Tsuyoshi Nakai

Abstract Platinum(IV) [Pt(IV)] compounds are being developed worldwide as next-generation platinated drugs for a broad range of cancers. Since Pt(IV) compounds show relatively stable structural features, they are applicable for oral administration. The anti-cancer mechanism of Pt(IV) compounds has been proposed as that Pt(II) generated from Pt(IV) by endogenous reductants binds to DNA, resulting in the inhibition of transcription and replication of cancer cells. Here, we investigated the interaction manner between DNA, Pt(IV) and endogenous reductants such as ascorbic acid (AsA) and glutathione (GSH). cis-Diammine-tetrachloro-Pt(IV) [cis-Pt(IV)], which is a prodrug of cisplatin [cis-diammine-dichloro-platinum(II), cis-Pt(II)], was incubated with calf thymus DNA in the presence of AsA or GSH. In the presence of AsA, cis-Pt(IV) induced oxidative damage in calf thymus DNA, in which elevated amount of 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG), a biomarker for DNA oxidation, was observed. Hydroxyl radical (HO·) scavengers (ethanol and dimethylsulfoxide) suppressed the AsA-associated oxidative damage in a dose-dependent manner, whereas superoxide dismutase and catalase did not, suggesting that HO· was involved in the DNA oxidation. CD spectral change and crosslink formation in calf thymus DNA were also observed during this DNA oxidation, suggesting the cis-Pt(IV) reduction by AsA and the DNA conformational change by cis-Pt(II)-DNA binding. However, GSH did not show any 8-oxodG formation in calf thymus DNA likely due to its own HO· scavenging capability. GSH also suppressed the cis-Pt(II)-like conformational change probably by cis-Pt(II) sequestering from DNA by GSH. This GSH-cis-Pt(II) complex formation was also suggested by agarose gel electrophoresis. In order to reveal the detailed mechanisms of the complex formation, three thiol compounds (2-mercaptoethanol, 2-mercaptoacetic acid, and 2-mercaptoethylamine) were compared with regard to the reductive complex formation capabilities with cis-Pt(II). As observed in GSH-cis-Pt(IV)-DNA system, 2-mercaptoacetic acid and 2-mercaptoethylamine inhibited cis-Pt(II)-induced CD spectral change and crosslink formation in calf thymus DNA, whereas 2-mercaptoethanol did not, suggesting that thiol- and amine-/carbonyl-groups are involved in GSH-cis-Pt(II) complex formation via ligand exchange between these groups and Cl of cis-Pt(II). In this study, we revealed that the interaction manner of cis-Pt(IV) with endogenous reductants. In the reactions, AsA reduced Pt(IV) to Pt(II) and induced DNA-crosslink formation and DNA oxidation, where HO· was produced; GSH scavenged the radical and interfered with the DNA-crosslink formation. These data strongly indicate the importance of the proper Pt(IV) ligands which are suitable for Pt(IV) reduction by endogenous reductants without complex formation with the generated Pt(II). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3646.