FoxO3a-FoxM1反馈调控环路在胃癌侵袭转移中的作用及分子机制研究

The death of patients with gastric cancer ascribe to invasion and metastasis.previous studies indicated that FOXO3a an FOXM1 played an pivotal role in invasion and metastasis. Our research found that the expression of FoxO3a and FoxM1 in gastric cancer tissue is negatively related. In addition, knock-down FoxO3a induces the expression of FoxM1 and overexpression of FoxM1 increases the ubiquitin-dependent degradation of FoxO3a via inhibiting the expression of USP9x. According to the result above, we hypothesis that in gastric cancer, the low level of FoxO3a increases the expression of FoxM1, and high level of FoxM1 ulteriorly accelerate the degradation of FoxO3a via depressing USP9x,through forming a FoxO3a-FoxM1feedback regulation loop, promote the invasion and metastasis of gastric cancer via induce the expression of metastasis-related genes. In this study, we will take Dual-luciferase report assay, EMSA,ChIP to verify whether FoxM1 is the target gene of FoxO3a and the mechanism that FoxM1 increases the ubiquitination of FoxO3a. Thereby, illuminate the role of FoxO3a-FoxM1feedback regulation loop in the invasion and metastasis of gastric cancer and provide a theoretical basis for the prevention and treatment of gastric cancer invasion and metastasis.

侵袭转移是胃癌患者死亡的重要原因,文献表明FoxO3a及FoxM1分别抑制和促进侵袭转移。课题组前期发现FoxO3a与FoxM1在胃癌组织中表达负相关,沉默FoxO3a或上调FoxM1均可促侵袭转移。干扰FoxO3a显著增加FoxM1转录活性,过表达FoxM1显著抑制去泛素化酶USP9x,从而促FoxO3a泛素化降解。课题组推测,沉默FoxO3a可削弱其对FoxM1的转录抑制,从而上调FoxM1,而上调的FoxM1可通过抑制USP9x促FoxO3a泛素化降解,形成FoxO3a-FoxM1反馈调控环路,最终上调转移相关基因表达促胃癌侵袭转移。本项目拟通过双萤光素酶,EMSA,ChIP等实验,证实FoxM1是否为FoxO3a直接靶基因,以及FoxM1促FoxO3a泛素化的机制,从而阐明FoxO3a-FoxM1环路在胃癌侵袭转移中的作用,为靶向该环路作为防治胃癌侵袭转移策略提供理论依据。

项目背景：最新研究表明胃癌的发病率及死亡率分别占我国恶性肿瘤的第二位，文献表明FOXO3a及FOXM1分别抑制和促进肿瘤发生发展。课题组主要研究结果发现：1.FOXO3a与FOXM1在胃癌组织中表达负相关，沉默FOXO3a或上调FOXM1均可促进胃癌发生发展；2.干扰FOXO3a显著增加FOXM1表达，而过表达FOXO3a显著抑制FOXM1表达；组构建了FoxM1 的启动子活性报告质粒，通过双萤光素酶报告基因实验发现干扰FOXO3a显著增强了FoxM1 的启动子活性，而过表达FOXO3a显著抑制FOXM1启动子活性。ChIP实验进一步表明FOXO3a可以直接与FOXM1启动子区结合；3. FOXM1可以与PDK1结合，从而促进PDK1与AKT结合，进而激活AKT1，而激活的AKT可进一步促进FOXO3a的磷酸化(P-FOXO3a(S253), P-FOXO3a(T32)), 进而导致FOXO3a的泛素化降解；4.体外激酶实验表明FOXM1可促进AKT激酶活性；5. FOXO3a抑制PD-L1表达，沉默FOXO3a促进PD-L1表达；而FOXM1可显著促进PD-L1表达，沉默FOXM1可以抑制PD-L1表达；6. 在胃癌组织中，FOXM1表达与PD-L1负相关，而FOXO3a与PD-L1正相关。上述研究结果提示FOXO3a-FOXM1-PD-L1信号轴可以调控免疫应答，从而影响肿瘤发生发展。阐明FOXO3a-FOXM1环路在胃癌侵袭转移中的作用,为靶向FOXO3a-FOXM1-PD-L1通路的抗胃癌策略提供新策略。