared with 16.2 mos in standard-of-care-treated patients (p=0.044; ClinicalTrials.gov NCT02507583). A Phase 2b study in ndGBM is currently underway (ClinicalTrials.gov NCT04485949), where a primary endpoint of progression-free survival and key secondary endpoints of overall survival and safety will be evaluated.

METHODS
The orthotopically implanted GBM (luciferase-expressing GL261-luc2) model was utilized to evaluate the efficacy of mIGV-001 (the murine version of IGV-001) alone or in combination with anti-PD-1 immunotherapy. The following four groups were compared: mIGV-001 or BDC placebo + isotype or anti-PD-1 antibody (4 doses total every 3-4 days). GL261-luc2 cells were treated with IGF-1R antisense IMV-001. The treated cell suspension was loaded into clinical-grade BDCs (0.1-µm pore size, polyvinylidene difluoride membrane) at a density of 1 × 106 cells per BDC and then irradiated with 5–6 Gy. mIGV-001 or BDCs filled with saline were administered 28 days prior to tumor challenges and tumor progression and survival were monitored for an additional 100 days.

RESULTS
Preventative treatment with mIGV-001 limited tumor progression and extended OS in mice orthotopically (i.e., intracranially) challenged with GL261-luc2 cells. The therapeutic benefit of mIGV-001 was enhanced with anti-PD-1 treatment and the combination therapy was superior to either monotherapy.
IMMU-32. DIPEPTIDYL PEPTIDASE-4 (DPP-4) MEDIATED CD8+ T-CELL REGULATION IN GLIOBLASTOMA
Authors: Teran Pumar Oriana Y , Ruiz Clara Lopez , Colon Bruno , Rafie Christine Isabelle , Khatwani Natasha Karishma , Stelekati Erietta , Watson Dionysios C , Bayik Defne 
Published: November 2024

Abstract
Glioblastoma (GBM) remains to be one of the deadliest cancers with poor prognosis and limited treatment options. While immunotherapies have revolutionized the field for other cancers, GBM remains resistant to these types of interventions partly due to its highly immunosuppressive tumor microenvironment, which is characterized by accumulation of exhausted CD8+ T cells. Therefore, there is a pressing need to identify molecular targets for improve anti-tumorigenic T cell response. Previous studies in our laboratory identified dipeptidyl peptidase-4 (DPP-4), an exopeptidase and a membrane-bound signaling receptor, as a potential marker of immunosuppression in GBM. While DPP-4 is well-drugged due to its role in insulin metabolism, the function of DPP-4 in immune regulation and cancer remains poorly defined. I observed that DPP-4 is highly expressed by CD8+ T cells GBM patients and preclinical models. Furthermore, DPP-4 levels increased with the exhaustion or memory differentiation of effector T cell. Pharmaceutical inhibition of DPP-4 enriched expression of genes related to lymphocyte activation and mitochondrial metabolism. These transcriptional changes were accompanied by enhanced T cell proliferation, cytotoxic function, and effector mediator production under exhaustion conditions. These findings suggest that pharmacological inhibition of DPP-4 promotes anti-tumorigenic function of CD8+ T-cell by preventing exhaustion and support future translational efforts to repurpose DPP-4 inhibitors as immunotherapy agents in GBM, given their favorable safety profiles.
IMMU-38. LEVERAGING INNATE IMMUNE SENSORS TO GENERATE DURABLE ANTI-GLIOMA ADAPTIVE IMMUNE RESPONSES
Authors: Lea Spencer , Najem Hinda , Chen Chao-Hsien , Wei Jun , William Ivana , Tripathi Shashwat , Hurley Lisa , Heimberger Amy , Curran Michael 
Published: November 2024

