DNMT1-miR-148a-Wnt1/10b调控轴介导的EMT促进胰腺癌“干性”转化和侵袭转移

Epithelial mesenchymal transition (EMT) is an important promoting factor on the stemness transformation and metastases of pancreatic cancer. Our previous studies indicated that up-regulation of miR-148a could inhibit the growth of pancreatic cancer, while its mechanism is still unkown. Wnt1 and Wnt10b can promote EMT process through activing Wnt/β-catenin signaling pathway. We found that Wnt1/10b were two direct targets of miR-148a by miRNA targets prediction softwares, luciferase reporter assay and pancreatic cancer cell in vitro study. The inhibition of DNMT1 activity could up-regulation the expression of miR-148a through demethylation of its gene promoter. Therefore, we deduce there may be an DNMT1-miR-148a-Wnt1/10b regulating axis to continuously downregulate miR-148a expression, upregulate Wnt1/10b expression, activate Wnt/β-catenin signaling pathyway which eventually result in the induction of EMT, promote stemness transformation and metastases in pancreatic cancer. To clarify this deduction, we plan to analysis the expression status and their relationship of each molecule of DNMT1-miR-148a-Wnt1/10b regulating axis in pancreatic cancer specimens. The regulatory relationships and the effects on induction of EMT and stemness phenotype in pancreatic cancer of DNMT1-miR-148a-Wnt1/10b regulating axis will also be determined in vitro and in vivo experiments. The study will provide new target for the treatment of pancreatic cancer by inhibition of EMT and stemness transformation.

上皮间质转化（EMT）是促进胰腺癌干性转化和侵袭转移的重要因素。课题组前期研究发现上调miR-148a可抑制胰腺癌生长，但机制尚不明确。Wnt1/10b可以激活Wnt/β-catenin信号通路而诱导EMT。我们预实验证实胰腺癌中miR-148a可靶向调控Wnt1/10b；抑制DNMT1活性可使miR-148a基因启动子去甲基化而上调其表达水平。故推测胰腺癌中存在DNMT1-miR-148a-Wnt1/10b调控轴持续下调miR-148a、上调Wnt1/10b的表达，激活Wnt/β-catenin信号通路介导EMT，促进干性转化和侵袭转移。本课题首先分析该调控轴中各分子在胰腺癌组织中表达的相关性，然后经体外实验干预调控轴中各分子的表达，明确其上下游调控关系及诱导EMT和干性表型的作用机制，最后经裸鼠移植瘤实验进一步体内验证。研究结果有望为抑制胰腺癌EMT和干性转化的靶向治疗提供新的切入点。

上皮间质转化（epithelial mesenchymal transition，EMT）是促进胰腺癌“干性”转化和侵袭转移的重要因素。近年来研究发现miR-148a可抑制胰腺癌的恶性表型，但其在胰腺癌EMT及“干性”转化中的作用及其自身表达调控机制尚不明确。本项目通过qRT-PCR检测miR-148a在胰腺癌组织和胰腺癌细胞株的表达，结果显示miR-148a在胰腺癌中表达显著降低，且与肿瘤大小、组织学分级、淋巴结转移、TNM分期及预后密切相关。进一步的体内外功能分析实验结果表明：上调miR-148a可通过Wnt1/10b介导的Wnt/β-catenin信号通路抑制胰腺癌细胞EMT和“干性”转化，同时靶向抑制DNMT1从而重新激活抑癌基因ppENK、p16和RASSF1A的表达，抑制胰腺癌细胞增殖、迁移和侵袭。研究同时证实，DNMT1高表达可导致胰腺癌细胞miR-148a基因启动子区域高甲基化，从而下调miR-148a的表达。总而言之，本研究明确了DNMT1-miR-148a-Wnt1/10b调控轴的存在及其介导的EMT对胰腺癌细胞“干性”转化和侵袭转移的影响，有助于发掘胰腺癌临床诊治的新靶点。