Interstrand cross-linking of DNA by FK317 and its deacetylated metabolites FR70496 and FR157471.
Interstrand cross-linking of DNA by FK317 and its deacetylated metabolites FR70496 and FR157471.
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FK317 及其脱乙酰代谢物 FR70496 和 FR157471 进行的 DNA 链间交联。
DOI：
10.1021/bi035202x
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发表时间：
2003
期刊：
影响因子：
2.9
通讯作者：
Ducept,Pascal
中科院分区：
文献类型：
--
作者：
Williams,RobertM;Ducept,Pascal
FR900482（1）和FR66979（2）是Fujisawa在1987年分离的结构新颖的天然产物，并且已被证明是与丝裂霉素结构相关的高效抗肿瘤抗生素。对作用模式的研究已经确定，这些新试剂在体外和体内都形成共价DNA链间交联，这是生物还原活化后产生的反应性线粒体中间体的结果。半合成类似物如FK 973（3）和FK 317（4）是在寻找潜在的上级临床候选药物时开发的。尽管FK 317已被证明是一种有效的化合物，但迄今为止尚未报道DNA链间交联序列特异性的直接证据。在本研究中，通过用FK317（4）及其脱乙酰代谢产物FR 70496（5）和FR 157471（6）处理合成双链体DNA底物，产生DNA链间交联。凝胶电泳分析显示，形成了取向异构体，其电泳迁移率远远大于FR 900482（1）产生的迁移率。尽管存在这些差异，但通过Fe（II）−EDTA足迹法确定，FK317（4）以及5和6在预期的5'CpG 3 '步骤内形成DNA链间交联，清楚地表明1和2中的酚氢不是FR类化合物有效DNA链间交联的先决条件。
FR900482 (1) and FR66979 (2) are structurally novel natural products isolated by Fujisawa in 1987 and have been shown to be highly potent antitumor antibiotics structurally related to the mitomycins. Studies on the mode of action have established that these new agents form covalent DNA interstrand cross-links bothin vitroandin vivoas a result of the reactive mitosene intermediate generated upon bioreductive activation. Semisynthetic analogues such as FK973 (3) and FK317 (4) were developed in the search for potentially superior clinical candidates. Although FK317 has been shown to be a potent compound, to date no direct evidence of DNA interstrand cross-link sequence specificity has been reported. In this study, DNA interstrand cross-links were generated by treatment of a synthetic duplex DNA substrate with FK317 (4) and its deacetylated metabolites FR70496 (5) and FR157471 (6). Analysis by gel electrophoresis revealed the formation of orientation isomers displaying electrophoretic mobility vastly greater than the mobilities of those generated from FR900482 (1). Despite these differences, it was established by Fe(II)−EDTA footprinting that FK317 (4) as well as5and6forms DNA interstrand cross-links within the expected 5‘CpG3‘ step, clearly demonstrating that the phenolic hydrogen in1and2is not a prerequisite for efficient DNA interstrand cross-linking by the FR class of compounds.