Abstract

Midazolam (MDZ) and triazolam (TRZ) hydroxylation, reactions considered to be cytochrome P-4503A (CYP3A)-mediated in humans, were examined in mouse and human liver microsomes. In both species, α- and 4-hydroxy metabolites were the principal products. Western blotting with anti-CYP3A1 antibody detected a single band of immunoreactive protein in both human and mouse samples: 0.45 ± 0.12 and 2.02 ± 0.24 pmol/mg protein (mean ± S.E., n = 3), respectively. Ketoconazole potently inhibited MDZ and TRZ metabolite formation in human liver microsomes (IC₅₀ range, 0.038–0.049 μM). Ketoconazole also inhibited the formation of both TRZ metabolites and of 4-OH-MDZ formation in mouse liver microsomes (IC₅₀ range, 0.0076–0.025 μM). However, ketoconazole (10 μM) did not produce 50% inhibition of α-OH-MDZ formation in mouse liver microsomes. Anti-CYP3A1 antibodies produced concentration-dependent inhibition of MDZ and TRZ metabolite formation in human liver microsomes and of TRZ metabolite and 4-OH-MDZ formation in mouse liver microsomes to less than 20% of control values but reduced α-OH-MDZ formation to only 66% of control values in mouse liver microsomes. Anti-CYP2C11 antibodies inhibited α-OH-MDZ metabolite formation in a concentration-dependent manner to 58% of control values in mouse liver microsomes but did not inhibit 4-OH-MDZ formation. Thus, TRZ hydroxylation appears to be CYP3A specific in mice and humans. α-Hydroxylation of MDZ has a major CYP2C component in addition to CYP3A in mice, demonstrating that metabolic profiles of drugs in animals cannot be assumed to reflect human metabolic patterns, even with closely related substrates.