病毒胸苷激酶基因治疗人胃癌的实验研究

THE EXPERIMENTAL STUDY ON GENE THERAPY FOR HUMAN GASTRIC CANCER WITH HERPES SIMLEX VIRUS THYMIDINE KINASE GENE

为观察单纯疱疹病毒脱氧胸苷激酶（ＨＳＶ－ＴＫ）／丙氧鸟苷（ＧＣＶ）基因治疗系统对胃癌细胞的杀伤作用，将ＨＳＶ－ＴＫｃＤＮＡ定向克隆入逆转录病毒载体ｐＤＯＲ－ｎｅｏ中，用ＬＩＰＯＦＥＣＴＡＭＩＮＥ法将该ＨＳＶ－ＴＫ基因转染胃癌细胞系ＳＧＣ－７９０１，测定阳性转染细胞（ＳＧＣ－７９０１／ＴＫ）在体内外对ＧＣＶ的敏感性。结果示ＧＣＶ在体外对ＳＧＣ－７９０１／ＴＫ细胞有明显的杀伤作用，其作用呈现剂量和时间依赖性特点，且同时表现出对周围亲代ＳＧＣ－７９０１细胞的杀伤效应（旁观者效应）。体内实验表明ＧＣＶ可抑制ＨＳＶ－ＴＫ阳性胃癌细胞的生长，使已形成的肿瘤逐渐消失。提示逆转录病毒载体介导的ＨＳＶ－ＴＫ基因转移胃癌细胞，配合以ＧＣＶ治疗，是胃癌基因治疗的又一途径。

To observe retrovirally mediated gene therapy using HSV TK gene/GCV (ganciclovir) system for human gastric carcinoma, HSV TK gene(1 7kb) was digested with restriction enzymes EcoRI and SAU I and isolated from plasmid pIC19R/MCl TK,then it was directionally inserted into retroviral vector pDOR neo. The HSV TK gene packaged in the vector was transfected into SGC 7901 gastric carcinoma cells(SGC 7901/TK cell) by LIPOFECTAMINE. Sensitivity of SGC 7901/TK cell to GCV was examinated in vitro. Moreover, SGC 7901/TK cells were transduced into nude mice subcutaneously,and the effects of GCV therapy was examinated in vivo. In vitro,SGC 7901/TK cells were sensitive to GCV in a dose and time dependent manner, IC 50 was 2 1 μg/ml in 72h,and toxic to nearby parental SGC 7901 cells that were resistant to GCV, this phenomenon was termed a “bystander effect”. Moreover, in vivo studies demonstrated that tumors containing HSV TK gene regressed when the animals were treated with GCV. The results may indicate that retrovirally transmitted HSV TK gene therapy with GCV may provide a new therapeutic approach for treatment of gastric carcinoma.