44: Pten deletion-induced murine model of cholangiocarcinoma: a pathologic and immune analysis

Introduction: Mouse models of cholangiocarcinoma (CCA) that reliably recapitulate the characteristic stroma and tumor immune microenvironment (TIME) of human disease are critical for the success of preclinical studies exploring novel therapeutic strategies. Here, we expound on the pathologic phenotype and immune infiltrate of a recently described murine model of intrahepatic CCA induced by Pten deletion and Kras activation.


Methods: C57BL/6 mice with deletion of one (AKP) or both (AKPP) Pten alleles in the setting of Kras activation were monitored via ultrasonography for onset of tumor development, ascites, and multifocal disease. At end of life, tumors were collected for flow cytometry analysis to characterize the TIME. Additionally, immunohistochemistry (IHC) staining and digital quantification of fibroinflammatory and biliary markers were performed. IHC staining was reviewed by a board-certified pathologist.


Results: AKPP and AKP mice developed tumors at 4 and 14 weeks, respectively. After tumor onset, AKPP mice developed ascites sooner than AKP mice (2 vs 15 weeks, p< 0.01). Multifocal disease was evident in AKP mice approximately 4.5 weeks after disease onset. Histological examination of AKPP tumors exhibited a periductal growth pattern, in comparison to the mass-forming nature of AKP tumors. IHC evaluation of AKP and AKPP tumors demonstrated a fibroinflammatory reaction that recapitulates human disease (Fig 1). AKP and AKPP tumors demonstrated increased activated fibroblast (p < 0.001) and collagen deposition (p < 0.01) signatures, as well as increased immune infiltrate (p < 0.05 and p< 0.001, respectively), compared to normal littermate controls. AKP tumors had a higher degree of collagen deposition than AKPP counterparts (p < 0.01). Flow cytometry analysis of AKP and AKPP tumors demonstrated a significant myeloid cell infiltrate (85-95% CD11b+) compared to normal littermate controls (p < 0.001). Of this population, granulocytic-myeloid derived suppressor cells (G-MDSC) outnumbered monocytic-MDSCs and tumor associated macrophages (TAM) (p < 0.001). Furthermore, AKPP tumors had a higher proportion of monocytic-MDSCs and TAMs than AKP tumors (p < 0.001).


Conclusions: Mice with Pten deletion and Kras activation represent a reliable spontaneous CCA model that recapitulates human disease, with prominent myeloid expansion within the TIME. AKPP mice quickly develop a periductal infiltrating type of CCA, while AKP mice develop mass-forming disease with more desmoplasia and a longer duration to tumor onset. This model may provide insight into management strategies to treat Pten-deficient CCA.