To establish a method for detecting genetic polymorphism of CYP2C9 1075A>C and VKORC1-1639 G>A and evaluate the correlation of SNP with Warfarin dose

Abstract

Objective To set up a method to detect the mutation of CYP2C9 1075A>C and VKORC1-1639 G>A in clinical practice, investigate the distribution of CYP2C9 1075A>C and VKORC1-1639 G>A in Beijing Han native, and evaluate the correlations between CYP2C9 1075A>C, VKORC1-1639 G>A and Warfarin maintenance dose, so that the results will contribute to Warfarin therapy. Methods A total of 145 Han Beijing patients received long-term Warfarin anticoagulant therapy orally in the people′s hospital between May to September in 2011 were enrolled in this study.The patients suffer from atrial fibrilation, thromboembolism and heart operation respectively.The ARMS-TaqMan real time PCR technique was used to detect the mutation of CYP2C9 1075A>C and VKORC1-1639 G>A.The data were collected on gender, age and BSA.The correlations between stable warfarin dosage and each factor were analysised. Results The detection limit of ARMS-TaqMan real time PCR technique was 105 copies/ml in this assay.The intra- and inter-coefficients of variations（CV）were less than 5.5％ and less than 9.0% respectively.The coincidence rate of ARMS-TaqMan real time PCR and direct sequencing was 100% for testing 50 paralled samples.Among the 145 patients, the frequencies of CYP2C9 1075 AA, AC and CC was 93.8%（136/145），6.2%（9/145），0（0/145） respectively.The frequencies of VKORC1-1639 GG, GA and AA was 0.7%（1/145）, 21.4%（31/145）, 77.9%（113/145） respectively.Allele frequencies of genotypes were in Hardy-Weinberg equilibrium.Every patients′ INR was within range 2.0-3.0.Among the 145 patients,dosage requirements were higher in CYP2C9 1075 AA phenotype than that in AC type (F=0.199，P<0.05) and were higher in VKORC1-1639 GA phenotype than that in AA type (F=1.745，P<0.001).Irrespective of other factors, age, BSA, CYP2C9 1075 and VKORC1-1639 was accounted for 11.9%, 12.9%, 4.4% and 16.7% of interindividual variation of Warfarin dosage respectively.Linear stepwise regression analysis was used to identify factors contributing to Warfarin stable does.The final equation accounted for 40.4% of interindividual variation of Warfarin dose. Conclusions The ARMS-TaqMan real time PCR technique was established to analyse CYP2C9 1075 and VKORC1-1639 genotype.The technique can achieve the needs of clinical practice.Warfarin dosing by multivariable regression equation in patients could increase efficiency for Warfarin dose adjustment，and could give suggestion for the individualization and rationalization for Warfarin dose.（Chin J Lab Med, 2012,35:1010-1015） Key words: Cytochrome; Warfarin; Polymorphism，single nucleotide