PXR polymorphisms interacted with CYP2B6 polymorphisms on methadone metabolites
Journal
Journal of Clinical Psychopharmacology
Journal Volume
33
Journal Issue
1
Pages
137-140
Date Issued
2013
Author(s)
Tsai H.-J.
Wang S.-C.
Tian J.-N.
Chang T.-K.
Ho I.-K.
Hsiao C.-F.
Chen C.-H.
Tan H.K.-L.
Lin L.
Su L.-W.
Huang C.-L.
Yang Y.-H.
Liu M.-L.
Lin K.-M.
Liu Y.-L.
Abstract
To the Editors
Methadone is well known as a maintenance drug to reduce heroin use and to prevent heroin addicts from experiencing withdrawal symptoms.1 The chemical composition of methadone includes a chiral center and a pair of enantiomers in either an R- or S-form.2 Previous results have suggested that the pharmacological activities of methadone in the plasma are dependent on the type of the enantiomer present.3 Methadone is almost exclusively metabolized in the liver through specific isoforms of the cytochrome P-450 (CYP) enzyme system.4 Previous in vitro studies have documented 2 CYP isoforms (CYP3A4 and CYP2B6) involved in methadone metabolism.4,5 It has been reported that the nuclear receptors pregnane X receptor (PXR), a xenobiotic nuclear receptor (MIM*603065), plays a role in regulating the expression of CYP enzymes.6 Pregnane X receptor is an essential regulator of a wide spectrum of drug disposition genes corresponding to all phases of drug metabolism, including CYP enzymes, phase II enzymes, and drug transporters such as the ABC family.7 Currently, the genetic influence of PXR on methadone metabolites, either individually or jointly via interaction with CYP isoforms, has remained largely unknown. The aim of this study was to examine whether PXR single nucleotide polymorphisms (SNPs) individually influenced and/or interacted with CYP2B6 genetic variants on plasma methadone metabolites.
The study cohort consisted of 366 heroin addicts from 6 participating hospitals who were undergoing methadone maintenance treatment (MMT) as outpatients in 2009. All the hospitals implemented similar low-threshold, once daily, outpatient MMT programs as described previously.8 Likewise, the inclusion/exclusion criteria have been described elsewhere.8 This project is registered with the National Institutes of Health Clinical Trial database (http://www.clinicaltrial.gov/ct/show/NCT01059747), and our study protocol was approved by the institutional review boards of the National Health Research Institutes and the 6 participating hospitals. Written informed consent was obtained from each participant.
Plasma methadone metabolites were measured using high-performance liquid chromatography, as described in our previous study.9 DNA was extracted from whole blood lymphocyte pellets from each subject using a Puregene kit (Gentra Systems, Minneapolis, MN). Detailed information for 25 selected PXR SNPs and the corresponding pairwise linkage disequilibrium (LD) patterns are provided in Supplemental Table A (Supplemental Digital Content 1, https://links.lww.com/JCP/A147) and Supplemental Figure A (Supplemental Digital Content 2, https://links.lww.com/JCP/A148). Detailed information on CYP2B6 gene polymorphisms is available elsewhere.9 All genotyping was performed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), which has been widely used in a broad variety of clinical applications.10,11 We tested whether the 25 PXR SNPs were in Hardy-Weinberg equilibrium (HWE). A pairwise LD quantity (R2) was computed for all subjects. Linkage disequilibrium blocks were identified using the Haploview program.12 The interactions between PXR and CYP2B6 on plasma methadone metabolites were examined using multiple linear regressions. Pregnane X receptor CYP2B6 interactions were tested by adding a product term in each regression model. Covariates included in all regression analyses were sex and body mass index. We applied a false discovery rate to correct for multiple testing.13 All data analyses were performed using the statistical packages R 2.10.0 (http://www.r-project.org) and the PLINK program.14
We first investigated whether any of 22 PXR SNPs (with a minor allele frequency of >5% among 25 genotyped PXR SNPs) was associated with plasma methadone metabolites but did not find such a relationship (data not shown). We speculated that PXR may affect plasma methadone metabolites through CYP2B6 gene polymorphisms. Therefore, we tested the interactive effects of PXR and CYP2B6 on plasma methadone metabolites. The results in Table 1 showed significant interactive effects of PXR-CYP2B6 on the plasma S-methadone concentration, dose-corrected plasma S-methadone concentration, S-EDDP/S-methadone ratio, and apparent clearance of S-methadone in subjects undergoing MMT. We then examined whether different numbers of unfavorable alleles for the 2 most identified SNPs (rs1464603 in PXR and rs10403955 in CYP2B6) would lead to different levels of plasma methadone metabolites in the carrier subjects. The alleles “C” for rs1464603 and “G” for rs10403955 were defined as unfavorable alleles because they were associated with increasing levels of plasma methadone metabolites, specifically the dose-corrected plasma S-methadone concentration. As shown in Supplementary Figure B (Supplemental Digital Content 3, https://links.lww.com/JCP/A149), the dose-corrected plasma S-methadone concentration tended to increase in subjects carrying a greater number of unfavorable alleles.
