Modeling of human hepatic CYP3A4 enzyme kinetics, protein, and mRNA indicates deviation from log-normal distribution in CYP3A4 gene expression

Aims: To determine whether the variability in cytochrome P-450 (CYP)3A4 metabolic function is exhibited at both transcription and translation levels and to examine the population distribution of CYP3A4 enzyme kinetics, protein, and mRNA. Methods: Enzyme kinetics of testosterone 6beta-hydroxylation, immunoblot CYP3A4 protein, and CYP3A4 mRNA were determined in a microsomal bank of human livers. The distribution of these determinations was analyzed using cumulative distribution (probit) plots and normality test variable (NTV) to detect deviation from normality. Results: Mean hepatic CYP3A4 protein and relative CYP3A4 mRNA were 35 +/- 23 pmol/mg and 79 59 (CYP3A4/beta-actin), respectively. Kinetic parameter estimates of testosterone 6beta-hydroxylation were 611 +/- 684 pmol/mg/min for maximum rate of the reaction (V-max) and 206 +/- 48 muM for the Michaelis constant (K-m). The CYP3A4 gene expression and its activity exhibited a relatively high degree of interindividual variability. Furthermore, significant correlation between CYP3A4 protein and V-max of testosterone 6beta-hydroxylation (r = 0.82, P < 0.001) as well as CYP3A4 protein and its mRNA (r=0.52, P<0.01) was observed. Cumulative distribution, plots and histograms for the CYP3A4 protein, its mRNA, and maximum testosterone 6beta-hydroxylation exhibited evidences of deviation from log-normal distribution. The minimum NTV value for the distribution of CYP3A4 protein corresponding to the inflection point in the probit occurred at approximately 10% cumulative frequency. The percentage of low CYP3A4 protein phenotype was consistent for CYP3A4 activity and its mRNA. In contrast, the distribution of K-m of testosterone 6beta-hydroxylation does not show evidence of bimodality. Conclusions: The distribution of CYP3A4 metabolic function, protein, and mRNA is non-normal and may represent a regulatory polymorphism in hepatic CYP3A4 gene expression. In contrast, assessment of the possibility of a structural variant of CYP3A4 through evaluation of the standard deviation relative to the mean kinetic constant value suggests that structural mutation of CYP3A4 may not be a major factor affecting interindividual variation in CYP3A4 metabolic function.