DNA聚合酶 epsilon (Polε)催化亚基及其下游驱动基因突变在上皮性卵巢癌中的作用及分子机制

The pathogenesis of epithelial ovarian carcinoma (EOC) is a complex process with mutations in multiple genes. DNA polymerase epsilon (Pol ε) is composed of a catalytic subunit POLE1 and three other regulatory subunits, its primary function involves the processes of DNA replication and DNA mismatch repair, dysfunction of this polymerase would promote the development of tumors. Recently, we identified a high frequency of POLE1 mutation in our EOC samples for the first time, the evolutionary conservation analysis suggested that POLE1 mutation was a damaging mutation and might cause the dysfunction of DNA mismatch repair in Pol ε. However, regarding the roles of the POLE1 mutation played in the carcinogenesis of EOC, three crucial questions remain to be answered, that is, whether POLE1 mutation could promote the carcinogenesis of EOC? Whether POLE1 mutation would cause DNA mutation rate increase in the cancer genome of the affected samples? Which driver mutations would play synergetic roles with POLE1 mutation in the development of EOC? In the current project, we will perform the following assays to answer these questions: we will continue to perform the mutational screen of the POLE1 gene in Chinese patients with EOC, and confirm the potential tumor-promoting roles of POLE1 mutation via functional assays; then we will search for the global somatic mutations in the cancer genomes of the POLE1-mutated EOC patients via whole genome sequencing, then select the potential driver mutations and confirm their potential tumor-promoting roles, to test whether these mutations will play synergetic roles with POLE1 mutations in the development of EOC. These results will help us to clarify the roles and molecular mechanisms of POLE1 mutation in the carcinogenesis of EOC, and provide theoretical bases for the molecular target therapy for the patients with POLE1 mutation.

上皮性卵巢癌发病是一个多基因突变的复杂过程。DNA聚合酶epsilon(Pol ε)由催化亚基POLE1和另外3个调控亚基组成，在细胞内行使DNA复制及错配修复功能，其功能异常能诱发肿瘤。近期，我们首次在上皮性卵巢癌中发现POLE1高频突变，进化保守性分析结果表明其为有害突变，可能影响Polε的DNA错配修复功能。然而对于POLE1突变在卵巢癌发病中的作用，有3个重要问题尚不清楚：该突变能否真正促进卵巢癌发病？该突变能否增加患者肿瘤基因组突变率？该突变与其中的哪些驱动基因突变协同作用，促进卵巢癌发病？本项目拟继续检测上皮性卵巢癌患者的POLE1突变，并明确其致癌性；全基因组测序找出POLE1突变患者肿瘤基因组的体细胞突变，甄别其中的驱动突变并证实其促癌性，以验证其能否与POLE1突变协同作用促进卵巢癌发病。研究结果将明确POLE1突变的促癌性及分子机制、为患者分子靶向治疗提供理论依据。

上皮性卵巢癌发病是一个多基因突变的复杂过程。DNA聚合酶ε(Polε)由催化亚基POLE1和另外3个调控亚基组成，在细胞内行使DNA复制及错配修复功能，其功能异常往往能诱发肿瘤。本项目首先通过DNA测序法，首次在上皮性卵巢癌中发现POLE1 p.S297F高频突变，随后分别构建POLE1野生型和突变型慢病毒并分别感染卵巢癌细胞系TOV-112D后，行细胞增殖、侵袭、迁移、凋亡等细胞功能实验，结果表明：POLE1 p.S297F突变可通过促进细胞增殖而促进卵巢癌发病。利用全基因组测序技术，进一步分析含POLE1 p.S297F突变卵巢癌患者的体细胞突变谱，发现1/3的含POLE1突变患者同时存在PPP2R1A基因突变；随后分别构建PPP2R1A野生型和突变型慢病毒，分别感染POLE1不同基因型稳转细胞株后，细胞生物学实验表明，PPP2R1A突变不仅能单独促进细胞增殖，且能与POLE1突变协同作用，共同促进卵巢癌细胞的增殖；进一步的整体动物实验也证实上述结果。综上，本项目研究结果表明POLE1突变可与PPP2R1A突变协同作用，共同促进上皮性卵巢癌发病，这些结果有望为相关患者的分子靶向药物研究设计提供新思路。