Associations Between CYP3A4/3A5、ABCB1Gene Polymorphisms And Carbamazepine、oxcarbazepine、their Main Metabolites Plasma Concentrations In Epileptic Patients

Objective:To establish the HPLC method for the simultaneous determination of carbamazepine, oxcarbazepine, and their main metabolites (carbamazepine-10,11-epoxide, monohydroxy derivative) in plasma samples and investigate the associations between CYP3A4> CYP3A5、ABCB1genetic polymorphisms and plasma concentrations of carbamazepine, carbamazepine-10,11-epoxide, oxcarbazepine, monohydroxy derivative.Methods:1. We recruited the epileptic patients who were treated with carbamazepine or oxcarbazepine from XiangYa hospital, they were all Han Chinese in Central south area and selected by inclusion and exclusion criteria. We registered their clinical data and extracted2ml venous blood in the morning. They were classified into monotherapy and polytherapy group, people took carbamazepine or oxcarbazepine only in the monotherapy group, and in polytherapy group people took carbamazepine (oxcarbazepine) with other antiepileptic drugs, monotherapy group and polytherapy group were included in the sum group.2. A HPLC method was estabolished to monitor carbamazepine、 carbamazepine-10,11-epoxide and oxcarbazepine、monohydroxy derivative in plasma samples of epileptic patients we recruited. Genetic DNA was extracted from whole blood by phenol-chloroform method, CYP3A4rs4646440、rs2242480, CYP3A5rs15524、rs776746, ABCB1rs1045642、rs2032582、rs10234411、rs11285038SNPs polymorphisms were determined by iMLDR.3. The distribution of genotypes in patients and the deviation of enumeration data were statistically analyzed with χ2test, the differences of measurement data and statistical description were performed by independent sample t test or analysis of variance,we used multiple linear regression to detect significant counfouding factors that affecting dose and plasma concentrations of antiepileptic drugs, PLINK were performed to analyze the association between SNPs、haplotypes and adjusted plasma concentrations of antiepileptic drugs.Results:1. We estabolished a rapid、simple HPLC method for the simultaneous determination of carbamazepine、oxcarbazepine carbamazepine-10,11-epoxide、monohydroxy derivative. They were not interfered by peaks of endogenous substances in blank plasma, the linearity of our method checked in their plasma concentration ranges was excellent, and their calibration curves were Y=0.0795254X-0.00128006、 Y=0.0519381X-0.00246967、Y=0.0255575X-0.00102140Y=0.0239854X-0.000662765, The RSDs of intra-and inter-day precision ranged from0.50%-16.67%, the extraction recovery ranged from90.0%-106.8%and the relative recovery ranged from90.7%-103.1%.2. We recruited88epileptic patients who were treated with carbamazepine and69epileptic patients who were treated with oxcarbazepine. allele frequencies of8SNPs fiting HWE (P>0.05) indicated that samples we recruited were group representive.3. In monotherapy group, adjusted carbamazepine concentrations were not associated with SNPs and haplotypes.In polytherapy group, rs776746、rs15524were associated with dose-adjusted carbamazepine concentrations and sum concentrations (P=0.004、0.013、0.007、0.012); rs776746、rs15524were significantly associated with weight, dose-adjusted carbamazepine concentrations and sum concentrations (P=0.004、0.009、0.010、0.009); rs1045642、 rs10234411、rs2032582-rs10234411AT haplotype were assocoated with CBZ-EP/CBZ ratio(P=0.042、0.014、0.010).In total group, rs776746showed association with dose-adjusted carbamazepine concentrations and sum concentrations (P=0.025、0.028), rs15524was associated with dose-adjusted carbamazepine concentrations(p=0.039); rsl5524-rs776746GT were associated with dose-adjusted carbamazepine concentrations and sum concentrations(P=0.028、0.036), rs15524-rs776746AC was associated with dose-adjusted sum concentrations (P=0.045); rs4646440-rs2242480GT was associated with the adjusted CBZ-EP/CBZ ratio(P=0.002).4. In monotherapy group, adjusted oxcarbazepine concentrations were not associated with SNPs and haplotypes. In polytherapy group, rs2032582-rs10234411-rs1045642GAA was associated with dose-adjusted monohydroxy derivative plasma concentrations and sum plasma concentrations (P=0.026、0.029). In total group, rs2032582was associated with age、dose-adjusted oxcarbazepine(P=0.016).Conclusion:1. Our method for the simultaneous determination of carbamazepine、oxcarbazepine、carbamazepine-10,11-epoxide、 monohydroxy derivative was very simple、accurate and suitable for antiepileptic drugs monitoring practice.2. CYP3A5rs776746、rs15524were associated with carbamazepine plasma concentrations, other genetic polymorphisms showed no significant association with carbamazepine and oxcarbazepine plasma concentrations.