Abstract
Aims
We investigated the CYP2C19*17 allelic frequency in Japanese subjects, and evaluated whether CYP2C19*17 is an important determinant of interindividual variability of CYP2C19 activity.
Methods
We enrolled 265 subjects to determine their CYP2C19 genotype and plasma metabolic ratio following a single dose of 40 mg omeprazole.
Results
Seven subjects heterozygous for CYP2C19*17 and no *17/*17 subjects resulted in the CYP2C19*17 frequency being 1.3%. These heterozygotes had moderate metabolic activities when compared with the metabolic ratio of the other subjects.
Conclusions
The low frequency of CYP2C19*17 and the absence of *17/*17 indicates that CYP2C19*17 plays a minor role in the Japanese population.
What is already known about this subject
A novel CYP2C19 gene variant, CYP2C19*17, is associated with increased metabolic activity.
Ethnic differences in the frequency of the variant allele have been reported. However, the frequency of the CYP2C19*17 allele has not been studied in the Japanese population.
What this study adds
In a population of 265 healthy Japanese subjects, a low frequency (1.3%) of the CYP2C19*17 allele was observed.
The limited frequency of the *17 allele and the absence of a subject homozygous for *17 indicated that CYP2C19*17 would play a minor role in a Japanese population.
Keywords: CYP2C19, genetic polymorphism, Japanese
Introduction
Cytochrome P450 (CYP) 2C19 metabolizes a wide variety of drugs, such as the proton pump inhibitors (PPIs) omeprazole and lansoprazole, and the anticonvulsant S-mephenytoin [1–3]. A marked interindividual difference has been reported in CYP2C19 activity and the bimodal distribution of its activity is known as a genetic polymorphism. A subject without the CYP2C19 activity is called a poor metabolizer (PM) of CYP2C19 [4], and ethnic difference is noted in its frequency [5]. A CYP2C19 PM subject carries two defective CYP2C19 alleles, and the frequency of mutant alleles causing CYP2C19 PM also shows ethnic differences [5].
A recent study reported a novel CYP2C19 gene variant (CYP2C19*17, −3402C > T and −806C > T), associated with increased gene transcription and thus ultrarapid metabolism of CYP2C19 substrates [6]. In the study of Sim et al.[6] the allelic frequency of CYP2C19*17 was different between Swedish and Chinese subjects being 18% and 4%, respectively [6], and their findings and previous studies about the frequency of mutant alleles suggest that the frequency of CYP2C19*17 in a Japanese population would be different from that in a Swedish or a Chinese population. In the present study, therefore, we investigated the allelic frequency of CYP2C19*17 in 265 Japanese subjects, and evaluated whether CYP2C19*17 is an important determinant of interindividual variability of CYP2C19 activity among them.
Methods
Two hundred and sixty-five unrelated healthy Japanese volunteers (183 men and 82 women) were enrolled in this study after giving their written informed consent. They were 5th year students (with a mean age of 25.2 years, ranging from 22 to 37 years and a mean body weight of 61.9 kg ranging from 42 to 100 kg) of Hirosaki University School of Medicine from 2004 to 2006, and they had no history of significant medical illness or hypersensitivity to any drugs. The protocol of the present study was approved by the ethics committee of Hirosaki University School of Medicine.
Each subject was administered two tablets of 20 mg omeprazole (Merck Ltd, Osaka, Japan) with 100 ml of tap water at 09.00 h (1–2 h after eating breakfast), and a blood sample (10 ml) for genotyping and phenotyping was obtained at 13.00–14.00 h (4–5 h after administering omeprazole). They did not take any medication or fruit juices for at least 7 days before the administration of omeprazole.
For CYP2C19 genotyping, genomic DNA was extracted from the venous blood using an extraction kit (AquaPure Genomic DNA Blood Kit, BIO-RAD, USA). The defective mutation alleles for CYP2C19, CYP2C19*3 and CYP2C19*2, were identified using the polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) assay [7]. Genotyping of the CYP2C19*17 allele was analyzed by PCR-RFLP (−3402C > T) and allele-specific amplification methods (−806C > T) described by Sim et al.[6]. A subject with two defective mutation alleles was categorized as a PM, and a subject without or with one defective mutation allele as an extensive metabolizer (EM).
Plasma concentrations of omeprazole and 5-hydroxyomeprazole were quantified using an HPLC method developed in our laboratory [8]. The limits of quantification were 3 ng ml−1 for both omeprazole and 5-hydroxyomeprazole, and the intra- and interday coefficient variations were less than 5.1 and 6.6% for omeprazole (range 4–1600 ng ml−1) and 4.6 and 5.0% for 5-hydroxyomeprazole (range 4–400 ng ml−1), respectively [8]. The plasma omeprazole metabolic ratio (MR) (omeprazole : 5-hydroxyomeprazole) 4–5 h after drug intake was used as a measure of CYP2C19 activity.
