Allele and genotype frequencies of CYP2C9, CYP2C19 and CYP2D6 in an Italian population
Introduction
Genetic polymorphisms are known to contribute to interindividual variations in the metabolism of numerous drugs in humans [1], [2]. Mutation in a gene coding for a drug metabolising enzyme can cause enzyme variants with high, low or no activity [3]. Pharmacogenetic polymorphisms divide the population into at least two phenotypes, poor (or slow) metabolisers (PM) and extensive (or rapid) metabolisers (EM), and, eventually, ultrarapid metabolisers (UM) [4]. The PM condition can lead to an excessive or prolonged therapeutic effect or drug-related toxicity after a normal dose, conferring a genetic predisposition to drug-induced adverse effects. Moreover, in case of compounds that need to be activated, the PM condition may result associated with decreased response. On the contrary, UMs may not achieve therapeutic levels of the drug given at a standard dose and this might account for lack of therapeutic effect. Furthermore, in cases where the enzyme bioactivates the drug, increased plasma levels of the active metabolite can be expected in UMs. Determination of an individual’s metabolic capacity by use of phenotyping or genotyping tests may become an important tool for a more rational and safe drug administration, especially for agents with a narrow therapeutic index. The major genetic polymorphisms affecting drug metabolising enzymes activity of potential clinical relevance are those related to drug oxidation by cytochrome P450 enzymes (CYP) 2C9, 2C19 and 2D6 [5], [6].
The human CYP2C subfamily consists of at least four isoforms, 2C8, 2C9, 2C18 and 2C19, whose genes are located together on chromosome 10. CYP2C9, the most abundant among human CYP2C isoforms, metabolises a number of therapeutically important drugs, including most nonsteroidal anti-inflammatory drugs, S-warfarin, phenytoin and losartan (Table 1), and is polymorphically expressed [7]. In vitro studies have shown that even relatively conservative changes in the aminoacid sequence may significantly alter both enzymatic activity and substrate specificity [8]. To date, three different allelic variants (CYP2C9*1, CYP2C9*2, CYP2C9*3), coding for enzymes with different catalytic activity, have been well characterised [7]. The frequencies of the detrimental alleles CYP2C9*2 and CYP2C9*3 vary between 8 and 12% and 3 and 8%, respectively, among Caucasians, while they are lower in Orientals and Black Africans [7], [9]. Additional rare defect alleles have been described, but so far their impact on the enzyme activity in vivo is unclear (http://www.imm.ki.se/CYPalleles).
CYP2C19, an isoform contributing to the clearance of S-mephenytoin, diazepam, omeprazole, proguanil, citalopram, R-warfarin and many antidepressants (Table 1), also exhibits genetic polymorphism [2]. About 3% of Caucasians have been found to be PMs of S-mephenytoin with very little variation noted between the studies [10], [11]. By contrast, several independent studies have shown a much higher incidence of PMs in Orientals, up to 18–23% in Japanese [12], [13]; 15–17% in Chinese [14], [15]; 12–16% in Koreans [16], [17]. In Black Africans, PM frequencies vary between 4 and 7% [18]. The PM condition is inherited as an autosomal recessive trait [11]. The best characterised defect CYP2C19 alleles responsible for the PM phenotype are CYP2C19*2 and CYP2C19*3 [19], [20]. CYP2C19*2 accounts for 75% of the defective alleles in Orientals [19], and 93% in Caucasians [21]. The other well characterised detrimental allele (CYP2C19*3), discovered in Japanese PMs [20], accounts for approximately 25% of all inactive forms in Orientals, being by converse extremely rare in non-Oriental populations [22]. Therefore, the defective forms are still uncharacterised in 15% of the Caucasian PMs. Additional rare defect alleles have been described (http://www.imm.ki.se/CYPalleles), but their impact on the enzyme activity has to be further clarified.
