Clinical relevance of VKORC1 (G-1639A and C1173T) and CYP2C9*3 among patients on warfarin

What is known and Objectives: Testing for cytochrome P450‐2C9 (CYP2C9) and vitamin K epoxide reductase complex subunit 1 (VKORC1) variant alleles is recommended by the FDA for dosing of warfarin. However, dose prediction models derived from data obtained in one population may not be applicable to another. We therefore studied the impact of genetic polymorphisms of CYP2C9 and VKORC1 on warfarin dose requirement in Malaysia. Methods: Patients who were attending clinics at our hospital and prescribed warfarin with stabilized INR levels of 2–4 were selected. DNA was extracted from blood samples and subsequently genotyped for CYP2C9*1, *2, *3, VKORC1 (G‐1639A) and VKORC1 C1173T. Linear regression modelling using age, CYP2C9 and VKORC1 genotypes, sex, weight and height was undertaken to define a warfarin dosing algorithm. An initial model was developed using data from one cohort of patients and validated using data from a second cohort. Results and Discussion: A model which included age and variants of CYP2C9 and VKORC1 account for about 37% of the variability in warfarin dose required to achieve INR of 2–4. Among the parameters evaluated, only VKORC1 (G‐1639A) and (C1173T) alleles, and age correlated with warfarin dose at 6 month. The mean dose predicted using the algorithm derived from cohort 1 was lower than the actual dose for cohort 2 (3·30 mg, SD 0·84 vs. 3·45 mg, SD 1·42). There was no relationship between INR values and the dose taken by the patients. Race, sex, weight and height did not correlate with dose. What is new and Conclusion: This study identifies factors which affect warfarin dosing in the Malaysia population. However, our best model does not account sufficiently for the variability in dose requirements for it to be used in dose prediction for the individual patient. Other important influential factors affecting warfarin dose requirement remain to be identified.     

Influence of genetics and non-genetic factors on acenocoumarol maintenance dose requirement in Moroccan patients

What is known and Objective: Coumarin derivatives such as acenocoumarol represent the therapy of choice for the long‐term treatment and prevention of thromboembolic diseases. Many genetic, clinical and demographic factors have been shown to influence the anticoagulant dosage. Our aim was to investigate the contribution of genetic and non‐genetic factors to variability in response to acenocoumarol in Moroccan patients. Methods: Our study included 114 adult Moroccan patients, receiving long‐term acenocoumarol therapy for various indications. Tests for VKORC1 ‐1639G>A promoter polymorphism (rs9923231), CYP2C9*2 rs1799853, CYP2C9*3 rs1057910, and CYP4F2 rs2108622 alleles were undertaken using Taq Man^® Pre‐Developed Assay Reagents for allelic discrimination. The statistical analysis was performed using the SAS V9 statistical package. Results and Discussion: Genotyping showed that the allele frequencies for the SNPs studied were no different to those found in Caucasians population. A significant association was observed between the weekly maintenance dose and the VKORC1 (P = 0·0027) and CYP2C9 variant genotypes (P = 0·0082). A final multivariate regression model that included the target International Normalized Ratio, VKORC1 and CYP2C9 genotypes explained 36·2% of the overall interindividual variability in acenocoumarol dose requirement. What is new and Conclusion: Our study shows large interindividual variability in acenocoumarol maintenance dose requirement in our population. VKORC1 and CYP2C9 variants significantly affected acenocoumarol dose, in‐line with results in other populations. For the Moroccan population, the SNPs that have the largest effect on acecoumarol dose are CYP2C9 rs1799853, CYP2C9 rs1057910 and VKORC1 rs9923231.     

Predicting the warfarin maintenance dose in elderly inpatients at treatment initiation: accuracy of dosing algorithms incorporating or not VKORC1/CYP2C9 genotypes

