Impact Of Genetic And Clinical Factors On Anticoagulation Therapy In Patients Undergoing Heart Valve Replacement Surgery

ObjectivesRoutine anticoagulation therapy is needed for patients undertaking mechanical heart valve replacement(MHVR).Warfarin,a vitamin k antagonist,as a classic oral anticoagulant,is the first choice for long-term anticoagulation in patients with MHVR.Warfarin has a slow effect and heparin bridging anticoagulation is needed in the early stage after operation,but there is a lack of related guidelines and studies on the selection and dose recommendation of heparin drugs in Chinese patients.A major limitation in patients treated with warfarin is the large inter individual variation in the dose required for a target anticoagulant effect.Genetic and clinical factors can affect warfarin maintenance dose(Maintenance dose,MD).A variety of models are used to predict MD.However,the sensitivity to warfarin increases in the early stage after MHVR,and the various factors influencing MD,related genes and specific clinical factors(perioperative physiological and pathological changes,perioperative medication,etc.)on the initial response of warfarin need to be further clarified.TTR reflects anticoagulant stability and is related to adverse events,but the factors influencing TTR in patients in our province are rarely reported.Cefoperazone-sulbactam can significantly affect the early response of warfarin,but the mechanism has not been elucidated.Therefore,this paper will enroll patients in Gansu as the research object to evaluate different bridging anticoagulation strategies in the early stage of MHVR.The factors most associated with the initial anticoagulant period and stable stage of warfarin were screened,and the calculation equations for predicting the initial dose and MD respectively were established and the proportions of variation explained by each factor in the two models were given.The effects of genetic andclinical factors on TTR were evaluated.What's more,we explored the mechanism of drug drug interaction between cefoperazone-sulbactam and warfarin based on transporters.Methods1.Comparison of different bridging anticoagulation therapies used after mechanical heart valve replacement in Chinese patientsWe performed a prospective,single-center,observational cohort study of 305 patients who underwent elective MHVR with different bridging anticoagulation regimens.Patients enrolled in the study were divided into three bridging therapy groups: the unfractionated heparin(UFH)group(n=109),the low-molecular-weight heparin(LMWH)group(n=97),and the UFH with sequential LMWH(UFH-LMWH)group(n=99).All patients were followed for 4 weeks.2.Impact of genetic and clinical factors on warfarin therapy in patients after heart valve replacement surgeryIn this observational study,289 patients starting warfarin therapy early after heart valve replacement surgery were enrolled.VKORC1-1639 G>A(rs9923231),CYP2C9*2(rs1799853),CYP2C9*5 C1080G(rs28371686),CYP2C9*3(rs105791 0),CYP2C19*2(rs4244285),CYP4F2 V433M(rs2108622),GGCX(rs11676382),ABCB1 C3425T(rs1045642)and ABCG2 C421A(rs2231142)genotypes,clinical characteristics,response to therapy,and bleeding and thrombosis events were collected.Patients were followed at least 30 days.The primary outcomes were the time to the first INR equal to or more than lower limit of therapeutic range(?LLTR)and the warfarin dose requirements.The second outcome were First INR?LLTR,first INR?3.5 and bleeding and thrombosis events.Time needed to reach the first INR?LLTR were compared among groups with the use of log-rank tests and cox regression analyses were performed to determine the hazard ratios.Logistic regression analyses were performed to determine the odds ratio for first INR?3.5.Patients were followed3 months and INRs were recorded.TTR were calculated and logistic regression was used to analyze the factors influencing TTR and their OR values.The maintenance dose of the patients was obtained during 1-year follow-up.The genes and clinical factors that significantly affect MD were analyzed by univariate and multivariate analysis.Stepwise multiple linear regression was performed to develop dosing algorithms to predict the warfarin dose requirement for reaching INR?LLTR and MD respectively.The two models were compared.3.Effect of cefoperazone-sulbactam on pharmacokinetics and pharmacodynamics of warfarin and preliminary mechanism study on the cefoperazone-sulbactam and warfarin interaction.In long-term pharmacodynamic experiments,Wistar rats were continuously treated with warfarin(0.125 mg/kg qd)for 15 days to make INR stable in the therapeutic range.From day 16-19,the rats were divided into single use and combination groups.The rats in the single use group were given warfarin only,and the rats in the combination group were intraperitoneally injected with cefoperazone-sulbactam(0.3 g/kg bid)immediately after administration of warfarin.The changes of INRs between the two groups were compared within 4 days.In the pharmacokinetic and tissue distribution experiments,the rats were divided into two groups: the single use group was intraperitoneally injected with normal saline(0.1ml/100g)twice a day for 4 days,and the combination group was pretreated with cefoperazone-sulbactam(0.3 g/kg bid)for 4 days.During this period,the changes of plasma vitamin K1 and K2 contents in the combination group were measured.On the fifth day,the two groups were divided into the low dose and high dose groups.The low dose single use group was given warfarin 0.125 mg/kg by intragastric administration,and the high dose single use group was given warfarin 0.5 mg/kg.In low dose and high dose combination groups,cefoperazone-sulbactam 0.3 g/kg was injected intraperitoneally immediately after administration of 0.125 mg/kg and 0.5mg/kg warfarin respectively.Blood samples were collected at different time points after femoral