Results
In our 265 Japanese subjects comprising 217 EMs and 48 PMs of CYP2C19 (Table 1), seven subjects were heterozygous for CYP2C19*17 and none was homozygous, resulting in the frequencies of CYP2C19*1, *2, *3 and *17 being 57.9%, 27.9%, 12.8% and 1.3%, respectively.
Table 1.
CYP2C19 genotype frequencies and metabolic ratio of omeprazole in 265 healthy Japanese volunteers
| Genotype | Number (% of total) | Metabolic ratio (95% CI interval) | |
|---|---|---|---|
| EM (n = 217) | *1/*1 | 94 | 2.18 ± 2.24 |
| (35.5) | (1.71, 2.65) | ||
| *1/*17 | 3 | 0.87 ± 0.25 | |
| (1.1) | (0.24, 1.49) | ||
| *1/*2 or *1/*3 | 116 | 3.97 ± 3.15 | |
| (43.8) | (3.38, 4.55) | ||
| *17/*2 or *17/*3 | 4 | 3.85 ± 2.70 | |
| (1.5) | (−0.45, 8.15) | ||
| PM (n = 48) | *2/*2, *2/*3 or | 48 | 32.32 ± 14.55 |
| *3/*3 | (18.8) | (28.00, 36.64) |
EM, extensive metabolizer; PM, poor metabolizer.
In the seven subjects heterozygous for CYP2C19*17, complete linkage existed between the −3402C > T and −806C > T mutations, and both −3402C > T and −806C > T mutations were absent in the CYP2C19*2 or CYP2C19*3 allele. These seven subjects showed moderate metabolic activities of CYP2C19 (Table 1), with the lowest MR among three subjects with *1/*17 or four subjects with *17/*2 or *17/*3 in 12th or 22nd place when compared with those with *1/*1 or *1/*2 or *1/*3, respectively.
Discussion
We investigated the frequency of CYP2C19*17 in our 265 healthy Japanese subjects, and evaluated whether CYP2C19*17 is an important determinant of interindividual variability of CYP2C19 activity among them. A novel CYP2C19 gene variant (CYP2C19*17) was reported to have ultrarapid metabolic activity of CYP2C19, and ethnic differences were noted in the allelic frequency of CYP2C19*17[6]. In our Japanese subjects, the frequencies of CYP2C19*1, *2, *3 and *17 were 57.9%, 27.9%, 12.8% and 1.3%, respectively. Since the *17 allele was previously genotyped as CYP2C19*1, these allelic frequencies were in line with other published results [5]. The *17 allele frequency in the Japanese subjects was 1.3% of CYP2C19 alleles, which is lower than the *17 frequency in Chinese individuals (4%) [6]. This difference is not unusual because some differences have been reported in CYP2C19 mutant allele frequencies in Asian populations: 39.9% and 6.1% for *2 and *3, respectively, in Chinese subjects [5]; 32% and 5.5% for *2 and *3, respectively, in a Chinese-Taiwanese population [9]; 20.9% and 11.7% for *2 and *3, respectively, in a Korean population [5].
A marked variation in the CYP2C19 activity and lower frequency of the *17 allele implied that the extent of linkage of −3402C > T with the −806C > T mutation affecting gene transcription activity [6] might be different in our Japanese subjects. However, complete linkage existed between the −3402C > T and −806C > T mutations in our Japanese subjects as reported in people of European and Ethiopian descent [6]. The low frequency of CYP2C19–806C > T mutation reported to be 0.8% in a Japanese population [10] was found to be in reasonable agreement with our present findings. In the CYP2C19*2 or *3 allele of our subjects, both −3402C > T and −806C > T mutations were absent, as reported previously [6].
Although the median omeprazole MR of the Swedish *1/*17 subjects was significantly less than that of the *1/*1 subjects [6], our three *1/*17 subjects had moderate metabolic activities compared with the EMs without the *2 or *3 allele, and similar findings were noted in our EMs with one of *2 and *3 alleles. In the present study, the metabolic activity of CYP2C19 was determined by the metabolic ratio of a single point plasma sample 4–5 h after administration of 40 mg omeprazole after breakfast. Since the omeprazole metabolic ratio in a single plasma sample 2–3 h postdose after an overnight fast is proposed as an appropriate phenotyping procedure [11], the differences in the sampling procedure could affect the ratio. However, the limited frequency of the *17 allele and the absence of a subject homozygous for *17 would suggest that the allele is not an important determinant of intersubject variability of CYP2C19 activity in a Japanese population.
