CYP3A4/5 activity probed with testosterone and midazolam: Correlation between two substrates at the microsomal and enzyme levels

Testosterone (TST) and Midazolam (MDZ) are widely used as probes to detect CYP3A4/5 activity but the data acquired with these two substrates does not correlate well at the microsomal level (per milligram of microsomal protein) and the reason is unclear. In this study, CYP3A4/5 activity was probed with TST and MDZ at the microsomal and enzyme levels (per picomole of CYP3A4/5) in 72 human liver samples. Correlation coefficients were lower in Vmax, CLint at the microsomal level, as compared with those at the enzyme level (Vmax 0.658 vs. 0.883; CLint no correlation vs. 0.796). Compared with TST, MDZ was found to correlate better with content of CYP3A4/5 (no correlation vs. 0.431), CYP3A5 (no correlation vs. 0.447) and huge variations in enzyme content existed among different genotypes, which explained the lower degree of correlation at the microsomal level. In addition, different genotypes had varying effects on activity at the enzyme level, whereas the difference between activity at the enzyme level probed w...