HMGA2和FOXL2调控胃癌EMT的机制研究

Epithelial-Mesenchymal Transition (EMT) is the pivotal malignant behavior of tumors. It has been recently suggested that EMT could promote the occurrence of multidrug resistance of tumors, and it is also widely known that EMT is the key step in the initiation of tumor metastases. In the previous studies, we have established a strain of multidrug resistant gastric cancer cell line which exhibits typical EMT characteristics. Meanwhile, the invasive and migrative potential of these cells are also enhanced. The subsquent gene profiling, bioinformatic analysis and in vitro functional studies showed that HMGA2 and FOXL2 might play critical roles in this process. The immunohistochemical study of gastric cancer specimen showed that either HMGA2 or FOXL2 was inversely correlated with E-cadherin, which indicated their potential association with EMT. As a protein with conditioned transcriptional activity, HMGA2 was reported to regulate Snail1. Meanwhile, several binding sites of FOXL2 in the promotor regions of Snail1 and Twist1 were predicted. In addition, we noticed that both HMGA2 and FOXL2 were the potential targets of miR-23a/b/c and exogenous 3'UTR region of HMGA2 could significantly increase the expression of FOXL2. These facts indicated that HMGA2 might regulate FOXL2 as a ceRNA. In the present study, we aim to clarify the mechanism of HMGA2 and FOXL2 in regulating EMT and explore whether HMGA2 could function as a "ceRNA" in modulating FOXL2. This program could deepen the current understanding of EMT in gastric cancer and provide novel targets in treating gastric cancer.

EMT可促进肿瘤发生多药耐药，同时也是肿瘤发生转移的重要启动步骤。本课题组已发现胃癌细胞系SGC7901/ADR发生了典型的EMT现象，同时其耐药性和转移能力也显著增强。通过基因芯片、体外功能学实验和免疫组化，我们发现HMGA2和FOXL2可能是调控胃癌EMT的关键分子。既往研究提示HMGA2可转录激活Snail1，同时生物信息学分析提示FOXL2在Snail1和ZEB1的启动子区具有多个结合位点。我们进一步发现，HMGA2和FOXL2的3'UTR区内均存在miR-23a/b/c的结合位点，且HMGA2的3'UTR区段可正性调控FOXL2的表达。因此，我们推测HMGA2可能以ceRNA的形式调控FOXL2。本项目拟进一步明确HMGA2和FOXL2调控胃癌EMT的机制，重点阐明HMGA2对FOXL2的调控机制。该项目有助于深化对胃癌EMT的认识，发现的相关分子可为胃癌的治疗提供新的靶标。

项目以胃癌EMT为核心研究点，通过体内外功能学实验、机制学研究、临床样本评价等研究证实HMGA2-FOXL2是调控胃癌EMT的重要通路。通过基因芯片筛选和生物信息学分析锁定了8个与基因转录相关的候选分子，结合KM-plotter数据库和胃癌组织免疫组化研究，发现HMGA2和FOXL2表达与E-cadherin表达呈负相关。下调高转移潜能胃癌细胞内HMGA2和FOXL2的表达显著抑制胃癌细胞EMT，而上调低转移潜能胃癌细胞内HMGA2和FOXL2的表达显著促进胃癌细胞发生EMT。E2F1在胃癌细胞中转录活性增强，通过启动子Luciferase报告基因等检测证实FOXL2启动子区域存在E2F1的直接结合位点，提示E2F1是FOXL2的转录激活调控因子。HMGA2可与Rb发生相互作用，这种作用可解除Rb对E2F1转录活性的抑制，从而增强E2F1对FOXL2的转录活性。免疫组化研究表明HMGA2和FOXL2在胃癌组织中的表达强度呈正相关，二者在胃癌淋巴结和远处器官转移灶中的表达水平显著高于原发灶的水平，而且在伴有转移的原发灶中的表达显著高于无转移的原发灶。HMGA2和FOXL2的高表达均与胃癌患者总生存期缩短相关，提示二者是胃癌重要的预后评估标志物。