放射诱导启动子介导Slug基因沉默与PUMA基因表达靶向诱导口腔鳞癌细胞凋亡

Poor target activity is one of the most important reasons that constraints the clinical application of gene therapy in the treatment of malignant tumour. PUMA is a new member of the Bcl-2 protein family which has powerful pro-apoptotic effect and a key protein in apoptosis induced by radiotherapy. The Slug gene belongs to the transcription factor Snail family and encodes zinc finger protein which is the inhibitor of PUMA protein. Slug-RNAi could relieve the the inhibition effect of Slug on PUMA and enhance the sensitivity of radiation. In our previous work, radiation -induced promoter had been synthesized and was successfully used as a molecular switch to regulate gene expression. Based on that, in this study, we intends to establish a Slug-RNAi system regulated by radiation-induced promoter and PUMA gene expression plasmid vector mediated by the promoter, transfect the oral squamous cell carcinoma with nanospheres as vector, silence Slug gene by radiation and enhance the PUMA gene expression at the same time. The two kinds of intervention pathways including inhibition effect of Slug on PUMA relieved by Slug-RNAi and gene expression of PUMA upregulated by transfection should promote apoptosis. We will combine gene therapy with radiotherapy organically, play cooperative role in regulating the two gene targets through exogenous radiation, induce cell apoptosis, improve the target activity and therapeutic effect of gene therapy and provide a new strategy for the treatment of oral squamous cell carcinoma.

靶向性差是制约肿瘤基因治疗临床应用的主要原因之一。PUMA是具有强大促凋亡作用Bcl-2家族新成员，是放疗引起细胞凋亡过程中的关键蛋白。Slug基因属转录因子Snail家族，其编码锌指蛋白是PUMA蛋白抑制剂，Slug-RNAi可解除Slug对PUMA的抑制，增强细胞放疗敏感性。本课题组已合成放射诱导启动子并证实可用作调控基因表达的分子开关。在此基础上，本研究拟构建由放射诱导启动子调控的Slug-RNAi 系统及由该启动子介导PUMA基因表达质粒，以纳米微球为载体转染口腔鳞癌，通过放射诱导Slug基因沉默同时增强PUMA基因表达。利用Slug-RNAi解除Slug对PUMA的抑制，PUMA基因转染上调PUMA表达，两种干预途径共同促进细胞凋亡。本研究将基因治疗与放疗有机结合，通过外源性放射调控双基因靶点发挥协同效应，诱导细胞凋亡，提高基因治疗的靶向性和疗效，为口腔鳞癌的治疗提供新策略。

靶向性差是制约肿瘤基因治疗临床应用的主要原因之一。PUMA是具有强大促凋亡作用Bcl-2家族新成员，是放疗引起细胞凋亡过程中的关键蛋白。Slug基因属转录因子Snail家族，其编码锌指蛋白是PUMA蛋白抑制剂，Slug-RNAi可解除Slug对PUMA的抑制，增强细胞放疗敏感性。本课题组已合成放射诱导启动子并证实可用作调控基因表达的分子开关。在此基础上，本课题组在国家自然科学基金的资助下，按计划开展实验完成以下工作：（1）成功构建由放射诱导启动子调控的Slug-RNAi 系统，放射诱导后沉默Slug基因表达；（2）成功构建放射诱导启动子介导PUMA基因表达质粒；（3）将上述系统和质粒载体转染口腔鳞癌，检测放射诱导后Slug基因和PUMA基因在口腔鳞癌中的表达，发现放射线照射沉默Slug基因表达同时增强PUMA基因表达，促进口腔鳞癌细胞凋亡。本研究将基因治疗与放疗有机结合，通过外源性放射调控双基因靶点发挥协同效应，诱导细胞凋亡，提高基因治疗的靶向性和疗效，为口腔鳞癌的治疗提供新策略。本研究部分相关结果发表科研论文7篇，其中外文4篇，SCI收录4篇，同时培养硕士研究生毕业两名。资助课题组成员参加国际会议一次，国内会议五次。