Association Of MDR1 Gene Polymorphism With Warfarin Maintenance Dose In An Eastern Han Chinese Population

BackgroundWarfarin,a vitamin K antagonist,has been the main drug in various anticoagulants since it was approved in 1954.Although there are a variety of the emergence of new anticlotting drugs,but because of warfarin with the characteristics of cheap and effective,under China's current economic development level,will remain as the main anticoagulant drugs,widely used in heart valve surgery,atrial fibrillation,deep vein thrombosis and other diseases which needs anticoagulant therapy.Warfarin narrow therapeutic window,however,the individual difference is big,need to monitor blood coagulation function,poor compliance in patients with clinical practice,prone to anticoagulation is insufficient or excessive anticoagulation,cause serious adverse reactions,embolism,hemorrhage and even lifethreatening.Studies have shown that reasons of the individual differences are divided into two categories,category can be referred to as environmental factors,such as diet differences?vitamin K intake?,age,gender and body weight,another kind is genetic factors,two important genes have been identified as being responsible for warfarin treatment,including cytochrome P450 2C9?CYP2C9?and the vitamin K epoxide reductase complex subunit 1?VKORC1?.Warfarin affects the speed limit enzyme VKORC1 in the metabolism of vitamin K.In pharmacodynamics aspect,VKOR transformed Vit K to KH₂,warfarin interfered the way which Vit K turned to KH₂ by inhibiting VKORC1,and reduced the activation of blood coagulation factor???????,then achieved it's anticoagulation.Therefore,the VKORC1 activity directly determines the effect of warfarin anticoagulation.Warfarin is R-and S-enantiomeric racemic mixture,of which S-warfarin anticoagulation activity is 3-5 times of R-warfarin,which provides 70%anticoagulant activity of warfarin.Warfarin was metabolized by Cytochrome oxidase P450 as a non-active product,of which s-enantiomer was by CYP2C9 to produce 6-and7-carboxylated,while CYP1A1,CYP1A2 and CYP3A4 catalyzed R-enantiomer.Therefore,the activity of CYP2C9 can significantly affect the anticoagulant effect of warfarin.The establishment of individualized drug model based on warfarin gene single nucleotide polymorphisms?SNPs?is a hot topic in current research.Studies have shown that the SNPs of warfarin related genes are related to the dose of warfarin,and the main genes affecting warfarin dose are CYP2C9 and VKORC1.In 2013,the New England Journal of Medicine published two randomized controlled studies on warfarin,trying to compare the advantages and disadvantages of the model based on the prediction model,but the results were quite the opposite.An important factor is that the current prediction model is not very good at explaining the individual differences.At present,most of the prediction model only included CYP2C9?rs1057910?gene polymorphism and VKORC1?rs9923231?gene polymorphisms,age and body surface area,etc.,nearly 50%of the individual differences can be explained by these factors.The improved model which could explain much more individual differences is the main direction of further research.Therefore,it is very important to find new genes which can affect the warfarin maintenance dose.In recent years,the role of p-glycoprotein?P-gp?has been paid more and more attention in pharmacokinetic research.P-gp belongs to ATP binding box membrane transport protein superfamily.It is widely involved in the absorption,distribution,metabolism and excretion of various drugs in the body.Human P-gp genes have two subtypes,MDR1 and MDR2.Studies have shown that MDR1 C3435T?rs1045642?gene polymorphism is associated with pharmacokinetics of multiple drugs.Therefore,some scholars studied the relationship between the polymorphism of MDR1 C3435T gene and the dose of warfarin in Brazil and Egypt,and obtained the positive result.But in a mixed-race study,the opposite is true.Among the Asian populations,MDR1 gene polymorphisms mainly include rs1128503,rs2032582 and rs1045642,and these three mutant sites have high interlocking.Therefore,it is necessary to further study whether there is a correlation between the polymorphism of MDR1 C3435T gene polymorphism and the warfarin dose of han population.Objective1.Overview the distribution of CYP2C9,VKORC1 and MDR1 gene polymorphism in han population in east China.2.Overview the relationship between age,gender,weight,height,body surface area,CYP2C9,VKORC1,MDR1 gene polymorphism and the maintenance dose of warfarin in the han population in eastern China.Materials and MethodsA total of 125 patients who underwent mechanical valve replacement at Changzheng hospital in Shanghai from March 2016 to January 2018 were collected.All patients were required to undergo long-term anticoagulant therapy in warfarin,and INR was stable between 2.0 and 3.0,and the informed consent was signed.1.Statistics of patients with relevant data,including age,sex,body weight,the body surface area?BSA??using Xu Wensheng's formula,namely BSA=0.0061×height?cm?+0.0128×weight?kg?