Cellular Recognition and Repair of Monofunctional鈥揑ntercalative Platinum鈥揇NA Adducts

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• 作者：Fang Liu ; Jimmy Suryadi ; Ulrich Bierbach
• 刊名：Chemical Research in Toxicology
• 出版年：2015
• 出版时间：November 16, 2015
• 年：2015
• 卷：28
• 期：11
• 页码：2170-2178
• 全文大小：351K
• ISSN：1520-5010

文摘

The cellular recognition and processing of monofunctional鈥搃ntercalative DNA adducts formed by [PtCl(en)(L)](NO₃)₂ (P1-A1; en = ethane-1,2-diamine; L = N-[2-(acridin-9-ylamino)ethyl]-N-methylpropionamidine, acridinium cation), a cytotoxic hybrid agent with potent anticancer activity, was studied. Excision of these adducts and subsequent DNA repair synthesis were monitored in plasmids modified with platinum using incubations with mammalian cell-free extract. On the basis of the levels of [伪-³²P]-dCTP incorporation, P1-A1鈥揇NA adducts were rapidly repaired with a rate approximately 8 times faster (t_1/2 鈮?18 min at 30 掳C) than the adducts (cross-links) formed by the drug cisplatin. Cellular responses to P1-A1 and cisplatin were also studied in NCI-H460 lung cancer cells using immunocytochemistry in conjunction with confocal fluorescence microscopy. At the same dose, P1-A1, but not cisplatin, elicited a distinct requirement for DNA double-strand break repair and stalled replication fork repair, which caused nuclear fluorescent staining related to high levels of MUS81, a specialized repair endonuclease, and phosphorylated histone protein 纬-H2AX. The results confirm previous observations in yeast-based chemical genomics assays. 纬-H2AX fluorescence is observed as a large number of discrete foci signaling DNA double-strand breaks, pan-nuclear preapoptotic staining, and unique circularly shaped staining around the nucleoli and nuclear rim. DNA cleavage assays indicate that P1-A1 does not act as a typical topoisomerase poison, suggesting the high level of DNA double-strand breaks in cells is more likely a result of topoisomerase-independent replication fork collapse. Overall, the cellular response to platinum鈥揳cridines shares striking similarities with that reported for DNA adduct-forming derivatives of the drug doxorubicin. The results of this study are discussed in light of the cellular mechanism of action of platinum鈥揳cridines and their ability to overcome resistance to cisplatin.