Ellipticine oxidation and DNA adduct formation in human hepatocytes is catalyzed by human cytochromes P450 and enhanced by cytochrome b5.
Ellipticine is an antineoplastic agent considered a pro-drug, the pharmacological and genotoxic effects of which are dependent on cytochrome P450 (CYP)- and/or peroxidase-mediated activation to covalent DNA adducts. We investigated whether ellipticine-DNA adducts are formed in human hepatic microsomes and human hepatocytes. We then identified which human CYPs oxidize ellipticine to metabolites forming DNA adducts and the effect of cytochrome b(5) on this oxidation. 13-Hydroxyellipticine, the metabolite forming the major ellipticine-DNA adduct, was generated mainly by CYP3A4 and 1A1, followed by CYP2D6>2C19>1B1>1A2>2E1 and >2C9. Cytochrome b(5) increased formation of this metabolite by human CYPs, predominantly by CYP1A1, 3A4, 1A2 and 2C19. Formation of 12-hydroxyellipticine is generated mainly by CYP2C19, followed by CYP2C9>3A4>2D6>2E1 and >2A6. Other CYPs were less active (CYP2C8 and 2B6) or did not oxidize ellipticine to this metabolite (CYP1A1, 1A2 and 1B1). CYP2D6 was the most efficient enzyme generating ellipticine N(2)-oxide. CYP3A4 and 1A1 in the presence of cytochrome b(5) are mainly responsible for bioactivation of ellipticine to DNA adduct 1 (formed by ellipticine-13-ylium from 13-hydroxyellipticine), while 12-hydroxyellipticine generated during the CYP2C19-mediated ellipticine oxidation is the predominant metabolite forming ellipticine-12-ylium that generates ellipticine-DNA adduct 2. These ellipticine-DNA adducts were also generated by human hepatic microsomes and in primary human hepatocytes exposed to ellipticine. Ellipticine is toxic to these hepatocytes, decreasing their viability; the IC(50) value of ellipticine in these cells was 0.7 μM. In liver CYP3A4 is the predominant ellipticine activating CYP species, which is expected to result in efficient metabolism after oral ingestion of ellipticine in humans.
Department of Biochemistry, Faculty of Science, Charles University, Albertov 2030, 128 40 Prague 2, Czech Republic. stiborov@natur.cuni.cz
Marie
M
Stiborová
Jitka
J
Poljaková
Eva
E
Martínková
Jitka
J
Ulrichová
Vilím
V
Simánek
Zdeněk
Z
Dvořák
Eva
E
Frei
D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D001189:Aryl Hydrocarbon Hydroxylases / Q000378:metabolism; D019363:Cytochrome P-450 CYP1A1 / Q000378:metabolism; D065731:Cytochrome P-450 CYP2C19; D065729:Cytochrome P-450 CYP2C9; D019389:Cytochrome P-450 CYP2D6 / Q000378:metabolism; D051544:Cytochrome P-450 CYP3A / Q000378:metabolism*; D015786:Cytochromes b5 / Q000378:metabolism*; D018736:DNA Adducts / Q000187:drug effects*; D004611:Ellipticines / Q000494:pharmacology*; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D008099:Liver / Q000187:drug effects / Q000378:metabolism; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism; D010084:Oxidation-Reduction / Q000187:drug effects; D011355:Prodrugs / Q000494:pharmacology
C517214:12-hydroxyellipticine; C517215:13-hydroxyellipticine; D000972:Antineoplastic Agents, Phytogenic; D018736:DNA Adducts; D004611:Ellipticines; D011355:Prodrugs; D015786:Cytochromes b5; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; C569463:CYP1A1 protein, human; C045793:CYP2C19 protein, human; D019363:Cytochrome P-450 CYP1A1; D065731:Cytochrome P-450 CYP2C19; D019389:Cytochrome P-450 CYP2D6; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human