Summary. Background: Initiating warfarin is challenging in frail elderly patients because of low‐dose requirements and interindividual variability. Objectives: We investigated whether incorporating VKORC1 and CYP2C9 genotype information in different models helped to predict the warfarin maintenance dose when added to clinical data and INR values at baseline (Day 0), and during warfarin induction. Patients: We prospectively enrolled 187 elderly inpatients (mean age, 85.6 years), all starting on warfarin using the same ‘geriatric dosing‐algorithm’ based on the INR value measured on the day after three 4‐mg warfarin doses (INR₃) and on INR_{6 ± 1}. Results: On Day 0, the clinical model failed to accurately predict the maintenance dose (R² < 0.10). Adding the VKORC1 and CYP2C9 genotypes to the model increased R² to 0.31. On Day 3, the INR₃ value was the strongest predictor, completely embedding the VKORC1 genotype, whereas the CYP2C9 genotype remained a significant predictor (model‐ R² 0.55). On Day 6 ± 1, none of the genotypes predicted the maintenance dose. Finally, the simple ‘geriatric dosing‐algorithm’ was the most accurate algorithm on Day 3 (R² 0.77) and Day 6 (R² 0.81), under‐estimating (≥ 1 mg) and over‐estimating the dose (≥ 1 mg) in fewer than 10% and 2% of patients, respectively. Clinical models and the ‘geriatric dosing‐algorithm’ were validated on an independent sample. Conclusions: Before starting warfarin therapy, the VKORC1 genotype is the best predictor of the maintenance dose. Once treatment is started using induction doses tailored for elderly patients, the contribution of VKORC1 and CYP2C9 genotypes in dose refinement is negligible compared with two INR values measured during the first week of treatment.     

基于华法林药物基因组学的临床用药模型建立

目的探讨CYP2C9、CYP4F2和VKORC1基因多态性对中国汉族人群中华法林稳定剂量的影响,评价华法林用药遗传背景,为临床个体化用药奠定基础。方法采用PCR．HRM法对181例接受华法林治疗并达到稳定剂量的中国汉族患者检测CYP2C9＊2、CYP2C9＊3、CYP4F2V433M和VKORC11173C／T四个SNP位点的多态性。基因分型数据结合临床资料（性别、年龄和华法林剂量等）进行统计学分析并建立数学模型。结果20岁以下年龄组与70岁以上年龄组间华法林剂量有统计学差异（P=0．041）,且随着年龄的增长,华法林剂量有降低趋势;CYP2C9＊3基因型CA（杂合突变型）华法林日平均剂量（1．93mg）比AA（野生型）日平均剂量（2．96mg）低1mg（P=0．001）;VKORC11173C／T基因型CT（杂合突变型）华法林日平均剂量（3．34mg）比TT（野生型）日平均剂量（2．77mg）高0．57mg（P=0．002）;CYP4F2V433M基因型TT（纯毹铡）糊日平均剂量（3．52mg）比CT（杂譬洌）高0．68mg（P=0．021）,比CC（野生型）高0．72mg（P=0．026）。结论华法林剂量受CYP2C9、CYP4F2、VKORCl基因多态性和年龄的影响,临床用药前进行CYP2C9、CYP4F2、VKORC1基因多态性检测有助于华法林抗凝治疗用药剂量的评钻。     

Estimation of warfarin maintenance dose based on VKORC1 (-1639 G>A) and CYP2C9 genotypes

BACKGROUND: CYP2C9 polymorphisms are associated with decreased S-warfarin clearance and lower maintenance dosage. Decreased expression of VKORC1 resulting from the -1639G>A substitution has also been implicated in lower warfarin dose requirements. We investigated the additional contribution of this polymorphism to the variance in warfarin dose. METHODS: Sixty-five patients with stable anticoagulation were genotyped for CYP2C9 and VKORC1 with Tag-It allele-specific primer extension technology. Plasma S-warfarin concentrations and warfarin maintenance dose were compared among patients on the basis of the VKORC1 -1639G>A genotype. RESULTS: Eighty percent of CYP2C9*1/*1 patients stabilized on <4.0 mg/day warfarin had at least 1 VKORC1 -1639A allele. Mean warfarin doses (SD) were 6.7 (3.3), 4.3 (2.2), and 2.7 (1.2) mg/day for patients with the VKORC1 -1639GG, GA, and AA genotypes, respectively. Steady-state plasma concentrations of S-warfarin were lowest in patients with the VKORC1 -1639AA genotype and demonstrated a positive association with the VKORC1 -1639G allele copy number (trend P = 0.012). A model including VKORC1 and CYP2C9 genotypes, age, sex, and body weight accounted for 61% of the variance in warfarin daily maintenance dose. CONCLUSIONS: The VKORC1 -1639A allele accounts for low dosage requirements of most patients without a CYP2C9 variant. Higher plasma S-warfarin concentrations corresponding to increased warfarin maintenance dosages support a hypothesis for increased expression of the VKORC1 -1639G allele. VKORC1 and CYP2C9 genotypes, age, sex, and body weight account for the majority of variance in warfarin dose among our study population.     

