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目的 提高原位转基因效率 ,建立高效自杀基因杀伤胃癌肿瘤的方法。方法 首先构建含胞嘧啶脱氨酶 (CD)基因的重组腺病毒载体并对载体进行包装、收获病毒上清及纯化。将重组病毒液直接转染至 615小鼠皮下已接种MFC胃癌细胞成瘤的肿瘤部位进行抑瘤实验。结果 含CD基因的重组腺病毒载体经酶切鉴定正确 ;包装纯化后 ,检测病毒滴度为 1.2× 10 12 颗粒 /ml。将重组病毒液直接转染至肿瘤部位后 ,在前药 5 氟胞嘧啶 ( 5 FC)作用下 ,肿瘤生长较对照组明显缩小 (P <0 .0 1)。病理学检查可见肿瘤细胞大片坏死 ,且未转基因部位亦存在坏死区。结论 腺病毒载体转导的CD基因 ,不仅提高了转染效率 ,而且可以大幅度杀伤肿瘤细胞以及产生强效的旁观者效应。
Objective To improve the efficiency of transgene in situ and establish a highly efficient suicide gene to kill gastric cancer. Methods Firstly, a recombinant adenovirus vector containing cytosine deaminase (CD) gene was constructed and the vector was packaged. The virus supernatant was harvested and purified. The recombinant virus liquid was directly transfected into 615 mice subcutaneously inoculated with MFC gastric cancer cells into the tumor site tumor inhibition experiments. Results The recombinant adenoviral vector containing CD gene was identified by restriction enzyme digestion. After packaging and purification, the titer of virus was 1.2 × 10 12 particles / ml. After transfection of the recombinant virus directly into the tumor site, the growth of the tumor was significantly reduced (P <0.01) under the action of the 5-fluorocytosine (5 FC). Pathological examination showed tumor necrosis, and non-transgenic sites also exist necrosis. Conclusion The CD gene transduced by adenovirus vector not only improves the transfection efficiency, but also can kill the tumor cells and produce a powerful bystander effect.