Yamashita Y, Fujii N, Murakata C, Ashizawa T, Okabe M, Nakano H
Tokyo Research Laboratories, Kyowa Hakko Kogyo Co. Ltd., Japan.
Biochemistry. 1992 Dec 8;31(48):12069-75. doi: 10.1021/bi00163a015.
DNA topoisomerases have been shown to be important therapeutic targets in cancer chemotherapy. We found that KT6006 and KT6528, synthetic antitumor derivatives of indolocarbazole antibiotic K252a, were potent inducers of a cleavable complex with topoisomerase I. In DNA cleavage assay using purified calf thymus DNA topoisomerase I and supercoiled pBR322 DNA, KT6006 induced topoisomerase I mediated DNA cleavage in a dose-dependent manner at drug concentrations up to 50 microM, while DNA cleavage induced by KT6528 was saturated at 5 microM. The maximal amount of nicked DNA produced by KT6006 was more than 50% of substrate DNA, which was comparable to that of camptothecin. Heat treatment (65 degrees C) of the reaction mixture containing these compounds and topoisomerase I resulted in a substantial reduction in DNA cleavage, suggesting that topoisomerase I mediated DNA cleavage induced by KT6006 and KT6528 is through the mechanism of stabilizing the reversible enzyme-DNA "cleavable complex". Both KT6006 and KT6528 did not induce topoisomerase II mediated DNA cleavage in vitro. KT6006 and KT6528 were found to induce nearly identical topoisomerase I mediated DNA cleavage patterns, which was distinctly different from that with camptothecin. In contrast to the similarity between KT6006 and KT6528 in their structures and the nature of their cleavable complex with topoisomerase I, these drugs have different properties with respect to their interaction with DNA: KT6006 is a very weak intercalator whereas KT6528 is a strong intercalator with potentials comparable to that of adriamycin. These results indicate that KT6006 and KT6528 represent a new distinct class of mammalian DNA topoisomerase I active antitumor drugs.
DNA拓扑异构酶已被证明是癌症化疗中的重要治疗靶点。我们发现，吲哚咔唑抗生素K252a的合成抗肿瘤衍生物KT6006和KT6528是拓扑异构酶I可裂解复合物的有效诱导剂。在使用纯化的小牛胸腺DNA拓扑异构酶I和超螺旋pBR322 DNA的DNA切割试验中，KT6006在高达50 microM的药物浓度下以剂量依赖方式诱导拓扑异构酶I介导的DNA切割，而KT6528诱导的DNA切割在5 microM时达到饱和。KT6006产生的最大切口DNA量超过底物DNA的50%，这与喜树碱相当。对含有这些化合物和拓扑异构酶I的反应混合物进行热处理（65摄氏度）会导致DNA切割大幅减少，这表明KT6006和KT6528诱导的拓扑异构酶I介导的DNA切割是通过稳定可逆的酶-DNA“可裂解复合物”的机制进行的。KT6006和KT6528在体外均未诱导拓扑异构酶II介导的DNA切割。发现KT6006和KT6528诱导的拓扑异构酶I介导的DNA切割模式几乎相同，这与喜树碱明显不同。与KT6006和KT6528在结构上以及与拓扑异构酶I的可裂解复合物性质上的相似性相反，这些药物在与DNA的相互作用方面具有不同的特性：KT6006是一种非常弱的嵌入剂，而KT6528是一种强嵌入剂，其潜力与阿霉素相当。这些结果表明，KT6006和KT6528代表了一类新型的哺乳动物DNA拓扑异构酶I活性抗肿瘤药物。
