CYP3A4-mediated hepatic metabolism of the HIV-1 protease inhibitor saquinavir in vitro.

1. The aim was to identify the major metabolites of saquinavir (SQV) from human hepatic microsomal incubations and the CYP isoform(s) responsible.2. Ten fractions containing various metabolites were separated by isocratic reversed- phase HPLC and characterized by HPLC, mass spectrometry and NMR.3. Metabolites were either mono- or di-hydroxylated derivatives of SQV. Fast-atom bombardment and electrospray MS showed that hydroxylation was predominantly situated on the decahydroisoquinoline ring. A major metabolite (M4) was rigorously identified as 6-equatorial-hydroxy SQV.4. Metabolism of saquinavir to all metabolites was inhibited by the CYP3A4-selective inhibitor ketoconazole (IC50 = 0.55 ± 0.12 µM). Other isoform-selective inhibitors were non-inhibitory. The protease inhibitors ritonavir, indinavir and nelfinavir potently inhibited SQV metabolism in hepatic microsomes with IC50 = 0.025 ± 0.004, 0.82 ± 0.26 and 0.58 ± 0:14 µM, respectively.5. Saquinavir metabolism correlated with immunochemically determine...