Vitamin K Epoxide Reductase Complex 1 Research Articles

Correlation between Metabolic Parameters and Warfarin Dose in Patients with Heart Valve Replacement of Different Genotypes.

Warfarin has become the first choice for anticoagulation in patients who need lifelong anticoagulation due to its clinical efficacy and low price. However, the anticoagulant effect of warfarin is affected by many drugs, foods, etc. accompanied by a high risk of bleeding and embolism. The Vitamin K epoxide reductase complex 1 (VKORC1) and Cytochrome P450 2C9 (CYP2C9) genotypic variation can influence the therapeutic dose of warfarin. However, it is not clear whether there is a correlation between warfarin dose and liver function, kidney function and metabolic markers such as uric acid (UA) in patients with different genotypes. We performed a single-center retrospective cohort study to evaluate the factors affecting warfarin dose and to establish a dose conversion model for warfarin patients undergoing heart valve replacement. We studied 343 patients with a mechanical heart valve replacement, compared the doses of warfarin in patients with different warfarin-related genotypes (CYP2C9 and VKORC1), and analyzed the correlation between liver function, kidney function, UA and other metabolic markers and warfarin dose in patients with different genotypes following heart valve replacement. Genotype analysis showed that 72.01% of patients had CYP2C9*1/*1 and VKORC1 mutant AA genotypes. Univariate regression analysis revealed that the warfarin maintenance dose was significantly correlated with gender, age, body surface area (BSA), UA and genotype. There was no correlation with liver or kidney function. Multiple linear regression analysis showed that BSA, genotype and UA were the independent factors influencing warfarin dose. There is a significant correlation between UA content and warfarin dose in patients with heart valve replacement genotypes CYP2C9*1/*1/VKORC1(GA+GG), CYP2C9*1/*1/VKORC1AA and CYP2C9*1/*1/VKORC1AA.

Sodium dehydroacetate-induced disorder of coagulation function in broiler chickens and the protective effect afforded by vitamin K

Sodium dehydroacetate (S-DHA) is used widely as a preservative in several products, including poultry feed. The anticoagulation effect of 200 mg/kg S-DHA in rats has been reported to accompany a reduction in hepatic expression of vitamin K epoxide reductase complex 1 (VKORC1). Poultry and mammals have different physiology and coagulation systems, and species differences in VKORC1 expression have been found. The effect of S-DHA on blood clotting of poultry has not been studies deeply. S-DHA was given to yellow-plumage broilers (YBs) as single and multiple administrations. Vitamin K3 (VK3) was injected into YBs 2 wk after S-DHA administration. Then, the prothrombin time (PT), partial activated prothrombin time (APTT), plasma levels of vitamin K (VK), factor IX (FIX), and S-DHA, and hepatic expression of VKORC1 were obtained. Chicken hepatocellular carcinoma (LMH) cells were also exposed to S-DHA, and the cell activity, VK level, and FIX level were measured. S-DHA prolonged the PT or APTT significantly, decreased levels of VK and FIX in blood, and inhibited hepatic expression of VKORC1. The maximum changes were 1.15-fold in the PT, 1.42-fold in the APTT, 0.8-fold in the VK level, 0.7-fold in the FIX level, and 0.35-fold in VKORC1 expression compared with controls. The cell activity, VK level, FIX level, and VKORC1/VKORC1L1 expression of LMH cells were reduced significantly at S-DHA doses of 2.0 to 10.0 mM. Prolongation of the PT/APTT and lower levels of VK/FIX in YBs or the lower cell activity and VK/FIX levels in LMH cells induced by S-DHA therapy were resisted significantly by VK3 treatment. We demonstrated that S-DHA could induce a disorder in coagulation function in YBs or in LMH cells via reduction of VKORC1/VKORC1L1 expression, and that VK could resist this anticoagulation effect.

