Influence Of Genetic Polymophism On MPA Pharmacokinetics And Impdh Activity In Chinese Renal Transplant Recipients

Mycophenolic acid(MPA)is a nonnucleoside,uncompetitive and reversible inosine monophosphate dehydrogenase(IMPDH)inhibitor,which participates in guanosine triphosphates de novo synthesis.Mycophenolate mofetil(MMF)and enteric-coated mycophenolate sodium(EC-MPS)are the dose form of MPA,which are the most widely used antiproliferative immunosuppressive drug in patients after solid organ transplantation.We established an LC-MS/MS method for the determination of IMPDH acitivity of Chinese renal transplant recipients.Firstly,peripheral blood mononuclear cells(PBMC)were separated from whole blood of renal transplant patients.Enzymatic reaction catalysed by IMPDH was carried out and solid phase extraction was used to purify the incubated PBMC samples.The formed XMP and AMP were determined.The latter was used for the estimation quantity of PBMC added.The extracted incubated resultant was eluted through Agilent Eclipse XDB-C18(5?m,4.6×150 mm)column by water(2 mmol·L-1 ammonium acetate,PH=8)and methanol(2 mmol L-1 ammonium acetate)through a gradient elution.The flow rate was set as 0.4 mL min-1.Electrospray ionization(ESI)source was applied and operated in the positive ion mode.Multiple reaction monitoring(MRM)mode with the transition of m/z 365.2?97.3(XMP),348.4?136.1(AMP)and 427.7?216.1(Br-AMP)was used for quantification.The method was proved to be linear in the range of 0.2-20?mol·L-1 for XMP,5-100?mol·L-1 for AMP(r2>0.99).The within-day and between-day variation were both lower than 15%.The extract recovery,matrix effect,process recovery of XMP and AMP was 22.9%-33.6%,94.8%-102.6%,23.0%-32.6%,respectively.The LC-MS/MS method we established was rapid,sensitive,precise and reliable for the determination of IMPDH in PBMC,which can be applied to study pharmacokinetic of MPA.We further studied MPA and its metabolites plasma concentration and IMPDH activity of renal transplant recipients 0-12 hours after administration of MMF or EC-MPS under steady state.The main pharmacokinetic and pharmacodynamic parameters were determined.IMPDH activity decreased with increasing MPA and mycophenolic acid acyl glucuronide(AcMPAG)plasma concentration,with maximum inhibition coinciding with maximum MPA concentration.AcMPAG AUC0-12h,and AEC0-12h,of the EC-MPS combined with cyclosporine A(CsA)group was higher than of MPA combined with TAC group.MPA concentration of 74 renal transplant patients was determined by enzyme multiplied immunoassay technique 0-12 hours after administration of MMF or EC-MPS under steady state.DNA of patients were extracted and genotypes of 11 SNPs of ABCB1,ABCC2,SLCO1B1,SLCO1B3,IMPDH ?and IMPDH ? were determined by a SNaPshot assay.According to the Hardy-Weinberg equilibrium equation,none of the observed frequencies of were significantly different from expected values.Renal transplant recipients with T carriers of ABCC2 C24T were associated with reduced MPA AUC under both of MMF and EC-MPS therapy(P=0.060,P=0.002).Genetic polymorphism of ABCC2 C24T and SLCO1B3 T334G was probably related to enterohepatic circulation(EHC).IMPDH ? G125A and IMPDH ? T3757C mutation was related to the increased AEC0-12h,which can explain the individual variation of IMPDH activity.