转录因子E2F1在胃癌免疫序贯化疗中的双调控作用及机制研究

Immune sequential chemotherapy is one of the most promising treatment for gastric cancer. In view of our previous studies, It have found that transcriptional factor E2F1 participate in the regulation of gastric cancer cells and dendritic cells function, we did the following design: (1) through the study patients serum, pathology and E2F1 expression in correlation with immune sequential chemotherapy;(2) various types of E2F1 carrier transfect gastric cancer cell lines and dendritic cells, to comparison of the effect of reverse drug resistance in gastric cancer cells by directly silent E2F1 and the effect of the immune response stimulates by E2F1 overexpression as a benefit of target protein, and respectively to observe the gastric cancer drug resistant cells and dendritic cells function change;(3) gastric cancer liver metastasis model, studies the E2F1 expression type of gastric cancer drug resistant cell lines in the immune status, tumor cell metastasis, drug resistance, the change of the respect such as immunity, and in vitro experiments;(4) to establish a DNA, protein, pathological tissues, serum specimens library, using chip technology and molecular biological informatics technology search the common channel of regulation drug-resistant tumor cells and dendritic cells, authentication key miRNA, clarify E2F1 mechanism in immune sequential chemotherapy for gastric cancer.

胃癌的免疫序贯化疗是最具希望的胃癌治疗手段之一，本项目针对前期研究发现转录因子E2F1同时参与调控胃癌细胞耐药和树突状细胞功能表达的特点：（1）通过临床病例研究E2F1表达同免疫序贯化疗的相关性；（2）构建各类型E2F1载体转染胃癌耐药细胞株、树突状细胞，比较在胃癌耐药细胞中直接沉默E2F1逆转耐药和过表达E2F1靶蛋白激发免疫反应的获益，分别观察胃癌耐药细胞和树突状细胞功能变化，了解E2F1在免疫序贯化疗体外实验中的作用；（3）建立胃癌肝转移模型，研究各E2F1表达类型的胃癌耐药细胞株在免疫状态下，细胞耐药、免疫等方面的变化，与体外实验相互验证；（4）建立临床病例和体内外实验的DNA、蛋白、病理组织的标本库，运用芯片技术、分子生物信息学技术寻找E2F1调控肿瘤细胞耐药和树突状细胞功能的共同通路和关键的miRNA，阐明其在免疫序贯化疗中的作用机制。

本研究通过构建E2F1过表达载体和RNA干扰的E2F1沉默型载体，转染胃癌细胞和树突状细胞，发现过表达E2F1的胃癌细胞抗原致敏的树突状细胞介导的胃癌细胞杀伤作用增强。另外，E2F1沉默可以加快树突状细胞的成熟。在动物实验中，经过E2F1沉默的树突状细胞作用的胃癌转移瘤，再续贯化疗后明显缩小，机制研究发现：E2F1通过调控凋亡、增殖相关等不同的基因通路影响树突状细胞成熟和对胃癌细胞杀伤的生物学行为。并结合数据库分析初步验证了，E2F1基因的表达差异对胃癌周围组织中的淋巴细胞浸润及免疫微环境有调控作用。本研究为进一步探索E2F1在胃癌生物免疫学中的作用及其潜在的治疗作用研究打下坚实的基础。