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ABSTRACT:The monkey CYP2C76 gene does not correspond to any of the human CYP2C genes, and its enzyme is at least partly responsible for the species difference occasionally seen in drug metabolism between monkeys and humans. To establish a line and/or lines of monkeys that are expected to show metabolic patterns highly similar to humans, we set out to find monkeys that lacked CYP2C76 activity. By genetic screening of 73 monkeys and a database search of expressed sequence tags, we found a total of 10 nonsynonymous genetic variants in the coding region of CYP2C76, including a null genotype (c.449TG>A). Some of the variants were differently distributed between two animal groups originating from different geographical regions (Indochina and Indonesia). After screening 170 additional genomic samples, we identified a total of eight animals (six males and two females) that were heterozygous for c.449TG>A, which could be used for establishing a homozygous line. If the homozygotes show drug-metabolizing properties more similar to humans than wild-type monkeys, the homozygotes may serve as a better animal model for drug metabolism. The data presented in this article provide the essential genetic information to perform a successful study by using cynomolgus monkeys and present a possible tool to generate a better animal model for drug metabolism.

The cynomolgus monkey is used to study drug metabolism because of its evolutionary closeness to humans. Despite their importance, regional distribution of cytochrome P450 (CYP) enzymes including CYP3As in the liver and small intestine, the major sites of drug metabolism, has not been fully investigated in cynomolgus monkeys. In this study, we measured mRNA expression levels of 14 CYPs in the CYP1, 2, and 3 subfamilies, including orthologs of human CYP3A4 and CYP3A5, in the liver and small intestine of cynomolgus monkeys. Expression levels of each CYP mRNA in various regions of the liver were quantified and comparisons were made between the right lobe, quadrate lobe, left medial lobe, left lateral lobe, and caudate lobe and with four different sections of the right lobe. In the small intestine, the same mRNAs were measured in the duodenum and six different sections from the proximal jejunum to the distal ileum. Expression levels of the CYP mRNAs were not substantially different between liver samples, but varied between the different sections of the small intestine, including CYP3A4. These results suggest that analysis of distinct sections is required for a better understanding of cynomolgus monkey CYPs in the small intestine.

The cynomolgus monkey is an animal species widely used to study drug metabolism because of its evolutionary closeness to humans. However, drug-metabolizing enzyme activities have not been compared in various parts of the liver and small intestine in cynomolgus monkeys. In this study, therefore, drug-metabolizing enzyme activities were analyzed in the liver (the five lobes) and small intestine (six sections from the duodenum to the distal ileum). 7-Ethoxyresorufin O-deethylation, coumarin 7-hydroxylation, paclitaxel 6α-hydroxylation, diclofenac 4'-hydroxylation, tolbutamide methylhydroxylation, S-mephenytoin 4'-hydroxylation, bufuralol 1'-hydroxylation, chlorzoxazone 6-hydroxylation, midazolam 1'-hydroxylation, and testosterone 6β-, 16α-, 16β-, and 2α-hydroxylation were used as the probe reactions for this investigation. In liver, all probe reactions were detected and enzyme activity levels were similar in all lobes, whereas, in the small intestine, all enzyme activities were detected (except for coumarin 7-hydroxylase and testosterone 16α-hydroxylase activity), but from jejunum to ileum there was a decrease in the level of enzyme activity. This includes midazolam 1'-hydroxylation and testosterone 6β-hydroxylation, which are catalyzed by cynomolgus monkey cytochrome P450 (CYP) 3A4/5, orthologs of human CYP3A4/5, which are important drug-metabolizing enzymes. The data presented in this study are expected to facilitate the use of cynomolgus monkeys in drug metabolism studies.

ABSTRACT. Cytochrome P450 2B6 (CYP2B6), an important drug-metabolizing enzyme, is involved in the metabolism of prescribed drugs in humans. Despite its importance, cDNA for a CYP2B6 ortholog has not been identified and characterized in cynomolgus macaques, which are frequently used in preclinical studies. In this study, cDNA highly homologous to human CYP2B6 was cloned from the cynomolgus macaque liver. This cDNA contained an open reading frame of 491 amino acids and functional domains characteristic for CYP protein, such as substrate recognition sites and a heme-binding region. Cynomolgus CYP2B6 was expressed predominantly in the liver with some extra-hepatic expression among 10 tissues. Moreover, cynomolgus CYP2B6 revealed activities toward testosterone 16-hydroxylation and bupropion hydroxylation. These results suggest that cynomolgus CYP2B6 has a functional role in the liver.