FoxM1c-miR-149-FoxM1b/1c负反馈环路在结肠癌血管生成中的调控作用及分子机制研究

The cross-regulation between target genes and microRNA is involved in many human physiological and pathological processes, whereas its roles in colon cancer angiogenesis remain further exploration. As we reported, the up-regulation of transcription factor forkhead box M1 (FoxM1) promoted the formation of malignant phenotypes of proliferation, invasion and metastasis in colon cancer, which was negatively regulated by miR-149. However, the functions and related molecular mechanisms of FoxM1 in colon cancer angiogenesis are still unclear. Our team has found out that the level of FoxM1 positively correlates with those of microvessel density, VEGF and MMP-2 in cancer tissues, and that two potential FoxM1c-binding but not FoxM1b-binding sites exist in the core promoter region of miR-149 by bioinformatics softwares. Therefore, a scientific hypothesis is raised that a negative feedback loop of FoxM1c-miR-149-FoxM1b/1c should play key roles in colon cancer angiogenesis. In this study, many techniques including site mutation, chromatin immunoprecipitation, gene over-expression and RNA interference are to be used in cultured cell lines and small live animal models, respectively, to deep elucidate the roles and molecular mechanisms of the mentioned negative feed back loop. Our study may help to provide theoretical evidences for further screening and developing novel therapeutic anti-angiogenesis targets against colon cancer.

靶基因-miRNA交互调控参与人多种生理和病理过程，其在结肠癌血管生成中的作用有待深入探索。本课题组前期已报道：转录因子叉头框蛋白M1(forkhead box M1,FoxM1)促进结肠癌增殖、侵袭和转移等恶性表型形成，并受miR-149负向调控。然而，FoxM1在结肠癌血管生成中的作用及分子机制并不清楚。课题组预实验发现：FoxM1与结肠癌组织微血管密度、VEGF和MMP-2表达正相关；软件分析显示miR-149核心启动子区含两个FoxM1c结合位点而没有FoxM1b结合位点。基于此，我们提出科学假说：FoxM1c-miR-149-FoxM1b/1c负反馈环路可能是结肠癌血管生成的重要分子机制。本研究拟采用位点突变、免疫共沉淀、基因过表达及RNA干扰等技术，分别从细胞和活体动物水平深入阐明上述负反馈环路在结肠癌血管生成中的作用和分子机制，为进一步筛选和开发抗血管治疗新靶标提供理论依据。

血管生成对肿瘤的生长和转移具有重要作用，而结肠癌血管生成的调控机制尚待探索。本项目通过分子生物学、细胞水平、动物水平等实验，筛选出了靶向FOXM1的关键miRNA--miR-6868-5p，发现miR-6868-5p通过靶向FOXM1进而抑制IL-8产生，从而抑制结肠癌血管生成。此外，进一步研究表明FOXM1通过促进EZH2表达，增加pri-miR-6868启动子区H3K27me3抑制miR-6868-5p表达，从而形成反馈环路。本课题揭示了miR-6868-5p与FOXM1间的调控环路，并阐明了该环路在结肠癌血管生成中的作用。研究结果为结肠癌血管生成提出了新的机制，并为结肠癌治疗提供了潜在的治疗靶点。本项目资助获得的研究成果已发表SCI论文6篇，其中影响因子5.0以上4篇；参加国内学术交流一次，获大会优秀论文；培养硕士研究生4名，博士研究生1名。