Uckert W, Kammertöns T, Haack K, Qin Z, Gebert J, Schendel D J, Blankenstein T
Max-Delbrück-Centrum für Molekulare Medizin, Berlin, Germany.
Hum Gene Ther. 1998 Apr 10;9(6):855-65. doi: 10.1089/hum.1998.9.6-855.
Suicide genes such as cytosine deaminase (CD) and herpes simplex virus thymidine kinase (TK) encode products that convert nontoxic substances (prodrugs) into toxic metabolites. Suicide gene transfer is currently being used in cancer therapy or can be used as a safety modality. To analyze the reliability of suicide genes as a safety modality for a vaccination study with viable cytokine/B7 gene-modified tumor cells, the individual and combined efficacy of the two suicide genes was compared for in vitro and in vivo cell killing of a murine mammary adenocarcinoma cell line (TS/A). To adapt the system to an in vivo gene delivery situation, bulk cultures cotransfected with the CD and TK gene were used instead of selected clones. In vitro, both CD and TK conferred sensitivity to the respective prodrug but the combined cytotoxic effects of both gene products were always superior. For in vivo analysis BALB/c mice were injected subcutaneously with CD- and TK-modified TS/A cells, treated with prodrugs, and tumor size was evaluated for a period of 100 days. In the in vivo situation the combination of both enzyme/prodrug systems was again most effective. The highest single concentration of 5-FC (500 mg/kg) or GCV (100 mg/kg) was not able to fully protect the animals from developing tumors, whereas a combination of 5-FC (250 mg/kg) and GCV (50 mg/kg) resulted in complete tumor eradication. In nude mice treated in the same way, most CD/TK tumors could not be eliminated. Furthermore, BALB/c mice cured of TS/A-CD/TK tumors developed a systemic tumor immunity against challenge with parental TS/A cells. These findings indicate that reliable tumor elimination by the suicide genes depends on T cells. The cooperative effect of both suicide genes was confirmed in vitro with the human renal cell carcinoma line RCC26. We conclude that TK and CD together, but neither gene alone, act as a safety mechanism for the elimination of tumor cells in a reliable fashion and suggest that a rapid and quantitative antigen release by effective TK- and CD-mediated tumor destruction is necessary for T cell immunity to develop.
自杀基因，如胞嘧啶脱氨酶（CD）和单纯疱疹病毒胸苷激酶（TK），编码的产物可将无毒物质（前体药物）转化为有毒代谢物。自杀基因转移目前正在癌症治疗中应用，或者可作为一种安全手段。为了分析自杀基因作为用活细胞因子/B7基因修饰的肿瘤细胞进行疫苗接种研究的安全手段的可靠性，比较了这两种自杀基因对小鼠乳腺腺癌细胞系（TS/A）的体外和体内细胞杀伤的个体及联合效果。为使该系统适应体内基因递送情况，使用了共转染CD和TK基因的大量培养物而非筛选出的克隆。在体外，CD和TK都赋予细胞对各自前体药物的敏感性，但两种基因产物的联合细胞毒性作用总是更优。为进行体内分析，将经CD和TK修饰的TS/A细胞皮下注射给BALB/c小鼠，用前体药物进行处理，并在100天内评估肿瘤大小。在体内情况下，两种酶/前体药物系统的组合再次最为有效。5-氟胞嘧啶（5-FC）的最高单剂量浓度（500 mg/kg）或更昔洛韦（GCV）（100 mg/kg）无法完全保护动物不发生肿瘤，而5-FC（250 mg/kg）和GCV（50 mg/kg）的组合则导致肿瘤完全消除。在用相同方法处理的裸鼠中，大多数CD/TK肿瘤无法被消除。此外，治愈TS/A-CD/TK肿瘤的BALB/c小鼠对亲本TS/A细胞的攻击产生了全身性肿瘤免疫。这些发现表明，自杀基因可靠地消除肿瘤依赖于T细胞。在人肾癌细胞系RCC26中体外证实了两种自杀基因的协同作用。我们得出结论，TK和CD共同作用，但单独任何一个基因都不能可靠地作为消除肿瘤细胞的安全机制，并且表明通过有效的TK和CD介导的肿瘤破坏实现快速和定量的抗原释放对于T细胞免疫的发展是必要的。
