Inhibition of cytochrome P-450 3A (CYP3A) in human intestinal and liver microsomes: Comparison of K-I values and impact of CYP3A5 expression

The purpose of this study was to compare the kinetics of intestinal and hep atic cytochrome P-450 3A (CYP3A) inhibition by using microsomal midazolam 1 '-hydroxylation as a marker of enzyme activity. The effect of two antifunga l agents commonly implicated in CYP3A drug-drug interactions was examined, Inhibition type and affinities were determined for human liver and intestin al microsomes screened for the presence or absence of CYP3A4 and CYP3A5, as well as for cDNA-expressed CYP3A4 and CYP3A5 microsomes. Ketoconazole and fluconazole were found to be noncompetitive inhibitors of both enzymes. Ket oconazole exhibited a K-i for cDNA-expressed CYP3A4 of 26.7 +/- 1.71 nM, wh ereas the K-i for cDNA expressed CYP3A5 was 109 +/- 19.7 nM, Corresponding K-i values for fluconazole were 9.21 +/- 0.51 mu M and 84.6 +/- 12.9 mu M. For liver and intestinal microsomes that contained only CYP3A4, the average ketoconazole K-i was found to be 14.9 +/- 6.7 nM and 17.0 +/- 7.9 nM, resp ectively, whereas fluconazole yielded mean respective K-i values of 10.7 +/ - 4.2 mu M and 10.4 +/- 2.9 mu M. Liver and intestinal microsomes that cont ained an equal or greater amount of CYP3A5, in addition to CYP3A4, were les s susceptible to inhibition by both ketoconazole and fluconazole. These fin dings suggest that there can be significant differences in the affinity of these two enzymes for inhibitors. This may further broaden interindividual variability with respect to the magnitude of in vivo drug-drug interactions . We also conclude that there is no significant difference in inhibition ty pe and affinity of ketoconazole and fluconazole for hepatic versus intestin al CYP3A4.