基于IL-2、NK4双功能肽的减毒沙门氏菌肿瘤疫苗抗肝癌的分子机制研究

以减毒沙门氏菌为载体的基因工程疫苗系统在肿瘤免疫及治疗领域备受关注。前期研究发现，基于IL-2与NK4单、双基因的肿瘤疫苗TPI、TPN及TPIN均具有抗癌作用；且双基因疫苗的抗癌效果显著优于单基因疫苗；因此推测，此创新型双基因疫苗在肿瘤免疫及治疗两方面具有协同作用，但其作用机制不明确。.课题组拟采用比较基因组学研究策略，基于表达谱芯片技术建立单基因、双基因抗肝癌的差异基因表达谱，采用qPCR、WB、IHC、免疫荧光、免疫共沉淀等方法对“差异细胞信号通路”进行分子、细胞及组织学水平的筛选验证；同时构建针对双基因的RNAi腺病毒表达载体，进一步采用上述方法检测抑制双基因表达后“差异分子”、细胞功能、瘤体生长及转移等分别在细胞、瘤体组织中的变化，确定双基因协同作用的细胞信号网络，阐明其抗肝癌作用的分子机制，评价其在设计思路及方法上对实现新型“抗癌疫苗”开发中的现实意义，并为其临床应用提供依据。

The gene engineering vaccine system based on Attenuated Salmonella has attracted much attention in the field of tumor immunity and treatment. Previous studies have found that TPI, TPN and TPI based on IL-2 and NK4 single and double gene tumor vaccines have antitumor effect, and the antitumor effect of the double gene vaccine is significantly better than that of the single gene vaccine. Therefore, it is speculated that this innovative double gene vaccine has synergistic effect in tumor immunity and tumor treatment, but its mechanism of action is not clear..Therefore, we intend to use the strategy of comparative genomics to establish the differential gene expression profiles of IL-2 and NK4 genes against hepatocellular carcinoma based on expression profiling chip technology. Then, we will use qPCR, WB, IHC, immunofluorescence, immunoprecipitation and other methods to screen and verify the molecular, cellular and histological levels of the "differential cell signaling pathway". At the same time, we will construct a dual-gene adenovirus RNA expression vector, and then use qPCR, WB, IHC, immunofluorescence, immunoprecipitation and other methods to detect the "difference", the changes of molecule, cell function, tumor growth and metastasis in hepatocellular carcinoma cells and mouse tumor tissues after inhibiting the expression of dual genes. Ultimately, we will determine the cellular signal network of double gene synergy, clarify the mechanism of tumor immunity and treatment, and evaluate the practical significance of the vaccine in the design ideas and methods for the development of a new "anti-cancer vaccine", and finally provide theoretical data for its clinical application.