We have previously shown the formation of covalent DNA adducts by ellipticine activated by microsomal cytochrome P450 (CYP) (Biochem.Pharmacol. 62, 1675-1684. 2001. Chem. Res. Toxicol. 16, 38-47, 2003). Ellipticine is an alkaloid which has antineoplastic properties and exerts its activity against different cellular targets. The discovery of the new mechanism of covalent ellipticine-DNA adduct formation detectable by 32P-postlabeling, has prompted us to investigate the metabolism of ellipticine, and to elucidate the structure of the active species binding to DNA. The pattern of adduct spots detected after activation by rat, rabbit or human hepatic microsomes is identical. Also in vivo in rats and in cultivated cells the same adducts were detected (Biochem. Pharmacol. 64, 289-295, 2002. Int. J. Cancer 107, 885-890, 2003. Collect. Czech. Chem. Commun. 69, 603-615, 2004). All species therefore contain enzymes converting ellipticine to metabolite(s) forming these DNA adducts. Supersomes® containing recombinant human CYP and microsomes isolated from human livers were used to oxidatively activate ellipticine. Ellipticine-DNA adducts were determined by 32P-postlabeling using the nuclease P1 enrichment version of the assay. Metabolites of ellipticine were isolated by reverse phase HPLC and analysed by mass spectrometry and NMR. For the studies investigating the persistence of DNA adducts male rats were treated with 80 mg ellipticine /kg, killed after 2 days, 2, 10 and 32 weeks and adducts determined in the DNA isolated from different organs. The active metabolites forming DNA adducts were identified as 13-hydroxy-ellipticine and the N2-oxide of ellipticine, each of which forms one of the two major ellipticine-deoxyguanosine adducts in DNA detected in vitro and in vivo by 32P-postlabeling. Ellipticine-DNA adducts were found in decreasing amounts in liver, spleen, lung, kidney, heart and brain, but not in testis, and showed a biphasic decrease over time. In female rats ellipticine-DNA adducts were also detected in the target tumor tissue. Ellipticine-DNA adduct levels in a methylnitrosourea-induced mammary tumor of a rat injected with ellipticine were higher than in the mammary gland of a healthy rat, which might explain the reported activity of ellipticine against breast cancer.
[Proc Amer Assoc Cancer Res, Volume 46, 2005]