The CYP3A4 intron 6 C>T polymorphism (CYP3A4*22) is associated with reduced CYP3A4 protein level and function in human liver microsomes

Showa Pharmaceutical University.
The Journal of Toxicological Sciences (Impact Factor: 1.29). 05/2013; 38(3):349-54. DOI: 10.2131/jts.38.349
Source: PubMed


Effects of the CYP3A4 intron 6 C>T (CYP3A4*22) polymorphism, which has recently been reported to have a critical role in vivo, were investigated by measuring CYP3A4 protein expression levels and CYP3A4-dependent drug oxidation activities in individual human liver microsomes in vitro. Prior to protein analysis, analysis of DNA samples indicated that 36 Caucasian subjects were genotyped as CYP3A4*1/*1 and five subjects were CYP3A4*1/*22, with a CYP3A4*22 allelic frequency of 6.1%. No CYP3A4*22 alleles were found in the Japanese samples (106 alleles). Individual differences in CYP2D6-dependent dextromethorphan O-demethylation activities in liver microsomes from Caucasians were not affected by either the CYP3A4*1/*22 or CYP3A5*1/*3 genotype. Liver microsomes genotyped as CYP3A4*1/*22 (n = 4) showed significantly lower CYP3A-dependent dextromethorphan N-demethylation, midazolam 1'-hydroxylation, and testosterone 6β-hydroxylation activities, as well as lower expression levels of CYP3A protein (28% of control), compared with those of the CYP3A4*1/*1 group (n = 19). The other polymorphism, CYP3A5*1/*3, did not show these differences (n = 4). The CYP3A4*22 polymorphism was associated with reduced CYP3A4 protein expression levels and resulted in decreased CYP3A4-dependent activities in human livers. The present results suggest an important role of low expression of CYP3A4 protein associated with the CYP3A4*22 allele in the individual differences in drug clearance.

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    • "In humans, CYP3A4 and minor CYP3A isoforms contribute to xenobiotic metabolism, owing to overlap in substrate specificity, inducers and inhibitors; moreover, accumulating evidence has revealed that CYP3As exhibit marked ethnic and individual variability, with obvious consequences (Daly, 2006; Liu et al., 2007; Okubo et al., 2013). So far, no data about the effects of physiological factors or prototypical CYP inducers on the expression of cattle hepatic CYP3A individual isoforms have been published. "
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    ABSTRACT: In humans and laboratory animals, knowledge about cytochrome P450 (CYP) regulation and function is detailed and very extensive. However, CYPs have still been incompletely characterized in veterinary species so far. In this study, mRNA levels of three CYP3A enzymes (CYP3A28, CYPA38 and CYPA48) were measured in cattle liver by using quantitative real-time RT-PCR (qPCR) assays and an absolute quantification approach. In particular, the possible presence of breed-differences in CYP3A expression was investigated in five different meat cattle breeds (Charolais, CH; Piedmontese, PM; Blonde d'Aquitaine, BA; Marchigiana, MA; Valdostana, VALD) and the potential transcriptional effect of the prototypical inducer phenobarbital (PB) upon the CYP3A isoforms was evaluated. Cytochrome P450 3A38 showed the highest amounts of gene copy numbers, followed by CYP3A48 and CYP3A28. Significant breed-differences in CYP3A gene abundances were found, and PB significantly up-regulated all the CYP3As isoforms. The data provide new information about CYP3A expression in cattle, particularly the heterogeneity in the pattern of expression of distinct hepatic CYP3As (CYP3A38>3A48>>3A28), the significant effect of breed, and their common up-regulation following the exposure to PB, although with different orders of magnitude.
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    • "Multiple enzymes participate in clopidogrel activation, which includes CYP3A4.1 The intronic variant CYP3A4*22 is associated with reduced protein expression, by unknown mechanism, and thus, reduced activity.27 Therefore, it is expected that the proportion of clopidogrel activation via CYP3A4 is diminished. "
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    ABSTRACT: Recent candidate gene studies using a human liver bank and in vivo validation in healthy volunteers identified polymorphisms in cytochrome P450 (CYP) 3A4 gene (CYP3A4*22), Ah-receptor nuclear translocator (ARNT), and peroxisome proliferator-activated receptor-α (PPAR-α) genes that are associated with the CYP3A4 phenotype. We hypothesized that the variants identified in these genes may be associated with altered clopidogrel response, since generation of clopidogrel active metabolite is, partially mediated by CYP3A activity. Blood samples from 211 subjects, of mixed racial background, with established coronary artery disease, who had received clopidogrel, were analyzed. Platelet aggregation was determined using light transmittance aggregometry (LTA). Genotyping for CYP2C19*2, CYP3A4*22, PPAR-α (rs4253728, rs4823613), and ARNT (rs2134688) variant alleles was performed using Taqman® assays. CYP2C19*2 genotype was associated with increased on-treatment platelet aggregation (adenosine diphosphate 20 μM; P=0.025). No significant difference in on-treatment platelet aggregation, as measured by LTA during therapy with clopidogrel, was demonstrated among the different genotypes of CYP3A4*22, PPAR-α, and ARNT. These findings suggest that clopidogrel platelet inhibition is not influenced by the genetic variants that have previously been associated with reduced CYP3A4 activity.
    Clinical Pharmacology: Advances and Applications 12/2013; 5:185-92. DOI:10.2147/CPAA.S53151
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    ABSTRACT: Several studies have shown that renal failure decreases cytochrome P450 (CYP)3A activity and that uremic toxins may play a role via transcriptional or translational modifications of CYP enzymes and direct inhibition of CYP-mediated metabolism. In this study, we evaluated the relationship between CYP3A activity using plasma concentration of 4β-hydroxycholesterol as a biomarker and clinical characteristics including plasma concentrations of indoxyl sulfate (3-INDS) and indole-3-acetic acid (3-IAA) in stable kidney transplant recipients. Forty-five Japanese kidney transplant recipients who underwent transplantation more than 90 days prior to the study were included. Morning blood samples were collected and plasma concentrations of 4β-hydroxycholesterol, 3-INDS and 3-IAA were measured. Plasma concentrations of 4β-hydroxycholesterol were 57.1 ± 11.2, 42.1 ± 11.8 and 34.5 ± 7.3 ng/mL in recipients with CYP3A5*1/*1 (n = 5), *1/*3 (n = 15) and *3/*3 (n = 25) genotypes, respectively, with significant differences between three genotypes. A significant correlation was observed between plasma concentrations of 4β-hydroxycholesterol and 3-INDS but not 3-IAA. Multiple regression analysis identified the number of CYP3A5*3 allele in genotype, plasma concentration of 3-INDS and body weight as independent variables associated with plasma concentration of 4β-hydroxycholesterol. In conclusion, these results suggest that CYP3A5 polymorphism and plasma concentration of 3-INDS may account for the inter-individual variability of CYP3A activity, and that plasma concentration of 3-INDS may partially explain the gap in CYP3A activity that cannot be explained by genetic contribution in patients with renal failure.
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