抑癌基因ECRG4在肾癌中的表达调控及功能研究

The promoter hypermethylation of novel candidate tumor suppressor gene ECRG4 is closely related with the development of a varity of tumors. However, expression and role of ECRG4 in renal cell carcinoma have not yet been studied, and effects of histone modification on ECRG4 expression have not yet been reported..Our previous work and preliminary results show that ECRG4 is highly expressed in normal renal tissue but is significantly reduced in renal carcinoma cell lines and renal clear cell carcinoma, and that its expression is negatively correlated with its promoter methylation levels. Immunohistochemical staining has demonstrated that ECRG4 protein in cancerous tissues is lower than that in adjacent normal tissues.Therefore, we come up with a hypothesis that ECRG4 is a candidate tumor suppressor gene in renal cell carcinoma. To attain this goal , we will investigate .(1) ECRG4 expression and its association with linicaopathologicalparameters in clinical samples by using real-time quantitative PCR, Western blot and immunohistochemistry;(2) its epigenetic regulation mechanisms of ECRG4 in renal cell carcinoma by bisulfite genomic sequencing and chromatin immunoprecipitation; (3) the function and role of ECRG4 in renal cell carcinoma by lentiviral vector system. All these investigations are to provide new clues for early diagnosis and gene therapy for renal cell carcinoma.

抑癌基因ECRG4异常甲基化与多种肿瘤的发生发展密切相关。但是，该基因与肾癌的关系尚无报道；也未见有关组蛋白修饰情况对ECRG4基因表达影响的研究。本项目前期工作及预实验结果显示：ECRG4基因在正常肾组织中高表达，但在肾癌细胞株和肾透明细胞癌组织中表达显著下调，且表达量与其启动子甲基化程度成负相关。免疫组化染色也显示ECRG4蛋白在癌组织中表达比癌旁正常组织低。因此，我们推测ECRG4在肾癌中是个抑癌基因。为此，本课题组拟：(1)采用RT-PCR、Western blot和免疫组化等技术，分析ECRG4基因表达及其与肾癌临床病理特征的相关性；（2 使用亚硫酸氢盐基因组测序和染色质免疫共沉淀等技术分析ECRG4基因表达的表观调控机制；（3）利用慢病毒表达系统等技术研究ECRG4基因的功能。以期初步揭示ECRG4在肾癌发生中的作用，为肾癌的早期辅助诊断和基因治疗提供新依据。

ECRG4是个候选的抑癌基因。但是其在肾癌中的表达及功能尚无研究。本课题的研究结果发现ECRG4在肾癌细胞株和肾癌组织中的表达是下调的。ECRG4表达下调同肾癌病理分级(p<0.001)、临床分期 (p=0.017)及肿瘤转移 (p=0.017)显著相关。ECRG4低表达是一个独立的肾癌不良预后因素。肾癌细胞株中ECRG4表达与其启动子甲基化状态呈反相关。过表达ECRG4可显著抑制肾癌细胞的增殖及浸润。我们的结果显示，ECRG4表达是受表观遗传调控，并且ECRG4在肾癌中可能是个抑癌基因，可作为肾癌不良预后的标志物。