title: Competitive inhibition of human liver microsomal cytochrome P450 3A-dependent steroid 6 beta-hydroxylation activity by cyclophosphamide and ifosfamide in vitro.
abstract: The prodrugs cyclophosphamide (CP) and ifosfamide (IF) are oxidized by hepatic cytochrome P450 (P450) to the active cytotoxic species, phosphoramide mustard. Acrolein (prop-2-enal) is also formed during CP and IF activation in rat liver and has been associated with P450 destruction. Analogous inactivation of human liver P450s by CP or IF could lead to pharmacokinetic interactions with coadministered drugs. The present study investigated the susceptibilities of human hepatic P450s to inhibition and inactivation by CP and IF in vitro. Unlike the situation in rat liver microsomes, total P450 was not decreased after incubation of CP or IF with NADPH and human fractions. However, CP and IF inhibited testosterone 6 beta-hydroxylation mediated by P450s 3A but not P450 1A2-dependent 7-ethylresorufin O-deethylation, P450 2C-dependent tolbutamide methyl hydroxylation or P450 2E1-mediated N-nitrosodimethylamine N-demethylation. Kinetic analysis indicated that the drugs were reversible (competitive) inhibitors of testosterone 6 beta-hydroxylation (Km, 94 +/- 8 microM) in human liver microsomes (KiS, 510 +/- 20 microM and 490 +/- 40 microM for CP and IF, respectively). Time-dependent intensification of the inhibition of the activity by CP or IF did not occur; this supports the observation that P450 was refractory to inactivation. The rates of acrolein formation from CP and IF in human hepatic microsomes (0.76 +/- 0.23 and 0.19 +/- 0.07 nmol min-1 mg-1 of protein, respectively) were only 18% and 10% of the rates estimated in fractions from untreated rat liver (4.20 +/- 0.04 and 1.96 +/- 0.12 nmol min-1 mg-1 of protein, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Medicine, University of Sydney, Westmead Hospital, Australia.
authors.forename: M
authors.initials: M
authors.lastname: Murray
authors.forename: A M
authors.initials: AM
authors.lastname: Butler
authors.forename: I
authors.initials: I
authors.lastname: Stupans
mesh_terms: D000171:Acrolein / Q000378:metabolism; D000818:Animals; D001667:Binding, Competitive; D003520:Cyclophosphamide / Q000494:pharmacology*; D019392:Cytochrome P-450 CYP2E1; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006801:Humans; D007069:Ifosfamide / Q000494:pharmacology*; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*; D006899:Mixed Function Oxygenases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013250:Steroid Hydroxylases / Q000378:metabolism*; D013739:Testosterone / Q000378:metabolism
chemical_list: D065607:Cytochrome P-450 Enzyme Inhibitors; D013739:Testosterone; D009249:NADP; D000171:Acrolein; D003520:Cyclophosphamide; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D013250:Steroid Hydroxylases; D019392:Cytochrome P-450 CYP2E1; C022657:steroid hormone 6-beta-hydroxylase; D007069:Ifosfamide