Autologous colorectal cancer-mesothelial co-culture model identifies stromal FGFR3 dependency driving peritoneal dissemination

Background: Peritoneal dissemination of colorectal cancer (CRC) remains a fatal disease with limited therapeutic options. Mesothelial cells, which line the peritoneal cavity, play critical roles in metastatic implantation and stromal remodeling, yet the mechanisms governing tumor-stroma interactions in this setting remain poorly understood. We aimed to establish a physiologically relevant model to elucidate these interactions and identify stromal vulnerabilities. Methods: We simultaneously derived a novel CRC cell line (OMUCR-1) and cancer-associated mesothelial cells (CAmeso) from the malignant ascites of a single patient with metastatic CRC. As a control, normal mesothelial cells (Nmeso) were established from a patient without dissemination. Cell morphology, growth kinetics, STR profile, and mutational status were characterized. The effects of CAmeso on CRC migration and invasion were evaluated using wound-healing and Matrigel assays. In vivo, OMUCR-1 cells were implanted alone or co-transplanted with CAmeso into nude mice. RNA sequencing was performed to analyze host stromal gene expression, and FGFR inhibition (BGJ398) was tested therapeutically. Results: OMUCR-1 cells exhibited robust tumorigenicity and harbored KRAS (G12D) and TP53 (R306) mutations. CAmeso significantly enhanced CRC cell migration and invasion in vitro. In xenografts, co-transplantation with CAmeso yielded markedly larger tumors enriched in alphaSMA-positive stroma compared with OMUCR-1 alone. Bulk RNA-seq revealed upregulation of murine Fgfr3 in tumors containing CAmeso. Pharmacologic FGFR blockade with BGJ398 reduced tumor size and depleted FGFR3-positive stromal components, indicating stromal FGFR3 dependency. Conclusions: We report the first concurrent establishment of CRC and mesothelial cell lines from the same patient, providing an autologous system for dissecting tumor-stroma crosstalk during peritoneal metastasis. Our findings highlight stromal FGFR3 signaling as a potential mediator of CRC progression and a promising target for therapeutic intervention. This unique platform offers a valuable resource for mechanistic and translational studies on the peritoneal microenvironment in advanced colorectal cancer.