The screening for anticoagulant rodenticide gene VKORC1 polymorphism in the rat Rattus norvegicus, Rattus tanezumi and Rattus losea in Hong Kong

Anticoagulants are a major component of rodenticides used worldwide, which function by effectively blocking the vitamin K cycle in rodents. The rat Vitamin K epoxide Reductase Complex (VKORC) subunit 1 is the enzyme responsible for recycling vitamin K, and five substitution mutations (Tyr139Cys, Tyr139Ser, Tyr139Phe and Leu128Gln and Leu120Gln) located in the VKORC1 could result in resistance to anticoagulant rodenticides. This study carried out a VKORC1-based survey to estimate the anticoagulant rodenticide resistance in three Rattus species (R. losea, R. norvegicus, and R. tanezumi) collected in Hong Kong. A total of 202 rats captured in Hong Kong between 2017 and 2021 were analysed. Sequencing of molecular marker cytochrome c oxidase subunit 1 (COX1) was carried out to assist the species identification, and the identities of 52 lesser ricefield rats (R. losea), 81 common rats (R. norvegicus) and 69 house rats (R. tanezumi) were confirmed. Three VKORC1 exons were amplified from individuals by PCR followed by Sanger sequencing. A total of 47 R. tanezumi (68.1%) contained Tyr139Cys mutation in VKORC1 gene, and half of them were homozygous. None of the collected R. losea and R. norvegicus were detected with the five known substitutions leading to anticoagulant rodenticides resistance, and previously undescribed missense mutations were revealed in each species. Whole genome sequencing was further carried out on some individuals, and single nucleotide polymorphisms (SNPs) were also identified in the introns. This is the first study investigating the situation of anticoagulant rodenticide resistance in the rats collected in Hong Kong. Given that the efficacy of rodenticides is crucial for effective rodent management, regular genetic testing as well as population genomic analyses will be required to both monitor the situation and understand the adaption of different rat haplotypes for integrated pest management. Susceptibility tests for individual rodenticides should also be conducted regularly to assess their effectiveness on local species.

Influence of Renal Impairment and Genetic Subtypes on Warfarin Control in Japanese Patients.

The genotypes of vitamin K epoxide reductase complex 1 (VKORC1) and cytochrome P450 2C9 (CYP2C9) can influence therapeutic warfarin doses. Conversely, nongenetic factors, especially renal function, are associated with warfarin maintenance doses; however, the optimal algorithm for considering genes and renal dysfunction has not been established. This single-center prospective cohort study aimed to evaluate the factors affecting warfarin maintenance doses and develop pharmacogenetics-guided algorithms, including the factors of renal impairment and others. To commence, 176 outpatients who were prescribed warfarin for thromboembolic stroke prophylaxis in the stroke center, were enrolled. Patient characteristics, blood test results, dietary vitamin K intake, and CYP2C9 and VKORC1 (-1639G>A) genotypes were recorded. CYP2C9 and VKORC1 (-1639G>A) genotyping revealed that 80% of the patients had CYP2C9 *1/*1 and VKORC1 mutant AA genotypes. Multiple linear regression analysis demonstrated that the optimal pharmacogenetics-based model comprised age, body surface area, estimated glomerular filtration rate (eGFR), genotypes, vitamin K intake, aspartate aminotransferase levels, and alcohol intake. eGFR exercised a significant impact on the maintenance doses, as an increase in eGFR of 10 mL/min/1.73 m2 escalated the warfarin maintenance dose by 0.6 mg. Reduced eGFR was related to lower warfarin maintenance doses, independent of VKORC1 and CYP2C9 genotypes in Japanese patients.

Sex Metabolic Differences and Effects on Blood Coagulation Among Rats Exposed to Sodium Dehydroacetate.

Sodium dehydroacetate (Na-DHA), a fungicide used in food, feed, cosmetics, and medicine, has been found to cause coagulation aberration accompanied by the inhibition of vitamin K epoxide reductase (VKOR) in the liver in rats. VKOR complex 1 (VKORC1) and VKORC1 like-1 (VKORC1L1) are two homologous VKOR proteins. Little information is available on the effect of Na-DHA on VKORC1L1 in the liver or VKORC1/VKORC1L1 in extrahepatic tissue and sex differences in Na-DHA metabolism. In the present study, after administration of 200 mg/kg Na-DHA by gavage, significant inhibition of VKORC1 or VKORC1L1 expression in tissues, as well as prolonged prothrombin time (PT) and activated partial thromboplastin time (APTT), were observed. The PT/APTT in the Na-DHA-exposed males were 1.27- to 1.48-fold/1.17- to 1.37-fold, while the corresponding values in the Na-DHA-exposed females were 1.36- to 2.02-fold/1.20- to 1.70-fold. Serum or tissue Na-DHA concentrations were significantly higher in females than in males. The pharmacokinetic parameters (t1/2, Cmax, AUC0∼24 h, and MRT0∼24 h) of Na-DHA in female rats were significantly higher than those in male rats. Furthermore, cytochrome P450 (CYP) activity was investigated using the cocktail probe method. The results revealed that Na-DHA exhibited an inductive effect on CYP1A2, 2D1/2, and 3A1/2 activities by changing the main pharmacokinetic parameters of probe drugs in male rats. However, no significant change in CYP2E1 activity was found. There were sex differences in the metabolism and coagulation in rats exposed to Na-DHA. The lower metabolism and higher blood Na-DHA concentration in females may be the reasons for higher coagulation sensitivity in female rats.

