Mutational landscape and evolutionary patterns of peritoneal metastases in colorectal cancer

Peritoneal metastasis (PM) of colorectal cancer (CRC) is associated with significant morbidity and poor clinical outcomes; however, its evolutionary pattern and molecular mechanisms remain elusive. Whole-exome sequencing was performed on 49 matched samples comprising primary tumors, peritoneal metastases, and lymph node metastases from 11 CRC patients. Single-cell RNA sequencing was performed on 14 paired samples from three patients. Phylogenetic analyses revealed that 8 of 11 patients exhibited polyclonal dissemination, and MACHINA-inferred metastasis-to-metastasis seeding was found in five patients. Lymphatic spread did not appear to be the predominant route of peritoneal dissemination. Metastatic dissemination occurred early before the primary tumor became clinically detectable in 9 out of 11 patients. PM-associated driver genes, including KRAS, SMAD4, and RNF43 , were identified, and potentially actionable alterations were detected across patients, including site- or region-specific events. Single-cell transcriptomic analyses revealed substantial heterogeneity among metastatic deposits within individual patients and an enrichment of COL11A1 + fibroblasts in PM lesions, suggesting their involvement in creating a metastasis-permissive microenvironment. These findings delineate the evolutionary patterns, genomic features, and microenvironmental heterogeneity of CRC peritoneal metastases and provide novel insights into their molecular basis and potential therapeutic vulnerabilities.