Cytochrome P450 2D6 (CYP2D6) is a highly polymorphic enzyme that metabolizes a large number of therapeutic drugs. To date, more than 100 CYP2D6 allelic variants have been reported. Among these variants, we recently identified 22 novel variants in the Chinese population. The aim of this study was to functionally characterize the enzymatic activity of these variants in vitro. A baculovirus-mediated expression system was used to express wild-type CYP2D6.1 and other variants (CYP2D6.2, CYP2D6.10 and 22 novel CYP2D6 variants) at high levels. Then, the insect microsomes containing expressed CYP2D6 proteins were incubated with bufuralol or dextromethorphan at 37°C for 20 or 25 min., respectively. After termination, the metabolites were extracted and used for the detection with high-performance liquid chromatography. Among the 24 CYP2D6 variants tested, two variants (CYP2D6.92 and CYP2D6.96) were found to be catalytically inactive. The remaining 22 variants exhibited significantly decreased intrinsic clearance values for bufuralol 1 0 -hydroxylation and 20 variants showed significantly lower intrinsic clearance values for dextromethorphan O-demethylation than those of the wild-type CYP2D6.1. Our in vitro results suggest that most of the variants exhibit significantly reduced catalytic activities compared with the wild-type, and these data provide valuable information for personalized medicine in Chinese and other Asian populations.The human cytochrome P450 (CYP450) superfamily contains the most important phase I drug-metabolizing enzymes that catalyse the oxidative metabolism of more than 90% of prescribed medications [1]. CYP2D6 (also referred to as debrisoquine 4-hydroxylase) belongs to the CYP2D subfamily and is an important member of the hepatic P450 enzymes. Despite accounting for only approximately 2% of the total human liver CYP protein content, CYP2D6 plays a major role in the metabolism of a wide variety of drugs [2]. It has been reported that approximately 20% of drugs in common clinical use are metabolized by CYP2D6 [3]. These drugs include the antidepressants fluoxetine, amitriptyline and venlafaxine; the antitussive dextromethorphan; the b-adrenergic antagonists bufuralol, propranolol and metoprolol; the opioid analgesics codeine, dihydrocodeine and tramadol; the antipsychotic agent risperidone; and the antiarrhythmic propafenone [4][5][6].

1. CYP2D6 is an important member of the cytochrome P450 (CYP450) enzyme superfamily, we recently identified 22 CYP2D6 alleles in the Han Chinese population. The aim of this study was to assess the catalytic activities of these allelic isoforms and their effects on the metabolism of venlafaxine in vitro. 2. The wild-type and 24 CYP2D6 variants were expressed in insect cells, and each variant was characterized using venlafaxine as the substrate. Reactions were performed at 37 °C with 5-500 μM substrate (three variants was adjusted to 1000 μM) for 50 min. By using high-performance liquid chromatography to detect the products, the kinetic parameters Km, Vmax, and intrinsic clearance (Vmax/Km) of O-desmethylvenlafaxine were determined. 3. Among the 22 CYP2D6 variants, the intrinsic clearance (Vmax/Km) values of all variants were significantly decreased (from 0.2% to 84.5%) compared with wild-type CYP2D6*1. In addition, the kinetic parameters of two CYP2D6 variants could not be detected because they have no detectable enzyme activity. 4. The comprehensive in vitro assessment of CYP2D6 variants provides significant insights into allele-specific activity towards venlafaxine in vivo.

CYP2D6 is an important member of the cytochrome P450 (CYP450) enzyme super family, with at least 100 CYP2D6 alleles being previously identified. Genetic polymorphisms of CYP2D6 significantly influence the efficacy and safety of some drugs, which might cause adverse effects and therapeutic failure. The aim of this study was to clarify the catalytic activities of 24 CYP2D6 alleles on the oxidative in vitro metabolism of methadone. Reactions were incubated with 50-2000  µM methadone for 30 min at 37 °C and terminated by cooling to -80 °C immediately. Methadone and the major metabolite EDDP were analyzed by an ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) system. Compared with wild-type CYP2D6*1, most variants showed significantly altered values in V and intrinsic clearance (V /K ). Only three variants (CYP2D6*88, *91 and E215K) exhibited markedly increased intrinsic clearance values, and one variant CYP2D6*94 showed no significant difference. On the other hand, the kinetic parameters of two CYP2D6 variants (CYP2D6*92 and *96) could not be determined because they had no detectable enzyme activity, whereas 18 variants exhibited significantly decreased values. To sum up, this study demonstrated that more attention should be paid in clinical administration of methadone to individuals carrying these CYP2D6 alleles. Copyright © 2016 John Wiley & Sons, Ltd.

Cytochrome P450 (P450) 2C9 is an important member of the P450 enzyme superfamily, with 58 CYP2C9 allelic variants previously reported. Genetic polymorphisms of CYP2C9 significantly influence the efficacy and safety of some drugs, which might cause adverse effects and therapeutic failure. The aim of this study was to assess the catalytic activities of 38 human CYP2C9 alleles, including 24 novel alleles (*36-*60) found in the Han Chinese population, toward bosentan (BOS) in vitro. Insect microsomes expressing the 38 CYP2C9 alleles were incubated with 10-625 mM bosentan for 30 minutes at 37°C and terminated by cooling to 280°C immediately. BOS and hydroxyl bosentan, the major metabolite of BOS, were analyzed by ultraperformance liquid chromatography-tandem mass spectrometry system. Thirty-eight defective alleles can be classified into three categories according to the relative clearance value compared with wild type: nine alleles exhibited significantly increased intrinsic clearance values (V max /K m ) compared with the wild type (1.5-fold-∼4.9-fold relative clearance); nine alleles exhibited significantly reduced intrinsic clearance values compared with the wild type (0.6-28.9% relative clearance). The remaining 20 alleles exhibited no significant difference (1-fold) in enzyme activity compared with the wild type. These findings suggest that more attention should be directed to subjects carrying these infrequent CYP2C9 alleles when administering BOS in the clinic. This is the first report of all these rare alleles for BOS metabolism, providing fundamental data for further clinical studies on CYP2C9 alleles.

1. The objective of this study were to investigate the effect of orally administered resveratrol on the pharmacokinetics of aripiprazole (APZ) in rat, and the inhibitory effects of resveratrol on APZ dehydrogenation activity in liver microsomes and human cytochrome P450 3A4 and 2D6. 2. Twenty-five healthy male Sprague-Dawley rats were randomly divided into five groups: A (control group), B (multiple dose of 200 mg/kg resveratrol), C (multiple dose of 100 mg/kg resveratrol), D (a single dose of 200 mg/kg resveratrol) and E (a single dose of 100 mg/kg resveratrol). A single dose of 3 mg/kg APZ administered orally 30 min after administration of resveratrol. In addition, CYP2D6*1, CYP3A4*1, human and rat liver microsomes were performed to determine the effect of resveratrol on the metabolism of APZ in vitro. 3. The multiple dose of 200 or 100 mg/kg resveratrol significantly increased the AUC and Cmax of APZ. The resveratrol also obviously decreased the CL, but without any significant difference on t1/2 in vivo. On the other hand, resveratrol showed inhibitory effect on CYP3A4*1, CYP2D6*1, human and rat microsomes, the IC50 of resveratrol was 6.771, 87.87, 45.11 and 35.59 μmol l(-1), respectively. 4. Those results indicated more attention should be paid when APZ was administrated combined with resveratrol.