Human gene polymorphisms and their possible impact on the clinical outcome of SARS-CoV-2 infection.

The SARS-CoV-2 pandemic has become one of the most serious health concerns globally. Although multiple vaccines have recently been approved for the prevention of coronavirus disease 2019 (COVID-19), an effective treatment is still lacking. Our knowledge of the pathogenicity of this virus is still incomplete. Studies have revealed that viral factors such as the viral load, duration of exposure to the virus, and viral mutations are important variables in COVID-19 outcome. Furthermore, host factors, including age, health condition, co-morbidities, and genetic background, might also be involved in clinical manifestations and infection outcome. This review focuses on the importance of variations in the host genetic background and pathogenesis of SARS-CoV-2. We will discuss the significance of polymorphisms in the ACE-2, TMPRSS2, vitamin D receptor, vitamin D binding protein, CD147, glucose‐regulated protein 78 kDa, dipeptidyl peptidase-4 (DPP4), neuropilin-1, heme oxygenase, apolipoprotein L1, vitamin K epoxide reductase complex 1 (VKORC1), and immune system genes for the clinical outcome of COVID-19.

AtreMorine Treatment Regulates DNA Methylation in Neurodegenerative Disorders: Epigenetic and Pharmacogenetic Studies

Background: Neurodegenerative disorders are one of the major health problems in Western countries. Genetic and epigenetic mechanisms play crucial roles in the origin and progression of these disorders. DNA methylation is the most widely studied epigenetic mark and is an important regulator of gene expression. Objective: Little is known about the influence of bioactive dietary components on epigenetic mechanisms in neurodegenerative diseases. In this study, we investigated the effects of E-PodoFavalin-15999 (AtreMorine®), a bioproduct with potent neuroprotective and dopamine enhancing capabilities, on DNA methylation patterns in Alzheimer’s (AD) and Parkinson’s Disease (PD). We also aimed to assess, in patients with PD, the effects that genetic variation across candidate pharmacogenes may have on dopamine synthesis and release in response to treatment with AtreMorine. Methods: We analyzed global DNA methylation and de novo DNA methyltransferase (DNMT) expression in a transgenic (3xTg) mouse model of AD, and further examined global DNA methylation in blood samples from patients with PD. Results: AtreMorine treatment increased global DNA methylation in 3xTg mice and in patients with Parkinson´s disease, and produced high DNMT3a expression in AD mice. We observed varied responses to AtreMorine across the following pharmacogenetic genophenotypes analyzed, cytochrome P450 oxidases (CYP2D6, CYP2C19, CYP2C9, CYP3A4, CYP3A5, CYP1A2), human arylamine N-acetyltransferase 2 (NAT2), the vitamin K epoxide reductase complex subunit 1 (VKORC1), ATP-binding cassette subfamily B member 1 (ABCB1), and solute carrier organic anion transporter family member 1B1 (SLCOB1). Conclusion: Our results suggest that AtreMorine regulates DNA methylation in neurodegenerative disorders and may constitute a new therapeutic option for the treatment of these pathologies.

Impact of CYP2C9-Interacting Drugs on Warfarin Pharmacogenomics.

Precise dosing of warfarin is important to achieve therapeutic benefit without adverse effects. Pharmacogenomics explains some interindividual variability in warfarin response, but less attention has been paid to drug‐drug interactions in the context of genetic factors. We investigated retrospectively the combined effects of cytochrome P450 (CYP)2C9 and vitamin K epoxide reductase complex (VKORC)1 genotypes and concurrent exposure to CYP2C9‐interacting drugs on long‐term measures of warfarin anticoagulation. Study participants predicted to be sensitive responders to warfarin based on CYP2C9 and VKORC1 genotypes, had significantly greater international normalized ratio (INR) variability over time. Participants who were concurrently taking CYP2C9‐interacting drugs were found to have greater INR variability and lesser time in therapeutic range. The associations of INR variability with genotype were driven by the subgroup not exposed to interacting drugs, whereas the effect of interacting drug exposure was driven by the subgroup categorized as normal responders. Our findings emphasize the importance of considering drug interactions in pharmacogenomic studies.