Pharmacogenetics of acenocoumarol in patients with extreme dose requirements

Summary. Background: There is currently intense debate as to whether pharmacogenetic algorithms for estimating the initial dose of coumarins provide a more accurate dose than the fixed‐dose approach. Recently, it has been suggested that the greatest benefit of pharmacogenetic algorithms is observed in patients with extreme dose requirements. Objectives: To identify clinical and genetic factors that better characterize patients who need extreme acenocoumarol doses for steady anticoagulation state. Patients/methods: We reviewed 9538 patients with a steady acenocoumarol dose from three Spanish hospitals, selecting 83 who took ≤ 5.00 mg week^?1 (percentile 5, p5) and 203 taking ≥ 30.00 mg week^?1 (p95). We also selected patients matched by gender and age taking 13.50–14.00 mg week^?1 (p50). We genotyped VKORC1 (rs9923231), CALU (rs1043550), GGCX (rs699664), CYP2C9 (rs1799853; rs1057910), CYP4F2 (rs2108622) and F7 (rs5742910) single‐nucleotide polymorphisms (SNPs). Results: Comparison between p5 and p95 revealed five parameters with significant differences: body surface area (BSA) (P = 0.006), age, VKORC1, CYP2C9 and CYP4F2 genotypes (all P < 0.001). First VKORC1, and second, CYP2C9 SNPs played a strong effect by determining extreme doses, particularly in p95. Only one out of 203 p95 had the VKORC1 A‐1639A genotype, but this subject was CYP2C9*1/*1. In contrast, nine out of 83 p5 carried the VKORC1 G‐1639G genotype, although six of them were CYP2C9*3 homozygotes and another two were heterozygotes. Surprisingly, CYP4F2 V433M SNP displayed prevalences that suggest that its influence might only be evident when patients are treated with high doses. Conclusion: Two clinical data, age and BSA, and three SNPs in the VKORC1, CYP2C9 and CYP4F2 genes strongly predict outlier patients treated with acenocoumarol.     

A genotyping method for VKORC1 1173C>T by Pyrosequencing® technology

Vitamin K epoxide reductase complex subunit 1 (VKORC1) is the site of inhibition by warfarin and other anti‐vitamin K drugs during oral anticoagulant therapy. The SNP rs9934438 in intron 1 of VKORC1 (c.173+1000C>T or 1173C>T) discriminating the VKORC1*2 haplotype is associated with low warfarin dose requirement and unstable prothrombin time – international normalized ratio. To genotype this SNP, we have developed a rapid method using Pyrosequencing^® technology. The proposed method takes a post‐PCR sample preparation of less than 1?h and a DNA sequencing time of less than 15?min to genotype 96 samples. The current method was compared with a dHPLC method that we reported previously. Genotype frequencies at VKORC1 1173C>T for our Swedish population were 38?% wild‐type, 40?% heterozygote and 22?% homozygote. The frequency of the T‐allele was 0.42, which exactly matches the frequency previously reported for Germans. The current method can be used to determine whether patients initiating warfarin therapy are carriers of SNP 1173 C>T that is strongly associated with low warfarin dose requirement.     

Comparative performance of gene‐based warfarin dosing algorithms in a multiethnic population

Summary. Background: Gene‐based warfarin dosing algorithms have largely been developed in homogeneous populations, and their generalizability has not been established. Objectives: We sought to assess the performance of published algorithms in a racially diverse and multiethnic sample, and determine if additional clinical variables or genetic variants associated with dose could enhance algorithm performance. Patients and methods: In 145 compliant patients on warfarin with a goal international normalized ratio (INR) of 2–3, stable, therapeutic doses were compared with predicted doses using 12 reported algorithms that incorporated CYP2C9 and VKORC1 variants. Additional covariates tested with each model included race, concurrent medications, medications known to interact with warfarin and previously described CYP4F2, CALU and GGCX variants. Results: The mean patient age was 67 ± 14 years; 90 (62%) were male. Eighty‐two (57%) were Caucasian, 28 (19%) African‐American, 20 (14%) Hispanic and 15 (10%) Asian. The median warfarin dose was 35 mg per week (interquartile range 23–53 mg per week). Gene‐based dosing algorithms explained 37–55% of the variation in warfarin dose requirements. Neither the addition of race, number of concurrent medications nor the number of concurrent medications interacting with warfarin enhanced algorithm performance. Similarly, consideration of CYP4F2, CALU or GGCX variant genotypes did not improve algorithms. Conclusions: Existing gene‐based dosing algorithms explained between approximately one‐third and one‐half of the variability in warfarin dose requirements in this racially and ethnically diverse cohort. Additional clinical and recently described genetic variants associated with warfarin dose did not enhance prediction in our patient population.     

