Effect Of Glycyrrhizin On CYP450s Activites And Its Molecular Mechanisms

BACKGROUND:Cytochrome P450s superfamily expressed widely in organisms are known to play an important role in the biotransformation of many endogenous and exogenous substances. Inhibition or induction of cytochrome P450 isozymes is one of the major causes for clinical drug-drug interactions. Licorice and glycyrrhizin is widely used in the treatment of various diseases such as chronic hepatitis. Although licorice and glycyrrhizin are widely used, there is little study on its effects on CYP450 in human beings. To investigate medicines effect on CYP450s activities has important clinical significance, it helps with clinical rational administration and reducing the drug adverse reaction inducing by drug interactions. Glycyrrhizin obviously affects the activities of CYP450s in rat and mouths. However, as our knowledge,This is the first study to investigate the effects of glycyrrhizin on CYP450s in vivo.OBJECTIVE:This study was designed to investigate the effects of glycyrrhizin on CYP3A, CYP1A2, CYP2E1, CYP2D6 activities in humans. The effect of glycyrrhizin on CYP activity in vivo was assessed by a four-drug cocktail approach useing CYP probe drugs midazolam(CYP3A), caffeine(CYP1A2), chlorzoxazone(CYP2E1) and metoprolol(CYP2D6).METHODS:Twelve healthy adult men were enrolled in a 2-phase randomized crossover design. In each phase the volunteers received placebo or glycyrrhizin for 14 days.Then probe drug cocktails of midazolam, caffeine, chlorzoxazone and metoprolol were administered to determine in vivo CYP activities.RESULTS:Glycyrrhizin administration significantly increased 1-hydroxymidazolam/midazolam serum ratios (1 hour sample) (p=0.021), Conversely, the plasma paraxanthine/caffeine ratios and 6-hydroxychlorzoxazone/chlorzoxazone plasma ratios were both significantly decreased by glycyrihizin treatment compared to placebo treatment (p=0.004, p=0.021).14 days glycyrrhizin treatment did not alter the urinary metoprolol/a-hydroxymetoprolol ratio (the index of CYP2D6 activity) (p= 0.248).CONCLUSIONS:By utilizing single time-point phenotypic ratios, administration of glycyrrhizin resulted in an induction of CYP3A and a significant inhibit of CYP1A2 and CYP2E1 activity, and showed no significant effects on CYP2D6. BACKGROUND:Glycyrrhizin is a major ingredient of licorice which is widely used in the treatment of various diseases such as chronic hepatitis. Licorice or glycyrrhizin have been shown to alter the activity of CYP3A in rodents. The influence of glycyrrhizin on CYP3A has not been elucidated in humans. This is the first report showing the effects of glycyrrhizin on CYP3A enzyme activity in humans.OBJECTIVE:To investigate the effects of repeated glycyrrhizin ingestion on the oral pharmacokinetics of midazolam, a probe drug for CYP3A activity in humans. METHODS:Sixteen healthy adult male subjects were enrolled in a 2-phase randomized crossover design. In each phase the volunteers received placebo or glycyrrhizin for 14 days. On the 15th day, midazolam was administered as a probe drug to determine in vivo CYP3 A activity in vivo.RESULTS:Glycyrrhizin administration decreased midazolam maximum plasma concentration(Cmax) by 12.4% (p<0.01; 95%CI:-28.86 to 4.03) and midazolam AUC0-∞by 20.2% (p<0.05; 95%CI:-31.76 to-8.66).Conversely, 1'-hydroxymidazolam Cmax was increased by 19.1% (p<0.01; 95%CI:10.6 to 27.7) and 1'-hydroxymidazolam AUC0-∞was increased by 20.3% (p<0.01; 95%CI:11.0 to 29.6) with glycyrrihizin. For AUC0-∞and Cmax of midzolam, the 90% CI for the geometric mean ratio of glycyrrhizin over placebo were both out of the no-effect boundaries of 0.80-1.25.CONCLUSIONS:Administration of glycyrrhizin resulted in a modest induction of CYP3A with clinically relevant according to the bioequivalence analysis.BACKGROUND:Traditional medicine licorice has been used as a medicinal plant for thousands of years.The active component of licorice, glycyrrhizin, is hydrolyzed in vivo to glycyrrhetinic acid, which is responsible for most of its pharmacological properties. Our previous clinical trial has shown that glycyrrhizin administration exerted inhibitive effects on the activities of CYP1A2 and CYP2E1, and inductive effect on CYP3A activity. However, the mechanism for glycyrrhizin's above effects has not been clarified.OBJECTIVE:To find out whether glycyrrhizin and glycyrrhetic acid have effect on CYPs activities including CYP1A2, CYP3A4, CYP2C19, CYP2E1, and CYP2D6 in pooled human liver microsomes.METHODS:The in vitro effects of glycyrrhizin and glycyrrhetic acid on CYP1A2 (caffeine deethylation), CYP3A (midazolam 1-hydroxylation), CYP2C19 (omeprazole 5'-hydroxylation), CYP3A4(omeprazole sulfoxidation), CYP2D6 (metoprololα-hydroxylation), and CYP2E1 (chlorzoxazone 6-hydroxylation) activities were examined using pooled human liver microsomes. The concentrations of the probe drugs and their metabolites were determined by HPLC and LC-MS/MS.RESULTS:With concentrations up to 200μM, Glycyrrhizin showed no appreciable effect on