Association between VKORC1 gene polymorphism and warfarin dose requirement and frequency of VKORC1 gene polymorphism in patients from Kerman province

Warfarin is an anticoagulant prescribed in the treatment and prevention of thrombosis. Variation in dose requirements is different for everyone, and genetic factors have an effect on dose variation. Polymorphism of vitamin K epoxide reductase complex 1 (VKORC1) gene is identified as the main genetic factor involved in warfarin dosage requirement variations. This study aims to determine the frequency of VKORC1 polymorphism in patients using warfarin from Kerman city and investigated association between VKORC1 gene polymorphism and patient characteristics with warfarin dose requirement. A total of 112 patients taking warfarin with stable dose requirements enrolled in the study. DNA samples from these patients were genotyped for VKORC1 gene polymorphism by using the polymerase chain reaction restriction fragment length polymorphism method (PCR-RFLP) and examined associations between demographic characteristics (e.g. age, sex, smoking, etc.) and genetic factors with maintenance dose of warfarin. The most common genotype was VKORC1 GA (48.2%). genotype frequency subjects carried VKORC1 GG and AA were 39.3% and 12.5%, respectively. In addition, a significant relationship was found between VKORC1-1639G>A and the daily dose of warfarin (P = 0.011, R2 = 0.080). The frequencies of the VKORC1-1639 A alleles were significantly lower than VKORC1-1639 G alleles and required fewer warfarin dose.

Effect of Vitamin K Epoxide Reductase Complex 1 Polymorphism on Warfarin Dose Requirement among Patients in Tertiary Care Hospital.

Background:Warfarin, anticoagulant is used for thromboembolic disorders. Inter-individual variation in clinical response to warfarin is due to various factors, including polymorphism of Vitamin K epoxide reductase complex 1 (VKORC1)-1639G>A. The aim of our study was to evaluate the effect of VKORC1 polymorphism on the maintenance dose of warfarin.Materials and Methods:Cross-sectional study conducted by the departments of Pharmacology, Cell Biology and Molecular Genetics on patients attending cardiology clinic, receiving warfarin for at least 2 months. Genomic deoxyribonucleic acid was extracted and genotyping was done by Polymerase Chain Reaction - Restriction Fragment Length Polymorphism. The correlation between VKORC1 gene polymorphism and warfarin maintenance dose was analyzed.Results:A total of 102 patients with a mean age of 47.72 ± 10.31 years, of which 58 (56.86%) were male. The frequency of VKORC1 G>A for GG, GA, and AA genotypes was 74.51%, 19.61%, and 5.88%, respectively. Variant allele AA was less frequent than the wild type. Mean weekly warfarin dose was 23.12 ± 8.08, 22.93 ± 8.21, and 15.6 ± 5.35 mg in patients with GG, GA, and AA genotypes, respectively. Patients with GG genotype required therapeutic dose compared to variant type (P = 0.001). Multiple stepwise regression model showed 26.3% variability in warfarin dose was due to VKORC1 genotype (R = 0.513, R2 = 0.263, adjusted R2 = 0.256, P = 0.0001).Conclusion:VKORC1 polymorphism alone influence 26.3% variability in warfarin dose and AA genotype patients required lower dose.

VKORC1 variants as significant predictors of warfarin dose in Emiratis.

