Development of genotyping method for functionally relevant variants of cytochromes P450 in cynomolgus macaques

Abstract

In cynomolgus macaques (Macaca fascicularis), widely used in drug metabolism studies, CYP2C9, CYP2C76, CYP2D6, CYP3A4, and CYP3A5, important drug‐metabolizing enzymes, are abundantly expressed in liver and metabolize cytochrome P450 substrates. CYP2C9 (c.334A>C), CYP2C76 (c.449TG>A), CYP2D6 (c.891A>G), CYP3A4 (IVS3 + 1G>del), and CYP3A5 (c.625A>T) substantially influence metabolic activity of enzymes, and thus are important variants in drug metabolism studies. In this study, a real‐time PCR method was developed for genotyping these variants. The validity of the methods was verified by genotyping two wild type, two heterozygous, and two homozygous DNAs and was used to genotype 41 cynomolgus macaques (from Cambodia, Indonesia, the Philippines, or Vietnam) for the five variants, along with another important variant CYP2C19 (c.308C>T). The CYP2C9 and CYP2C19 variants were found only in Cambodian and Vietnamese animals, while the CYP2C76 and CYP2D6 variants were found only in Indonesian and Philippine animals. The CYP3A4 and CYP3A5 variants were not found in any of the animals analyzed. Mauritian animals, genotyped using next‐generation sequencing data for comparison, possessed the CYP2C19 and CYP2D6 variants, but not the other variants. These results indicated differences in prevalence of these important variants among animal groups. Therefore, the genotyping tool developed is useful for drug metabolism studies using cynomolgus macaques.

Does Geographical Origin of Long-Tailed Macaques (Macaca fascicularis) Matter in Drug Safety Assessment?: A Literature Review and Proposed Conclusion

“…26 Previous single-nucleotide polymorphism also indicated an Indochinese animal origin from Chinese breeding farms. 26 This observation is further supported by cytochrome P450 variant analysis, since the CYP2C9 and CYP2C19 variants-predominant in Cambodian and Vietnamese origin longtailed macaques 27 -are also prevalent in long-tailed macaques bred in China. 28 Long-tailed macaques carry some relatedness to rhesus monkeys, who have the second-largest habitat distribution among NHPs.…”

Cytochrome P450 1A1, 2C9, 2C19, and 3A4 Polymorphisms Account for Interindividual Variability of Toxicological Drug Metabolism in Cynomolgus Macaques

“…Such a variable metabolism might be accounted for by genetic variants of cynomolgus CYP2C9 or CYP2C19 analyzed in this study because these enzymes catalyze S -mephenytoin 4′-hydroxylation. ,, Moreover, the CYP2C9 p.I112L and CYP2C19 p.[F100N; A103V; I112L] relevant to the enzyme function are responsible for metabolism variations. − Because numerous variants of CYP2C9 and CYP2C19 remain to be characterized, further study of those variants might reveal more variants relevant to the enzyme function. For such important variants, a genotyping tool could be developed to aid drug metabolism studies using cynomolgus macaques, as was done for the CYP2C9 and CYP2C19 variants. , …”

“…For such important variants, a genotyping tool could be developed to aid drug metabolism studies using cynomolgus macaques, as was done for the CYP2C9 and CYP2C19 variants. 20,53 It is established that multiple forms of cynomolgus P450 enzymes have generally similar substrate recognition functionality to human P450 enzymes. 51 Genetic/acquired individual differences in cynomolgus P450 enzymes involved in drug oxidation could be associated with toxicological effects.…”