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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. [ABSTRACT FROM AUTHOR]

Abstract: Introduction: Multiple warfarin pharmacogenetic algorithms have been confirmed to predict warfarin dose more accurately than clinical algorithm or the fixed-dose approach. However, their performance has never been objectively evaluated in patients under low intensity warfarin anticoagulation, which is optimal for prevention of thromboembolism in Asian patients. Material and methods: We sought to compare the performances of 8 eligible pharmacogenetic algorithms in a cohort of Chinese patients (n=282) under low intensity warfarin anticoagulation with target international normalized ratio (INR) ranged from 1.6 to 2.5. The performance of each algorithm was evaluated by calculating the percentage of patients whose predicted dose fell within 20% of their actual therapeutic dose (percentage within 20%), and the mean absolute error (MAE) between each predicted dose and actual stable dose. Results: In the entire cohort, the pharmacogenetic algorithms could predict warfarin dose with the average MAE of 0.87±0.17mg/day (0.73-1.17mg/day), and the average percentage within 20% of 43.8%±8.1% (29.1% - 52.1%). By pairwise comparison, warfarin dose prediction was significantly more accurate with the algorithms derived from Asian patients (48.6% – 50.0%) than those from Caucasian patients (29.1% - 39.7%; odds ratio [OR]: 1.61-3.36, p ≤0.02). Algorithms with additional covariates of INR values or CYP4F2*3 performed better than those without the covariates (adding INR: OR: 1.71 (1.08-2.72), p =0.029; adding CYP4F2*3: OR: 2.67(1.41-5.05), p =0.004). When the patients were stratified according to the dose range, the algorithms from Caucasian and racially mixed populations tended to perform better in higher dose group (≥4.5mg/day), and algorithms from Asian populations performed better in intermediate dose group (1.5-4.5mg/day). None of the algorithms performed well in lower dose group (≤1.5mg/day). Conclusions: No eligible pharmacogenetic algorithm could perform the best for all dosing range in the Chinese patients under low intensity warfarin anticoagulation. Construction of a refinement pharmacogenetic algorithm integrating 3 genotypes (CYP2C9, VKORC1 and CYP4F2) and INR data should be warranted to improve the warfarin dose prediction in such patients. [Copyright &y& Elsevier]

What is known and Objective: Polymorphisms in the gene encoding CYP4F2 may partly explain the variability in warfarin maintenance dose by altering the metabolism of vitamin K. To determine the genetic factors that cause large inter-patient variability in warfarin efficacy, we investigated the relationship between serum warfarin concentration and CYP4F2 V433M (1347C>T, rs2108622) polymorphism in Japanese subjects. Methods: Gene variations in VKORC1, CYP2C9 and CYP4F2 were analysed in 126 Japanese patients treated with warfarin. The daily dosage of warfarin, concentration of S- and R-warfarin in plasma, and prothrombin time international normalized ratio (PT-INR) was used as the pharmacokinetic and pharmacodynamic indices. Results and Discussion: The maintenance dose of warfarin was larger in the CYP4F2 1347 CT genotype group (3·59 ± 1·80 mg/day, P = 0·027) than in the CYP4F2 CC genotype group (2·88 ± 1·00 mg/day). CYP4F2 1347C>T polymorphism significantly affected serum R-warfarin concentration when the VKORC1-1639 genotypes are AG and GG. What is new and Conclusion: Although a significant inter-patient difference in warfarin maintenance dose was observed between the CYP4F2 CC and CT genotypes, serum S-warfarin concentration was not significantly different between them. An effect of CYP4F2 V433M polymorphism on warfarin maintenance dose was observed but was relatively small when compared to the effects of CYP2C9 and VKOR polymorphism. [ABSTRACT FROM AUTHOR]

Abstract: Introduction: Warfarin is a commonly used oral anticoagulant and the dosage is individually adjusted on the basis of the international normalized ratio (INR) monitoring. It is well known that gene polymorphisms of CytochromeP450 (CYP) 2C9 gene and the vitamin K epoxide reductase complex 1 (VKORC1) were significantly associated with warfarin dose. However, the association between Cytochrome P450 4F2 (CYP4F2) polymorphism and warfarin dose requirement is still controversial. This study was to investigate the influence of the CYP4F2 polymorphism, V433M (rs2108622) on warfarin dose for patients by meta-analysis. Methods: Strict inclusion and exclusion criteria were set, and the studies prior to December 19, 2010 were searched in PubMed, EMBASE and CNKI. References were examined and experts of primary studies were consulted for additional information. Revman 5.0.2 software was used to analyze the relationship between warfarin maintenance dose and CYP4F2 polymorphism Results: Thirteen studies were included in the meta-analysis which consisted of Caucasian, Asian and African populations. Compared to individuals with the homozygous CYP4F2 genotype (CC), carriers of CT, TT genotypes required 10.0% (95% confidence interval(CI) 4.0-15.0) and 21.0% (95% CI 9.0-33.0) higher warfarin doses respectively (P value <0.05). In addition, T carriers required 11.0% (95% CI 6.0-17.0) higher warfarin dose than CC genotype. Conclusions: Our study showed that polymorphism of CYP4F2 had a moderate but statistically significant association with the variation of interindividual warfarin dose. However, whether CYP4F2 can improve the prediction of warfarin dose warrants need further investigation when combined with environmental factors. [Copyright &y& Elsevier]