Purpose: Variability in response to warfarin is one of the main obstacles challenging its use in clinical practice. Vitamin K epoxide reductase complex (VKORC) is the target enzyme of warfarin, and variations in the form of single nucleotide polymorphisms (SNPs) in VKORC1, coding for this enzyme, are known to cause resistance to warfarin treatment. This study aimed to explore VKORC1 variants in Emirati patients receiving warfarin treatment and to correlate their genotypes at the studied SNPs to their maintenance warfarin dose.Patients and methods: Sanger sequencing of the majority of the VKORC1 gene was applied to samples from 90 patients and 117 normal individuals recruited from Tawam Hospital, Al-Ain, UAE. Genotypes at the following variants were determined (rs9923231, rs188009042, rs61742245, rs17708472, rs9934438, rs8050894, rs2359612, rs7294). Statistical analysis was applied, including ANOVA, cross-tabulation, and multiple linear regression analysis, to determine the ability of nongenetic factors (age and gender) and genetic factors (VKORC1 genotypes) to explain variability in warfarin dose in patients.Results: Different frequencies of minor alleles were detected in the selected SNPs. Significant variation among genotypes at six VKORC1 variants were identified (rs9923231, rs9934438, rs8050894, rs2359612, rs7294). The main predictors for warfarin dose were rs9923231, age, and rs61742245 with 50.7% of the average warfarin dose in our sample could be explained by a regression model built on these three factors.Conclusion: This is the first report of the explanatory power of VKORC1 genotypes and nongenetic factors (age and gender) on warfarin dose among Emiratis. Also, this study highlighted the positive effect of considering rare pharmacogenomic variants on explaining warfarin dose variability.

Verification of pharmacogenomics-based algorithms to predict warfarin maintenance dose using registered data of Japanese patients

Large inter-individual differences in warfarin maintenance dose are mostly due to the effect of genetic polymorphisms in multiple genes, including vitamin K epoxide reductase complex 1 (VKORC1), cytochromes P450 2C9 (CYP2C9), and cytochrome P450 4F2 (CYP4F2). Thus, several algorithms for predicting the warfarin dose based on pharmacogenomics data with clinical characteristics have been proposed. Although these algorithms consider these genetic polymorphisms, the formulas have different coefficient values that are critical in this context. In this study, we assessed the mutual validity among these algorithms by specifically considering racial differences. Clinical data including actual warfarin dose (AWD) of 125 Japanese patients from our previous study (Eur J Clin Pharmacol 65(11):1097-1103, 2009) were used as registered data that provided patient characteristics, including age, sex, height, weight, and concomitant medications, as well as the genotypes of CYP2C9 and VKORC1. Genotyping for CYP4F2*3 was performed by the PCR method. Five algorithms that included these factors were selected from peer-reviewed articles. The selection covered four populations, Japanese, Chinese, Caucasian, and African-American, and the International Warfarin Pharmacogenetics Consortium (IWPC). For each algorithm, we calculated individual warfarin doses for 125 subjects and statistically evaluated its performance. The algorithm from the IWPC had the statistically highest correlation with the AWD. Importantly, the calculated warfarin dose (CWD) using the algorithm from African-Americans was less correlated with the AWD as compared to those using the other algorithms. The integration of CYP4F2 data into the algorithm did not improve the prediction accuracy. The racial difference is a critical factor for warfarin dose predictions based on pharmacogenomics.

Evaluation of the effects of ontogenetic or maturation functions and chronic heart failure on the model analysis for the dose-response relationship of warfarin in Japanese children.

We previously demonstrated that the rational pediatric dosage of warfarin can be well-described by a SIZE parameter that includes an allometry exponent of weight. On the other hand, allometry alone is considered to be insufficient to predict drug clearance in neonates and infants. The primary purpose of the present study was to evaluate the effects of incorporation of the maturation process into the analysis model for the dose-response relationship of warfarin in Japanese children. In addition, we evaluated the effect of chronic heart failure (CHF) on the response to warfarin as an independent risk factor for increased anticoagulant effects. Thirty-eight patients with stable anticoagulation by warfarin were enrolled. During a mean follow-up period of 4.74 ± 3.51years, 1092 data points including prothrombin time-international normalized ratio (PT-INR) were obtained. The data were subjected to multiple regression analysis to identify covariates related to the anticoagulant effects. Two different models describing the maturation process did not improve the predictive performance for the dose-response relationship in pediatric patients. In addition to the SIZE-normalized daily dose, the vitamin K epoxide reductase complex 1 (VKORC1) genotype, and concomitant use of bosentan, CHF was identified as a covariate increasing the anticoagulant effects of warfarin to 118%. The SIZE parameter was useful even without incorporation of maturation models to describe the response to warfarin in pediatric patients, and our longitudinal follow-up study design with multiple observations was beneficial to detect changes within individual subjects.

Mutations in CYP2C9 and/or VKORC1 haplotype are associated with higher bleeding complications in patients with Budd-Chiari syndrome on warfarin.

