ROLE OF CYTOCHROME-P450 IN OXAZAPHOSPHORINE METABOLISM - DEACTIVATIONVIA N-DECHLOROETHYLATION AND ACTIVATION VIA 4-HYDROXYLATION CATALYZEDBY DISTINCT SUBSETS OF RAT-LIVER CYTOCHROMES P450

The roles of individual liver cytochrome P450 (P450) enzymes in N-dech loroethylation leading to deactivation and; neurotoxification of the i someric alkylating agent prodrugs ifosfamide (IF) and cyclophosphamide (CPA) were investigated using an in vitro rat liver model. Rats were pretreated with a panel of drugs, including phenobarbital (a strong in ducer of liver P450 2B1/2B2) and dexamethasone (a strong inducer of P4 50 3A enzymes), to examine the effects of these P450-inducing agents o n IF and CPA N-dechloro-ethylation catalyzed by rat hepatic microsomes . The P450 3A-specific inhibitor troleandomycin and inhibitory monoclo nal antibodies reactive with P450 2B and 2C enzymes were used to ident ify the individual P450 subfamilies involved in microsomal N-dechloroe thylation of IF and CPA. It was found that dexamethasone pretreatment preferentially elevated microsomal CPA N-dechloroethylation activity ( 12-fold increase) and that P450 3A enzymes catalyzed up to >95% of thi s reaction in both uninduced and drug-induced liver, In contrast, IF N -dechloroethylation activity was stimulated (similar to 8-fold increas e) in liver microsomes by phenobarbital pretreatment, and P450 2B1/2B2 were responsible for the majority of this activity, In addition, P450 2C11 catalyzed similar to 50% of IF N-dechloroethylation in uninduced male rat liver microsomes, Inducers of P450 1A and 4A enzymes had mo effect on N-dechloroethylation of IF or CPA. These P450 enzyme pattern s for the N-dechloroethylation reaction are distinct from those previo usly determined for IF and CPA activation via 4-hydraxylation. In acco rd with this observation, the balance between oxazaphosphorine activat ion (4-hydroxylation pathway) and deactivation/neurotoxication (N-dech loroethylation pathway) could be modulated by P450 form-selective indu cers and inhibitors. Thus, dexamethasone pretreatment substantially de creased the extent of IF N-dechloroethylation, from 47% to 24% of tota l metabolism, whereas it increased CPA N-dechloroethylation from 29% t o 84% of total metabolism. Moreover, troleandomycin selectively inhibi ted CPA N-dechloroethylation, thereby increasing net metabolism of the drug via the therapeutically productive 4-hydroxylation pathway, Oxaz aphosphorine activation and deactivation/neurotoxication are thus cata lyzed by distinct subsets of liver P450 enzymes, in a manner that may allow for improvements in therapeutic: indices for this class of drugs by using P450 form-selective modulators.