AKT/Yap协同促进肝内胆管癌形成的分子作用机制研究

Intrahepatic cholangiocarcinoma (ICC) is the second most common malignant tumor in the liver. However, the precise molecular genetic mechanisms in ICC remain to be better defined. The present study shows that the gene mutation or abnormal expression can cause cell signaling pathways out of control, resulting in abnormal cell proliferation and apoptosis, and induce the formation of liver cancer. While increasing evidence indicates that proto-oncogenes AKT and Yap are frequently overexpressed or activated in ICCs. Our previous study found that overexpression of AKT or Yap did not result in any liver anomaly in mice up to 20 weeks post injection. In striking contrast, co-expression of AKT and Yap triggered rapid ICC development in mice, and all mice developed lethal burden of liver tumor within 8 to 10 weeks post-injection.. Based on the previous research, we will set up AKT/Yap ICC mode in mice by hydrodynamic transfection, overexpression AKT/Yap in ICC cell lines by stable transfection. We will study the molecular mechanism of AKT synergizes with Yap to promote ICC development, which how to effect in AKT/mTOR and Hippo/Yap signaling pathway, fatty acid and glucose metabolism. The research results will be to further improve the pathogenesis of the ICC, and provide a new research idea for targeted against this deadly disease.

肝内胆管细胞癌（ICC）是肝脏中第二大常见的恶性肿瘤，但其分子遗传学基础仍不清楚。研究表明，基因突变或表达异常会引起肝细胞信号通路失控，从而导致肝细胞增殖与凋亡异常，最终形成肝癌。实验证据显示，致癌基因AKT与Yap经常在ICC患者中过表达或激活。我们前期预实验发现：小鼠肝脏中单独过表达AKT或Yap，转染20周均无法诱导肝癌的形成；但小鼠肝内同时过表达AKT和Yap则可协同促进ICC的快速形成，并在8--10周后发展成致命的肝肿瘤负荷。.本项目拟在前期研究基础上，采用高压尾静脉转染技术构建AKT/Yap小鼠ICC模型；采用稳定转染技术构建AKT/Yap过表达ICC细胞株；借助现代分子生物学手段，开展AKT/mTOR、Hippo/Yap信号通路及葡萄糖和脂肪酸代谢途径在AKT/Yap协同促进ICC形成中的分子作用机制研究，进一步完善ICC发病机理，为ICC靶向治疗提供新的研究思路。

肝内胆管细胞癌（ICC）是肝脏中第二大常见的恶性肿瘤，其发病机制仍未完全阐述清楚。研究表明，癌基因AKT与Yap经常在ICC中过表达或激活，并存在一定的相互作用。但是，AKT与Yap在ICC中具体发挥的作用及分子机制仍然有待深入研究，特别是进行在体实验研究。因此，本项目以临床ICC标本、小鼠和人源ICC细胞系为研究对象，采用多种现代分子生物学手段，全面系统的研究AKT和Yap在ICC形成中的作用和分子机制。研究结果表明，癌基因AKT/Yap共表达与临床ICC发生存在相关性；AKT和Yap在小鼠体内共表达可以协同促进ICC快速形成；AKT/Yap促进ICC形成的机制可能与AKT/mTOR、Hippo/Yap、MARK信号通路和糖酵解激活有关，并且mTORC1和mTORC2在ICC形成中发挥关键作用；mTOR抑制剂MLN0128在体内体外对ICC均有明显的抑制作用。本项目首次发现AKT和Yap共表达可以协同促进小鼠ICC快速形成，并阐明其形成机制，从而完善了ICC发病机理。构建的AKT/Yap小鼠ICC模型简单可靠、重复性强，可以用来系统研究ICC的体内发病机制和药物抗肿瘤作用评价。最终，本项目研究成果将帮助我们为今后的肝癌发病机制和靶向治疗奠定研究基础。