Anticoagulation is universally recommended in Budd-Chiari syndrome [BCS]. Vitamin K epoxide reductase complex 1 (VKORC1) and CYP2C9 are involved in the metabolism of warfarin. The present study was done to assess whether these mutations are associated with the risk of bleeding in patients with BCS receiving warfarin. Patients diagnosed with BCS underwent genotyping for three single nucleotide polymorphisms [SNPs]-two for the CYP2C9 and one for the VKORC1 haplotype. The patients were followed up for at least 12months and all bleeding episodes were recorded. Patients with and without mutations were compared for bleeding complications and a crude odds ratio [crude OR] was derived for the association between bleeding and presence or absence of mutant alleles. Eighty patients [mean (SD) age 27.47 (8.93) years, 35 male] with BCS underwent genetic testing. 37/80 (46.2%) patients had mutation of CYP2C9 and/or VKORC1; 22/80 (27.5%) had either of the mutant alleles of CYP2C9 and, similarly, 22/80 (27.5%) had the VKORC mutation. Over a median follow-up of 20 (range 12-96) months, 21/80 (26.3%) patients had bleeding complications. Patients with mutant SNPs had a higher risk of bleeding than those without [14/37 vs. 7/43, p = 0.04, crude OR (95% CI) 3.13 (1.1-8.9)]. The presence of mutations in VKORC1 or CYP2C9 is associated with increased risk of bleeding in patients with BCS on warfarin. Such patients with SNPs of CY2C9 or VKORC1 haplotype should be monitored intensively while receiving warfarin.

The pediatric acenocoumarol dosing algorithm: the Children Anticoagulation and Pharmacogenetics Study

Background The large variability in dose requirement of vitaminK antagonists is well known. For warfarin, pediatric dosing algorithms have been developed to predict the correct dose for a patient;however,this is not the case for acenocoumarol. Objectives To develop dosing algorithms for pediatric patients receiving acenocoumarol with and without genetic information. Methods The Children Anticoagulation and Pharmacogenetics Study was designed as a multicenter retrospective follow-up study in Dutch anticoagulation clinics and children's hospitals. Pediatric patients who used acenocoumarol between 1995 and 2014 were selected for inclusion. Clinical information and saliva samples for genotyping of the genes encoding cytochromeP450 (CYP)2C9, vitaminK epoxide reductase complex subunit1 (VKORC1), CYP4F2, CYP2C18 and CYP3A4 were collected. Linear regression was used to analyze their association with the log mean stable dose. A stable period was defined as three or more consecutive International Normalized Ratio measurements within the therapeutic range over a period of ≥3weeks. Results In total, 175 patients were included in the study, of whom 86 had a stable period and no missing clinical information (clinical cohort; median age 8.9years, and 49% female). For 80 of these 86 patients, genetic information was also available (genetic cohort). The clinical algorithm, containing body surface area and indication, explained 45.0% of the variability in dose requirement of acenocoumarol. After addition of the VKORC1, CYP2C9, and CYP2C18 genotypes to the algorithm, this increased to 61.8%. Conclusions These findings show that clinical factors had the largest impact on the required dose of acenocoumarol in pediatric patients. Nevertheless, genetic factors, and especially VKORC1, also explained a significant part of the variability.

Pharmacogenetics of vitamin K antagonists and bleeding risk prediction in atrial fibrillation

Polymorphisms in the vitamin K epoxide reductase complex 1 (VKORC1) and cytochrome P450 2C9 (CYP2C9) genes increase the bleeding risk in anticoagulated atrial fibrillation (AF) patients. Here, we aimed to investigate whether VKORC1 and CYP2C9 polymorphisms improved the predictive performance for major bleeding using the HAS-BLED score. We recruited 652 consecutive AF patients stable on vitamin K antagonist (INR 2.0-3.0) during at least the previous 6 months. A baseline venous blood sample was obtained for DNA extraction. We gave an extra point to the HAS-BLED score if the patient was a simultaneous carrier of the VKORC1 and CYP2C9 polymorphisms related to bleeding, and we called this modified score "GEN|HAS-BLED." During a median follow-up of 7.6 years (IQR 5.6-8.0), all major bleeding events were recorded. During follow-up, 106 (16.2%) patients experienced a major bleeding (2.81%/y; 42 intracranial haemorrhages and 44 gastrointestinal bleeding) and 24 (3.7%) died from major bleeding (0.48%/y). Cox regression analyses demonstrated a significant association between HAS-BLED or GEN|HAS-BLED and major bleeds, both as continuous or categorical scores. Comparison of receiver operating characteristic (ROC) curves shows that original HAS-BLED clinical score had better predictive ability than GEN|HAS-BLED (0.660, 95% CI 0.622-0.696 vs 0.645, 95% CI 0.607-0.682; P = .030). Discrimination and reclassification analyses showed that GEN|HAS-BLED did not improve sensitivity compared with the original score and even showed significant negative reclassification. Adding pharmacogenetic factors (ie polymorphisms of the VKORC1 and CYP2C9 genes) to the HAS-BLED score does not improve the prediction or discrimination performance for major bleeding.

