Novel syngeneic cell lines for studying high-risk BRAFV600E-driven colorectal cancer in vivo [RNA-seq_organoids]

Combining anti-EGFR antibodies with BRAF/MEK/ERK-pathway blockade opened new treatment options for BRAFV600E driven colorectal cancer (CRC). Nevertheless, responses of these poor-prognostic cancers are short-lived and heterogeneous, thereby highlighting the unmet need for novel concepts integrating targeted therapy and immuno-oncology. To this end, an immunocompetent mouse model with reliable disease onset and responding to clinically relevant drugs is essential but unavailable. Here, we generated three cell lines from peritoneal metastases originating from orthotopically transplanted organoids carrying BRAFV600E, Trp53R172H and Apc mutations and characterized their ground state and responses to targeted therapy compounds in detail by RNAseq, WES and various functional assays. Interestingly, these NaJa lines displayed distinct differentiation states and responses to the clinically relevant RAF inhibitors (RAFi) encorafenib and exarafenib, thereby resembling the clinical heterogeneity of BRAFV600E driven CRC. RAFi resistance was overcome by the EGFR-family inhibitor afatinib. RAFi also influenced the expression of the antigen presentation machinery, cytokines and other immunomodulatory factors. Upon re-transplantation into syngeneic mice, all NaJa lines established aggressive tumors with distinct tumor microenvironments correlated with their differentiation states. Thus, the NaJa lines provide a unique tool to study tumor heterogeneity, drug resistance and the interplay between tumor, stroma and immune cells in BRAFV600E driven CRC.