原癌基因c-Met转录调控因子的识别及其在肺癌发生中的作用机理研究

Lung cancer is a high incidence cancer in Yunnan province, and its incidence rate is two times higher than national average. The inefficiency of the present treatment urgently asks for improvement in probing the pathogenesis of lung cancer and identifying new therapeutic targets. c-Met, a proto-oncogene, is abnormally activated in cells and tissues of lung cancer, and its activation is capable of promoting proliferation, differentiation, invasion and angiogenesis. We found that c-Met was activated by the transcriptional factor Sp1 and Smad1. But there is little known about the panorama of potential transcriptional factors of c-Met. In this study, we will systematically screen the transcriptional factors of c-Met, and explore its roles in cell proliferation, diferentiation, DNA synthesis and invasion through interference of the related transcriptional factors. We will also examine the impact of transcriptional regulation of c-Met on Xenograft model of lung cancer. This study will help to clarify as the function of transcriptional factors of c-Met in lung carcinogenesis, and also provide a new clue and the treatment for c-Met related diseases, especially lung cancer.

肺癌是云南省的高发病，其发病率是全国的两倍。目前市售的肺癌治疗药物疗效不明显，急需对肺癌的发病机理进行研究，发现新的治疗靶点。原癌基因c-Met在肺癌组织、细胞中被异常激活，并能够促进肺癌细胞的增殖、分化、侵袭和血管生成。我们的前期研究发现，转录因子Sp1和Smad1能激活c-Met的基因表达。然而至今对c-Met诸多潜在的转录激活因子仍缺乏系统研究。本研究将在申请人前期工作基础上，对c-Met的转录调控因子进行系统筛查，并对筛查到的转录因子进行干扰，研究转录调控c-Met对肺癌细胞增殖、分化、DNA合成、凋亡、侵袭转移的影响，同时在动物模型上考察转录调控c-Met对肺癌发生的影响。本研究将为阐明转录因子调控c-Met在肺癌发生中的作用奠定基础，为诠释转录激活c-Met的致癌机理提供新线索，同时为治疗与c-Met相关的疾病，特别是肺癌提供全新的靶标和方法。

原癌基因c-Met在肺癌组织和细胞中被异常激活，并能够促进肿瘤细胞的增殖、分化、侵袭转移和血管生成等过程。c-Met的异常活化通常是被转录激活所致，在c-Met基因大约2.6kb的启动子上存在多个不同的顺式作用元件，可以与多种转录因子作用。本项目首先建立了双荧光素酶报告基因检测和染色质免疫共沉淀法联合用于转录因子的筛查平台，对c-Met基因的转录因子进行系统筛查，共筛查到Sp1、Ap1（c-jun/c-fos）、Smad2和Smad3四个转录因子。之后，成功构建了稳定的过表达和沉默表达Sp1、c-jun、c-fos和Smad3的细胞系。细胞功能实验阐明了Smad3转录负调控c-Met的表达和激活，抑制肺癌H1299细胞EMT转化、血管生成、侵袭和转移的机理。明确了Sp1转录正调控c-Met的表达和激活，促进肺癌A549细胞增殖、抑制A549细胞凋亡的具体机制。诠释了c-jun转录正调控c-Met的表达和激活，促进肺癌H1299细胞增殖、抑制H1299细胞凋亡的机理。裸鼠皮下移植瘤实验表明Smad3转录负调控c-Met，能够抑制肺癌的发生、发展过程。本研究将为治疗与c-Met相关的疾病，特别是肺癌提供全新的靶标和方法。