Abstract
Glioblastoma harbors a tumor microenvironment (TME) that is mostly devoid of T cell effector infiltration but is dominated by immune suppressive microglia, macrophages, and myeloid-derived suppresser cells. The innate immune cells in the TME have impaired phagocytosis and antigen presentation and are inadequate for triggering the subsequent immune activating signals needed for anti-tumor effector responses. Phagocytic activity can also be blocked by the upregulation of CD47 on the tumor cells. In the immune checkpoint resistant Qki-/- Pten-/- P53-/- (QPP) glioma model, intratumoral treatment with the synthetic cyclic di-nucleotide STING agonist IACS-8803 (8803) induces global immunological reprogramming and long-term survival resistant to tumor rechallenge. More specifically, in the infiltrating myeloid stroma 8803 increases costimulatory protein CD86 while reducing the expression of suppressive markers CD163 and CD206. Conventional type 1 dendritic cells are increased in both the TME (p&lt;0.05) and the draining cervical lymph node (p&lt;0.01). Intratumoral infiltration of both CD8 T (p&lt;0.05) and NK (p&lt;0.001) cells increases. However, only T cells, but not NK cells, are necessary for 8803 therapeutic activity when assessed with in vivo NK cell depletion studies and treatment in the RAG-/- background. STING activation increases microglial phagocytic capacity, but also induces a compensatory increase in SIRPα expression (p&lt;0.001) – the ligand for CD47. 8803 synergizes with anti-CD47 blockade to enhance microglia phagocytosis of QPP glioblastoma cells in vitro. Finally, the combination of 8803 and anti-CD47 synergizes to extend both median survival and the fraction of long-term survivors in orthotopic QPP-bearing mice (QPP8v; vehicle control MS=39d, anti-CD47 MS=40d, 8803 MS=67d 20% long-term survival, 8803 + anti-CD47 MS=107.5d 40% long-term survival). These data demonstrate that 8803 induces proinflammatory conversion of the glioblastoma TME, generates T cell-dependent survival benefits in QPP-bearing mice, and synergizes with anti-CD47 blockade to enhance microglia phagocytosis of glioblastoma cells and extend survival in QPP-bearing mice.
IMMU-46. CROSS-TALK OF INNATE IMMUNE CELLS IN GLIOBLASTOMA PATHOPHYSIOLOGY: UNDERSTANDING NLRP3 AND NLRP12
Authors: Rajkhowa Sushmita , Meena Durgesh , Jha Sushmita 
Published: November 2024

Abstract
Glioblastoma (GBM) are malignant gliomas with an average survival of less than 14 months after diagnosis, despite surgery, chemotherapy, and radiotherapy. Tumor survival is aided by the heterogeneous cell populations in the tumor microenvironment. Two crucial innate immune cells of the brain, namely microglia and macrophages, favor a tumor-promoting microenvironment IN GBM. The nucleotide-binding domain leucine-rich repeat-containing receptor (NLR) family in astrocytes and microglia cells recognizes pathogen- and damage-associated molecular patterns that induce inflammation. NLRP3, upon activation, leads to cleavage and activation of Caspase-1, causing the proteolytic cleavage-mediated activation and conversion of pro-IL-1ß and pro-IL-18 to IL-1ß and IL-18, leading to pyroptosis. NLRP12 downregulates inflammatory responses in microglia and macrophages by regulating the NF-kB pathway. NLRP3 and NLRP12 have tumor-promoting and anti-tumour functions in various cancers. NLRP12 is a prognostic marker for glioblastoma, and its increased expression correlates with poor survival in GBM patients. However, the cellular and molecular mechanisms of NLRP3 and NLRP12 in GBM pathophysiology are largely unknown. This study aims to understand the effect of NLRP3 and NLRP12 expression in GBM, to interpret the pathophysiology, and to help develop future targeted therapeutic interventions. To understand the baseline expression of NLRP3 and NLRP12 in cell lines and patient-derived GBM cells, PCR, western blotting, and Immunocytochemistry were performed comparing WT and nlrp3-/- and nlrp12-/- cells. Differential expression of NLRP3 and NLRP12 in patient-derived cells can be compared to GBM, astrocyte, and microglia cell lines. This may be the underlying cause for intra and inter-tumor heterogeneity and resultant differential therapy outcomes. To understand cell-to-cell communication in GBM-like tumor microenvironments, heterocellular 3D organoids, and patient-derived tumor organoids were developed. Cell viability in 3D patient-derived tumor organoids decreased by up to 10% with Doxorubicin and 40% with Carboplatin treatment decreases compared to untreated controls. This suggests the scope of developing drugs targeting inflammation regulatory genes.