Akt/Notch activation drives rapid cholangiocarcinoma development originating from mature hepatocytes in the mouse

Abstract

Introduction: Intrahepatic cholangiocarcinomas (ICCs) are primary liver tumors with a dismal prognosis. Activated AKT/mTOR pathway promotes both HCC and ICC development in the mouse liver. Notch signaling is known to play a critical role during bile duct development. Here we hypothesized that activated Notch signaling converted AKT-overexpressing preneoplastic hepatocytes into ICC in vivo. Methods: Activated Notch (NICD) alone or together with the activated form of AKT (myr-AKT) were stably transfected into the mouse liver via hydrodynamic injection. Dominant negative form of RBP-J (dnRBP-J) was co-injected with AKT to block canonical Notch pathway in the mice. Notch and AKT pathway status was determined in human ICC samples by Western blotting and immunohistochemical staining. Results: Overexpression of NICD alone induced cystic cholangiocellular tumors, including cystic ICCs, over long latency. Concomitant activation of Notch and AKT signaling synergized to promote rapid cholangiocarcinogenesis in mice. Intriguingly, we found that the ICC tumor cells originated from mature hepatocytes. Using a mouse model of hepatocyte fate tracing in combination with morphological and ultrastrutural evaluation by light and electron microscopy, we demonstrated that Notch and AKT coexpression transformed normal hepatocytes into ICCs. The hepatocyte-cholangiocyte conversion occurred at the earliest stages of the tumorigenic process, supporting that it is the critical step leading to the malignant initiation and subsequent progression. Mechanistic studies revealed the increase in both cell proliferation and glycolysis in AKT/NICD induced ICC lesions. Furthermore, blocking canonical Notch signaling with dnRBP-J in AKT-injected mice prevented ICC, but not HCC development in these mice, supporting that AKT-induced ICC development was dependent on Notch signaling. Finally, coordinated activation of AKT and Notch signaling was detected in a subset of human ICC samples. Conclusion: Our studies suggest that hepatocytes can be the cellular origin of ICC in mice and possibly also in humans. Activated AKT and Notch signaling pathways have critical roles during ICC development, and targeted therapy against these pathways may be useful in treating human ICC.