Identification of P450 enzymes involved in metabolism of verapamil in humans

Abstract

The calcium channel blocker verapamil[2,8-bis-(3,4-dimethoxyphenyl)-6-methyl-2-isopropyl-6- azaoctanitrile] is widely used in the treatment of hypertension, angina pectoris and cardiac arrhythmias. The drug undergoes extensive and variable hepatic metabolism in man with the major metabolic steps comprising formation of D-617 [2-(3,4-dimethoxyphenyl)-5-methylamino-2-isopropylvaleronitrile] and norverapamil [2,8-bis-(3,4-dimethoxyphenyl)-2-isopropyl-6-azaoctanitrile]. The enzymes involved in metabolism of verapamil have not been characterized so far. Identification of these enzymes would enable estimation of both interindividual variability in verapamil metabolism introduced by the respective pathway and potential for metabolic interactions. We therefore characterized the enzymes involved in formation of D-617 and norverapamil. The maximum rate of formation of D-617 and norverapamil was 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 CYP3A3/4) by means of Western blotting. Specific antibodies directed against CYP3A were used to inhibit formation of D-617 and norverapamil. Finally, formation of both metabolites was investigated in microsomes obtained from yeast cells which were genetically engineered for stable expression of human P450. Formation of D-617 was correlated with the expression of CYP3A (r = 0.85; P < 0.001) and CYP1A2 (r = 0.57; P < 0.01) in the microsomal fraction of 21 human livers after incubation with racemic verapamil.(ABSTRACT TRUNCATED AT 250 WORDS)