Midazolam and triazolam biotransformation in mouse and human liver microsomes: Relative contribution of CYP3A and CYP2C isoforms

Midazolam (MDZ) and triazolam (TRZ) hydroxylation, reactions considered to be cytochrome P-4503A (CYP3A)-mediated in humans, were examined in mouse an d human liver microsomes. In both species, alpha- and 4-hydroxy metabolites were the principal products. Western blotting with anti-CYP3A1 antibody de tected a single band of immunoreactive protein in both human and mouse samp les: 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 form ation in human liver microsomes (IC50 range, 0.038-0.049 mu M). Ketoconazol e also inhibited the formation of both TRZ metabolites and of 4-OH-MDZ form ation in mouse liver microsomes (IC50 range, 0.0076-0.025 mu M). However, k etoconazole (10 mu M) did not produce 50% inhibition of alpha-OH-MDZ format ion in mouse liver microsomes. Anti-CYP3A1 antibodies produced concentratio n-dependent inhibition of MDZ and TRZ metabolite formation in human liver m icrosomes and of TRZ metabolite and 4-OH-MDZ formation in mouse liver micro somes to less than 20% of control values but reduced alpha-OH-MDZ formation to only 66% of control values in mouse liver microsomes. Anti-CYP2C11 anti bodies inhibited alpha-OH-MDZ metabolite formation in a concentration-depen dent 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 spe cific in mice and humans. alpha-Hydroxylation of MDZ has a major CYP2C comp onent in addition to CYP3A in mice, demonstrating that metabolic profiles o f drugs in animals cannot be assumed to reflect human metabolic patterns, e ven with closely related substrates.