LncRNA MALAT1通过抑制miR-140上调Aurora-A促进肝癌索拉非尼耐药的机制研究

Sorafenib leads to survival benefit in patients with advanced hepatocellular carcinoma but its use is hampered by the occurrence of drug resistance. The overespression of Aurora-A, an important member of the serine/threonine kinase family, is closely correlated with the sorafenib resistance in hepatocellular carcinoma. In our previous studies, we have developed a sorafenib-resistant human liver cell lines, in which we found that the expression levels of Aurora-A and lncRNA MALAT1 were both significantly increased, while the expression of miR-140-5p targeting Aurora-A was dramatically reduced. Inhibition of MALAT1 could lead to downregulation of Aurora-A. However, their relationship is still unclear. In this study, we will explore the role of MALAT1 in sorafenib resistance in vitro and in vivo. Moreover, we will investigate the precise mechanisms by which MALAT1 interacts with miR-140-5p on the basis of bioinformatic analysis using dual-luciferase reprorter gene analysis, RIP-seq and RNA pull-down. Our works will help to further clarify and improve the molecular regulatory networks of Aurora-A and provide new strategies for overcoming sorafenib resistance in hepatocellular carcinoma.

索拉非尼是晚期肝癌患者的一线用药，但耐药的出现已成为阻碍肝癌患者总生存期进一步延长的主要因素。Aurora-A是丝氨酸/苏氨酸激酶家族的一个重要分子，其表达紊乱与肝癌索拉非尼耐药密切相关。本课题组前期成功建立了肝细胞癌索拉非尼耐药细胞模型，与亲本细胞相比Aurora-A和lncRNA MALAT1表达明显增高，而靶向Aurora-A的miR-140-5p表达则降低，干扰MALAT1可抑制Aurora-A表达，但三者的关系目前尚不清楚。本课题拟从细胞、活体动物和组织水平探讨MALAT1在肝细胞癌索拉非尼耐药中的作用，并在前期生物信息学分析的基础上，通过荧光素酶报告基因实验、RNA免疫共沉淀及RNA pull down验证MALAT1通过抑制miR-140-5p上调Aurora-A的分子机制，从而进一步阐明肝细胞癌中Aurora-A过表达的分子调控网络，为临床逆转肝癌索拉非尼耐药提供新的思路。

长期非编码RNA（lncRNAs）已被证实在包括肝细胞癌（HCC）的多种恶性肿瘤的发生和发展中发挥关键作用。研究表明lncRNA MALTA1是肝癌的致癌基因和预后生物标志物。本实验中，我们通过分析lncRNA差异表达谱，证明在索拉非尼耐药的HCC细胞中MALAT1的表达明显升高。敲除MALAT1可促进肝癌细胞耐药株对索拉非尼的敏感性，而MALAT1过表达则促进肝癌细胞对索拉非尼的耐药。功能学实验表明，MALAT1可促进癌细胞的增殖、迁移和EMT过程。此外，动物学实验也表明敲除MALAT1表达增强了索拉非尼的体内抗肿瘤作用。进一步研究表明，MALTA1通过与miR-140-5p结合调控Aurora-A的表达，从而促进肝癌细胞对索拉非尼的抵抗。在肝癌患者中，MALAT1的表达与miR-140-5p的表达呈负相关，但与Aurora-A的表达呈正相关，MALAT1过表达与肝癌患者的不良预后和索拉非尼反应密切相关。这些结果表明，MALAT1可能是肝癌患者的预后指标和逆转索拉非尼耐药的治疗新靶点。