Grignet-Debrus C, Calberg-Bacq C M
Laboratory of Fundamental Virology and Immunology, University of Liège, Sart-Tilman, Belgium.
Gene Ther. 1997 Jun;4(6):560-9. doi: 10.1038/sj.gt.3300435.
To investigate the potential of the thymidine kinase gene from Varicella zoster virus (VZVtk) to act as a suicide gene, VZVtk was transferred via a dicistronic retroviral construct into MCF7, T-47D and MDA-MB-435 human breast cancer cells. The cytotoxicity of antiviral drugs was then evaluated in vitro on the wild-type and transduced cells. Acyclovir and ganciclovir did not show any selective toxicity for the modified cells. In contrast, (E)-5-(2-bromovinyl)-2'-deoxyuridine (BVDU) was extremely toxic for the VZVtk expressing cells, with IC50 values of 0.6 microM, 0.1 microM and 0.06 microM for MCF7, T-47D and MDA-MB-435 cells, respectively. The selectivity index of BVDU (ie the IC50 value ratio of the wild-type to the VZVtk cells) was 400 for MCF7, 750 for T-47D and 2000 for MDA-MB-435 cells. To test the system in vivo, VZVtk carrying MDA-MB-435 cells were inoculated subcutaneously into nude mice. An intraperitoneal treatment with BVDU administered at the emergence of the tumors, led to a prolonged arrest of the tumor growth and a reduced tumor mass. This effect was BVDU dose-dependent. No bystander effect of BVDU killing could be demonstrated in vitro on mixed populations of VZVtk positive and negative MDA-MB-435 cells. However, an important bystander effect was observed in identical experiments performed on 9L rat gliosarcoma cells infected with the VZVtk-carrying vector. These results demonstrate the efficiency of VZVtk as a suicide gene when BVDU is used as prodrug. The bystander effect measured in vitro, depends however on the tumoral cell type used.
为研究水痘带状疱疹病毒胸苷激酶基因（VZVtk）作为自杀基因的潜力，通过双顺反子逆转录病毒构建体将VZVtk转入MCF7、T - 47D和MDA - MB - 435人乳腺癌细胞。然后在体外评估抗病毒药物对野生型和转导细胞的细胞毒性。阿昔洛韦和更昔洛韦对修饰细胞未显示出任何选择性毒性。相比之下，（E）-5 -（2 - 溴乙烯基）-2'-脱氧尿苷（BVDU）对表达VZVtk的细胞具有极强毒性，MCF7、T - 47D和MDA - MB - 435细胞的IC50值分别为0.6微摩尔/升、0.1微摩尔/升和0.06微摩尔/升。BVDU的选择性指数（即野生型与VZVtk细胞的IC50值之比），MCF7细胞为400，T - 47D细胞为750，MDA - MB - 435细胞为2000。为在体内测试该系统，将携带VZVtk的MDA - MB - 435细胞皮下接种到裸鼠体内。在肿瘤出现时腹腔注射BVDU进行治疗，导致肿瘤生长长期停滞且肿瘤体积减小。这种效应呈BVDU剂量依赖性。在体外对VZVtk阳性和阴性MDA - MB - 435细胞混合群体进行实验时，未证明BVDU杀伤存在旁观者效应。然而，在用携带VZVtk载体感染的9L大鼠胶质肉瘤细胞进行的相同实验中观察到了重要的旁观者效应。这些结果表明，当使用BVDU作为前药时，VZVtk作为自杀基因的有效性。然而，体外测量的旁观者效应取决于所使用的肿瘤细胞类型。
