STAT3调控miR-125a-5p与肝癌对sorafenib治疗敏感性的机制研究

Sorafenib is the only effective molecular targeted drug for advanced hepatocellular carcinoma, however,no molecular marker is available to predict effecacy of sorafenib.The interplay between STAT3 and microRNA has gain more and more attention in terms of resistance to chemotherapy and molecular targeted therapy. STAT3 is one of the main target of sorafenib. Our previous study demonstrated that downregulation of STAT3 and HTATIP2 contributed to resistance to sorafenib by promoting tumor invasiveness and metastatic potential. However, the undelying mechanism by which STATE regulated HTATIP2 and other anti-oncogene is still unclear. The relationship between STAT3 and microRNA has become a hotspot and the interplay between STAT3 and microRNA play important role in resistance to molecular targeted therapy and chemotherapy. By using high-throughout techinique, we screeened a number of microRNAs which are upregulated in sorafenib-resistant tumor. And by applying bioinformatic mathod, we identified potential microRNAs that can be regulated by STAT3, and can regulate HTATIP2 and P53. microRNA-125a-5p was identified as microRNA of interest. The current study will focus on the regulatory function of STAT3/pSTAT3 on miR-125a-5p and its downstream molecular pathway. By up-regulating STAT3 and knockdown of STAT3, we examined expression of microRNA-125a-5p; and by up-regulating microRNA-125a-5p by pre-microRNA-125a-5p and down-regulating microRNA-125a-5p by anti-microRNA-125a-5p, we confirm the regulatory function of microRNA-125a-5p to HTATIP2/P53 and their downstream pathway. The cellular function assay and in vivo xenograft model were applied to evaluated invasiveness of hepatocellular carcinoma and confirm the pro-tumoral effect of microRNA-125a-5p.In terms of translational significance, we will finally classify HCC patients by specific molecular markers including STAT3/microRNA-125a-5p/HTATIP2/P53 and downstream molecular of HTATIP2/P53. By predicting more precisely the sensitivity and resistance of hepatocellular carcinoma to sorafenib, we will treat patients with hepatocellular carcinoma individually.

肝癌唯一有效的全身靶向药物Sorafenib缺乏有效分子标志物指导个体化治疗。目前STAT3调控微小RNA(microRNA)与恶性肿瘤化疗及靶向治疗敏感性已成研究热点。本课题组前期发现STAT3下调HTATIP2导致肝癌侵袭转移潜能增强参与sorafenib耐药机制，但STAT3通过何种机制调控HTATIP2并引起肝癌生物学行为改变的深入机制尚不清楚。我们推测STAT3通过microRNA调控HTATIP2等抑癌基因导致肝癌对sorafenib耐药。本课题通过高通量手段并借助生物信息学方法筛选并锁定调控HTATIP2的miR-125a-5p，深入研究STAT3对miR-125a-5p及下游通路的调节机制。通过过表达和下调STAT3证实STAT3对miR-125a-5p表达的调控作用；通过上调/下调miR-125a-5p证实对HTATIP2、P53及下游分子的调控作用。

一、.研究背景：. 索拉非尼是晚期肝癌的标准全身治疗药物，目前仍缺乏有效预测sorafenib疗效的分子标志物。微小RNA（MicroRNA，miRNA）是一种小的内源性非编码RNA分子，大约由21-25个核苷酸组成，与其靶基因在生物体内形成复杂的调控网络。以microRNA为突破点进行干预为克服肿瘤对靶向药物耐药提供新的途径。从sorafenib治疗的肝癌患者中筛选出16种与sorafenib治疗敏感性相关的microRNA分子，通过生物信息学分析和RT-PCR验证认为microRNA-125a-5p可预测sorafenib疗效。.二、.研究内容：.1.扩大样本量验证miR-125a-5p低表达与sorafenib治疗敏感有关；.2.分别敲减和过表达miR-125a-5p，在体外检测Sorafenib对肝癌细胞增殖能力、侵袭运动能力和凋亡的胰腺。在体内实验中证实；.3.探索miR-125a-5p调控STAT3与sorafenib治疗敏感性的机制。.三、.重要结果.1.在临床病例中证实miR-125a-5p低表达与sorafenib治疗敏感有关。.2.在SMMC7721和LM3，敲低miR-125a-5p后SMMC7721-KD对sorafenib治疗敏感性升高；而过表达miR-125a-5p后LM3-OE对sorafenib治疗敏感性降低。.3.通过高通量的全基因组芯片，筛选miR-125a-5p过表达后的mRNA表达水平变化，发现miR-125a-5p可抑制IL-1B/STAT3/IL6R并调节肿瘤微环境中的炎症细胞和炎症因子。.四、.关键数据及科学意义.1.从临床、细胞、动物模型三个层面，证实证miR-125a-5p低表达与sorafenib治疗敏感有关。阐明miR-125a-5p调控STAT3与sorafenib治疗敏感性；认为miR-125a-5p抑制IL-1B/STAT3/IL6R并调节炎症细胞和炎症因子。.2.揭示miR-125a-5p作为sorafenib疗效的分子标志物，为其他肝癌相关分子靶向药物的个体化治疗提供科学依据。