Abstract 5092: Characterization of mismatch repair status in colorectal cancer using single cell and spatial transcriptomics

Abstract Background: Mismatch repair (MMR) status plays a critical role in colorectal cancer (CRC), influencing tumor biology and response to therapies. MMR-deficient (MMR-d) tumors, characterized by microsatellite instability (MSI), exhibit high mutation burdens and increased T cell infiltration, whereas MMR-proficient (MMR-p) tumors typically demonstrate low T cell density and poor immune checkpoint inhibitor response. Despite well-documented clinical implications, the molecular and cellular differences of T cells in the tumor microenvironment between MMR-d and MMR-p CRC remain incompletely understood. Methods: Single-cell RNA sequencing (scRNA-seq) data were analyzed using the CellChat R package to identify ligand-receptor interactions enriched in MMR-p colorectal cancer. Differentially enriched pathways involving T cells were identified and subjected to Gene Ontology (GO) enrichment analysis to uncover biological processes specific to these interactions. Validation was performed using spatial transcriptomic data from the Human Tumor Atlas Network (HTAN) to confirm ligand-receptor co-expression in a spatial context. Results: CellChat analysis revealed significant enrichment of TNFRSF14-associated signaling pathways (BTLA-TNFRSF14, LTA-TNFRSF14, and TNFS14-TNFRSF14) in MMR-p CRC. Functional analysis of TNFRSF14+ T cells in MMR-p tumors demonstrated upregulation of pathways related to leukocyte migration, negative regulation of T cell activation, and immune effector process regulation, as identified through GO enrichment analysis. Validation using spatial transcriptomic data confirmed co-expression of the BTLA-TNFRSF14 ligand-receptor pair in a single MSS sample but did not support broader enrichment across MMR-p samples. Conclusion: These findings highlight distinct ligand-receptor interactions and immune regulatory processes in MMR-p CRC, particularly involving TNFRSF14 signaling. While functional analysis underscores the role of these interactions in shaping T cell activity, spatial transcriptomic validation suggests heterogeneity in their enrichment across samples. This underscores the complexity of immune microenvironment regulation in MMR-p CRC tumors. Citation Format: Dimitri Joseph, Alexandra Guillaume. Characterization of mismatch repair status in colorectal cancer using single cell and spatial transcriptomics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5092.