To evaluate the impact of gene polymorphisms of Cytochrome P450 2C9 (CYP2C9), Vitamin K epoxide reductase complex subunit 1 (VKORC1) and Cytochrome P450 4F2 (CYP4F2) and clinical factors on warfarin maintenance dose in Han-Chinese patients from main land. DNA was extracted from 115 patients taking warfarin for more than 3 months with a stable international normalized ratio (INR) and genotyped for CYP2C9*3, VKORC1-1639 and CYP4F2 (rs2108622) polymorphisms using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Univariate analysis and multiple regression analysis were undertaken to assess the effect of genetic and clinical factors on the warfarin maintenance dose. Our study demonstrated that patients carrying CYP4F2 CT or TT allele needed a significantly higher warfarin dose compared to those carrying CC ((3.36 ± 0.14 mg/d vs. 2.77 ± 0.14 mg/d), P = 0.004). We also confirmed CYP2C9 *3 variant was related to lower warfarin dose (2.01 ± 0.23 mg/d) requirement compared to wild type (3.21 ± 0.11 mg/d) ( P = 0.001). VKORC1-1639 AG genotype was associated with a higher maintenance dose compared to those with the AA genotype (4.06 ± 0.21 mg/d vs. 2.95 ± 0.11 mg/d, P < 0.001). The multiple linear regression model including VKORC1-1639G>A, CYP2C9, CYP4F2 and clinical factors (body surface area (BSA) and age) could explain 42 % of the variance in the warfarin maintenance dose. We developed a dose algorithm based on genetic polymorphism and clinical variables for Han-Chinese patients and evaluated its performance. CYP4F2 rs2108622 has a small but significant association with warfarin stable dose in Han-Chinese population. [ABSTRACT FROM AUTHOR]

Abstract: Background and aim: The genetic basis of Metabolic syndrome (MetS) is largely unknown but a link with salt sensitivity is recognized. The cytochrome P450 isoform 4F2 (CYP4F2) is involved in renal production of 20-hydroxyeicosatethraenoic acid (20-HETE), a natriuretic substance associated with salt sensitivity. The same enzyme is implicated in ω-hydroxylation of very long and medium chain fatty acids in the liver suggesting its possible influence on gluco-metabolic components of MetS. The aim of the present study was to evaluate the effect of CYP4F2 V433M, a functional polymorphism previously associated with hypertension via renal salt reabsorption, on the individual components of MetS and MetS itself. Methods: The polymorphism was genotyped in the cardiovascular cohort of the Malmö Diet and Cancer (MDC-CVA) study and successively in the Malmö Preventive Project (MPP) cohort. Different definitions of the MetS were applied. Results: In the MDC-CVA, male, but not female, CYP4F2 M433 carriers had significantly higher levels of waist, triglycerides, BP and a composite sum of MetS phenotypes (MetS score) beside lower HDL-cholesterol respect to V-homozygotes. MetS, as defined in the ATPIII and the AHA/NHLBI definitions, was more prevalent in M-carriers with respect to V-homozygotes. In the MPP cohort, significant association was detectable only for triglycerides at baseline and for Diastolic BP at reinvestigation in male M-carriers. Conclusion: The initial positive association of the CYP4F2 V433M polymorphism with components of MetS and MetS itself, found in MDC-CVA, was partially denied in another large cohort. The first association either could be due to a false positive result or alternatively, different genetic background or population stratification could have hidden the effect of the polymorphism in the replication cohort. [Copyright &y& Elsevier]

Abstract: Background: The cytochrome P450 (CYP) 4F2 isozyme has been reported to metabolize vitamin K1 in vitro, and the V433M polymorphism in the CYP4F2 gene has been associated with reduced vitamin K1 metabolism and the need for a higher maintenance dosage in patients receiving warfarin. Objective: The purpose of the present study was to evaluate the effects of V433M polymorphism on warfarin response during the induction phase. Methods: Warfarin-naive white patients in whom warfarin was scheduled to be initiated with a target INR of 2 to 3 were enrolled into the study. On enrollment, a single blood sample for the genotyping of CYP4F2, CYP2C9, and VKORC1 was drawn. The international normalized ratio (INR) was followed daily during induction and twice weekly until stable anticoagulation was reached. The relationships between several markers of warfarin response during induction and CYP4F2 polymorphism were determined. Results: The cohort consisted of 241 patients (115 men; mean [SD] age, 55.2 [19.4] years; weight, 79.5 [18.3] kg). Most of the patients were carriers of the CYP4F2 CC genotype (112 patients) or the CT genotype (104 patients). In carriers of the TT genotype (25 patients), INR >3 was >4-fold lower compared with that in carriers of the CC or CT genotype, suggesting that patients with the TT genotype were less sensitive to warfarin during induction. Also in TT carriers, the extent of excessive anticoagulation was >10-fold lower than in the other carriers. Both of these findings had a nominal P value of <0.05. After adjustment for false discovery rate, none of the findings remained significant at a threshold q value of <0.05. Among CC carriers, the concurrent use of a statin was associated with a 1-mg/d reduction in warfarin maintenance dosage. No similar effect was noted in the CT or TT carriers, suggesting a possible genetic influence on warfarin-statin interaction. Conclusions: These preliminary findings suggest that among white patients treated with warfarin, CYP4F2 polymorphism had a measurable effect on warfarin responsiveness during induction; however, the observed differences failed to reach the level of statistical significance. The possibility that the effect of statins on warfarin anticoagulation varies among carriers of different CYP4F2 genotypes could not be excluded and should be evaluated further in a larger patient sample. [Copyright &y& Elsevier]

