肿瘤转移相关基因MST4调控ERM细胞运动信号通路促进肝癌侵袭转移的机制研究

Metastatic recurrence of hepatocellular carcinoma(HCC) is a key limiting factor of patients’ prognosis. In order to improve the curative effect of HCC, it becomes more and more crucial to select driven genes related to metastases, reveal its molecular mechanism and find effective approaches of prevention and treatment. In the preliminary work of this group, MST4 had been approved to enhance the potential of metastases in HCC cell lines, in which feasible mechanism is that MST4 could regulate the modeling of cytoskeletion via interacting with ezrin(belongs to ERM family). Accordingly, this project is designed to demonstrate the detailed signaling pathway and molecular mechanism for the purpose of making MST4 function clear and confirm both its direct physical interaction with ezrin and influention on HCC metastasis utilizing genomic and proteomic methods, as an example, gene mutation, co-immunoprecipitation(CO-IP) and mass spectrometry(MS). Meanwhile, functional assays will be performed in vivo and in vitro. Aimed to discover the axis of MST4-ERM, the proteins which could possess the interaction with active status or inactive status of ezrin all will be investigated by IP-MS/Ms and analized by bioinformatic programme. A series of experiments will be done on Candidated protein for validation and conduction of the axis of MST4-ERM. Furthermore, applied of prospective and retrospective study on the expression level of the axis of MST4-ERM in a large scale of HCC tissues, it is intended to uncover the relationship between the axis and clinical index including metastatic recurrence and patients’ prognosis. And a prediction model about MST4-ERM axis of HCC will be constructed under this condition. After illuminating the molecular mechanism of MST modulating HCC metastasis, it will supply a slice of approvement that it is promissing to break new ground for predict the metastasis of HCC by the expression level of MST4.

转移复发是制约肝癌预后的关键因素，筛选肝癌转移相关基因、探明其分子机制是进行肝癌早期预测和干预的关键。本课题组前期工作已证实MST4可增强肝癌细胞侵袭转移能力。本项目预实验发现MST4可能与ERM家族蛋白ezrin有相互作用，故本项目将进一步通过体外、体内功能研究及基因突变技术验证二者的相互作用，明确二者在肝癌转移中的作用。进而应用免疫共沉淀、蛋白质谱技术和信号通路数据库研究分析寻找MST4-ERM下游靶蛋白，并通过体内外功能实验对其功能进行验证，阐明MST4在肝癌转移中的信号通路和作用机制。同时分别回顾性、前瞻性研究MST4、ERM及下游靶蛋白在肝癌患者肿瘤组织中的表达水平及其与肝癌转移复发及病人预后间的关系，构建肝癌转移预测模型。通过上述研究，阐明MST4调控肝癌转移潜能的分子机制及其进行肝细胞癌转移预测的可行性。

转移是制约肝细胞癌预后的关键因素，筛选肝癌转移相关基因、探明其分子机制是进行肝癌早期预测和干预的关键。本课题组前期工作已证实MST4可增强肝癌细胞侵袭转移能力。在本次研究中，课题从体外、体内和临床病理三个层面，应用PCR、蛋白印迹、免疫共沉淀等分子生物学技术和细胞工程及培养技术、肿瘤异种移植模型技术结合信息生物技术和临床病理统计的多种不同技术开展课题研究。课题组证实：① ezrin 的高表达促进肝癌细胞侵袭转移和增殖的作用；②MST4和ezrin存在相互作用且共同促进肝癌细胞的侵袭转移； ③CD44和CD54是MST4/ezrin下游的功能分子，承接MST4和ezrin的作用促进肝癌细胞的侵袭转移；④在临床肝癌标本中发现CD44和CD54、MST4和ezrin具有密切的表达相关性；⑤CD44和CD54促进肝癌侵袭转移的能力受到MST4和ezrin的抑制调节；⑥MST4可以通过MAPK-pERK信号通路活化完成对肿瘤细胞分泌炎症因子及肿瘤微环境的调节，促进肝癌的侵袭转移。通过本次研究，我们证实了肝癌细胞中MST4和ezrin是促进肝癌侵袭转移的重要分子，且他们的作用是通过下游CD44/CD54来完成。因此，我们认为MST4/EZRIN/CD44或CD54是重要的促进肝癌侵袭转移的信号通路，如过阻断该通路活化，可以有效减少肝癌肿瘤的侵袭转移，为寻找新的肝癌治疗靶向药物和预防肿瘤复发筛查模型提供理论依据。