基于RNA-Seq卵巢癌高易感基因EPS8L1的验证及研究

we previously used the next generation sequencing technology (NGS) to perform transcriptome sequencing (RNA-Seq) on tumors and normal ovarian tissues in 31 patients with epithelial ovarian cancer. Bioinformatics analysis display that epidermal growth factor receptor pathway 8-related protein 1 (EPS8L1) was significantly up-regulated in epithelial ovarian cancer tissues (20.266-fold) compared with normal ovarian tissues. Through database comparison and document retrieval, we speculated that EPS8L1 may be closely related to the invasion and metastasis of ovarian cancer，and conducted a preliminary verification. On the basis of the previous research, this project is planned to detect the expression level of EPS8L1 by collecting clinical specimens from ovarian and normal ovarian tissue (included paraffin-embedded specimens ),and combined with follow-up of patients, to explore the correlation between EPS8L1 and clinicopathological features of ovarian cancer. To investigate the effects of EPS8L1 on the invasion, metastasis and tumorigenesis of ovarian cancer and related mechanisms by gene editing techniques to construct overexpression or Knockdown EPS8L1 Gene Cells and Animal Models.Using the in vitro and in vivo models constructed to detect the sensitivity of commonly used chemotherapy drugs for ovarian cancer, and to explore the potential value of EPS8L1 in the clinical drug selection of ovarian cancer..This study is of great significance for clarifying the function and mechanism of EPS8L1 gene in the development of ovarian cancer, exploring its application value in clinical chemotherapy of ovarian cancer, optimizing the existing chemotherapy regimen, and improving the status of ovarian cancer treatment.

本课题组前期利用二代测序对31例上皮性卵巢癌患者的肿瘤和正常卵巢组织进行转录组测序（RNA-Seq），生物信息学分析显示，表皮生长因子受体通路底物8相关蛋白1（EPS8L1）在上皮性卵巢癌组织中较正常卵巢组织明显上调（20.266倍）。通过数据库及文献检索，推测EPS8L1与卵巢癌侵袭转移密切相关，并进行了初步验证。本课题拟收集临床卵巢癌和正常组织标本，检测EPS8L1表达水平，结合随访探讨EPS8L1与卵巢癌临床特征相关性；应用基因编辑技术构建EPS8L1基因过表达或敲低细胞和动物模型，探索EPS8L1对卵巢癌侵袭、转移及肿瘤发展的影响及机制；利用构建体内外模型，对卵巢癌临床常用化疗药进行药敏检测，探讨EPS8L1在卵巢癌临床药物选择方面的潜在价值。本研究对于明确EPS8L1在卵巢癌发展中的功能及机制、探讨其在卵巢癌临床化疗中的应用价值，优化现有化疗方案、改善卵巢癌治疗现状具有重要意义。

卵巢癌（OC）是女性生殖系统三大恶性肿瘤之一，死亡率居妇科肿瘤之首。上皮性卵巢癌（EOC）是最常见类型，预后最差。其发病隐匿，临床现有标记物在其早期诊断、预后判断等方面的作用仍有限。因此发现新的EOC生物标记物并探讨其机制，将为早期诊断、治疗、预后评估提供帮助。.课题组前期利用二代测序对31例EOC患者的肿瘤和正常卵巢组织进行RNA-Seq，结果显示EPS8L1在EOC中较正常卵巢组织上调20.266倍。数据库及文献检索显示EPS8L1与卵巢癌侵袭转移密切相关。.本课题对EPS8L1在卵巢癌中的表达及与患者临床病理学指标之间的关系，构建体内外模型对EPS8L1在OC细胞增殖、迁移、转移中的生物学功能及机制，对化疗药物敏感性影响等方面进行了较为深入的研究，取得以下成果：1. EPS8L1表达与患者年龄、肿瘤部位（单/双侧）等无关，而与类型、淋巴结转移和FIGO 分期等病理指标相关。高表达组OC患者生存期明显降低。同时外周血中EPS8L1-cfDNA 的水平亦与肿瘤发生发展相关，EPS8L1有望成为EOC新的生物标志物。2. EPS8L1在OC细胞株中普遍高表达；促进OC细胞ES-2和SKOV3的细胞周期、增殖、迁移；明显增强SKOV3细胞的体内成瘤能力；促进OC细胞ES-2和SKOV3对OC一线化疗药物铂类和紫杉醇的耐药。3. EPS8L1促进OC细胞的体内转移，此功能主要通过胞外信号如EGF.增强EPS8L1磷酸化，激活下游TIAM2功能，使其发挥特异GEF活性，进而激活RAC1-GDP向RAC1-GTP转化，调节下游MAPK级联信号，直接影响细胞骨架重塑，最终促进OC细胞转移。此外，EPS8L1促进OC细胞迁移转移的作用可能还与影响基质金属蛋白酶2和9（MMP2 和MMP9）的表达有关。.这些结果对于认识EPS8L1基因在卵巢癌进展、转移及化疗耐药中的作用，寻找卵巢癌诊断、治疗和预后的生物标记物提供科学依据，是对卵巢癌研究的重要补充。