miR-506靶向UHRF1抑制肾癌细胞增殖与转移的作用机制研究

Renal cell carcinoma (RCC) is the most common urologic malignancy all over the world and clear cell RCC (ccRCC) is the largest subtype of RCC. microRNAs (miRNAs) are a family of single stranded non-coding RNAs that play critical roles in carcinogenesis. However, the association between RCC and miRNAs are still largely unknown. Our previous study revealed that miR-506 decreased expression in ccRCC and enforced expression of miR-506 could inhibit renal cancer cell proliferation and invasion. Further studies showed that UHRF1 is a direct target of miR-506 which is an essential regulator for DNA methylation. mRNA array showed that 6 non-clustered PCDHs was collectively up-regulated in 786-O-miR-506 group. Therefore, we hypothesis that miR-506 may promote re-expression of non-clustered PCDHs through demethylation of its promotor by directly targeting UHRF1. To test the hypothesis, we will establish miR-506 over-expression model in renal cancer cell lines and combined with clinical samples to investigate the biological function of miR-506 in RCC. Clarify the underline mechanisms of miR-506 regulating UHRF1 on RCC tumorigenesis by regulation the promoter methylation of non-clustered PCDHs. Clarify the clinical significance of miR-506-UHRF1 axis in RCC. Our study will provide a new insight into renal cancer diagnosis and therapy.

肾透明细胞癌(ccRCC)占转移性肾癌的90%，是肾癌死亡的主要病理类型。目前miR-506在肾癌中的作用机制仍不清楚。本课题组前期研究发现miR-506在ccRCC中低表达，过表达miR-506能降低肾癌细胞的增殖与侵袭能力，并进一步确定了甲基化调节基因UHRF1为其直接靶基因。接下来芯片结果表明过表达miR-506促进非成簇PCDHs高表达。我们以此为基础提出：miR-506靶向作用UHRF1，从而降低非成簇PCDHs启动子甲基化，促进其再表达，从而抑制肾癌细胞的增殖与转移。为验证假说，本课题拟建立miR-506过表达肾癌细胞模型，结合临床样本，探讨miR-506在肾癌中的生物学功能，阐明miR-506调节UHRF1介导非成簇PCDHs启动子甲基化在肾癌发生中的分子机制，明确miR-506-UHRF1轴在肾癌发生中的临床意义，为肾癌诊治提供新思路。

肾细胞癌是泌尿系统常见的恶性肿瘤。由于肾癌对放疗、化疗等常规肿瘤治疗手段反应差，因此肾癌的早期诊断及治疗具有重要的临床意义。本项目按原计划进行，进展顺利。我们首先构建了稳定miR-506过表达的肾癌细胞模型，通过细胞学实验（MMT,迁移，侵袭等实验）研究了miR-506对RCC生物学功能的影响。进一步建立肾癌裸鼠模型，研究其对RCC体内生物学功能的作用。结果显示miR-506在肾癌中发挥抑癌基因的作用。建立UHRF1稳定低表达肾癌细胞模型，研究UHRF1对肾癌细胞增殖、迁移，侵袭等生物学功能的影响以及对PCDH10/PCDH17表达及其甲基化的影响，结果显示UHRF1在肾癌中发挥癌基因的作用，并且降低肾癌细胞中的UHRF1表达可以降低PCDH10/PCDH17的甲基化水平，从而使其高表达。通过荧光素酶报告基因等实验方法进一步明确了miR-506对UHRF1的直接靶向调控作用（miR-506-UHRF1轴的确立）。并进一步研究了miR-506-UHRF1轴对PCDH10/PCDH17甲基化的影响，结果显示miR-506高表达能够降低PCDH10/PCDH17甲基化水平，从而增高其表达影响肾癌的增殖和侵袭能力。最后扩大肾癌患者数量，完善病人信息，进一步研究miR-506-UHRF1-PCDH10/PCDH17轴在肾癌患者中的临床意义。结果显示，miR-506能够通过直接调控UHRF1，影响PCDH10/PCDH17甲基化，从而对肾癌的增值及侵袭过程进行调控，提示miR-506-UHRF1-PCDHs轴可望成为肾癌治疗的重要靶点，值得进一步研究。