This is the first study to evaluate the spectrum and prevalence of dose-predictive genetic polymorphisms of the CYP2C9, CYP4F2 and VKORC1 loci together, in a geographically defined, ethnically admixed healthy adult Omani population sharing common lifestyle/environmental factors. Since the present-day Omani population is the result of an admixture of Caucasian, African and Asian ancestries, we compared the pharmacogenetic profile of these three loci in this population. Interestingly, the Omani pharmacogenetic profile, in terms of allele and genotype distribution, has values that are intermediate between Caucasians and African Americans, the African admixture further substantiated by the presence of the CYP2C9*8 allele. However, limitations and usefulness of such comparisons warrant caution, as the data from pharmacogenetic literature do not always represent bona fide population categories. Furthermore, definition of study population based on microgeographical scale would be more appropriate in pharmacogenetic research rather than the flawed racial, ethnic, or social categorizations since pharmacogenetic variation is clinal, and genetic influences will be further altered by lifestyle and environmental factors. [ABSTRACT FROM AUTHOR]

Background: Individuals with implanted mechanical valve prostheses require lifelong anticoagulation therapy with warfarin. The narrow therapeutic index of warfarin makes it difficult to dose and maintain proper anticoagulation. A number of single nucleotide polymorphisms (SNPs) affecting vitamin K or warfarin metabolism have been shown to affect warfar in dosing. Our aim was to study the effect of the CYP4F2 rs2108622-1347 (C > T) variant on warfarin dosing in Chinese patients. Methods: We studied 352 patients after heart valve replacement surgery. Warfarin dosing for patients was adjusted to achieve 1.8 ? INR ? 2.5. We determined the presence of SNPs in CYP4F2 in these patients and investigated their association with warfarin dosing. Results: We found the frequency of the CYP4F2 rs2108622 C allele was 79.5% and T-allele frequency was 20.5%. The warfarin dose requirement for CC individuals was significantly lower than that for CT or TT individuals (P < 0.05). TT-homozygous individuals required a 0.56 mg/day higher dose of warfarin than their CC counterparts. Conclusions: This study demonstrates that CYP4F2 rs2108622 significantly affects the warfarin dose requirement to achieve adequate anticoagulant activity in Chinese individuals. Genotyping of this SNP may allow clinicians to determine the initiation dose for patients following valve-replacement surgery in China. [ABSTRACT FROM AUTHOR]

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 theapeutic 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. [ABSTRACT FROM AUTHOR]

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT • Warfarin is an oral anticoagulant widely used for thromboembolic disease prophylaxis. The pharmacokinetics and pharmacodynamics of this drug vary with environmental and genetic factors. Several algorithms including genetic and non-genetic factors have been developed for different populations. These investigations have shown that polymorphisms in CYP2C9 and VKORC1 genes are related to warfarin dose variation. WHAT THIS STUDY ADDS • This study confirms that CYP2C9 and VKORC1 genes are the main predictors of warfarin dose, but there are other genes with smaller effects that can improve the algorithm for better dose prediction. • This study observed for the first time an effect of the F2 gene on warfarin dose prediction. • The inclusion for the first time of co-medication with amlodipine, carbamazepine, β-adrenoceptor blockers and diuretics refined the construction of an algorithm for dose prediction. AIMS To investigate the influence of polymorphisms in CYP2C9, VKORC1, CYP4F2 and F2 genes on warfarin dose-response and develop a model including genetic and non-genetic factors for warfarin dose prediction needed for each patient. METHODS A total of 279 patients of European ancestry on warfarin medication were investigated. Genotypes for −1639G>A, 1173C>T, and 3730G>A SNPs in the VKORC1 gene, CYP2C9*2 and CYP2C9*3, 1347C>T in the CYP4F2 gene and 494C>T in the F2 gene were determined by allelic discrimination with Taqman 5'-nuclease assays. RESULTS The CYP2C9*2 and CYP2C9*3 polymorphisms in the CYP2C9 gene, −1639G>A and 1173C>T in the VKORC1 gene and 494C>T in the F2 gene are responsible for lower anticoagulant doses. In contrast, 1347C>T in the CYP4F2 gene and 3730G>A in the VKORC1 gene are responsible for higher doses of warfarin. An algorithm including genetic, biological and pharmacological factors that explains 63.3% of warfarin dose variation was developed. CONCLUSION The model suggested has one of the highest coefficients of determination among those described in the literature. [ABSTRACT FROM AUTHOR]

