KRAS G12D inhibition stimulates antigen presentation and potentiates mRNA-based immunotherapy in pancreatic cancer

Inhibition of constitutively-active KRAS G12D , a powerful oncogenic driver in pancreatic ductal adenocarcinoma (PDAC), has been shown to sensitize tumors to T-cell-mediated immunotherapy in preclinical models. However, the influence of KRAS G12D inhibition on tumor cell antigen presentation remains unclear. Therefore, we employed a multi-omic profiling workflow to investigate alterations in the proteome and peptide MHC-I ligandome (immunopeptidome) following the treatment of HPAC human cancer cells with a RAS(ON) G12D-selective inhibitor RMC-9945 (representative of investigational agent zoldonrasib), and/or IFNɣ in vitro . KRAS G12D inhibition induced a unique IFN response signature and potentiated the expression of proteins involved in antigen processing and presentation. Immunopeptidomic analysis revealed a two-fold increase in both ligand abundance and diversity, reflecting the differential expression of proteins induced by KRAS G12D inhibition. We hypothesized that KRAS G12D signaling inhibition in preclinical models could augment immune responses initiated by immunization targeting tumor antigens in vivo via the following mechanism: the release of antigens from KRAS G12D -induced cell death, coupled with increased MHC-I expression on remaining tumor cells, would enhance interactions with T cells activated by immunization. To test this, we orthotopically inoculated C57Bl/6 mice with a murine PDAC-KRAS G12D cell line engineered to express the model self/tumor-associated antigen gp100. Mice were then immunized with either empty ionizable lipid nanoparticles (e-iLNPs) or iLNPs packaged with mRNA encoding full-length gp100 (gp100-iLNP). Prior to and during subsequent vaccine boost doses, mice were treated continuously with either KRAS G12D inhibitor RMC-9945 or vehicle. Mice treated with both gp100 mRNA-iLNP immunization and KRAS G12D inhibition exhibited significant tumor regression that was sustained over time compared to either treatment modality alone. This anti-tumor response was associated with a superior antigen-specific T cell response to subsequent immunizations, the retention of functional tumor antigen-specific cytotoxic T lymphocytes in both the tumor microenvironment and local secondary lymphoid organs, and sustained MHC-I and MHC-II surface expression on tumor cells. These preclinical findings underscore the dynamic nature by which PDAC antigen presentation can be modulated by KRAS signaling inhibition and IFNɣ. Combining allele-specific KRAS inhibition with tumor antigen immunization resulted in sustained tumor regression in vivo and was associated with the retention and function of tumor antigen-specific T cells. This rationalizes the combination of both approaches for enhanced anti-tumor activity, offering a strategy that is worth exploring clinically for improving outcomes in pancreatic cancer treatment.