CT26.MtC3, a new preclinical model to investigate immune resistance in metastatic colorectal cancer.

Preclinical syngeneic tumor models are widely used to evaluate therapeutic responses in immunocompetent hosts, particularly for radiotherapy and radio-immunotherapy. Standard models, such as CT26.WT mouse colorectal carcinoma, have provided valuable insights into treatment efficacy and immune modulation. However, they often fail to reproduce the complexity of advanced human cancers, especially metastases, which differ from primary tumors in immunogenicity, antigen presentation, and tumor microenvironment (TME). Metastatic lesions are typically characterized by poor cytotoxic T cell infiltration and enrichment of immunosuppressive populations such as regulatory T cells and myeloid-derived suppressor cells, limiting the translational relevance of conventional approaches. To better model these features, we developed CT26.MtC3, a novel cell line derived from pulmonary metastases generated by systemic injection of CT26.WT cells into BALB/c mice. When implanted subcutaneously, CT26.MtC3 tumors displayed greater aggressiveness than parental CT26.WT. Immune profiling revealed a strongly immunosuppressive TME, with reduced CD8⁺ T cell infiltration and increased myeloid populations. Complementary in vitro analyses confirmed intrinsic differences, including higher migratory capacity, and significant upregulation of CAECAM-1a. Upon irradiation, CT26.MtC3 cells also showed altered expression of immune-regulatory molecules such as CD47, Fas, PD-L1, and PD-L2, potentially contributing to their immunoevasive phenotype in vivo. Together, these findings establish CT26.MtC3 as a clinically relevant model of metastatic, immune-resistant colorectal cancer. This model provides a robust platform for evaluating strategies to overcome tumor immunoresistance and for rigorously testing the potential of radio-immunotherapy combinations in settings that better reflect clinical reality. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12935-026-04200-x.