miR-184在肾癌糖酵解代谢中的调节作用及机制研究

Cancer cells exhibit metabolism alterations characterized by increased glycolysis, but the modulation of this feature is still largely unclear. In recent years, studies have revealed important functions for miRNAs in the modulation of cellular signaling process in cancer, however, their role in the modulation of glycolysis metabolism remains largely unknown. In our prior study, we observed that aerobic glycolysis was significantly increased in the process of tumorigenesis and metastasis in the patients of renal cell carcinoma (RCC); miR-184 was shown to be significantly downregulated on tumor tissue compared with normal precancerous tissue in RCC, on the contrary, pyruvate kinase M2 (PKM2) was overexpressed; In addition, forced overexpression of miR-184 by miR-184 mimic led to a reduction of PKM2 and inhibited glycolysis in 786-O cells; Meanwhile, according to bioinformatics techniques, PKM2 is the target gene of miR-184; In the other hand, miR-184 was reported to be regulated by N-Myc in a recent study, but the mechanism is not clear. Based on these findings, in this project our study is to investigate the role of miR-184 in the modulation of glycolysis metabolism and the modulation relationship between miR-184, c-Myc and PKM2 through transfection of miR-184 mimic and inhibitors, Luciferase gene reporter gene techniques, gene overexpression and silence. Moreover, we will study the clinical and prognostic significances of these molecular markers in human RCC using our tissue and data bank of Renji Hospital.

细胞中糖酵解增加是肿瘤细胞重要的代谢特征之一，目前其调控通路未明。近年来发现miRNA在肿瘤信号传导通路中起重要调控作用，但其在糖酵解代谢中的作用有待研究。我们前期研究发现：肾癌在生长和转移过程中，无氧糖酵解代谢显著增加；在肾癌组织中miR-184明显低表达，M2型丙酮酸激酶 (PKM2)高表达，两者存在负相关；进一步在肾癌细胞株786-O中发现miR-184可以抑制糖酵解代谢，下调PKM2表达水平；另外根据生物信息学提示PKM2为miR-184靶基因；而文献报道miR-184受N-Myc调控，但具体调控及机制不明。据此我们拟应用仁济医院肾癌组织数据库，在细胞及组织水平，通过基因过表达及沉默，转染和敲低miR-184及荧光素酶报告基因方法，研究其对肾癌糖代谢的调节作用，明确miR-184与c-Myc, PKM2之间的分子调控机制，并揭示其在肾癌诊断、治疗和预后中的价值。

本项目主要从糖酵解代谢的角度角度探讨miR-184与c-Myc, PKM2之间的分子调控机制及其在肾癌诊断、治疗和预后中的价值的作用。研究内容主要包括：研究肾癌中c-Myc、PKM2蛋白、miR-184与肿瘤病理分期分级、患者预后相关性；miR-184及PKM2表达水平对肾癌细胞株糖酵解代谢及增殖、侵袭、凋亡等的影响；c-Myc-miR-184-PKM2 细胞信号调控通路肾癌糖代谢调控机制。主要研究结果：1）c-Myc在肾癌组织中表达上调，而miR-184在肾癌组织中表达下调；2）miR-184表达水平变化可以调节肾癌细胞系786-O及RCC4中糖代谢及肿瘤细胞增殖；3）miR-184表达可以特异性抑制PKM2的表达；4）敲除PKM2可以抑制肾癌细胞的糖酵解及细胞增殖；5）透明细胞癌组织标本中PKM2的mRNA表达水平和蛋白表达水平增加；本项目揭示了肾癌中miR-184与c-Myc, PKM2之间的分子调控机制，为寻找新的肾癌治疗药物靶点提供了实验依据，已发表标注受本项目资助的SCI收录期刊论文10 篇，中文核心2篇。培养博士研究生1名与硕士1名。参与国际学术交流1次并且参与特邀报告。