Warfarin is a commonly used oral anticoagulant with a narrow therapeutic index and various genetic and clinical factors that influence interpatient variability in dose requirements. This study investigated the impact of genetic and nongenetic factors on warfarin dose requirements in Egyptians.DNA was extracted from 207 patients taking warfarin for more than 2 months and genotyped for VKORC1 (3673 G>A), CYP2C9 ∗2∗3∗4∗5∗8, CYP4F2 (V33M; rs2108622), APOE (rs429358, rs7412), and CALU(rs339097) gene polymorphisms. Linear regression modeling was conducted to identify the genetic and nongenetic factors that independently influence warfarin dose requirements.VKORC1 3673 AA or GA genotype (P<0.0001), one or two variant alleles of CYP2C9 gene (P=0.0004), APOE ε2 haplotype (P=0.01), and increasing age (P<0.0001) were all associated with lower warfarin dose, whereas smoking (P=0.025) and pulmonary embolism (P=0.0059) showed association with higher warfarin doses. These factors explained 31% of the warfarin dose variability. This is the first independent confirmation of the association of the CALU rs339097 variant with higher warfarin dose requirement, although inclusion of thissinglenucleotide polymorphism in the multiple regressionmodel failed to achieve significance (P=0.066). CYP4F2 (V33M) polymorphism was not significant (P=0.314), despite itshigh frequency in the studied population (42%).The study shows that VKORC1, CYP2C9 polymorphisms, APOE ε2 variant, and several clinical/demographic variables are important determinants of warfarin dose requirements in Egyptian patients. The percentage of variability explained by these factors is lowerthan in those of European ancestry, but similar tothevariability explained in Asians and African ancestry. [ABSTRACT FROM AUTHOR]

Genome-wide association studies (GWAS) on warfarin and acenocoumarol showed that interindividual dosage variation is mainly associated with single nucleotide polymorphisms (SNPs) in VKORC1 and to a lesser extent in CYP2C9 and CYP4F2. For phenprocoumon dosage, the genes encoding CYP3A4 and ApoE might play a role.To assess the association between common genetic variants within VKORC1, CYP2C9, CYP4F2, CYP3A4, and ApoE and phenprocoumon maintenance dosage, and to identify novel signals using GWAS.We selected all participants from the Rotterdam study who were treated with phenprocoumon. For each SNP, we tested the association between the above-mentioned genotypes and age, sex, body mass index, and target INR adjusted-phenprocoumon maintenance dosage.Within our study population (N=244), VKORC1, CYP2C9, CYP4F2 genotypes together explained 46% of phenprocoumon maintenance dosage variation. Each additional VKORC1 variant allele reduced phenprocoumon maintenance dosage by 4.8 mg/week (P<0.0001) and each additional CYP2C9 variant allele by 2.2 mg/week (P=0.002). Each additional variant allele of CYP4F2 increased phenprocoumon dosage by 1.5 mg/week (P=0.022). Variant alleles of CYP3A41∗B and ApoE showed no association with phenprocoumon dosage. Genome-wide significant SNPs were all related to VKORC1 activity. Best associated were two SNPs in complete linkage disequilibrium with each other and with SNPs within VKORC1: rs10871454 [Syntaxin 4A (STX4A)] and rs11150604 (ZNF646), each with a P value of 2.1×10-22. Each reduced phenprocoumon maintenance dosage weekly by 4.9 mg per variant allele.Similar to earlier findings with warfarin and acenocoumarol, phenprocoumon maintenance dosage depended on polymorphisms in the VKORC1 gene. CYP2C9 and CYP4F2 were of modest relevance. [ABSTRACT FROM AUTHOR]

Warfarin anticoagulation is complicated by the highly variable inter-individual response. Approximately 50% of the dose variability arises from clinical factors and variants in two genes, CYP2C9 (*2 and *3 variants) and VKORC1 -1173 C > T. We tested variants in five additional genes ( EPHX1, PROC, APOE, CYP4F2, CALU and a new variant in VKORC1 in an attempt to further reduce the variability in predicted stable warfarin dose. Consecutive consenting outpatients requiring anticoagulation on stable warfarin dose (target INR 2-3) were genotyped; the association of SNP genotypes with stable warfarin dose was evaluated using the test of linear contrasts in analysis of variance (ANOVA). Study participants were 71 ± 13 years, 53% female, 85 ± 23 kg, body mass index 29 ± 7 kg/m. Genotypes were in Hardy-Weinberg equilibrium with the exception of VKORC1 -1639. Weekly stable dosages were 31.7 ± 13.9 mg/week; median: 30 mg/week, range: 11-70 mg/week. Significant associations with dose were seen for VKORC1 -1639 ( P < 0.001), CYP2C9*2 ( P = 0.005) and *3 ( P = 0.003), the CYP4F2 SNP ( P-trend = 0.00037), and VKORC1 3730 (p-trend = 0.042). In linear regression, age, sex, weight, and CYP2C9 *2 and *3 and VKORC1-1639 genotype explained 42% of variance. The addition of CYP4F2 genotype to the regression model increased the degree of variance explained to 47%. Addition of VKORC1 SNP -1639 to a model eliminated the association of VKORC1 3730 with warfarin dose ( P-trend = 0.74), but -1639 remained highly significant. No impact on dose was observed for the other tested genetic variants. [ABSTRACT FROM AUTHOR]

