mTERT启动子调控SEA/CD80基因治疗小鼠肝癌

Gene therapy of mouse hepatoma by staphylococcal enterotoxin A and CD80 genes driven by mTERT promoter

目的观察我们以前构建的端粒酶反转录酶启动子调控的葡萄球菌肠毒素A(SEA)/CD80基因重组腺病毒载体对肝癌的疗效和诱导的免疫学效应。方法重组腺病毒采用瘤体内直接注射的方式对小鼠皮下移植性肝癌进行治疗,采用RT-PCR和Western blot方法检测腺病毒注射部位的SEA和CD80 mRNA和蛋白的表达情况;采用ELISpot方法和LDH释放实验分别检测脾脏淋巴细胞中肝癌特异性IFN-γ分泌细胞的频数和细胞毒性T细胞(CTLs)对Hepa1-6细胞的特异杀伤活性;通过观察荷瘤小鼠经治疗后肿瘤体积的变化及生存时间,评价重组腺病毒对肝癌的治疗作用。结果我们构建的腺病毒能够使SEA和/或CD80mRNA和蛋白靶向地在肝癌组织中表达;与空载体组和PBS对照组相比,双基因组和单基因组分泌IFN-γ的T细胞数量均明显增多,CTL对Hepa1-6细胞的特异性杀伤作用均明显增强,荷瘤小鼠肿瘤体积明显减小,生存期明显延长;双基因组的疗效和对免疫系统的激活作用明显高于单基因组。结论我们制备的肿瘤靶向性重组腺病毒对肝癌有良好的治疗作用,联合基因治疗优于单个基因治疗。

We previously constructed the recombinant adenovirus vectors of staphylococcal enterotoxin A（SEA） and/or CD80 gene driven by mouse telomerase reverse transcriptase（mTERT） promoter.In this study,we aimed to investigate therapeutic effects of the recombinant adenoviruses on hepatoma and its immune efficacy.We set up animal model of hepatoma with Hepa1-6 cells,and then injected the adenoviruses into tumors.After intratumoral therapy,SEA and/or CD80 mRNA and protein were detected by RT-PCR and Western blot.IFN-γ-producing cell frequency and cytotoxicity to Hepa1-6 cells of T lymphocytes in the spleens were detected by ELISpot and LDH-released assay,respectively.The therapeutic effects of the recombinant adenoviruses on hepatoma were assessed with tumor growth speed and mice survival time.The results showed the recombinant adenoviruses could make SEA and/or CD80 mRNA and protein targetedly express in hepatoma tissues.When compared with the empty vector and PBS groups,the IFN-γ-producing cell frequency and cytotoxicity of T lymphocytes increased,the growth speed of tumors decreased,and the survival time prolonged in the double-gene or single-gene groups.In addition,double genes elicited better antitumor effects and stronger immune responses.The results indicate that the recombinant adenoviruses previously constructed by us can elicit effective antitumor effects on hepatoma and the effects of double genes are better than that of single gene.