逆转录病毒介导的HyTK基因转移及杀伤恶性黑色素瘤细胞的研究

Study of the “killing” effect of retrovirus mediated HyTK gene on melanoma cell line

用逆转录病毒载体将单纯疱疹病毒胸苷激酶（ＨＳＶｔｋ）基因导人恶性肿瘤细胞，随后应用药物丙氧鸟苷（ＧＣＶ）可选择性地杀死肿瘤细胞．本研究中我们将ＨｙＴＫ基因克隆到逆转录病毒载体ＬＸＳＮ中，并切除其中的ＳＶ４０早期启动子，构建成重组逆转录病毒载体ＬＨｙＴＫ／Ｎ，并用该重组病毒转染小鼠恶性黑色素瘤细胞系Ｂ１６细胞，经ＰＣＲ方法检测证明ＨｙＴＫ基因已成功地导入肿瘤细胞中，且不含可复制的辅助病毒．分别用不同浓度的ＧＣＶ作用于ＨｙＴＫ－及ＨｙＴＫ＋的Ｂ１６细胞，４８ｈ后，在光镜下观察细胞形态及进行活细胞计数．结果表明，ＧＣＶ浓度大于０．

Recent research showed that transfer of the Herpes simplex virus thymidine kinase (HSV tk) gene into malignant tumor cells could confer the tumor cells susceptibility to the antiviral drug ganciclovir (GCV), and thus produce “killing” effect on the tumor cells exposed to GCV. We constructed a recombinant retroviral vector LHyTK/N by inserting HyTK gene into the retroviral vector LXSN and cutting out the SV40 early promoter. The recombinant virus was exployed to transfect mouse melanoma cell line B16. PCR results showed that the HyTK gene was transferred successfully into B16 cells and free of duplication competent virus. B16 cells of HyTK - and HyTK + were exposed to GCV of different concentration. After 48 h, morphological alteration was observed under light microscope and live cell counting was also conducted. The results show that GCV (>0.1 μmol/L) has evident “killing” effect on B16/HyTK + cells.