CYP1A2, CYP3A, CYP2C19,CYP3A4, CYP2E1, and CYP2D6 activities. Glycyrrhetic acid potent inhibited CYP3A(midazolam 1-hydroxylation) activity with a IC50 value of 32.94±9.51μM. Glycyrrhetic acid exhibited somewhat smaller inhibitory effects on CYP2C19 (omeprazole 5'-hydroxylation) and CYP3A4(omeprazole sulfoxidation) activities, with IC50 values of 176.35±22.46μM and 141.28±22.46μM, respectively. Km and Vmax for midazolam 1'-hydroxylation by microsomes were 3.59±0.85μM and 2.78±0.18 nmol/min.mg, respectively. Formation of 1'-OH MDZ by CYP3A4 was noncompetitively inhibited by glycyrrhetic acid (GA), with a Ki of 7.38±0.93μM.CONCLUSIONS:.Glycyrrhizin showed no effect on CYP1A2, CYP3A, CYP2C19, CYP3A4, CYP2E1, and CYP2D6 activities in human liver microsomes. Glycyrrhetic acid noncompetitively inhibits Formation of 1'-OH MDZ by CYP3A4, and also inhibit metabolism of omeprazole to 5'-Hydroxyomeprazole by CYP2C19 and inhibit metabolism of omeprazole to omeprazole sulfone by CYP3A4.BACKGROUND:Research has found that licorice(Gan Cao) could induce total CYP450s in rodents. And previous reports have demonstrated that licorice(Gan Cao) total extracts activated PXR and induced the expression of drug-metabolism, however, the constituents responsible for licorice-mediated CYP450s induction and the underlying molecular mechanisms remains unknown. Our previous work has demonstrated that administration of glycyrrhizin in healthy males resulted in an increased metabolism of Midazolam, By contrast, glycyrrhetic acid presented play an inhibitive action in metabolism of midazolam in pooled human liver microsomes. We also observed inhibition metabolism of omeprazole to 5'-Hydroxyomeprazole by CYP2C19 in human liver microsomes. The discovery of a family of nuclear receptors such as pregnane X recepor (PXR) and constitutive androstane receptor(CAR) gives insight into the molecular explanation of CYP3A induction by glycyrrhizin. In the present study, interactions between glycyrrhizin and glycyrrhetic acid and human CAR and PXR were evaluated using a reporter gene assay.OBJECTIVE:In our study, we tested the hypothesis whether glycyrrhizin or glycyrrhetic acid in therapeutic concentrations has potential to affect expression of CYP3A4 and CYP2C19 via constitutive androstane receptor(CAR) and pregnane X receptor(PXR) pathways.METHODS:Interaction of glycyrrhizin and glycyrrhetic acid with CAR and PXR nuclear receptors was studied using luciferase reporter assays, real-time reverse transcriptase chain reaction (RT-PCR), and analysis of CYP3A4 catalytic activity.RESULTS:Using transient transfection reporter assays in Caco2 cells, Glycyrrhetic acid was recognized to activate CYP3A4 promoter via CAR and PXR pathways. A significant effect of glycyrrhetic acid on CYP3A4 promoter activation in HepG2 cells was observed only by CAR pathway. Glycyrrhizin showed no impact on CYP3A4 promotor activation by CAR or PXR pathways. Neither glycyrrhizin nor glycyrrhetic acid have impacts on CYP2C19 promoter via CAR or PXR pathways. These data well correlated with up-regulation of CYP3A4 mRNA analyzed by real-time RT-PCR in cells with expression vectors encoding CAR or PXR and treated with glycyrrhetic acid. In addition, analysis of specific CYP3A4 catalytic activity revealed its significant increase in glycyrrhetic acid-treated LS174T transfected with CAR.CONCLUSIONS:.In conclusions,we provide novel insight into the mechanism by which GA affects gene expression of CYP3A4. Our results demonstrate that GA has potential to up-regulate CYP3A4 through direct activation of CAR and/or PXR pathways.OBJECTIVE:To investigate the interaction between glycyrrhizin and omeprazole and observe the effects of glycyrrhizin on CYP2C19 and CYP3A4 activities in healthy Chinese male volunteers with different CYP2C19 genotypes..METHODS:Eighteen healthy subjects (six CYP2C19*1/*1, five CYP2C19*1/*2, one CYP2C19*1/*3, five CYP2C19*2/*2 and one CYP2C19*2/*3) were enrolled in a two-phase randomized crossover trial. In each phase, all subjects received placebo or glycyrrhizin salt tablet 150mg twice daily for 14 consecutive days. The pharmacokinetics of omeprazole(20mg orally on day 15) was determined for up to 12h following administration by HPLC.RESULTS:After 14-day treatment of glycyrrhizin, plasma omeprazole significantly decreased, and those of omeprazole sulfone significantly increased. However, plasma concenetrations of 5-hydroxyomeprazole did not significantly change. The ratio of AUC0-∞of omeprazole to omeprazole sulfone decreased by 43.93±13.56%(p=0.009) in CYP2C19*1/*1,44.85±14.84%(p=0.002) in CYP2C19*1/*2or*3 and 36.16±7.52%(p＜0.001) in CYP2C19*2/*2 or*3 while those of omeprazole to 5-hydroxyomeprazole did not change significantly in all three genotypes. No significant differences in glycyrrhizin response were found among CYP2C19 genotypes..CONCLUSIONS:Glycyrrhizin induces CYP3A4-catalyzed sulfoxidation of omeprazole and leads to decreased omeprazole plasma concentrations, but has no significant impact on CYP2C19-dependent hydroxylation of omeprazole.