Abstract: Introduction: Several factors responsible for inter-individual differences in response to warfarin have been confirmed; however, unidentified factors appear to remain. The purpose of this study was to examine a simple method to evaluate whether optional variables are appropriate as factors to improve dosing algorithms. Materials and Methods: All patients were Japanese. Genotyping of selected genes was conducted, and other information was obtained from medical record. Dosing algorithms were constructed by multivariate linear regression analyses and were evaluated by the Akaike Information Criterion (AIC). Results and Conclusions: Multivariate analysis showed that white blood-cell count (WBC), concomitant use of allopurinol, and CYP4F2 genotype are apparently involved in warfarin dose variation, in addition to well-known factors, such as age and VKORC1 genotype. We evaluated the adequacy of these variables as factors to improve the dosing algorithm using the AIC. Addition of WBC, allopurinol administration and CYP4F2 genotype to the basal algorithm resulted in decreased AIC, suggesting that these factor candidates may contribute to improving the prediction of warfarin maintenance dose. This study is the first to evaluate the warfarin dosing algorithm by AIC. To further improve the dosing algorithm, AIC may be a simple and useful tool to evaluate both the model itself and factors to be incorporated into the algorithm. [ABSTRACT FROM AUTHOR]

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT • Genetic polymorphisms of VKORC1 and CYP2C9 are known to influence warfarin dosage. • Recent studies among Caucasians showed that polymorphisms of CYP4F2 also play a role in warfarin pharmacogenetics. • The contribution of CYP4F2 variants to the variability inwarfarin dose requirement in Chinese subjects remains to be investigated. WHAT THIS STUDY ADDS • This research was to study the effect of CYP4F2 variants on warfarin requirements in the Han Chinese population. • This study developed a multiple regression model including CYP2C9, VKORC1 3673G>A, CYP4F2 genotypes and age, weight, combination use of amiodarone which could explain 56.1% of the individual variability in warfarin dose CYP4F2 could explain 4% of the variance in warfarin dose. • We found that one novel genotypic polymorphism 5417G>T for Asp36Tyr, which was identified as an important marker of warfarin resistance, was absent in the Han Chinese population in our study. AIMS The objective of this study was to assess the effect of the CYP4F2 on the daily stable warfarin dose requirement in Han Chinese patients with mechanical heart valve replacement (MHVR). METHODS From March 2007 to November 2008, 222 Han Chinese MHVR patients were recruited in our study. VKORC1 3673G>A, 5417G>T, CYP2C9*3 and CYP4F2 rs2108622 were genotyped by using the polymerase chain reaction restriction fragment length polymorphism method (PCR-RFLP). Polymorphisms of VKORC1 9041G>A were detected by direct sequencing. Multiple linear regression analysis was used to investigate the contribution of CYP4F2. RESULTS The CYP4F2 rs2108622 CT/TT group took a significantly higher stable warfarin dose (3.2 mg day−1) than the CC group (2.9 mg day−1, 95% CI 0.2, 1.0, P= 0.033). The multiple linear regression model included VKORC1 3673G>A, CYP2C9, CYP4F2 genotypes and clinical characteristics. The model could explain 56.1% of the variance in stable warfarin dose in Han Chinese patients with MHVR. CYP4F2 contributed about 4% to the variance in the warfarin dose. There was no variation in the SNPs of VKORC1 5417G>T. CONCLUSION CYP4F2 is a minor significant factor of individual variability in the stable warfarin dose in Han Chinese patients with MHVR. The effect of CYP2C9 and VKORC1 genotypes on variability in the stable warfarin dose had also been confirmed. [ABSTRACT FROM AUTHOR]

Warfarin dosing remains challenging because of its narrow therapeutic window and large variability in dose response. We sought to analyze new factors involved in its dosing and to evaluate eight dosing algorithms, including two developed by the International Warfarin Pharmacogenetics Consortium (IWPC).we enrolled 108 patients on chronic warfarin therapy and obtained complete clinical and pharmacy records; we genotyped single nucleotide polymorphisms relevant to the VKORC1, CYP2C9, and CYP4F2 genes using integrated fluidic circuits made by Fluidigm.When applying the IWPC pharmacogenetic algorithm to our cohort of patients, the percentage of patients within 1 mg/d of the therapeutic warfarin dose increases from 54% to 63% using clinical factors only, or from 38% using a fixed-dose approach. CYP4F2 adds 4% to the fraction of the variability in dose (R2) explained by the IWPC pharmacogenetic algorithm (P<0.05). Importantly, we show that pooling rare variants substantially increases the R2 for CYP2C9 (rare variants: P=0.0065, R2=6%; common variants: P=0.0034, R2=7%; rare and common variants: P=0.00018; R2=12%), indicating that relatively rare variants not genotyped in genome-wide association studies may be important. In addition, the IWPC pharmacogenetic algorithm and the Gage (2008) algorithm perform best (IWPC: R2=50%; Gage: R2=49%), and all pharmacogenetic algorithms outperform the IWPC clinical equation (R2=22%). VKORC1 and CYP2C9 genotypes did not affect long-term variability in dose. Finally, the Fluidigm platform, a novel warfarin genotyping method, showed 99.65% concordance between different operators and instruments.CYP4F2 and pooled rare variants of CYP2C9 significantly improve the ability to estimate warfarin dose. [ABSTRACT FROM AUTHOR]

