CSs经由钠钾ATP酶/ERK信号轴抑制肝癌术后复发转移的活性和相关机理研究

。 . It has been shown that several cytokines associated with liver regeneration after surgical resection promote recurrence and metastasis and have poor prognosis through activation of extracellular signal regulated kinase(ERK) in hepatocellular carcinoma (HCC), especially hepatitis virus associated HCC. Previously, we found that natural cardiotonic steroids (CSs) such as ouabain and cinobufagin markedly inhibited activation of ERK and induced anoikis through their receptor Na+/K+-ATPase, suggesting that CSs may suppress the postsurgical recurrence and metastasis of HCC by targeting the Na+-K+-ATPase/ERK signaling axis. In this project, using different metastatic and hepatitis virus associated human HCC cell lines and a luciferase-labeled orthotopic xenograft model of HCC, we will screen the effective CSs on increase of anoikis and decrease of recurrence and metastasis of HCC after surgical resection from a large number of CSs. Furthermore, we will further determine the role of Na+-K+-ATPase α1, β1 subunit or ERK in CSs-mediated inhibition of recurrence and metastasis of HCC after surgical resection by overexpression or knockdown technology. Finally, we will also analyze the related signaling network of CSs-mediated inhibition of recurrence and metastasis of HCC using quantitative proteomics. CSs are the main components of the Chinese traditional anticancer drug Huachansu and two new anticancer agents CSs have also entered the Phase II clinical trial in Europe. The goal of the present project provides a basis for investigating the mechanisms underlying the action of CSs and finding novel drugs to control the recurrence and metastasis of HCC.

肝癌（尤其肝炎病毒相关肝癌）术后肝再生相关细胞因子通过激活ERK导致其复发转移，且预后差。我们前期发现天然强心甾类固醇类（CSs）如哇巴因和华蟾毒配基等经由其受体钠钾ATP酶介导，强力抑制ERK通路，诱导细胞失巢凋亡。提示CSs可通过靶向钠钾ATP酶/ERK信号轴抑制肝癌术后复发和转移。本项目应用不同转移力的和肝炎病毒相关的人肝癌细胞株及荧光素酶标记的异种原位移植肝癌裸鼠模型，进一步从数十种CSs中筛选能有效诱导肝癌细胞失巢凋亡和抑制肝癌术后复发转移的活性CSs。应用基因过表达或敲除技术深入探讨钠钾ATP酶α1、β1亚单位和ERK在介导CSs抗肝癌复发和转移中的作用，并以定量蛋白质组学方法结合生物信息学分析CSs抗肝癌复发转移的信号网络。CSs资源丰富，我国抗癌药华蟾素主成分即为CSs,欧洲2个CS单体成分新药进入II期临床，本研究为阐明CSs作用机理和研发抗肝癌复发转移新型药物奠定基础。

肝癌（尤其肝炎病毒相关肝癌）术后肝再生相关细胞因子通过激活ERK导致其复发转移，且预后差，因此，筛选能有效抑制肝癌细胞增殖和转移的天然药物并探究其作用机制具有非常重要的意义。我们前期发现天然强心甾类固醇类（CSs）如哇巴因和华蟾毒配基等经由其受体钠钾ATP酶介导，强力抑制ERK通路，诱导细胞失巢凋亡。提示CSs可通过靶向钠钾ATP酶/ERK信号轴抑制肝癌术后复发和转移。本项目应用体外实验和肝癌裸鼠实验，从数十种CSs中筛选了能有效诱导肝癌细胞凋亡和抑制肝癌转移的天然活性CS。应用基因过表达或敲除技术研究证实，钠钾ATP酶α1、β1亚单位和ERK在介导CSs抗肝癌复发和转移中发挥重要作用。以定量蛋白质组学方法结合生物信息学分析CSs抗肝癌复发转移的信号网络，发现CS新的作用靶点如极光激酶家族的极光激酶A（AURKA）,影响肿瘤细胞染色体分离和DNA损伤修复，从而影响细胞周期，抑制肿瘤生长；发现CS抑制肿瘤细胞增殖转移与Akt/mTOR信号通路有关；发现哇巴因抑制肿瘤细胞增殖与雌激素受体（ER）亚型介导的信号通路有关。本研究为阐明CSs作用机理和研发抗肝癌复发转移新型药物奠定基础。