The Effect Of The P450Oxidoreductase*28Polymorphism On Individual Variability Of Tacrolimus In Clinic

Objectives: In order to assess the influence of the P450oxidoreductase*28（POR*28）polymorphism on individual variability of tacrolimus pharmacokinetics and explore therole of POR*28polymorphism in tacrolimus population pharmacokinetic model, the studyanalyzed the pharmacokinetic characteristics of tacrolimus in Chinese healthy volunteersand renal transplant recipients. Additionally, we analyzed the association of the geneticpolymorphisms with the incidence of acute rejection after renal transplant, expecting toprovide reference for tacrolimus individualized therapy in clinic.Method:（1） Seventy-one healthy Chinese male volunteers enrolled in this studyreceived an oral dose of2mg of tacrolimus after providing written informed consent.Blood samples were collected immediately before and0.25,0.5,0.75,1,1.5,2,3,4,6,8,10,12and24h after drug administration. Tacrolimus whole blood concentrations weredetermined by ultra performance liquid chromatography-tandem mass spectrometry andthe pharmacokinetics analysis was evaluated by nonparametric methods.（2） Tacrolimus concentrations and clinical details were retrospectively collected from83renal transplant recipients （male,60; female,21） who received tacrolimus as the basicimmunosuppressive agent after the transplant,13（male,11; female,2） of whom sufferedfrom actue rejection （AR） within1year after renal transplant.（3） The tacrolimus pharmacokinetic parameters and the intra or interindividualvariabilities of83renal transplant recipients were estimated using NONMEM program.The covariates, including demographics, liver functions, concurrent drugs and thegenotypes of POR, CYP3A4and CYP3A5were evaluated quantitatively. Additionally, thestability of the final model was validated by a nonparametric bootstrap procedure. （4） CYP3A4*1G and CYP3A5*3were genotyped by polymerase chainreaction-restriction fragment length polymorphism （PCR-RFLP） and POR*28wasgenotyped by PCR-direct sequencing.Result:（1） The frequencies of the POR*28, CYP3A4*1G and CYP3A5*3allele were29.6%,22.5%,73.2%and39.8%,31.3%,66.3%in healthy Chinese volunteers and renaltransplant recipients respectively. There was no significant difference between the twogroups （P>0.05）.（2） The result of the effect of POR*28polymorphism on CYP3A-mediated tacrolimusmetabolism in71healthy volunteers showed that the CYP3A4*1G and CYP3A5*3polymorphisms but not POR*28polymorphism were closely related with the markedinterindividual variations of tacrolimus pharmacokinetics. However, subgroup analysisshowed the mean tacrolimus AUC0–24, AUC0-∞and C_maxfor the POR*28CTheterozygotes in CYP3A5expressers （CYP3A5*1allele carriers） were46.7±24.9ng.mL^-1.h^-1,57.4±33.9ng.mL^-1.h^-1and11.2±6.4ng.mL-1, which were much lower than thePOR*28CC homozygotes of71.5±38.9ng.mL^-1.h^-1,94.3±58.3ng.mL^-1.h^-1and17.6±9.8ng.mL-1respectively （P<0.05）. We found that the POR*28polymorphism had no effect ontacrolimus pharmacokinetics either in CYP3A5non-expressers or in differentCYP3A4*1G allele carriers.（3） A significant difference was found in C0/D of tacrolimus in initial stage betweendifferent POR*28genotypes in83renal transplant recipients. Furthermore, subgroupanalysis showed that the effect of POR*28SNP on the metabolism of tacrolimus existedonly in CYP3A5-expressing recipients. In CYP3A5-expressing recipients, the C0/D oftacrolimus was1.42-1.75and1.78-2.21fold higher in POR*28CC carriers than inPOR*28CT and TT carriers respectively （P<0.05）. Similar to the result of healthy subjects,we found that the POR*28polymorphism had no effect on tacrolimus pharmacokineticseither in CYP3A5non-expressing recipients or in different CYP3A4*1G allele carriers.（4）246tacrolimus trough concentrations obtained from83renal transplant recipientswere analyzed using NONMEM program. Minimum steady-state concentration method was considered as the best method for fitting the base model. The combined genotype ofCYP3A5-POR rather than the demographics （age, sex, weight and so on）, liver functions,concurrent drugs of the renal recipients was confirmed as the only covariant for theapparent clearance （CL/F） of tacrolimus. In the final model, the CL/F was23.3L.h-1fortotal83recipients,24.2L.h-1for CYP3A5*1-POR*28CC genotype which was1.62foldhigher than CYP3A5*1-POR*28T genotype （CL/F=19.3L.h-1, P<0.05） and1.62foldhigher than CYP3A5*3/*3genotype （CL/F=14.9L.h-1, P<0.01）. The interindividualvariability of CL/F was34.8%in basic model which decreased to26.3%in final model.The validation showed that the final model was stable.（5） The result of case-control study showed that the incidence of acute rejection （AR）within1year after renal transplant in CYP3A5*1/*1genotype was40.0%, which wassignificant higher than CYP3A5*3/*3genotype （8.1%, P<0.05）. The other geneticpolymorphisms had no effect on the incidence of AR.Conclusion:（1） The POR*28mutation could increase the metabolism of tacrolimusin both healthy subjects and renal transplant recipients who carried CYP3A5*1allele（CYP3A5expressers）. However, the polymorphism had no effect on tacrolimuspharmacokinetics either in CYP3A5non-expressing recipients or in different CYP3A4*1Gallele carriers. The CYP3A4*1G, CYP3A5*3and POR*28SNPs were closely related tothe marked interindividual variations of tacrolimus.（2） The combined genotype of CYP3A5-POR was associated with the CL/F oftacrolimus which could provide references for personalized use of tacrolimus in clinic.（3） The CYP3A5*3polymorphism might be associated with the incidence of ARwithin1year after renal transplant.