Warfarin remains the drug of choice for long-term anticoagulation management in a variety of conditions. Despite an established role in prevention of thromboembolic events such as stroke, warfarin continues to be underutilized because of its association with serious drug-related adverse events. Lacking alternative therapeutic approaches, intensive research in the past decade has focused on making anticoagulation with warfarin safer. Much emphasis has been placed on defining factors associated with the wide individual variability in warfarin dose. Polymorphic sites in three genes, cytochrome P450 (CYP) 2C9, vitamin K 2,3 epoxide reductase complex 1 (VKORC1), and CYP4F2, have been shown to affect stable warfarin dose. An overview of the persistent issues related to warfarin therapy and our current understanding of the genetic and clinical factors affecting warfarin dosing is presented. Finally, unresolved issues in improving clinical care of warfarin patients and future directions are provided. [ABSTRACT FROM AUTHOR]

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A variant in the CYP4F2 gene, rs2108622, has been recently shown to determine stable warfarin dose requirements. CYP4F2 has also been shown recently to metabolize vitamin K.Three hundred and eleven patients were recruited prospectively from two UK hospitals and followed-up for 6 months. Fine mapping of the whole CYP4F2 region was performed to try and define the haplotype structure of CYP4F2. Genotyping was performed on the Sequenom platform. Univariate and multiple regression analyses were undertaken to assess the effect of CYP4F2 on predefined clinical outcomes of warfarin response.Fifty-nine single nucleotide polymorphisms in the CYP4F2 gene were analyzed. There was a high degree of linkage disequilibrium in the gene with two haplotype blocks. No association was found with warfarin stable dose and rs2108622 in our prospective cohort of patients even after adjustments to reduce patient heterogeneity. Interestingly, a single nucleotide polymorphism (rs2189784), which is in strong linkage disequilibrium with rs2108622, showed an association with time-to-therapeutic international normalized ratio which remained significant after the correction for multiple testing (Pc = 0.03). No association was shown with the haplotypes after false discovery rate correction.Although we were unable to demonstrate an association between rs2108622 and stable warfarin dose, our finding of an association between rs2189784 and time-to-therapeutic international normalized ratio is consistent with the recent finding that CYP4F2 plays a role in vitamin K metabolism. However, the effect of CYP4F2 is relatively small in all studies undertaken so far, and thus seems unlikely to be of clinical relevance. [ABSTRACT FROM AUTHOR]

Aiming to elucidate the system-wide effects of the alcohol-induced increase in the content of cytochrome P450 2E1 (CYP2E1) on drug metabolism, we explored the array of its protein-protein interactions (interactome) in human liver microsomes (HLM) with chemical crosslinking mass spectrometry (CXMS). Our strategy employs membrane incorporation of purified CYP2E1 modified with photoreactive crosslinkers benzophenone-4-maleimide and 4-(N-succinimidylcarboxy)benzophenone. Exposure of bait-incorporated HLM samples to light was followed by isolating the His-tagged bait protein and its crosslinked aggregates on Ni-NTA agarose. Analyzing the individual bands of SDS-PAGE slabs of thereby isolated protein with the toolset of untargeted proteomics, we detected the crosslinked dimeric and trimeric complexes of CYP2E1 with other drug-metabolizing enzymes. Among the most extensively crosslinked partners of CYP2E1 are the cytochromes P450 2A6, 2C8, 3A4, 4A11, and 4F2, UDP-glucuronosyltransferases (UGTs) 1A and 2B, fatty aldehyde dehydrogenase (ALDH3A2), epoxide hydrolase 1 (EPHX1), disulfide oxidase 1α (ERO1L), and ribophorin II (RPN2). These results demonstrate the exploratory power of the proposed CXMS strategy and corroborate the concept of tight functional integration in the human drug-metabolizing ensemble through protein-protein interactions of the constituting enzymes. [ABSTRACT FROM AUTHOR]