Acknowledgments
We thank Drs Makiko Shimizu and Norie Murayama, Laboratory of Drug Metabolism and Pharmacokinetics, Showa Pharmaceutical University, for their skilful assistance.
REFERENCES
- 1.Wrighton SA, Stevens JC, Becker GW, VandenBranden M. Isolation and characterization of human liver cytochrome P450 2C19: correlation between 2C19 and S-mephenytoin 4′-hydroxylation. Arch Biochem Biophys. 1993;306:240–5. doi: 10.1006/abbi.1993.1506. [DOI] [PubMed] [Google Scholar]
- 2.Klotz U, Schwab M, Treiber G. CYP2C19 polymorphism and proton pump inhibitors. Basic Clin Pharmacol Toxicol. 2004;95:2–8. doi: 10.1111/j.1600-0773.2004.pto950102.x. [DOI] [PubMed] [Google Scholar]
- 3.Desta Z, Zhao X, Shin JG, Flockhart DA. Clinical significance of the cytochrome P450 2C19 genetic polymorphism. Clin Pharmacokinet. 2002;41:913–58. doi: 10.2165/00003088-200241120-00002. [DOI] [PubMed] [Google Scholar]
- 4.Kupfer A, Preisig R. Pharmacogenetics of mephenytoin: a new drug hydroxylation polymorphism in man. Eur J Clin Pharmacol. 1984;26:753–9. doi: 10.1007/BF00541938. [DOI] [PubMed] [Google Scholar]
- 5.Kimura M, Ieiri I, Mamiya K, Urae A, Higuchi S. Genetic polymorphism of cytochrome P450s, CYP2C19, and CYP2C9 in a Japanese population. Ther Drug Monit. 1998;20:243–7. doi: 10.1097/00007691-199806000-00001. [DOI] [PubMed] [Google Scholar]
- 6.Sim SC, Risinger C, Dahl ML, Aklillu E, Christensen M, Bertilsson L, Ingelman-Sundberg M. A common novel CYP2C19 gene variant causes ultrarapid drug metabolism relevant for the drug response to proton pump inhibitors and antidepressants. Clin Pharmacol Ther. 2006;79:103–13. doi: 10.1016/j.clpt.2005.10.002. [DOI] [PubMed] [Google Scholar]
- 7.De Morais SM, Wilkinson GR, Blaisdell J, Meyer UA, Nakamura K, Goldstein JA. Identification of a new genetic defect responsible for the polymorphism of (S)-mephenytoin metabolism in Japanese. Mol Pharmacol. 1994;46:594–8. [PubMed] [Google Scholar]
- 8.Shimizu M, Uno T, Niioka T, Yaui-Furukori N, Takahata T, Sugawara K, Tateishi T. Sensitive determination of omeprazole and its two main metabolites in human plasma by column-switching high-performance liquid chromatography: application to pharmacokinetic study in relation to CYP2C19 genotypes. J Chromatogr B Analyt Technol Biomed Life Sci. 2006;832:241–8. doi: 10.1016/j.jchromb.2006.01.022. [DOI] [PubMed] [Google Scholar]
- 9.Goldstein JA, Ishizaki T, Chiba K, de Morais SM, Bell D, Krahn PM, Evans DA. Frequencies of the defective CYP2C19 alleles responsible for the mephenytoin poor metabolizer phenotype in various Oriental, Caucasian, Saudi Arabian and American black populations. Pharmacogenetics. 1997;7:59–64. doi: 10.1097/00008571-199702000-00008. [DOI] [PubMed] [Google Scholar]
- 10.Fukushima-Uesaka H, Saito Y, Maekawa K, Ozawa S, Hasegawa R, Kajio H, Kuzuya N, Yasuda K, Kawamoto M, Kamatani N, Suzuki K, Yanagawa T, Tohkin M, Sawada J. Genetic variations and haplotypes of CYP2C19 in a Japanese population. Drug Metab Pharmacokinet. 2005;20:300–7. doi: 10.2133/dmpk.20.300. [DOI] [PubMed] [Google Scholar]
- 11.Fuhr U, Jetter A, Kirchheiner J. Appropriate phenotyping procedures for drug metabolizing enzymes and transporters in humans and their simultaneous use in the ‘cocktail’ approach. Clin Pharmacol Ther. 2007;81:270–83. doi: 10.1038/sj.clpt.6100050. [DOI] [PubMed] [Google Scholar]