| Cytochrome P4502C9 (CYP2C9) is a polymorphic enzyme responsible for the metabolism of many clinically important drugs, particularly nonsteroidal anti-inflammatory drugs, antidepressants, antidiabetics, lipid-lowering drugs and oral anticoagulants. There is much inter-individual variation in CYP2C9 activity in the population, due in large part to non-synonymous single-nucleotide polymorphisms (SNPs) in CYP2C9. CYP2C9*3 and CYP2C9*13 are the principal variant alleles found in the Chinese population, the frequency of the two alleles is approximately 3.3% and 1.02%, respectively. CYP2C9*3 has been reported to reduce the metabolism of diclofenac, and thus can alter the extent of drug-drug interactions (DDIs) between diclofenac and other drugs, potentially causing adverse drug reactions (ADRs) in individuals carrying this allele. The effects of CYP2C9*13 on diclofenac metabolism are not well studied, and the influences of CYP2C9*13 on DDIs between diclofenac and clinical drugs are still unknown. Unlike CYP2C9*3, which has been extensively studied in humans, clinical studies of CYP2C9*13 have been limited by the difficulty in finding subjects carrying this low-frequency allele. Therefore, it would be very helpful to be able to predict the pharmacokinetic changes caused by CYP2C9*13 from in vitro data. Methods to predict the effects of CYP2C9 polymorphic alleles on the pharmacokinetics of CYP2C9-metabolized drugs from in vitro data have been successfully established. However, there is little in vitro data currently available for predicting the effects of CYP2C9*13 on in vivo drug metabolism.Studies of CYP2C9 genetic polymorphisms increasingly indicate that the degree of CYP2C9-mediated DDIs can be extensively influenced by variant alleles. For instance, the CYP2C9.3 allelic protein can alter the inhibitory potencies of many drugs toward CYP2C9-mediated metabolism in vitro, and the expression of the CYP2C9*3 allele can also alter the degree of DDIs between flurbiprofen and fluconazole in humans. To date, no studies have reported the effect of CYP2C9*13 on the interaction between diclofenac and other drugs. Understanding the effects of variant CYP2C9 alleles on DDIs can help prevent ADRs in individuals with different CYP2C9 genotypes when co-adminstering diclofenac with other drugs.In this study, CYP2C9.1 (the wildtype enzyme), CYP2C9.3 and CYP2C9.13 were expressed in yeast, and their metabolic kinetics for diclofenac 4'-hydroxylation examined. From the in vitro data, we then predicted the ratio of diclofenac oral clearance (CLoralR) in subjects carrying CYP2C9*3 or CYP2C9*13 alleles. Expression of either variant allele was predicted to decrease diclofenac oral clearance. Furthermore, we investigated the effects of these two alleles on diclofenac-drug interactions. The potentials of nine clinically used drugs to inhibit diclofenac 4'-hydroxylation catalyzed by either CYP2C9.3, CYP2C9.13 or CYP2C9.1 were compared. Our results indicated that CYP2C9.3 and CYP2C9.13 can alter the CYP-inhibitory potencies of some tested drugs. In particular, CYP2C9.13 significantly weakened the inhibitory potencies of sulfaphenazole, fluvastatin, fluvoxamine and tranylcypromine. These data provide helpful guidelines for co-administration of diclofenac with other drugs in individuals carrying CYP2C9*3 and CYP2C9*13 alleles. In the study of genotyping, we detected CYP2C9*3 genotypic distribution in 74 Chinese. The genotype analysis of CYP2C9*3 was based on polymerase chain reaetion (PCR)-restrietion fragment length polymorphism (RFLP), and the result of PCR was verified by direct sequencing analysis. The result indicated that three Chinese carried CYP2C9*1/*3,71 Chinese carried CYP2C9*1/*1, and no CYP2C9*3/*3 was found. The method appeared to be efficient, specific and accurate. It is suitable for the genotype analysis of CYP2C9*3. |
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