IDENTIFICATION OF P450 ENZYMES INVOLVED IN METABOLISM OF VERAPAMIL INHUMANS

The calcium channel blocker hoxyphenyl)-6-methyl-2-isopropyl-6-azaocta nitrile] is widely used in the treatment of hypertension, angina pecto ris and cardiac arrythmias. The drug undergoes extensive and variable hepatic metabolism in man with the major metabolic steps comprising fo rmation of D-617 oxyphenyl)-5-methylamino-2-isopropylvaleronitrile] an d norverapamil ,4-dimethoxyphenyl)-2-isopropyl-6-azaoxtanitrile]. The enzymes involved in metabolism of verapamil have not been characterize d so far. Identification of these enzymes would enable estimation of b oth interindividual variability in verapamil metabolism introduced by the respective pathway and potential for metabolic interactions. We th erefore characterized the enzymes involved in formation of D-617 and n orverapamil. The maximum rate of formation of D-617 and norverapamil w as determined in the microsomal fraction of 21 human livers which had been previously characterized for the individual expression of various P450 enzymes (CYP1A2, CYP2C, CYP2D6, CYP2E1 and CYP 3 A 3/4) by means of Western blotting. Specific antibodies directed against CYP3A were used to inhibit formation of D-617 and norverapamil. Finally, formatio n of both metabolites was investigated in microsomes obtained from yea st cells which were genetically engineered for stable expression of hu man P450. Formation of D-617 was correlated with the expression of CYP 3A (r = 0.85; P < 0.001) and CYP1A2 (r = 0.57; P < 0.0 1) in the micro somal fraction of 21 human livers after incubation with racemic verapa mil. Formation of norverapamil was correlated with the expression of C YP3A (r = 0.58; P < 0.01) and CYP1A2 (r = 0.5; P < 0.05) in the same p reparations after incubation with racemic verapamil. Antibodies agains t CYP3A reduced maximum rate of formation of D-617 (to 37.1 +/- 11% an d 40.6 +/- 6.8% of control after incubation with S- and R-verapamil, r espectively) and norverapamil (to 38.2 +/- 4.5% and 29.2 +/- 5.5% of c ontrol after incubation with S- and R-vempamil, respectively). Both D- 617 and norverapamil were formed by stable expressed CYP3A4 (16.6 pmol /mg protein/min and 22.6 pmol/mg protein/min, respectively). In summar y, formation of D-617 and norverapamil is catalyzed mainly by CYP3A4. D-617 is also formed by CYP1A2. Verapamil therefore has the potential to interact with other drugs which are substrates or inducers of CYP3A and CYP1A2.