KRAS withdrawal in Cholangiocarcinoma leads to immune infiltration and tumor regression
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Background and Aims
Cholangiocarcinoma (CCA) is a liver cancer with poor survival rates. Current treatments, including targeted therapies for specific mutations, are limited and benefit only a small subset of patients. KRAS mutations are found in 15-40% of CCA, representing a new potential treatment target. Whether KRAS inhibition leads to CCA tumor regression is unknown partly due to the lack of conditional animal models.
Approach and Results
We engineered a conditional Kras G12D -driven CCA mouse model by co-delivering plasmids encoding the Sleeping Beauty transposase with a luciferase reporter, a transposon-borne inducible Kras G12D transgene and Cas9 and Trp53 guide RNA into mouse liver by hydrodynamic tail-vein injection. In vivo bioluminescent imaging showed that Kras G12D withdrawal resulted in 99% tumor regression by day 7. Kras G12D withdrawal resulted in infiltration of activated CD8 + T cells by IHC and IF staining. Single cell RNA-Seq result also validated the enrichment of activated CD8 + T cells subpopulation in Kras G12D -withdrawn tumor. RNA-Seq suggested that Kras G12D withdrawal stimulated transforming growth factor beta pathway and induced senescence. We used cytokine array to characterize the secretion of pro-inflammatory factors, including IL-15 and Ccl17, upon Kras G12D withdrawal. Lentiviral overexpression of murine CCL17 delayed CCA tumor progression in a xenograft model, and overexpression of murine IL-15 resulted in tumor regression in a transplant model. Flow cytometry analysis revealed that IL-15 and CCL17 recruited and activated CD8 + T cells in CCA tumor. Expression of IL-15 resulted in blockade of tumor progression in our TKP CCA model.
Conclusions
Kras G12D withdrawal results in rapid tumor regression, highlighting the importance of oncogenic Kras in CCA tumor maintenance. Kras G12D withdrawal induces p53-independent senescence, secretion of pro-inflammatory factors, and immune surveillance by activated CD8 + T cells. We identified two secreted factors IL-15 and CCL17 that could recruit and activate T cells and control CCA tumor progression. This study underscores KRAS inhibition as a potential therapeutic approach for CCA.
