INTERINDIVIDUAL VARIABILITY IN CATALYTIC ACTIVITY AND IMMUNOREACTIVITY OF 3 MAJOR HUMAN LIVER CYTOCHROME-P450 ISOZYMES

Objective: Interindividual variations in immunoreactivity and function of three major human drug metabolising P450 monooxygenases has been i nvestigated in liver microsomes from 42 Caucasians (kidney donors or l iver biopsies). Methods: Diclofenac 4'-hydroxylation, dextromethorphan O-demethylation and midazolam 1'-hydroxylation, measured by HPLC in i ncubates, were used as probes to determine CYP2C9, CYP2D6 and CYP3A4 f unction kinetics, respectively. Immunoquantification of the three isof orms was achieved by Western blotting, using rabbit polyclonal antibod ies raised against human CYP2C9 and human CYP3A4, and mouse monoclonal antibody raised against human CYP2D6. Results: Diclofenac 4'-hydroxyl ation exhibited Michaelis-Menten kinetics with k(M) = 3.4 mu mol . l(- 1) and V-max = 45 nmole . mg(-1)P . h(-1). Relative immunoreactivity o f CYP2C9 was correlated with V-max and CL(int). Dextromethorphan O-dem ethylation in EM (extensive metabolisers) liver microsomes also showed Michaelis-Menten kinetics, with k(M) = 4.4 mu mol . l(-1) and V-max = 5.0 nmol . mg(-1)P . h(-1). Relative immunoreactivity of CYP2D6 was c orrelated with V-max and CL(int). Midazolam 1'-hydroxylation also exhi bited Michaelis-Menten kinetics with k(M) = 3.3 mu mol . l(-1) and V-m ax = 35 nmol mg(-1)P . h(-1). Relative immunoreactivity of CYP3A4 was correlated with V-max and CL(int). Immunoreactivity and function were correlated for each isozyme, but there was no cross correlation betwee n isozymes. Conclusion: The velocity of metabolite formation (V-max) b y the three major human drug metabolising P450 monooxygenases is corre lated with their immunoreactivity in liver microsomes. Interindividual variation was much larger for V-max than k(M). Interindividual variab ility was more pronounced for CYP2D6, probably due to the presence of several different functional alleles in the population of extensive me tabolisers.