ROS Related Genes Polymorphisms And Susceptibility To VPA-induced Liver Injury By SNaPshot Assay

The incidence of drug-induced liver injury(DILI)has an increasing tendency year by year.More than 1100 kinds of chemical drugs or biological drugs have a potential to cause liver injury,which is one of the most important potential threats to human health.Valproic acid(VPA)is the most widely used antiepileptic drug in clinical practice.It is mainly metabolized by liver.There is poor association between VPA serum concentration and maintenance dose.Liver injury is one of the main side effects of VPA with narrow therapy window.Metabolic activation of the drug to a reactive intermediate is an important step in the generation of the idiosyncratic reaction for drug-induced liver injury.In the case of VPA,this is illustrated by cytochrome P450-catalyzed metabolism of VPA to highly reactive metabolites,4-ene VPA and 2,4-diene VPA,which may be involved in the inhibition of mitochondrial fatty acid ?-oxidation,the enhancement of reactive oxygen species(ROS)generation,the accumulation of triglycerides and the pathogenesis of VPA-induced hepatotoxicity.It causes a variety of liver diseases including inflammation,hepatic steatosis and liver injury.The underlying mechanism responsible for VPA hepatotoxicity has been postulated to involve reactive metabolites of VPA and oxidative stress.Due to the unpredictability of liver injury caused by VPA,it is imperative for us to find markers for predicting VPA-induced liver injury.Recent studies suggest that genetic factors contribute to VPA-induced liver injury.A large number of researches focused on exploring the associations between genetic polymorphisms of VPA-metabolizing enzymes and liver injury.Although oxidative stress plays an important role in DILI,little is known about ROS-detoxifying enzyme genotypes and VPA-associated hepatotoxicity.The aim of this study was to comprehensively investigate the associations of glutathione S-transferase P1(GSTP1),NAD(P)H oxime reductase(NQO1),metal thioprotease(MT2A),paraoxonase1(PON1)and glutathione peroxidase1(GPX1)genetic polymorphisms with VPA-induced hepatotoxicity.Firstly,we established a SNaPshot method by multiple-step optimums,which may simultaneously analyze seven SNPs including GSTP1 rs1695,NQO1 rs1800566,MT2 A rs10636,PON1 rs662 or rs854560,and GPX1 rs1050450 or rs1800668.Then we explored the associations between the 7 SNPs and VPA-induced live injury in 96 patients with epilepsy.No significant differences at the genotype frequencies of the 7 SNPs were observed between normal liver function group and abnormal liver function group.To further explore the effects of the 7 SNPs on VPA metabolism,we analyzed the associations of the 7 SNPs with VPA maintenance doses,serum concentrations,and dose-adjusted serum concentrations.The results showed that patients carrying the mutant genotypes of GPX1 rs1800668 had an increased dose-adjusted VPA serum concentration relative to those carrying the wild-type genotypes of GPX1 rs1800668 in normal liver function group(P = 0.035),while patients carrying the mutant genotypes of PON1 rs854560 had a decreased VPA serum concentration(P = 0.038).Otherwise,GSTP1 rs1695 mutation significantly decreased the VPA serum concentration in normal liver function group(P = 0.043).It suggests that GPX1 rs1800668,PON1 rs854560 or GSTP1 rs1695 have influences on VPA metabolism.Due to a limited sample size,the association needs to be further investigated in a larger cohort of patients.Because there were gender-associated differences on VPA-induced liver injury,we next explored the associations of the antioxidant enzymes gene polymorphisms and VPA-induced liver injury after stratifying by gender.Likewise,no significant differences at the genotype frequencies of the 7 SNPs were observed between normal liver function group and abnormal liver function group.Interestingly,the frequencies of MT2 A rs10636 homozygous mutation in female patients with epilepsy was significantly higher than those of MT2 A rs10636 wild-type and heterozygous mutation(P = 0.000 and 0.034,respectively).In conclusion,we successfully established a SNaPshot method for simultaneously analyzing 7 SNPs of GPX1,GSTP1,NQO1,MT2 A,and PON1.No significant associations of these antioxidant enzymes gene polymorphisms and VPA-induced liver injury were observed in patients with epilepsy,even after stratifying by gender.Concerning the effects of these antioxidant enzymes SNPs on VPA metabolism,the results need to be verified in a larger cohort of patients.