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Modeling of human hepatic CYP3A4 enzyme kinetics, protein, and mRNA indicates deviation from log-normal distribution
Sherwin K B Sy1, Antonio Ciaccia, Wei Li
1Department of Pharmacology, University of Toronto, Medical Sciences Building, Toronto, Ontario, Canada. Sherwin.Sy@utoronto.ca
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 micro M 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.