-0.1529?,prothrombin time?PT?,INR,liver function,renal function before and after taking warfarin.2.Using pyrosequencing method,CYP2C9?rs1057910?,VKORC1?rs9923231?and MDR1?rs1045642?gene single nucleotide polymorphisms were detected by extraction of peripheral venous blood?5 ml?.3.In combination with the clinical data of patients and the distribution of genetic polymorphism,the differences of the warfarin dose in patients with stable INR were comprehensively analyzed,and the warfarin dose required for different genotypes was speculated.ResultsBetween different genotype groups of patients,gender,age,body mass index?BMI?,BSA,drinking history,smoking history,basic heart disease,the types of replacement valves are according with Hardy-Weinberg balance laws.125 patients including 62 male cases and 63 female cases,the average age is53.3±9.24 years.Additionally,the average height and weight is 164±8.08 cm and60.77±11.34 kg,BSA is 1.63±0.18 m²,preoperative INR is 1.26±0.27,prothrombin time is14.29±2.78 s,alanine aminotransferase is 25.82±8.93 U/L,aspartate transaminase is26.02±8.25 U/L.After 3 months with warfarin treatment,INR is 2.27±0.27,prothrombin time is 24.33±3.90 s,alanine aminotransferase is 24.83±10.44 U/L,aspartate transaminase is 26.41±6.33 U/L.Of the 125 patients,male is 62 cases?49.6%?,their average warfarin stable dose is3.44±0.12 mg/d,female is 63 cases?50.4%?,their average warfarin stable dose is2.95±1.07 mg/d,the warfarin stable dose in female patients is significantly lower than male patients,its variance has statistical significance?P<0.05?.CYP2C9 AA genotype is found in 114 patients?91.2%?,CYP2C9 AC genotype is found in 11 cases?8.8%?,CYP2C9 CC genotype is not found?0.0%?.VKORC1 AA genotype is found in 110 patients?88.0%?,VKORC1 GA genotype is found in 14 patients?11.2%?,and VKORC1 GG genotype is found in 1 case?0.8%?.MDR1 CC genotype is found in 37 patients?29.6%?,MDR1 CT genotype is found in 65 patients?52.0%?,and MDR1 TT genotype is found in 23 cases?18.4%?.The results were in accordance with the law of genetic equilibrium of Hardy-Weinberg?P>0.05?.The mean stable warfarin dose for those subjects carrying CYP2C9 AA genotype is3.00±0.80 mg/d,and the mean stable warfarin dose for those subjects carrying CYP2C9AC genotype is 1.82±0.90 mg/d.The mean stable warfarin dose for those subjects carrying CYP2C9 AC genotype is significantly lower than that for patients carrying CYP2C9 AA genotype?P<0.05?.The mean stable warfarin dose for those subjects carrying VKORC1AA genotype is 2.81±0.85 mg/d,and the mean stable warfarin dose for those subjects carrying VKORC1 GA genotype is 3.53±0.80 mg/d.There is only 1 patient carries VKORC1 GG genotype,and the warfarin dose is 3.75 mg/d.The mean stable warfarin dose for those subjects carrying VKORC1 AA genotype is significantly lower than that for patients carrying VKORC1 GA genotype?P<0.05?.The mean stable warfarin dose for those subjects carrying MDR1 CC genotype is 2.89±0.98 mg/d,the mean stable warfarin dose for those subjects carrying MDR1 CT genotype is 2.85±0.81 mg/d,and the mean stable warfarin dose for those subjects carrying MDR1 TT genotype is 3.07±0.86 mg/d,and there are no difference between patients carrying different MDR1 genotypes.Through covariance analysis,there is also no significant relationship between MDR1gene polymorphism and warfarin stable dose.based on the differences of stable warfarin dose,all patients are divided into three groups,including the low-dose group?warfarin dose of 2.5 mg/d?,the medium-dose group?<2.5 mg/d stable warfarin dose<5 mg/d?and the high-dose group?stable warfarin dose of 5 mg/d or higher?.MDR1 CC genotype is found in 41.3%patients in the low-dose group,MDR1 CT genotype is 54.3%and MDR1 TT genotype is 4.3%.In the medium-dose group,MDR1 CC genotype is found in 24.6%patients,MDR1 CT genotype is found in 50.8%patients,and MDR1 TT genotype is found in 24.6%patients.In the high-dose group,MDR1 CC genotype is found in 16.7%patients,MDR1 CT genotype is found in 50.0%patients,and MDR1 TT genotype is found in 33.3%patients.The proportion of patients carrying MDR1 TT genotype in high-dose group was obviously higher than that in the low-dose group,and the difference is statistically significant?P<0.05?.ConclusionsIn 125 cases of patients who received warfarin treatment after artificial valve replacement in east China,the average daily maintenance warfarin dose of female patients is lower than that of male patients,and the difference was statistically significant.The mean stable warfarin dose for those subjects carrying CYP2C9 AC genotype is significantly lower than that for patients carrying CYP2C9 AA genotype?P<0.05?.The mean stable warfarin dose for those subjects carrying VKORC1 AA genotype is significantly lower than that for patients carrying VKORC1 GA genotype?P<0.05?.There is no statistically significant difference in the stable warfarin dose for patients with different MDR1 genotypes.But the proportion of patients carrying MDR1 TT genotype in high-dose group was obviously higher than that in the low-dose group,and the difference is statistically significant?P<0.05?.Therefore,we conclude that in order to obtain better treatment effect,and reduce the side-effect of warfarin,it is necessary to detect CYP2C9?rs1057910?,VKORC1?rs9923231?and MDR1?rs1045642?gene single nucleotide polymorphisms at the beginning of warfarin therapy.