miR-15a/16在胃癌中的功能及作为分子诊断标志物的应用研究

Gastric cancer is one of the most common malignancies worldwide and is the second most frequent cause of cancer related death. A variety of genetic and epigenetic aberrations underlie development abnormality of gastric cancer. In our previous study, we have validated that YAP1 (Yes-associated protein 1) is involved in gastric tumorigenesis and functions as an oncogene. Recently we found that miR-15a and miR-16 could down-regulate YAP1 expression in gastric cancer cell lines, AGS and MKN1. However, the expression level and function of miR-15a/16 together with its application value for clinical gastric cancer patients are still unclear...In this study, we will focus on the following issues on the former basic research. First, we will explore the expression level of miR-15a/16 in gastric cancer cell lines and clinical samples. We will evaluate the correlation between miR-15a/16 expression and other clinical parameters to check if miR-15a/16 could be a prognostic maker for gastric cancer. Also, the downregulation expression mechanism of miR-15a/16 in gastric tumors should be investigated. Second, we will overexpress or knockdown miR-15a/16 in gastric cancer cell lines to observe the influence of its expression on cell proliferation, monolayer colony formation ability, cell invasion ability, cell distribution in cell cycle, xenograft proliferation in animal model. Also, we would investigate the possible signaling pathways miR-15a/16 might be involved in. Third, we will evaluate the expression correlation between YAP1 and miR-15a/16 and confirm YAP1 could be regulated by miR-15a/16 through the seed region in its 3' untranslated region (3'UTR), which has two complementary binding sites for miR-15a/16 recognize. Finally, we will continue to investigate other putative target genes of miR-15a/16, such as cyclin D1, Cyclin D3 and Cyclin E1. ..All these will enrich the understanding on microRNAs in gastric cancer and the regulation mechanism of YAP1.

胃癌是我国常见肿瘤，YAP1在胃癌发生过程具备癌基因功能。我们最新发现miR-15a/16 可调控YAP1 表达，但miR-15a/16在临床胃癌标本中的表达水平、应用价值及在胃癌发生过程中的功能尚不清楚。本课题拟在已有实验基础上进行如下研究：1）探讨miR-15a/16 在胃癌中的的表达，分析与胃癌临床参数的相关性；研究miR-15a/16的表达下调机制；2)在胃癌细胞中过表达或沉默miR-15a/16，观察对细胞增殖及凋亡、集落形成能力、侵袭能力、细胞周期及对裸鼠皮下成瘤性的影响，从而揭示其功能；3）在胃癌细胞与标本中研究miR-15a/16 和YAP1 的表达关系，确定miR-15a/16结合YAP1的3'非翻译区识别位点并下调YAP1的表达；4）验证miR-15a/16其它可能靶基因（CCND1、CCND3和CCNE1），再用共转染试验证实miR-15a/16和可能靶基因的关系。

胃癌是一种世界性恶性肿瘤同时也是导致死亡率第二位的癌症，该癌症好发于中国、日本、韩国及东亚地区。在胃癌的发病机制中，微小RNA的表达失调扮演了重要角色。在该项国家自然科学基金的资助下，我们对微小RNA-15a/16在胃癌的的表达以及在胃癌发病过程的功能进行了深入研究。我们发现miR-15a/16在胃癌组织中相对于癌旁相对正常组织表达下调；外源性过表达miR-15a/16 可通过负调控YAP1发挥抑癌功能，而且两者在胃癌中的表达呈负相关；同时miR-15a/16尚可靶向CCND1、CCND3和CCNE1等细胞周期蛋白。我们的研究结果首次证实miR-15a/16为抑癌微小RNA，其表达下调促进了胃癌的发生，这一发现将为临床转化及治疗干预提供实验基础。