TMIC-22. MYELOID CELLS INDUCE IMMUNOSUPPRESSION AND HYPORESPONSIVENESS OF CHIMERIC ANTIGEN RECEPTOR T CELLS IN THE NEURONAL MICROENVIRONMENT OF GLIOBLASTOMA

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.