VKORC1 gene variations are the major contributors of variation in warfarin dose in Japanese patients

OBJECTIVES: To compare the genetic and clinical factors that cause large interpatient variability and ethnic differences in warfarin efficacy, we investigated variations of the VKORC1, CYP2C9, and CYP2C19 genes in Japanese subjects. Furthermore, we evaluated the genetic variations and clinical data as contributors of variation in warfarin maintenance dose. METHODS: Gene variations of VKORC1, CYP2C9, and CYP2C19 in 125 patients treated with warfarin and 114 healthy subjects were analyzed. The daily dose of warfarin, concentrations of S- and R-warfarin in plasma, and prothrombin time expressed as the international normalized ratio were used as the pharmacokinetic and pharmacodynamic indices. Data were evaluated by a multivariate analysis method. RESULTS: Three missense mutations (47 G>C, 113 A>C, and 1338 A>G) in VKORC1 were newly identified in the Japanese population. The 113 A>C (Asp38Ser) variant decreased the warfarin dose requirement from 3.33 +/- 1.54 mg/d (n = 122) to 1.5 mg/d (n = 1). The variants -1639 G>A in the 5'-upstream region, 1173 C>T in intron 1, and 1542 G>C in intron 2 were in complete linkage disequilibrium, and the frequency of the -1639 G>A variation was only 0.8%, which contrasts with the frequency (39.8%-45.8%) reported previously for white persons. The dose of warfarin was larger in the VKORC1 -1639 GA genotype group (4.55 +/- 1.75 mg/d, P < .001) than in the -1639 AA group (2.94 +/- 1.15 mg/d). The mean daily dose of warfarin was lower in subjects with CYP2C9*1/*3 (1.86 +/- 0.80 mg/d, P = .007) than in subjects with CYP2C9*1/*1 (3.36 +/- 1.43 mg/d). When the relative contributions of the VKORC1 variants, CYP2C9*2, CYP2C9*3, CYP2C19*2, and CYP2C19*3, as well as the clinical characteristics of the patients, diagnoses, and concurrent medications, were compared, the VKORC1 -1639 GA genotype group accounted for 16.5% and CYP2C9 variants accounted for 13.4% of variation in warfarin dose. CONCLUSION: The ethnic difference in warfarin maintenance dose was mainly dependent on the linked VKORC1 variants. Genotyping of -1639 G>A of the VKORC1 gene could be clinically important for predicting individual variability in anticoagulant responses to warfarin.     

A warfarin-dosing model in Asians that uses single-nucleotide polymorphisms in vitamin K epoxide reductase complex and cytochrome P450 2C9

INTRODUCTION: Because of the unique lack of genetic diversity despite the multiethnicity in the Asian population, we hypothesize that single-nucleotide polymorphisms in cytochrome P450 (CYP) 2C9 (CYP2C9*3) and vitamin K epoxide reductase complex subunit 1 (VKORC1) at position 381, used to infer VKORC1haplotype in combination with demographic factors, can accurately predict warfarin doses. The aims of this study were to derive a pharmacogenetics-based dosing algorithm by use of retrospective information and to validate it through a data-splitting method in a separate cohort of equal size. METHODS: We used 215 records of warfarin patients recruited into a CYP2C9/VKORC1 genotyping study to perform this analysis. Univariate analyses for individual predictors, including age, weight, gender, serum albumin concentration, ethnic group, international normalized ratio, and CYP2C9 and VKORC1 381 genotypes, were conducted to select variables with P < .1 for further inclusion into the multivariate regression analysis. In the final model only predictors reaching a statistical significance of P < .05 were retained. RESULTS: Data from 107 subjects undergoing maintenance warfarin therapy with an international normalized ratio stabilized between 2 and 3 were used to derive the final model, as an exponential function of age, weight, CYP2C9*3 allele, and VKORC1 381 CC and TC genotypes, and this model accounted for 60.2% of the variability in daily warfarin dose requirement. The model was validated in a separate cohort of 108 subjects and showed a mean underestimation of 0.23 +/- 1.21 mg/d. CONCLUSION: Warfarin dose requirements in Asians can be accurately predicted by use of a combination of patient demographics and a simplified genotyping approach for single variants in CYP2C9 and VKORC1.     