The sphingosine-1-phosphate receptor modulator fingolimod is predominantly hydroxylated by cytochrome CYP4F2. In vitro experiments showed that ketoconazole significantly inhibited the oxidative metabolism of fingolimod by human liver microsomes and by recombinant CYP4F2. The authors used ketoconazole as a putative CYP4F2 inhibitor to quantify its influence on fingolimod pharmacokinetics in healthy subjects. In a 2-period, singlesequence, crossover study, 22 healthy subjects received a single 5-mg dose of fingolimod in period 1. In period 2, subjects received ketoconazole 200 mg twice daily for 9 days and a single 5-mg dose of fingolimod coadministered on the 4th day of ketoconazole treatment. Ketoconazole did not affect fingolimod t[submax] or half-life, but there was a weak average increase in C[submax] of 1.22-fold (90% confidence interval, 1.15-1.30). The AUC over the 5 days of ketoconazole coadministration increased 1.40-fold (1.31-1.50), and the full AUC to infinity increased 1.71-fold (1.53-1.91). The AUC of the active metabolite fingolimod-phosphate was increased to a similar extent by 1.67-fold (1.50-1.85). Ketoconazole predose plasma levels were not altered by fingolimod. The magnitude of this interaction suggests that a proactive dose reduction of fingolimod is not necessary when adding ketoconazole to a fingolimod regimen. The clinician, however, should be aware of this interaction and bear in mind the possibility of a fingolimod dose reduction based on clinical monitoring. [ABSTRACT FROM AUTHOR]

Introduction: It is known that warfarin treatment is problematic, due to its narrow therapeutic range and to the great interindividual variability. Numerous papers have shown the important contribution of CYP2C9 and VKORC1 genetic variants to this variability. Recently, a new SNP within the CYP4F2 gene was found associated with warfarin dose in the USA. Aims: The aim of our work was to replicate this study in the Italian population and to assess the new CYP4F2 variant relative contribution in explaining warfarin dose variability with respect to CYP2C9 and VKORC1 genetic variants together with age and weight. Materials & methods: CYP4F2 rs2108622 genotyping was performed by allelic discrimination assay by TaqMan® technology. Analysis of variance and multiple linear regression analyses were carried out to examine the contribution of genetic and nongenetic factors. Results: Our TT patients require 5.49 mg/day versus 2.93 mg/day of our CC patients. Analysis of variance indicates that about 7% of mean weekly warfarin dose variance is explained by CYP4F2 genotype. Our linear regression model including CYP4F2, CYP2C9 and VKORC1 genetic variants, age and weight, explains 60.5% of the interindividual variability. Conclusion: Our data confirm and strengthen the role of this variant. [ABSTRACT FROM AUTHOR]

Abstract: P450 enzymes comprising the human CYP4F gene subfamily are catalysts of eicosanoid (e.g., 20-HETE and leukotriene B4) formation and degradation, although the role that individual CYP4F proteins play in these metabolic processes is not well defined. Thus, we developed antibodies to assess the tissue-specific expression and function of CYP4F2, one of four CYP4F P450s found in human liver and kidney. Peptide antibodies elicited in rabbits to CYP4F2 amino acid residues 61–74 (WGHQGMVNPTEEG) and 65–77 (GMVNPTEEGMRVL) recognized on immunoblots only CYP4F2 and not CYP4F3b, CYP4F11 or CYP4F12. Immunoquantitation with anti-CYP4F2 peptide IgG showed highly variable CYP4F2 expression in liver (16.4±18.6pmol/mg microsomal protein; n =29) and kidney cortex (3.9±3.8pmol/mg; n =10), with two subjects lacking the hepatic or renal enzyme entirely. CYP4F2 content in liver microsomes was significantly correlated (r ⩾0.63; p <0.05) with leukotriene B4 and arachidonate ω-hydroxylase activities, which are both CYP4F2-catalyzed. Our study provides the first example of a peptide antibody that recognizes a single CYP4F P450 expressed in human liver and kidney, namely CYP4F2. Immunoquantitation and correlation analyses performed with this antibody suggest that CYP4F2 functions as a predominant LTB4 and arachidonate ω-hydroxylase in human liver. [Copyright &y& Elsevier]

Abstract: The oxidative metabolism of tocopherols and tocotrienols by monooxygenases is a key factor in the plasma and tissue clearance of forms of vitamin E other than α-tocopherol. It is well known that a commonly ingested form of vitamin E, γ-tocopherol, has greatly reduced plasma half-life (faster clearance) than α-tocopherol. The tocotrienols are metabolized even faster than γ-tocopherol. Both γ-tocopherol and α- and δ-tocotrienol possess intriguing biological activities that are different from α-tocopherol, making them potentially of interest for therapeutic use. Unfortunately, the fast clearance of non-α-tocopherols from animal tissues is a significant hurdle to maximizing their effect(s) as dietary supplements. We report here the design and synthesis of N-heterocycle-containing analogues of α-tocopherol that act as inhibitors of Cyp4F2, the key monooxygenase responsible for ω-hydroxylation of the side chain of tocols. In particular, an ω-imidazole containing compound, 1, [(R)-2-(9-(1H-imidazol-1-yl)nonyl)-2,5,7,8-tetramethylchroman-6-ol] had an ED50 for inhibition of γ-CEHC production from γ-tocopherol of ∼1nM when tested in HepG2 cells in culture. Furthermore, feeding of 1 to mice along with rapidly metabolized δ-tocopherol, resulted in a doubling of the δ-tocopherol/α-tocopherol ratio in liver (P <0.05). Thus, 1 may be a useful adjuvant to the therapeutic use of non-α-tocopherols. [Copyright &y& Elsevier]

