MiR-424-503簇在肝细胞癌中的协同抑癌功能及其上下游调节机制研究

Our previous studies showed that miRNA-mediated network plays an important role in development and progression of hepatocellular carcinoma (HCC). According to data from the miRbase resources，we found that about 30% miRNAs are polycistron. They frequently synergetically work together to drive a specific biological process. However, their regulatory mechanisms and synergetic functions are still elusive. Our preliminary data showed the existence of putative DNA binding motif of liver enriched transcriptional factor C/EBPa and CpG islands on the upstream of miR-424-503 cluster in the genome. These results indicate that loss of C/EBPa expression and DNA hypermethylation may be the important reasons for low levels of miR-424-503 in HCC. Our further bioinformatic analysis revealed that on the one side miR-424 and miR-503 have interaction target genes, and on the other side these two miRNAs can target different oncogenic pathways. In the present study, the following specific aims are proposed, 1). To identify that if C/EBPa is a direct transcriptional factor increasing miR-424-503 cluster in HCC; 2). To identify that if DNA hypermethylation is a major reason for low levels of miR-424-503 cluster in HCC. 3). To determine the roles and unknown mechanisms of miR-424-503 in HCC cells. 4). To confirm the significance of miR-424-503 cluster for prognosis in clinic. The objective of this proposal is to reveal regulatory mechanisms and synergetic functions of miR-424-503 cluster on the inhibition of hepatocellualr carcinoma. The proposed work is anticipated to define miR-424-503 cluster as a novel diagnostic marker and/or a therapeutic drug target for the treatment of advanced liver cancer.

申请人既往研究证实miRNA在肝癌的发生发展中扮演重要角色，提示以miRNA为切入点是阐明肝癌发病机制的重要环节。检索miRBase数据库，我们发现人体约30%的miRNA以成簇的多顺反子形式存在于基因组中，在进化中通过协同增效机制调控基因表达，但其在肿瘤发生发展中的功能和机制尚待深入研究。课题组前期研究结果显示：miR-424-503簇的基因组上游含有肝富集转录因子c/EBPa的DNA结合基序，同时具有CpG岛，提示DNA高甲基化和肝富集转录因子的缺失可能是该簇在肝癌中低表达的原因。生物信息学分析结果显示miR-424和miR-503存在共有下游靶点，而各自独立的靶点在调控肿瘤信号网络中又具有互补性的特点。据此，本课题希望从分子、细胞、动物模型和临床样品等层面阐明该miRNA簇在肝癌中的协同抑癌功能及其上下游调节机制，揭示成簇表达miRNA在肝癌发生发展中的信号网络并挖掘新的治疗靶标。

本项目明确了miR-424/miR-503等成簇表达miRNA分子在肝癌中的协同抑癌功能，并阐明其上下游调节机制，揭示成簇表达miRNA在肝癌发生发展中的信号网络，并探讨了其作为新的治疗靶标的可行性。在本项目系统阐明了miR-424/miR-503在肿瘤中的功能及其机制（Cancer Letters-2020），鉴定出FASN、PHF19等分子是miRNA簇可能的下游直接靶分子。同时，证明miR-29/miR-30、miR-124和miR-205等miRNA分子通过靶向赖氨酰氧化酶LOXL4、整合素ITGA3和溶血磷脂酰胆碱酰基转移酶LPCAT1等分子调控肿瘤免疫微环境重塑、肿瘤细胞抗脱落凋亡和肿瘤细胞存活。相关研究发表Theranostics（2020，封面文章）、Biochem Biophys Res Commun（2018，2020）等杂志。本项目拓展了研究内容，发现叉头转录因子FoxM1通过转录激活微管相关蛋白STMN1的表达调节细胞有丝分裂（Signal Transduct Target Ther. 2021）；明确巨噬细胞中m6A修饰动态变化是调控肿瘤免疫微环境的关键表观遗传学事件（Nat Commun. 2021）。本项目的研究共发表论文7篇，其中IF大于10的论文3篇，IF大于5的论文1篇。期间共招收博士研究生7人、硕士研究生3人、博士后2人。1人获2021年度国家博士生奖学金，1人获2021年度校级优秀博士生课题资助。