Cancer-associated fibroblasts metabolic targeting overcomes T-cell exclusion and chemoresistance [Mouse]

T cell-based immunotherapies have exhibited promising outcomes in tumor control; however, their efficacy is limited in immune-excluded tumors. Cancer-associated fibroblasts (CAFs) play a pivotal role in shaping the tumor microenvironment and modulating T cell infiltration. Despite the identification of distinct CAF subtypes using single-cell RNA-sequencing, their specific impact on hindering T cell infiltration remains unclear, particularly in soft-tissue sarcomas (STS) characterized by low response rates to T cell-based therapies. In this study, we comprehensively characterized the STS microenvironment using murine models with distinct immune composition and scRNA-seq, leading to the identification of a unique subset of CAFs termed glycolytic cancer-associated fibroblasts (glyCAFs). GlyCAFs rely on glycolysis to impede cytotoxic T cell infiltration into the tumor parenchyma. Targeting glycolysis in glyCAFs enhances T-cell infiltration and augments the efficacy of chemotherapy. These findings highlight new avenues for combinatorial therapeutic interventions in sarcomas and other solid tumors. Further investigations and clinical trials are needed to validate these potential strategies and translate them into clinical practice. Overall design: Mouse Undifferentiated Pleiomorphic Sarcoma (UPS) tumors bearing one of two tumor genetics (p53-/-Ccne1+ or p53-/-Vgll3+) were collected for single-cell RNA-sequencing. Digested tumor cell suspensions were enriched for specific cell types of interest. In experiments where the abundance of the cell type of interest was low (cancer-associated fibroblasts), multiple mouse tumors were first pooled together into a single sample, and processed only in aggregate. Where the abudance of the targeted cell type was sufficiently high, individual mouse tumors were processed as separate biological replicates, and tagged with an oligonucleotide hashtag to preserve sample identity. Hashtagged samples were finally pooled together prior to capture. Where indicated, technical replicates represent capture of additional cells and in a separate channel of the 10X Chromium chip, but drawn from the same overall pool of cells.