索拉非尼对肝癌细胞RAS-MAPK通路动态作用机制及其耐药逆转新策略的研究

Sorafenib is the only targeted drug for the treatment of advanced hepatocellular carcinoma(HCC) in China. However, drug resistance limits its clinical benefit, and the clinically translational progression about overcoming the resistance is few. Sorafenib was primarily designed to target the RAF proteins, while the underlying mechanisms about the dynamic effect of sorafenib on the RAF-MAPK pathway remain elusive in the HCC cells, and this might be the molecular basis of drug resistance. Our previous studies accidentally found that the treatment of sorafenib presented different effects at different time: short-time(≦2 Days) sorafenib treatment could inhibit RAF-MAPK signaling, while longer-time(≧3Days) treatment caused the rebound of the RAF-MAPK pathway. The previous results also indicated that the reactivation of RAS might be the initial point of the rebound process. In this study, we will further investigate the underlying mechanism about the dynamic process of the dysregulated RAS, the RAF dimerization and the reactivated MEK-ERK signaling, which was induced by the continuous sorafenib treatment in HCC cells, through utilizing several signaling-transduction related technologies. We will also validate several novel combined therapeutic strategies, which could overcome the resistance, via reversing these phenomenon. This project will provide new evidences for our deeply understanding the dynamic effect of sorafenib on HCC cells, and present novel directions and therapeutic strategies about overcoming the early-stage sorafenib resistance during the treatment of advanced HCC.

索拉非尼是目前国内治疗进展期肝癌唯一的分子靶向药，肿瘤耐药极大的限制了其临床疗效，克服其耐药的临床转化研究也鲜有进展。索拉非尼最初是针对RAF分子靶点设计的，然而其作用于肝癌细胞后RAF-MAPK通路相关分子活性的动态变化机制并不清楚，而这恰可能是其引起耐药的分子基础。项目组的前期研究意外发现，索拉非尼在肝癌中的作用随时相出现完全不同的效果：短时间处理(≦2天)可以抑制RAF-MAPK通路；较长时间处理(≧3天)却使RAF-MAPK通路出现反弹激活，而RAS的活化可能是该过程的始动节点。本课题拟进一步通过系列信号转导实验技术深入研究索拉非尼动态作用导致的肝癌细胞中RAS的异常活化，RAF的二聚化及MEK-ERK的反常激活等分子事件发生的动态过程及其机制，验证逆转该现象而克服耐药的多种全新的联合治疗方式。该研究将为我们深入认识索拉非尼在肝癌中的动态作用机制、克服其早期耐药提供新的方向和策略。

索拉非尼是晚期肝癌患者唯一可选择用于一线治疗的靶向药物，然而肿瘤耐药极大的限制了其临床疗效，克服其耐药的临床转化研究也鲜有进展。索拉非尼最初是针对RAF分子靶点设计的，然而其作用于肝癌细胞后RAF-MAPK通路相关分子活性的动态变化机制并不清楚，而这恰可能是其引起耐药的分子基础。项目组的研究显示短时间索拉非尼处理可以抑制MAPK通路的活性，而长时间索拉非尼处理则会导致MAPK通路的反弹激活，进一步的研究显示长时间索拉非尼处理会导致MAPK通路上游的RAS，RAF及MEK的活化，我们通过筛选各种不同类型的RAF抑制剂证实CRAF的同源二聚体可能主要介导了其MAPK的反弹激活。长时间索拉非尼处理引发上游KRAS的活化也介导了下游MAPK通路的反弹激活。我们利用RAF/MEK的双重抑制剂RO5126766，MEK抑制剂曲美替尼和司美替尼，及辛伐他汀分别联合索拉非尼治疗肝癌细胞，其均显示出良好的联合治疗效果。该项目将为我们深入认识索拉非尼对MAPK通路的动态作用机制提供新的见解，同时也为肝癌的治疗提供了多种新的联合治疗策略。在该项目的资助下，我们发表标注SCI论文4篇；参与申报专利1项；协助培养研究生4名（其中博士研究生2名，硕士研究生2名）；以上成果均已达到预期目标。总之，我们较好的完成了该项目的预期研究内容。