Drug interaction between St John's wort and zolpidem in healthy subjects

What is known and objective: St John’s wort (SJW, Hypericum perforatum) is one of the most commonly used herbal antidepressants for treatment of mild to moderate depression. SJW enhances CYP3A4 activity and alters the pharmacokinetics of CYP3A4 substrates. This study investigated the effect of SJW on the pharmacokinetics of zolpidem in healthy subjects. Methods: A controlled, open‐label, non‐randomized, fixed‐dose schedule design was used. Fourteen healthy male subjects received a single 10 mg oral dose of zolpidem followed by SJW administration (300 mg orally, three times a day) for 14 days; the last dose of SJW was coadministered with a single dose of zolpidem. Blood samples were obtained over a 24‐h period after zolpidem administration. Pharmacokinetic data for zolpidem alone and in combination with SJW were analysed by high‐performance liquid chromatography. Results: After repeated administration of SJW, the mean values of AUC and C_max for zolpidem significantly decreased (380·3 ± 181·4 vs. 265·4 ± 134·2 ng h/mL, P = 0·001; 83·1 ± 30·1 vs. 55·1 ± 24·8 ng/mL, P = 0·000 respectively) and the mean value of CL/F for zolpidem significantly increased (38·4 ± 31·5 vs. 56·9 ± 57·2 mL/min, P = 0·040). However, in three subjects, the AUC showed a small increase after SJW treatment. What is new and conclusion: The effect of SJW on the pharmacokinetics of zolpidem has not previously been reported. Repeated administration of SJW decreases the plasma concentration of zolpidem, probably by enhancing CYP3A4 activity. Given the wide inter‐subject variability observed, for personalized medicine, advice on the use of the combination should be individualized, based on the circumstances of the patient.     

Differences in genotype and allele frequency distributions of polymorphic drug metabolizing enzymes CYP2C19 and CYP2D6 in mainland Chinese Mongolian, Hui and Han populations

What is known and Objective: Cytochrome P450 2C19 (CYP2C19) and CYP2D6 are important xenobiotic metabolic enzymes and both show considerable genetic variability between Orientals and Caucasians. There are known marked heterogeneity in susceptibility to various cancers and hypertension among Chinese Mongolian, Hui and Han ethnic groups, but the molecular mechanisms are unknown. Our objective was to investigate the patterns of distribution of CYP2C19 and CYP2D6 polymorphisms among healthy Chinese subjects to determine whether any observed inter‐ethnic variability might be worth further investigation as possible contributors to the known differences in disease prevalence. Methods: Blood samples were collected from 454 unrelated Chinese healthy subjects (214 Han, 111 Hui, 129 Mongolian) for genotyping analysis. The single nucleotide polymorphisms (SNPs) CYP2C19*2 (681G>A in exon 5), CYP2C19*3 (636G>A in exon 4) and CYP2D6*10 (188C>T in exon 1) were determined by the polymerase chain reaction–restriction fragment length polymorphism (PCR‐RFLP) method. Results and Discussion: Significantly higher frequencies of the CYP2C19 poor metabolic genotypes were observed in Chinese Han (18·7%), Chinese Hui (25·0%) and Chinese Mongolian (10·9%) subjects than has been reported for Caucasians (1·7–3·0%, P < 0·01). The prevalent defective allele CYP2C19*2 occurred more frequently in both Chinese Hui (32·4%) and Han (29·7%) than in Chinese Mongolian (18·2%, P < 0·01) subjects. The CYP2C19*2 and CYP2C19*3 defective alleles were significantly more frequent in Chinese Han and Chinese Hui ethnic groups than have been reported for Caucasians (11·1–16·3% and 0–0·2%, P < 0·01). CYP2D6*1/*10 heterozygotes and CYP2D6*10/*10 homozygotes were observed more frequently in Chinese Han (43·1% and 27·2%), Hui (40·6% and 30·7%) and Mongolian subjects (31·3% and 9·6%, both P < 0·01) than have been reported for Caucasians (5·5% and 0·3%, P < 0·01). In Chinese Mongolians, the CYP2D6*10 allele occurred at a frequency (25·2%, P < 0·01) intermediate between those reported for Caucasians and the other two Chinese ethnic populations. What is new and Conclusions: This is first report of interethnic differences in frequencies of functional CYP2C19 and CYP2D6 genes among Chinese Mongolian, Hui and Han populations. These differences may be important in explaining reported inter‐ethnic differences in disease prevalence and response to drugs.     

