Hsiang Y H, Lihou M G, Liu L F
Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
Cancer Res. 1989 Sep 15;49(18):5077-82.
Camptothecin, which induces an unusual type of DNA damage by trapping cellular topoisomerase I on chromosomal DNA in the form of drug-enzyme-DNA cleavable complexes, inhibits DNA synthesis and specifically kills S-phase cells. Cotreatment of L1210 cells with aphidicolin, which is an inhibitor of replicative DNA polymerases, completely abolished camptothecin cytotoxicity, suggesting the involvement of DNA replication in camptothecin cytotoxicity. In order to study the role of DNA replication in drug action, a cell-free SV40 DNA replication system was used in the present study. Camptothecin inhibited SV40 DNA replication in this cell-free system only in the presence of topoisomerase I. Addition of excess purified calf thymus DNA topoisomerase I to this extract system in the presence of camptothecin resulted in severe inhibition of SV40 DNA replication and the accumulation of linearized replication products, which contained covalently bound DNA topoisomerase I. We propose that the collision between moving replication forks and camptothecin-stabilized topoisomerase I-DNA cleavable complexes results in fork arrest and possibly fork breakage, which are lethal to proliferating cells.
喜树碱通过以药物 - 酶 - DNA可裂解复合物的形式将细胞拓扑异构酶I捕获在染色体DNA上，诱导一种不寻常的DNA损伤类型，抑制DNA合成并特异性杀死S期细胞。用作为复制性DNA聚合酶抑制剂的阿非科林对L1210细胞进行共处理，完全消除了喜树碱的细胞毒性，这表明DNA复制参与了喜树碱的细胞毒性作用。为了研究DNA复制在药物作用中的作用，本研究使用了无细胞的SV40 DNA复制系统。喜树碱仅在拓扑异构酶I存在的情况下，才在该无细胞系统中抑制SV40 DNA复制。在喜树碱存在下，向该提取物系统中添加过量纯化的小牛胸腺DNA拓扑异构酶I，导致SV40 DNA复制受到严重抑制，并积累了线性化的复制产物，这些产物包含共价结合的DNA拓扑异构酶I。我们提出，移动的复制叉与喜树碱稳定的拓扑异构酶I - DNA可裂解复合物之间的碰撞导致叉停滞，并可能导致叉断裂，这对增殖细胞是致命的。