CYP2D6, although expressed at rather low levels compared with other human CYPs, plays an important role in drug metabolism, being partially or to a major extent responsible for the oxidative biotransformation of a variety of psychoactive and cardiovascular drugs (Table 1) [6]. The gene encoding the CYP2D6 enzyme is located on the long arm of chromosome 22. CYP2D6 activity ranges from complete deficiency to ultrarapid metabolism, depending on the existence of a large number of allelic variants of the CYP2D6 gene, causing either absent, decreased or increased enzyme activity in relation to the functional wild type allele [23]. The activity of CYP2D6 is bimodally distributed in Caucasian populations. Individuals with deficient enzyme activity (PM) represent approximately 3–10% of Caucasians, but only 1–2% of Orientals [6]. Among the EMs, 2–10% carry multiple copies of a functional CYP2D6 allele [6]. This genotype has been shown to be associated with extremely high CYP2D6 activity (ultrarapid metabolisers, UM) [24], [25]. The CYP2D6 gene is extremely polymorphic. To date, more than 70 allelic variants have been described (http://www.imm.ki.se/CYPalleles). Four major mutated alleles, CYP2D6*3, CYP2D6*4, CYP2D6*5 and CYP2D6*6, account for 90–95% of the PM alleles in Caucasians. The most common allele associated with the PM phenotype is CYP2D6*4, with an allele frequency of ∼21% in Caucasian populations [26]. The frequency of the CYP2D6*5 allele, with deletion of the entire CYP2D6 gene, is ∼4–6%, very similar in different ethnic populations [26]. Other detrimental alleles frequently (∼2–3%) found among Caucasians are CYP2D6*3, and CYP2D6*6 [6], [26]. Furthermore, alleles with duplication or multiduplication of a functional CYP2D6 gene (*1 or *2), causing increased CYP2D6 activity, have been described. The incidence of CYP2D6 gene duplication is only 1% in Sweden [25], 3% in Germany [23], but as high as 7–10% in Spain [27], [28], 9% in Turkey [29] and 10% in Italy [30]. There is thus a clear north-south gradient of the frequency within Europe. The highest incidence of gene duplication has been found in Ethiopia in Northern Africa with 29% [31].
Considering the pharmacological implications of CYP2C9, CYP2C19 or CYP2D6 genetic polymorphism, the definition of allele distribution pattern might represent a helpful support in the optimisation of pharmacological therapies. Therefore, in this study we first determined the CYP2C9, CYP2C19 and CYP2D6 genotype profile of a random Italian population by screening for the main allelic variants and compared their frequencies with previous findings in other Caucasian, as well African and Oriental populations.
Section snippets
Genotype evaluation
Three hundred and sixty unrelated Italian healthy volunteers (210 males and 150 females, aged 19–52 years, mean±S.D.: 30±9 years) of Caucasian origin, most students or employees at the Medical School of the University of Messina, Italy, were enrolled in the study. The study protocol was approved by the Ethics Committee at the University of Messina, Italy, and written informed consent to participate in the study was obtained from the volunteers. Five milliliters venous blood was obtained from
Results
The CYP2C9, CYP2C19 and CYP2D6 alleles as well as the genotypes frequencies in the Italian population are summarised in Table 2.
Twenty-three subjects (6.4%) carried two detrimental CYP2C9 alleles, while 114 (31.7%) carried one detrimental allele.
Six individuals (1.7%) were carrying two CYP2C19 mutated alleles, being homozygous for CYP2C19*2, and could therefore be classified as PM, while 68 subjects (18.9%) were carrying one mutated allele (CYP2C19*1/*2). CYP2C19*3 was not detected in the
Discussion
Inter-individual variation in CYP expression can lead to marked variability in drug response, drug activity or detoxification and therefore it is important to understand the genetic factors that influence CYP levels and activities.
Three major variants of the CYP2C9 gene have been so far well characterised in Caucasian populations [7]. Allelic variants CYP2C9*2 and CYP2C9*3 code for enzymes with approximately 10–40% and 5–15% of the activity of CYP2C9*1, for different substrates, respectively
Acknowledgements
This study has been supported by a grant from the Italian Society of Pharmacology (SIF).
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