An evaluation of gene–gene interaction between the CYP2C9 and VKORC1 genotypes affecting the anticoagulant effect of phenprocoumon and acenocoumarol

Background:  Previous studies have provided contradictory results regarding the interaction between the CYP2C9 and VKORC1 genotypes affecting various outcome measures. Objectives:  We aimed to provide a definite answer regarding the question whether there exists a gene–gene interaction between the CYP2C9 and VKORC1 genotypes affecting the anticoagulant effect of phenprocoumon and acenocoumarol. Patients/Methods:  The EU‐PACT cohort dataset, which contains data on 624 phenprocoumon and 471 acenocoumarol patients, was used. Patient characteristics, pharmacogenetic data, International Normalized Ratios (INRs) and dosages were available. We investigated whether there was an interaction between the CYP2C9 and VKORC1 genotypes affecting the maintenance dose, time to severe over‐anticoagulation and time to achieve stability during the first 180 days of phenprocoumon and acenocoumarol therapy, in addition to the effect of the separate genotypes. The interaction effect was investigated by adding the product term of the CYP2C9 and VKORC1 genotype classes for four different commonly used CYP2C9 classifications to the linear regression model – for the outcome measure maintenance dose – or to the Cox regression models – for the outcome measures time to severe over‐anticoagulation and time to achieve stability. Results:  No significant interactions – all P‐values above 0.23 for phenprocoumon and 0.30 for acenocoumarol – were observed for all outcome measures. Conclusions:  There are no interactions between the CYP2C9 and VKORC1 genotypes affecting the maintenance dose, time to severe over‐anticoagulation and time to achieve stability for phenprocoumon and acenocoumarol.     

Interethnic variability of warfarin maintenance requirement is explained by VKORC1 genotype in an Asian population

BACKGROUND: Chinese and Malay subjects have been reported to require less maintenance warfarin than Indians that could not be accounted for by cytochrome P450 (CYP) 2C9 variants. Vitamin K epoxide reductase complex 1 (VKORC1) is the target enzyme of warfarin, and VKORC1 intronic variants and haplotypes have recently been shown to influence VKORC1 activity and warfarin requirements. METHODS: We sequenced the coding regions of CYP2C9 and VKORC1 and inferred VKORC1 haplotype from 10 intronic variants in 147 Chinese, 85 Malay, and 43 Indian patients receiving maintenance warfarin. RESULTS: The mean weight-normalized warfarin dose was lower for Chinese and Malays than for Indians (0.058 +/- 0.025 mg/kg, 0.059 +/- 0.023 mg/kg, and 0.089 +/- 0.036 mg/kg, respectively; P < .001 for comparisons between Chinese and Malays with Indians). CYP2C9*2 and VKORC1 coding region variants were rare (<2%), whereas CYP2C9*3 associated with lower warfarin requirements was less common in Chinese and Malays (7% and 9%, respectively) than in Indians (18%) and could not account for their lower warfarin requirements. VKORC1 H1 and H7/H8/H9 haplotypes were associated with lower and higher warfarin requirements, respectively (0.050 +/- 0.019 mg/kg and 0.092 +/- 0.057 mg/kg, respectively; P < .001). VKORC1 H1 haplotype (requiring low warfarin doses) was common in Chinese (87%) and Malays (65%) but uncommon in Indians (12%), whereas H7, H8, and H9 haplotypes (requiring high warfarin doses) were rare in Chinese (9%), intermediate in Malays (30%), and common in Indians (82%). The interethnic difference in warfarin requirements became nonsignificant when adjusted for VKORC1 haplotype. CONCLUSIONS: Interethnic difference in VKORC1 haplotypes accounts for the difference in warfarin requirements between Chinese, Malays, and Indians, providing interesting insights into genetic variation between ethnogeographically distinct Asian groups.     