The effect of ‘cytochrome P-450 2C9’ and ‘vitamin K epoxide reductase complex 1’ genetic polymorphism upon oral anticoagulation requirements

Background Polymorphisms in the gene encoding the cytochrome P-450 2C9 enzyme (CYP2C9) are known to contribute to variability in sensitivity to Marevan. CYP2C9 is the enzyme primarily responsible for the metabolic clearance of s-enantiomer of Marevan. The vitamin K epoxide reductase complex 1 (VKORC1) is the target of coumarin anticoagulants, and its common genetic variants result in altered sensitivity to Marevan. VKORC1 polymorphisms are associated with a need for lower doses of Marevan during long-term therapy. Aim of work This study aimed to assess the allelic frequencies and to investigate the relationship between ‘CYP2C9’ and ‘VKORC1’ genotype and vitamin K antagonist anticoagulation. Patients and methods This study was conducted on 40 patients. There were 24 females and 16 males, with a male to female ratio of 2 : 3. Their ages ranged from 28 to 72 years. All the studied patients were laboratory investigated with international normalized ratio, complete blood count, and detection of VKORC1 and CYP2C9 by PCR reverse hybridization method using PGX thrombo strip assay (Vienna Lab.). Results Regarding the distribution of patients according to frequency of deep venous thrombosis (DVT) attacks, 16 (40%) patients showed single attack of DVT and 24 (60%) patients showed recurrent attacks. Patients with single attack of DVT comprised 12 (75%) females and four (25%) males, with male to female ratio of 1 : 3. As for the patients with recurrent attacks of DVT, there were 13 (54.2%) female and 11(45.8%) male patients, with a male to female ratio of 1 : 1.2. Conclusion Detection of genetic polymorphisms in CYP2C9 and VKORC1 genes before onset of warfarin therapy greatly influenced response to warfarin and shortened the time required to reach target international normalized ratio, and hence reduced the risk of recurrence of DVT.

Effects of VKORC1 and CYP2C9 gene polymorphisms on the stable dosage of warfarin and anticoagulation in patients with hypertrophic cardiomyopathy complicated with atrial fibrillation

Objective To investigate the gene polymorphisms and clinical features of vitamin K epoxide reductase complex 1(VKORC1) and cytochrome P450 2C9(CYP2C9), and their correlation with stable dosage of warfarin, the risk of hemorrhage and over anticoagulation for patients with hypertrophic cardiomyopathy. Methods A retrospective study was performed on 60 patients with hypertrophic cardiomyopathy(HCM) combined with atrial fibrillation(AF) who were admitted and regularly took oral warfarin as anticoagulation from January 2011 to December 2016.The clinical data such as age, body height, body weight, stable warfarin dosage and the situation of hemorrhage and over anticoagulation were recorded.Genotypes of CYP2C9 and VKORC1 were detected with fluorescence PCR-capillary electrophoresis sequencing and the relationship between the genotype and clinical features and the stable dose of warfarin, hemorrhage and over anticoagulation were analyzed by statistical methods. Results Among the 60 patients, VKORC1-1639G>A detection showed that 50 patients were AA type, 9 patients were AG type, 1 patient was GG type, the stabledosage of AA type patients were significant lower than that of AG type patients(P 0.05); CYP2C9 detection showed that 51 patients were *1/*1 type, 6 patients were *1/*3 type, 3 patients were *3/*3 type, *2 allele was not found, the stable dosage of patients who carry the *3 mutation were significant lower than that of *1/*1 type patients, and the more *3 mutation they own, the lower dosage they require(P 0.05). The incidence of hemorrhage and anticoagulation was significantly higher in patients aged 65 years or older than those who were younger than 65 years(P<0.05). Conclusion Stable warfarin dosage has relationship with different genotypes of VKORC1 and CYP2C9 in HCM patients, age may be a factor of warfarin over anticoagulation and hemorrhage. Key words: Hypertrophic cardiomyopathy; Atrial fibrillation; Warfarin; Vitamin K epoxide reductase complex 1; cytochrome P450 2C9; Gene polymorphisms; Hemorrhage; Over anticoagulation