Abstract: Approximately 12% of Americans do not consume the estimated average requirement for zinc and could be at risk for zinc deficiency. Since zinc has proposed antioxidant function, inadequate zinc consumption may lead to an enhanced susceptibility to oxidative stress through several mechanisms, including altered antioxidant defenses. In this study, we hypothesized that dietary zinc restriction would result in lower antioxidant status and increased oxidative damage. We fed weanling Sprague–Dawley rats (n=12 per group) a zinc-adequate (50 mg/kg of zinc) diet, a zinc-deficient (<0.05 mg/kg of zinc) diet or a pair-fed diet for 3 weeks and then assessed their antioxidant status and oxidative stress parameters. Rats were zinc deficient as indicated by a significant (P<.05) reduction in body weight (49%) and 19% lower (P<.05) hepatic zinc (20.6±2.1 mg/kg) as compared with zinc-adequate rats (24.6±2.2 mg/kg). Zinc deficiency resulted in elevated (P<.05) plasma F2 isoprostanes. Zinc deficiency-mediated oxidative stress was accompanied by a 20% decrease (P<.05) in the ferritin-reducing ability of plasma assay and a 50% reduction in plasma uric acid (P<.05). No significant change in plasma ascorbic acid or in plasma α-tocopherol and γ-tocopherol was observed. However, hepatic α-tocopherol and γ-tocopherol concentrations were decreased by 38% and 27% (P<.05), respectively, as compared with those in zinc-adequate rats. Hepatic α-tocopherol transfer protein levels were unaltered (P>.05) by zinc deficiency, but cytochrome P450 (CYP) 4F2 protein levels were elevated (P<.05) as compared with those in zinc-adequate rats. Collectively, zinc deficiency increased oxidative stress, which may be partially explained by increased CYP activity and reductions in hepatic α-tocopherol and γ-tocopherol and in plasma uric acid. [Copyright &y& Elsevier]

Abstract: The present study examined accumulation of the metal toxins cadmium (Cd) and lead (Pb) in relation to the abundance of cytochrome P450 4F2 (CYP4F2), CYP2E1 and concentrations of zinc and copper in liver and kidney samples using immunoblotting coupled with metal analysis. The post mortem liver and kidney cortex samples were from 23 males and 8 females aged 3–89 years. All were Caucasians who had not been exposed to metals in the workplace. Average kidney cortex Cd load of 17.4μg/g w.w. was 17 times greater than average liver Cd load (1.1μg/g w.w.). In contrast, average kidney cortex Pb load of 0.09μg/g w.w. was two times lower than liver Pb load of 0.19μg/g w.w. Average Zn and Cu concentrations in the kidney cortex samples were 67% and 33% lower than those in the liver. Liver and kidney Cd loads, but not liver or kidney Pb loads, correlated positively with donors’ age. After controlling for liver Cd load, an inverse correlation was seen between Zn and age (partial r =−0.39, P =0.02), suggesting reduction in liver Zn levels in old age. Liver CYP2E1 protein abundance correlated with age-adjusted Cd load (partial r =0.37, P =0.02) whereas kidney CYP4F2 protein abundance showed a positive correlation with age-adjusted Cd loads (partial r =0.40, P =0.02). These findings suggest that Cd may be an inducer of renal CYP4F2 and hepatic CYP2E1 and that increased renal CYP4F2 expression may implicate in Cd-linked renal tubular dysfunction and high blood pressure, involving CYP4F2-dependent arachidonic acid metabolism. [Copyright &y& Elsevier]

We previously reported the cloning of a human liver leukotriene B4 (LTB4) ω-hydroxy-lase P450 designated CYP 4F2 [Kikuta et al. (1994) FEBS Lett. 348, 70–74]. However, the properties of CYP 4F2 remain poorly defined. The preparation solubilized using n-octyl-β-D-glucopyranoside from microsomes of CYP 4F2-expressing yeast cells catalyzes ω- hydroxylation of LTB4, 6-trans-LTB4, lipoxin A4, 8-hydroxyeicosatetraenoate, 12-hydroxye-icosatetraenoate, and 12-hydroxystearate in the presence of rabbit liver NADPH-P450 reductase. In addition, the enzyme shows ethoxycoumarin O-deethylase and p-nitroanisole O-demethylase activities. The enzyme was purified to apparent electrophoretic homogeneity from yeast cells by sequential chromatography of solubilized microsomes through amino-n-hexyl-Sepharose 4B, DEAE-HPLC, and hydroxylapatite HPLC columns. The final preparation showed a specific content of 11.1 nmol of P450/mg of protein, with an apparent molecular mass of 56.3 kDa. CYP 4F2 was distinguished from the closely homologous CYP 4F3 (human neutrophil LTB4 to-hydroxylase) by its much higher K4 for LTB4, inability to ω-hydroxylate lipoxin B4; and extreme instability. [ABSTRACT FROM AUTHOR]