Pharmacodynamic resistance to warfarin is associated with nucleotide substitutions in VKORC1

Summary. Background: Vitamin K epoxide reductase subunit 1 (VKORC1) is the molecular target of coumarin anticoagulants and mutations in VKORC1 have been identified previously in individuals who required high warfarin doses. Objective: Detailed characterization of the relationship between variation in VKORC1 and the warfarin resistance phenotype. Patients and methods: Serum warfarin concentration and coagulation parameters were determined in 289 subjects who required warfarin doses >20 mg day^?1. The VKORC1 sequence was studied in selected study subjects. Results: Twenty‐eight out of 289 (10%) subjects had serum warfarin >2.3 mg L^?1 during stable therapeutic anticoagulation indicating pharmacodynamic warfarin resistance. Detailed analysis of 15 subjects from this group showed that eight out of 15 (53%) had nucleotide substitutions in VKORC1 predictive of p.V66M, p.L128R, p.V54L or p.D36Y. VKORC1 was normal in the remaining seven out of 15 (47%) subjects and in nine out of nine (100%) subjects with high warfarin dose requirement not caused by pharmacodynamic resistance. At referral, subjects with VKORC1 mutations received a median warfarin dose of 32 mg day^?1 (range 22–55) and had a median serum warfarin concentration of 4.6 mg L^?1 (range 2.6–9.0). VKORC1 substitutions were associated with a requirement for high warfarin doses but not with adverse clinical events. Family members with VKORC1 nucleotide substitutions and not receiving warfarin had undetectable PIVKA‐II and K₁ epoxide (K₁O). Conclusions: Nucleotide variations in VKORC1 are a common cause of pharmacodynamic warfarin resistance but are not associated with adverse outcome during anticoagulation. Mutations associated with warfarin resistance do not cause a discernible defect in VKORC1 reductase function.     

Standard warfarin dose in a patient with the CYP2C9*3/*3 genotype leads to hematuria

BackgroundPatients with certain CYP2C9 genetic variants have increased sensitivity to warfarin and are at increased risk of over-coagulation with standard warfarin dose. We report over-anticoagulation and hematuria manifest as a slow increase in the international normalized ratio (INR) due to warfarin treatment in a patient with the CYP2C9*3/*3 allele.CaseA 58-y-old man with paroxysmal atrial fibrillation received a standard warfarin dose of 2.0 mg/day. Because INR was 2.00 one week after treatment initiation, he was discharged from the hospital. One month later, hematuria was present and INR had increased to 7.26. Although in normal cases (R)-warfarin plasma concentrations are higher than (S)-warfarin, this patient had the opposite warfarin enantiomer plasma concentration profile.ConclusionsIncreased anticoagulation was due to an increased concentration of (S)-warfarin, the more active warfarin enantiomer. INR response to warfarin in this CYP2C9*3/*3 patient was slow. The later INR response appears to be strongly affected by CYP2C9 variants. He also had the VKORC1 ?1639G > A AA genotype, requiring a lower warfarin dose. In this case, increased risk of bleeding could have been identified by prospective genotyping of CYP2C9 and VKORC1 prior to initiating warfarin therapy.     

Absence of novel CYP4F2 and VKORC1 coding region DNA variants in patients requiring high warfarin doses

ObjECTIVE: Warfarin is an FDA-approved oral anticoagulant for long-term prevention of thromboembolism. Substantial inter-individual variation in dosing requirements and the narrow therapeutic index of this widely-prescribed drug make safe initiation and dose stabilization challenging. Single nucleotide polymorphisms (SNPs) occurring in CYP2C9, VKORC1, and CYP4F2 genes are known to impact dose, and VKORC1 and CYP4F2 polymorphisms are associated with higher therapeutic dose requirements in our cohort. However, the most advanced regression models using personal, clinical, and genetic factors to predict individual stable dose account for only 50% to 60% of the observed variability in stable therapeutic dose in Caucasians. DESIGN AND METHODS: In this study, we used DNA sequence analysis to determine whether additional variants in CYP4F2 and VKORC1 gene coding regions contribute to variable dosing requirements among individuals for whom the actual dose was the highest relative to regression model- predicted dose. RESULTS AND CONCLUSIONS: No novel DNA variants in the coding regions of these genes were identified among subjects requiring high warfarin doses, suggesting that other factors yet to be defined contribute to variability in warfarin dose requirements in this subset of our cohort.     

