Hippo-YAP信号通路相关基因遗传变异与肝癌预后的关系及其机制研究

Hippo-YAP signaling pathway is a novel tumor suppressor pathway, which is dysregulated in hepatocellular carcinoma (HCC) and promotes cell proliferation, invasion, and drug resistance. Dysregulation of Hippo-YAP signaling pathway is associated with poor prognosis in HCC. To further investigate the biological mechanisms of its dysregulation, genomic DNA from HCC was genotyped by using the Affymetrix Genome-Wide Human SNP Array 6.0 and multiple functional SNPs related with this pathway were revealed to be associated with the prognosis. Based on these findings, we will carry out a systematic research on the associations of Hippo-YAP pathway related functional SNPs with HCC clinical outcome, in order to identify the SNPs which are significantly related with HCC progression and chemoresistance. Furthermore, these SNPs’ effects on genes expression and functions of Hippo-YAP pathway will be investigated in HCC tissue and cells to assess the associations between genotypes and phenotypes. Finally, the biologic functions of each SNPs in HCC progression and chemoresistance, as well as the underlying mechanisms will be studied in the in vitro and in vivo experiments. This project aims to clarify the biological mechanisms of genetic variations in Hippo-YAP pathway in HCC, and to provide important references on TACE efficacy and prognosis prediction.

Hippo-YAP 信号转导通路是一条新型的抑癌通路，在肝癌中该信号通路失调促使肝癌细胞获得增强的侵袭、迁移和耐药能力，与肝癌患者预后不佳、阿霉素耐药有关。为进一步研究Hippo-YAP信号通路在肝癌中失调的生物学机制，我们通过SNPs Array发现该通路相关基因多个功能性SNPs与肝癌患者预后相关。本课题将在此基础上，系统性筛选Hippo-YAP信号通路相关基因功能性SNPs，分析并获得对应基因表达水平及功能性SNPs 单独或联合与肝癌预后、TACE疗效之间关系的证据；继而通过观察组织和细胞内相关基因表达水平，综合评价基因型-表型相互关系；并联合化疗药物，对阳性关联SNPs及对应基因进行生物学研究，最终阐明其影响肝癌预后的机制。本研究将发现一些新的可影响Hippo-YAP信号通路功能的调节因子，并为肝癌预后及TACE疗效预测提供新的生物标记物。

Hippo-YAP 信号转导通路是一条抑癌通路，在本研究中我们从功能基因组学角度，鉴定并探讨Hippo-YAP信号通路相关基因功能性SNPs（single nucleotide polymorphisms）与肝癌预后、TACE（transcatheter hepatic arterial chemoembolization）治疗疗效的预测价值和其参与调控细胞表型的机制。研究内容：Hippo信号通路中各基因功能性SNPs与肝癌预后和TACE治疗疗效的关系； Hippo信号通路中功能性SNPs与对应基因在癌及癌旁组织中表达水平的相关性分析（基因型-表型分析）；阳性关联SNP 的对应基因在肝癌中的生物功能学研究。结果：利用Sequenom MassARRAY iPLEX平台对331例肝癌患者外周血DNA进行基因分型发现：（1）LATS2 rs7317471 C>T 多态性与预后显著相关(HR=0.63, 95% CI =0.46–0.87, P=0.004)；在年龄<53岁、女性、无化疗或TACE治疗史等亚组中，LATS2 rs7317471 CT/TT 基因型预后较好；（2）TEAD3 rs2076173 C和rs11756089 T是提示预后较好的保护性等位基因，具有大于1个保护性等位基因与不具有保护性等位基因的患者相比，总体生存时间显著延长(MST=19.25 vs 12.85月), 特别是在无饮酒史亚组患者中，显著性更为明显 (HR=0.48, 95%CI=0.32-0.74, P=0.001)；（3）ITGA2 rs28095 T>C与肝癌预后较差相关，Integrin α2β1在肝癌细胞和组织中广泛表达，通过与胶原细胞外基质结合，抑制MST1 激酶磷酸化并活化YAP的癌基因功能；抑制ITGA2可以抑制YAP下游靶基因表达；体外磷酸酯酶实验证明 MST1 S1180磷酸化受到ITGA2直接调控；228例肝癌组织基因表达芯片分析显示ITGA2高表达与肿瘤进展显著相关，具有ITGA2与YAP靶基因共同高表达的患者预后显著较差。科学意义：（1）建立对肝癌病人预后有预测作用的风险基因型模型；（2）率先报道了Integrin α2β1参与调控Hippo-YAP信号通路的生物学功能和调控机制，为针对该通路失调的肝癌治疗提供了新靶点。