artery intubation,and the plasma concentration of warfarin was determined.The INR values were measured at different time points.The contents of warfarin and S/R-warfarin were determined by LC/MS/MS,and the pharmacokinetic parameters were calculated by non-compartment model.The liver of rats in low dose single use group and combination group was collected at different time points,and the ileum,kidney,spleen,lung and pancreas were also collected at 2h after administration.The content of S/R-warfarin in each tissue was determined.the expression of P-gp in liver and ileum was determined by Western blotting method,and the expression of Oat2 mRNA in liver was determined by RT-qPCR.For exploring the mechanism of impact of cefoperazone-sulbactam on hepatic uptake of warfarin,the uptake of R/S warfarin in HepaRG cells cultured with 1 ?M warfarin was compared with that in HepaRG cells cultured with 1 ?M warfarin and350 ?M cefoperazone-sulbactam at different time points.ResultsTwo patients experienced thromboembolic stroke events in the UFH group.The LMWH group was associated with an increase in the incidence of bleeding events compared with the UFH group(10.3% vs 2.8%,P=0.03).With a comparison of LMWH and UFH group in secondary endpoints,LMWH group indicated a trend of reduced ICU length of stay(P=0.08),postoperative length of stay(P=0.08)and time of achieving target INR(P=0.06).The creatinine level [(odds ratio 1.03,95%confidence interval(1.01 to 1.05,P=0.02)] and hypertension [(odds ratio 3.72,95%confidence interval(1.35 to 10.28,P=0.01)] were risk factors for bleeding events.The results of univariate analysis showed lone VKORC1-1639 G>A,CYP2C9*1/*3,cefazolin,cefoperazone-sulbactam,increased BMI,?hemoglobin,and white blood cell count could significantly affect patient responsiveness to warfarin in the initial period of anticoagulation.Multivariate analysis resulted in an equation:Accumulated warfarin doses(mg)=14.376 VKORC1-1639G>A–2.865 hypertension+0.468 BMI–0.108 age+2.366 ABCB1 CT/TT–4.050 CYP2C9 *1/*3–4.843cefoperazone-sulbactam–0.062 ?Hb–2.530 cefazolin+0.323 pre-albumin–16.200,which could explain 43.9% of the variability in warfarin dose needed to reach the first INR?LLTR.The results of external validation showed that there was a strong correlation between predicted and observed warfarin accumulated doses(Pearson r=0.6404,P<0.0001).VKORC1 GA/GG,cefoperazone-sulbactam and amiodarone significantly affected the INR values of reaching first INR?LLTR.Patients with cefoperazone-sulbactam had a higher risk of first INR?3.5 than those who did not take cefoperazone-sulbactam [OR 1.565,95% CI(1.156-2.119),P=0.004)].Age is another independent influencing factor [OR 1.082,95% CI(1.035-1.130),P<0.001].During the follow-up period,patients with VKORC1 AA had a higher risk of first INR?3.5 than patients with VKORC1 GA/GG [OR 5.487,95% CI(1.223-24.621),P=0.026)].Univariate analysis showed that age,BMI,amiodarone,VKORC1-1639 G>A and CYP2C9 *1/*3 were significantly correlated with MD.MD(mg/day)=1.436VKORC1-1639 G>A-0.967 CYP2C9 *1/*3-0.043 amiodarone+0.033 plasma albumin-0.010 age + 0.032 BMI+0.688,this algorithm can explain the variation of 40.2%MD.Among them,VKORC1-1639G>A accounts for 27.8%,and the proportion is the largest.Age and concomitant use of cefazolin were risk factors for TTR <70%.After INR got to the target range,cefoperazone-sulbactam was added.Theincrease of INR on the first day of combination was statistically significant,and gradually increased.With the increase of cefoperazone-sulbactam pretreatment days,the contents of plasma vitamin K1 and K2 decreased gradually,and there was significant difference on the 4th day.The results of pharmacokinetic experiment showed that compared with low dose single group,the AUC0-t and Cmax of warfarin and S-warfarin in the low dose combination group were significantly higher,while the CL/F was significantly decreased;The value of INR in the combination group significantly increased 8 hours after adminstration;The expression of P-gp in ileum and liver was significantly decreased in the combination group,and the expression of Oat2 mRNA in the liver in the combination group was significantly lower;The liver distribution of S-warfarin in the combination group was significantly lower at 2 h,6 h and 10 h;The distribution of warfarin enantiomers in ileum and kidney at 2 h in the combination group was significantly higher.When cultured with cefoperazonesulbactam and warfarin at different time points,the uptake of warfarin enantiomers in HepaRG cells was significantly lower than that in warfarin cultured alone.ConclusionFor MHVR patients,LMWH bridging anticoagulant therapy helps patients to achieve anticoagulant effect in the early stage but increases the incidence of bleeding events.There was no embolic event in the UFH-LMWH group,and there was no difference in the incidence of bleeding between the UFH-LMWH group and the UFH group,so UFH-LMWH were more easily accepted among the three groups.At the initial stage of warfarin treatment,both genetic(VKORC1,CYP2C9 and ABCB1 genotype and clinical factors(age,BMI,antibiotics,hypertension,?Hb and plasma albumin level)affect the anticoagulant effect of warfarin.The factors of influencing initial stage and maintenance period of warfarin are different,and the proportion of the same factors is different,so it is not suitable to directly use MD as the initial dose.Age and cefazolin affect anticoagulant stability,and genes seem to have little contribution.Cefoperazone-sulbactam can quickly and significantly affect the efficacy of warfarin,which related to reducing the synthesis of vitamin K by cefoperazonesulbactam and may be associated with increasing the absorption rate and degree of warfarin by its inhibiting the expression of P-gp,reducing the distribution and clearance of S-warfarin in the liver by its inhibiting the expression of Oat2 mRNA.