miR-126/ADAM9/DNMT1正反馈环路介导的EMT促进胃癌的恶性表型和“干性”转化

Metastasis is the main cause of high mortality of gastric cancer. There is still a lack of effective therapeutic method to prevent metastasis of gastric cancer till now. It was well documented that epithelial-mesenchymal transition (EMT) played a key role in tumor malignant progression and “stemness” transformation by which gastric cancer could obtain the ability of migration and invasion to form metastases. Our previous work showed that down-regulation of miR-126 could promote the growth and metastasis of gastric cancer, but the mechanism is not clear. It is reported that A disintegrin and metalloproteinase 9 (ADAM9) could induce EMT through its degradation effect on E-cadherin (E-cad). We found that ADAM9 was a negative regulative target of miR-126 by using biological information analysis, luciferase reporter assay and gastric cancer cell in vitro study. ADAM9 has been confirmed to inhibit the degradation of DNA methyltransferase 1 (DNMT1) through EGFR/Akt signaling pathway. DNMT1 can inhibit the expression of mature miR-126 through its host gene Egfl7 promoter methylation. We deduce there may be an miR-126/ADAM9/DNMT1 positive feedback loop to continuously downregulate miR-126 expression and upregulate ADAM9 expression which eventually result in the induction of EMT and “stemness” transformation in gastric cancer. To clarify this deduction, we plan to analysis the expression status and their relationship of each molecule of miR-126/ADAM9/DNMT1 loop in gastric cancer specimens. The regulatory relationships and the effects on induction of EMT and “stemness” in gastric cancer of miR-126/ADAM9/DNMT1 loop will also be determined in in vitro experiments and the effects will be further verified by nude mice experiments. The experimental results of the current project will give clues to the treatment for gastric cancer by targeting reversion of EMT and “stemness” of gastric cancer.

上皮间质转化（EMT）在胃癌等肿瘤恶性进展乃至“干性”转化中发挥关键作用。课题组前期研究发现下调miR-126可促进胃癌的生长和转移，但机制不明确。去整合素金属蛋白酶9(ADAM9)可降解E-cadherin而诱导EMT。我们预实验证实miR-126可靶向调控ADAM9。已知ADAM9可抑制DNA甲基转移酶1（DNMT1）的降解，DNMT1可甲基化miR-126的宿主基因Egfl7的启动子而抑制miR-126的表达。故推测胃癌中存在miR-126/ADAM9/DNMT1正反馈环路持续下调miR-126、上调ADAM9的表达以促进胃癌的EMT乃至“干性”转化。本课题拟从分析环路分子在胃癌表达的相关性入手，体外实验对环路各分子实施干预，明确其上下游调控关系及诱导EMT和干性表型的作用机制，然后经裸鼠实验进一步验证。可望为胃癌的靶向治疗、抑制/逆转EMT和“干性”转化提供新的理论依据和实验支持。

胃癌是发展中国家最常见的恶性肿瘤，我国是胃癌的高发国家，其发病率和死亡率居消化道恶性肿瘤之首，据统计，大约每三分钟就有一位中国人死于胃癌。迄今尚未找到理想的治疗靶点控制胃癌的恶性进程。大量关于转移机制的研究基于实验室研究体系，例如建立胃癌细胞系并对其体内外细胞分子生物学特性进行研究，由于胃癌的浸润转移发生过程复杂、建立胃癌恶性进展模型困难，对胃癌恶行性的许多基础机制仍缺乏充分了解。上皮间质转化（EMT）在胃癌等肿瘤恶性进展乃至“干性”转化中发挥关键作用。课题组前期研究发现下调miR-126可促进胃癌的生长和转移，但机制不明确。去整合素金属蛋白酶9(ADAM9)可降解E-cadherin而诱导EMT。我们预实验证实miR-126可靶向调控ADAM9。已知ADAM9可抑制DNA甲基转移酶1（DNMT1）的降解，DNMT1可甲基化miR-126的宿主基因Egfl7的启动子而抑制miR-126的表达。故推测胃癌中存在miR-126/ADAM9/DNMT1正反馈环路持续下调miR-126、上调ADAM9的表达以促进胃癌的EMT乃至“干性”转化。本项目通过人体胃癌组织标本及体外培养的胃癌细胞研究发现，胃癌中存在miR-126/ADAM9/DNMT1反馈环路，并可下调miR-126的表达，促进胃癌的增殖、侵袭力等恶性表型，能够促进胃癌的上皮间质转化，促进干性转化。胃癌荷瘤裸鼠实验进一步验证了上述信号通路的存在，通过下调miR-126对裸鼠移植胃癌具有治疗效果。该项研究成果可望为胃癌的靶向治疗、抑制/逆转EMT和“干性”转化提供新的理论依据和实验支持。