Cross-Validation of High-Resolution Melting Analysis-Based Genotyping Platform.

Developing genetic and pharmacogenetic panels enhances genetic testing in clinical molecular diagnostics and precision medicine. This study was designed to cross-validate the performance of Canon's multiplex high-resolution DNA melting analysis platform with the Applied Biosystems TaqMan®-based Quant Studio Real-Time PCR System and Pyrosequencing® genotyping platforms for common genetic polymorphisms of the vitamin K epoxide reductase complex 1 (VKORC1) and CYP2C9. Genomic DNA isolated from 240 blood and saliva samples was used to genotype the VKORC1-1639 G/A (rs9923231), CYP2C9*2 (430C>T, rs28371674), and CYP2C9*3 (1075A>C, rs1057910) single-nucleotide polymorphisms (SNPs) on the three above-mentioned genotyping platforms. There was 99.2%, 100%, and 100% concordance among the Canon DNA analyzer, the TaqMan-based QuantStudio, and the Pyrosequencing genotyping results for the VKORC1 (rs9923231), CYP2C9*2, and CYP2C9*3 SNPs, respectively, in DNA samples isolated from blood. The DNA samples isolated from saliva showed 100% concordance among the three test platforms for the three tested SNPs. These results show that, the DNA analyzer performed very well when compared with two commonly used genotyping platforms. The reliability, multiple genetic variant testing capability, and short turnaround time for up to eight samples make the DNA analyzer an ideal genotyping platform for genetic testing in the clinical practice setting, where efficient genotyping is important to prevent delays in optimizing drug therapy.

Genetic and Non-Genetic Factors Association with Warfarin Long Term Therapy Stability in Sudan

Anticoagulation with warfarin is characterized by a wide inter-individual variations in dose requirements and INR (International Normalised Ratio) stability, as there are evidences that warfarin response variability is associated with CYP2C9 (cytochrome P450, family 2, subfamily C, polypeptide 9) and VKORC1 (Vitamin K epoxide reductase complex1) genetic polymorphisms. Carriers of CYP2C9*2 and VKORC11639G&amp;gt;A variant alleles are at greater risk of unstable anticoagulation therapy. Objectives: This retrospective case control study was directed to analyze the impact of genetic and non-genetic factors on warfarin therapy in Sudanese out-patients who were on long term warfarin therapy. Method: 118 Sudanese outpatients receiving warfarin treatment for at least six months, were interviewed for their non-genetic factors that included age, sex, indication for warfarin therapy, compliance, Vitamin K rich foods intake and concomitant drug therapy, in addition to their blood samples which were taken for DNA extraction and genotyping of CYP2C9*2 and VKORC11639G&amp;gt;A gene polymorphisms to study the genetic factors. INR stability % index was calculated, accordingly patients were classified into 2 groups, stable and unstable groups. Results: The frequencies of VKORC11639G&amp;gt;A alleles in Sudanese out-patients who were on long term warfarin therapy were 70.3% and 29.7% for the VKORC1/G and VKORC1/A alleles respectively. The frequencies of CYP2C9*2 alleles in Sudanese out-patients were 92.4% and 7.6% for CYP2C9*1 and CYP2C9*2 alleles respectively. Variables associated with low INR stability were VKCOR1/AA genotype (p-value = 0.028) and sex (p = 0.017). Variables that showed no association with INR stability were age (p-value = 0.259), compliance (p-value = 0.058). Vitamin K rich foods intake (p- value = 0.743), and mean stable warfarin dose (p-value = 0.439). Conclusion: Polymorphism in warfarin drug target gene VKORC1-11639G&amp;gt;A and sex are important elements of INR stability in Sudanese out- patients on long term warfarin therapy.