Abstract 5176: NaJa cells - the first syngeneic cell line model to study BRAFV600E-driven colorectal cancer

Abstract Background: Due to the lack of suitable syngeneic murine models to study malignant BRAFV600E-driven colorectal cancers (CRC), the aim of this study was to establish new murine models that recapitulate this aggressive CRC subtype both in vitro and in vivo in an immunocompetent syngeneic context. Methods: A murine colonic organoid model with VillinCreERT2-inducible BRAFV600E, TRP53R172H and APC∆ mutations (designated BPAC) was generated. Orthotopic transplantations with BPAC organoids resulted in tumor growth and spontaneous metastasis into the peritoneal cavity of immunocompetent syngeneic C57/Bl6 recipient mice. Through subsequent peritoneal lavages, three new murine cell lines (NaJa-D, NaJa-F and NaJa-G) originating from the same mutational background were established. These cell lines were thoroughly characterized by RNA- and whole exome sequencing. Furthermore, their response to kinase inhibitors was tested and validated by colony formation assays and Western blotting. To test their invasive potential in vivo, the NaJa cells were injected via the portal vein into immunocompetent syngeneic mice and the tumor microenvironment was subsequently analyzed by immunohistochemistry (IHC). Results: The three established NaJa cell lines all expressed the aforementioned driver mutations and microsatellite stability was confirmed through whole exome sequencing. They differed greatly in their morphology despite the common genetic background, with NaJa-D displaying an epithelial morphology, NaJa-G adopting a mesenchymal phenotype and NaJa-F presenting an intermediate phenotype. Furthermore, they exhibited differential responses to treatment with clinically relevant kinase inhibitors, resulting in differentially altered signaling pathways and overall survival. Upon re-transplantation via the portal vein into immunocompetent syngeneic mice, all three NaJa cell lines induced fast and aggressive tumor growth in the liver while retaining their distinct morphological characteristics, and promoted metastasis, particularly to the peritoneal cavity and surrounding tissues. The tumor microenvironment was analyzed in terms of immune cell composition and distribution, revealing notable differences between the cell lines. These findings establish the NaJa cell-induced tumor model as a valuable tool for exploring the combinatory use of targeted therapies with immune checkpoint inhibition in future studies. Conclusion: Given the general lack of murine cell lines to model BRAFV600E-driven CRC, the NaJa cells provide a unique and valuable tool to research cancer heterogeneity, test clinically relevant kinase inhibitors and research the immune cell environment and immune escape of BRAFV600E-driven metastatic CRC. Citation Format: Jasmin Traichel, Ariane Lehmann, Nadine Wohlfeil, Ricarda Griffin, Maria Walz, Reinhild Feuerstein, Célia Asdih, Rebecca Kesselring, Melanie Boerries, Tilman Brummer. NaJa cells - the first syngeneic cell line model to study BRAFV600E-driven colorectal cancer [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 5176.