IDENTIFICATION OF THE MAJOR HUMAN HEPATIC CYTOCHROME-P450 INVOLVED INACTIVATION AND N-DECHLOROETHYLATION OF IFOSFAMIDE

Two NADPH-dependent metabolic routes for the anticancer drug ifosfamid e, 4-hydroxylation (activation) and N-dechloroethylation (a detoxicati on pathway),were studied in human liver microsomes to identify the cyt ochrome P450 enzymes involved. Naringenin, a grapefruit aglycone and a n inhibitor of cytochrome P450 3A4 (CYP3A4)-catalysed reactions, was f ound to inhibit ifosfamide activation and N-dechloroethylation by huma n liver microsomes. IC50 for both reactions was of the order of 70 mu M. The CYP3A4-specific inhibitor triacetyloleandomycin inhibited ifosf amide N-dechloroethylation by human Liver microsomes with an IC50 Of a pproximately 10 mu M. Furthermore, anti-human CYP3A4 antiserum inhibit ed by about 80% N-dechloroethylation of ifosfamide by human liver micr osomes. The relative levels of cytochromes P450 1A, 2C, 2E and 3A4 in 12 human livers were determined by western blotting analysis. A strong correlation (P < 0.001) was observed between CYP3A4 expression and bo th activation and N-dechloroethylation of ifosfamide. A role for human CYP3A4 in both pathways of ifosfamide metabolism was thus demonstrate d. This was substantiated by the observation that the nifedipine oxida se activities of the 12 samples of human liver microsomes correlated w ith ifosfamide activation (P < 0.009) and N-dechloroethylation (P < 0. 001). These findings have important clinical implications. The involve ment of the same key cytochrome P450 enzyme in both reactions prohibit s selective inhibition of the N-dechloroethylation pathway, as might b e desirable to reduce toxic side effects. They also demonstrate the ne ed to consider interaction with co-administered drugs that are CYP3A4 substrates.