The Effect of Compound Danshen Dripping Pills on the Dose and Concentration of Warfarin in Patients with Various Genetic Polymorphisms

PurposeThe combination of warfarin and compound Danshen dripping pill (CDDP) is helpful for patients with both coronary heart disease (CHD) and atrial fibrillation (AF). The main adverse drug reaction of warfarin is bleeding because of its narrow therapeutic index. The safety of a combination therapy with warfarin and CDDP is always a concern. Our previous research showed that the combination of warfarin and CDDP improved the quality of life for patients with both CHD and AF. This study describes the changes in dose and concentration of warfarin necessary and evaluates bleeding risk when warfarin is given concomitantly with CDDP.MethodsAn ultra-performance liquid chromatography–MS/MS method with a chiral column was developed to assay the concentration of S-warfarin and R-warfarin in human plasma simultaneously. The method was applied to compare the concentration of warfarin in patients taking warfarin combined with CDDP and without CDDP. International normalized ratio (INR) values were monitored to evaluate bleeding risk. Paired t tests were then used to compare the dose and the concentration in 2 periods. Moreover, patients with VKORC1, CYP2C9*3, CYP4F2, EPHX1, and PROC gene polymorphisms were evaluated to determine interactions.FindingsThe results indicate that the dose of warfarin had no significant change with or without CDDP. Also, the peak concentrations of S-warfarin and total warfarin were significantly different in CYP4F2 C/C patients, but there was no significant difference identified in other genetic groups. No bleeding occurred in the study.ImplicationsThe dose of warfarin would be sustainable when combined with CDDP, because CDDP did not affect concentration of warfarin significantly in most patients and the change of INR was not significant.China Clinical Trial Registry identifierChiCTR-ONRC-13003523.     

中国汉族人群CYP2C9及VKORC1基因多态性及其对华法林用药剂量的相关性研究

目的分析中国汉族人群CYP2C9和VKORC1的基因多态性及其与华法林稳定维持剂量的相关性。方法回顾性研究。收集2017年10月至2018年4月在北京大学人民医院进行凝血分析检测的458例中国汉族患者,男性213例,女性245例,年龄范围26~94岁。采用PCR-荧光探针法检测CYP2C9*3位点和VKORC1-1639A>G位点基因多态性,458例患者中服用华法林进行抗凝治疗且凝血酶原时间国际标准化比值(INR)达标(在2.0~3.0范围内)的患者130例,记录患者基本信息、华法林用药剂量及凝血酶原时间国际标准化比值(INR),应用SPSS统计分析数据,与美国FDA推荐的不同基因型患者华法林推荐剂量的参考表格进行对比,并且对华法林预测剂量公式进行简单验证。结果458例抗凝患者中CYP2C9*1/*1(AA)基因型频率90.8%,CYP2C9*1/*3(AC)基因型频率8.5%,CYP2C9*3/*3(CC)基因型频率0.7%;VKORC1-1639GG基因型频率0.9%,VKORC1-1639AG基因型频率14.2%;VKORC1-1639AA基因型频率84.9%。在达到抗凝指标(国际标准化比值INR 2.0~3.0)后,结果显示CYP2C9*1/*3与*3/*3基因型患者平均每日剂量为(2.92±1.29);3(2.75,3.375)mg,低于野生型CYP2C9*1/*1基因型患者所需的平均每日华法林剂量(3.91±1.63);3(3,5)mg,差异有统计学意义(P=0.018)。VKORC1纯合突变基因型VKORC1-AA患者平均每日剂量为(3.68±1.64);3(3,4.31)mg,低于杂合基因突变型的平均每日剂量(4.54±1.29);4.5(3.28,6)mg,差异有统计学意义(P=0.001)。不同VKORC1+CYP2C9基因型患者的华法林应用剂量与美国FDA参考表格的推荐剂量具有一致性。Miao2007公式的预测准确度较IWPC公式低,且94.1%的患者华法林剂量被低估。结论携带CYP2C9*3突变基因或VKORC1-AA纯合突变基因型的患者所需华法林剂量较低,CYP2C9和VKORC1基因多态性与华法林稳定维持剂量具有一定的相关性。