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The cynomolgus monkey is widely used as a primate model in preclinical studies because of its evolutionary closeness to humans. Despite their importance in drug metabolism, the content of each cytochrome P450 (P450) enzyme has not been systematically determined in cynomolgus monkey livers. In this study, liver microsomes of 27 cynomolgus monkeys were analyzed by immunoblotting using selective P450 antibodies. The specificity of each antibody was confirmed by analyzing the cross-reactivity against 19 CYP1-3 subfamily enzymes using recombinant proteins. CYP2A, CYP2B6, CYP2C9/19, CYP2C76, CYP2D, CYP2E, CYP3A4, and CYP3A5 were detected in all 27 animals. In contrast, CYP1A, CYP1D, and CYP2J were below detectable levels in all liver samples. The average content of each P450 showed that among the P450s analyzed CYP3A (3A4 and 3A5) was the most abundant (40% of total immunoquantified P450), followed by CYP2A (25%), CYP2C (14%), CYP2B6 (13%), CYP2E1 (11%), and CYP2D (3%). No apparent sex differences were found for any P450. Interanimal variations ranged from 2.6-fold (CYP3A) to 11-fold (CYP2C9/19), and most P450s (CYP2A, CYP2D, CYP2E, CYP3A4, and CYP3A5) varied 3- to 4-fold. To examine the correlations of P450 content with enzyme activities, metabolic assays were performed in 27 cynomolgus monkey livers using 7-ethoxyresorufin, coumarin, pentoxyresorufin, flurbiprofen, bufuralol, dextromethorphan, and midazolam. CYP2D and CYP3A4 contents were significantly correlated with typical reactions of human CYP2D (bufuralol 1'-hydroxylation and dextromethorphan O-deethylation) and CYP3A (midazolam 1'-hydroxylation and 4-hydroxylation). The results presented in this study provide useful information for drug metabolism studies using cynomolgus monkeys.

Fetal liver CYP3A7 plays an important role in placental estriol synthesis during pregnancy, yet little is known concerning the extent or consequences of variability in expression. The purpose of this investigation was to characterize the variability in CYP3A7 expression using several phenotypic measures in a panel of 54 fetal livers ranging in age from 76 days to 32 weeks gestation. CYP3A7 mRNA expression was measured using quantitative polymerase chain reaction, whereas immunoreactive CYP3A7 was determined using an affinity-purified antipeptide antibody. Variability in catalytic activity was evaluated using testosterone and dehydroepiandrosterone (DHEA) as substrates. Across the entire panel, CYP3A7 was the most abundant CYP3A mRNA species present and varied 634-fold from 151 to 95,700 transcripts/ng total RNA, corrected for 18S ribosomal RNA. CYP3A4 expression was minimal based on mRNA expression (1000-fold lower than CYP3A7) and the ratio of testosterone 2alpha- (T2alphaH) to 6beta- (T6betaH) hydroxylation. T2alphaH and T6betaH were highly correlated (r(2) = 0.859), and the correlation increased (r(2) = 0.974) in livers with CYP3A5*3/*3 genotypes implying that the same enzyme (CYP3A7) generated both products. Overall, T2alphaH and DHEA16alphaH activities varied 175- and 250-fold, respectively. A subset of five samples had extremely low mRNA, protein, and catalytic activity, possibly due to pathology affecting fetal viability (anencephaly, porencephaly). In the remaining samples, T2alphaH activity varied 6.7-fold (358 +/- 142, range 97 to 643 pmol/min/mg) and DHEA16alphaH activity varied 6.2-fold (8.07 +/- 2.87, range 2.41 to 14.9 nmol/min/mg). Observed variability in CYP3A7 activity was not related to CYP3A7*2, and alternative regulatory mechanisms require further investigation.