Methadone is well known as a maintenance drug to reduce heroin use and to prevent heroin addicts from experiencing withdrawal symptoms.1 The chemical composition of methadone includes a chiral center and a pair of enantiomers in either an R- or S-form.2 Previous results have suggested that the pharmacological activities of methadone in the plasma are dependent on the type of the enantiomer present.3 Methadone is almost exclusively metabolized in the liver through specific isoforms of the cytochrome P-450 (CYP) enzyme system.4 Previous in vitro studies have documented 2 CYP isoforms (CYP3A4 and CYP2B6) involved in methadone metabolism.4,5 It has been reported that the nuclear receptors pregnane X receptor (PXR), a xenobiotic nuclear receptor (MIM*603065), plays a role in regulating the expression of CYP enzymes.6 Pregnane X receptor is an essential regulator of a wide spectrum of drug disposition genes corresponding to all phases of drug metabolism, including CYP enzymes, phase II enzymes, and drug transporters such as the ABC family.7 Currently, the genetic influence of PXR on methadone metabolites, either individually or jointly via interaction with CYP isoforms, has remained largely unknown. The aim of this study was to examine whether PXR single nucleotide polymorphisms (SNPs) individually influenced and/or interacted with CYP2B6 genetic variants on plasma methadone metabolites.
The study cohort consisted of 366 heroin addicts from 6 participating hospitals who were undergoing methadone maintenance treatment (MMT) as outpatients in 2009. All the hospitals implemented similar low-threshold, once daily, outpatient MMT programs as described previously.8 Likewise, the inclusion/exclusion criteria have been described elsewhere.8 This project is registered with the National Institutes of Health Clinical Trial database (http://www.clinicaltrial.gov/ct/show/NCT01059747), and our study protocol was approved by the institutional review boards of the National Health Research Institutes and the 6 participating hospitals. Written informed consent was obtained from each participant.
Plasma methadone metabolites were measured using high-performance liquid chromatography, as described in our previous study.9 DNA was extracted from whole blood lymphocyte pellets from each subject using a Puregene kit (Gentra Systems, Minneapolis, MN). Detailed information for 25 selected PXR SNPs and the corresponding pairwise linkage disequilibrium (LD) patterns are provided in Supplemental Table A (Supplemental Digital Content 1, https://links.lww.com/JCP/A147) and Supplemental Figure A (Supplemental Digital Content 2, https://links.lww.com/JCP/A148). Detailed information on CYP2B6 gene polymorphisms is available elsewhere.9 All genotyping was performed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), which has been widely used in a broad variety of clinical applications.10,11 We tested whether the 25 PXR SNPs were in Hardy-Weinberg equilibrium (HWE). A pairwise LD quantity (R2) was computed for all subjects. Linkage disequilibrium blocks were identified using the Haploview program.12 The interactions between PXR and CYP2B6 on plasma methadone metabolites were examined using multiple linear regressions. Pregnane X receptor CYP2B6 interactions were tested by adding a product term in each regression model. Covariates included in all regression analyses were sex and body mass index. We applied a false discovery rate to correct for multiple testing.13 All data analyses were performed using the statistical packages R 2.10.0 (http://www.r-project.org) and the PLINK program.14
We first investigated whether any of 22 PXR SNPs (with a minor allele frequency of >5% among 25 genotyped PXR SNPs) was associated with plasma methadone metabolites but did not find such a relationship (data not shown). We speculated that PXR may affect plasma methadone metabolites through CYP2B6 gene polymorphisms. Therefore, we tested the interactive effects of PXR and CYP2B6 on plasma methadone metabolites. The results in Table 1 showed significant interactive effects of PXR-CYP2B6 on the plasma S-methadone concentration, dose-corrected plasma S-methadone concentration, S-EDDP/S-methadone ratio, and apparent clearance of S-methadone in subjects undergoing MMT. We then examined whether different numbers of unfavorable alleles for the 2 most identified SNPs (rs1464603 in PXR and rs10403955 in CYP2B6) would lead to different levels of plasma methadone metabolites in the carrier subjects. The alleles “C” for rs1464603 and “G” for rs10403955 were defined as unfavorable alleles because they were associated with increasing levels of plasma methadone metabolites, specifically the dose-corrected plasma S-methadone concentration. As shown in Supplementary Figure B (Supplemental Digital Content 3, https://links.lww.com/JCP/A149), the dose-corrected plasma S-methadone concentration tended to increase in subjects carrying a greater number of unfavorable alleles.
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