Genetic diversity of cytochrome P450 3A with different metabolic activity in domestic cats.
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ABSTRACT:
SUBMITTER: Sugiyama S
PROVIDER: S-EPMC6483910 | biostudies-literature | 2019 Apr
REPOSITORIES: biostudies-literature
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Project description:Knowledge of genetic polymorphisms of metabolizing enzymes of medical drugs and xenobiotics including cytochrome P450 (CYP) is very limited in cats. We investigated polymorphisms in CYP1A2, one of the major CYP isoforms in the feline liver. Wild-type and three non-synonymous polymorphic variants, but no synonymous variant, were identified in feline CYP1A2 in 50 alleles of domestic cats in Japan. Metabolic parameters, Km and Vmax, of ethoxyresorufin hydroxylation by CYP1A2 were shown to range within two times for identified non-synonymous variants by using a heterologous coexpression system. The results confirmed the polymorphic nature of CYP1A2 as a basis for effective application of medicines and prevention of adverse reactions in the treatment of domestic cats as well as for hereditary disorders.
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Project description:Knowledge of genetic polymorphisms of cytochrome P450 (CYP), the most important xenobiotic metabolizing enzyme, is very limited in cats. Preliminarily, we investigated genetic polymorphisms in CYP2A13, one of the major CYP isoforms in the liver and lung. Four synonymous and three non-synonymous polymorphic variants were identified in feline CYP2A13 in domestic cats in Japan, without an obvious major type. Metabolic parameters, Km and Vmax, of coumarin hydroxylation of CYP2A13 were shown to range within two times for the identified non-synonymous polymorphic variants by using heterologous coexpression system in Escherichia coli. The results confirmed the polymorphic nature of CYP2A13 as a basis for effective application of medicines and prevention of adverse reactions in treatment of domestic cats.
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Project description:Background and Aim: Liver cirrhosis is associated with decreased hepatic cytochrome P4503A (CYP3A) activity but the pathogenesis of this phenomenon is not well elucidated. In this study, we examined if certain microRNAs (miRNA) are associated with decreased hepatic CYP3A activity in cirrhosis. Hepatic CYP3A activity and miRNA microarray expression profiles were measured in cirrhotic (n=22) and normal (n=12) liver tissue. Hepatic CYP3A activity was measured via midazolam hydroxylation in human liver microsomes. Additionally, hepatic CYP3A4 protein concentration and the expression of CYP3A4 mRNA were measured. Analyses were conducted to identify miRNAs which were differentially expressed between two groups but also were significantly associated with lower hepatic CYP3A activity. Liver tissue was collected from individuals with established cirrhosis who were awaiting liver transplantation (n=22) at the time of their liver transplantation procedure in the operating room. Normal liver tissue (n=12) was obtained from patients undergoing liver resection for metastatic/benign liver lesions with no underlying chronic liver disease
2013-08-15 | E-GEOD-49012 | biostudies-arrayexpress
Project description:Background and Aim: Liver cirrhosis is associated with decreased hepatic cytochrome P4503A (CYP3A) activity but the pathogenesis of this phenomenon is not well elucidated. In this study, we examined if certain microRNAs (miRNA) are associated with decreased hepatic CYP3A activity in cirrhosis. Hepatic CYP3A activity and miRNA microarray expression profiles were measured in cirrhotic (n=22) and normal (n=12) liver tissue. Hepatic CYP3A activity was measured via midazolam hydroxylation in human liver microsomes. Additionally, hepatic CYP3A4 protein concentration and the expression of CYP3A4 mRNA were measured. Analyses were conducted to identify miRNAs which were differentially expressed between two groups but also were significantly associated with lower hepatic CYP3A activity.
2013-08-15 | GSE49012 | GEO
Project description:Sunitinib is an orally administered tyrosine kinase inhibitor associated with idiosyncratic hepatotoxicity; however, the mechanisms of this toxicity remain unclear. We have previously shown that cytochromes P450 1A2 and 3A4 catalyze sunitinib metabolic activation via oxidative defluorination leading to a chemically reactive, potentially toxic quinoneimine, trapped as a glutathione (GSH) conjugate (M5). The goals of this study were to determine the impact of interindividual variability in P450 1A and 3A activity on sunitinib bioactivation to the reactive quinoneimine and sunitinib N-dealkylation to the primary active metabolite N-desethylsunitinib (M1). Experiments were conducted in vitro using single-donor human liver microsomes and human hepatocytes. Relative sunitinib metabolite levels were measured by liquid chromatography-tandem mass spectrometry. In human liver microsomes, the P450 3A inhibitor ketoconazole significantly reduced M1 formation compared to the control. The P450 1A2 inhibitor furafylline significantly reduced defluorosunitinib (M3) and M5 formation compared to the control but had minimal effect on M1. In CYP3A5-genotyped human liver microsomes from 12 individual donors, M1 formation was highly correlated with P450 3A activity measured by midazolam 1'-hydroxylation, and M3 and M5 formation was correlated with P450 1A2 activity estimated by phenacetin O-deethylation. M3 and M5 formation was also associated with P450 3A5-selective activity. In sandwich-cultured human hepatocytes, the P450 3A inducer rifampicin significantly increased M1 levels. P450 1A induction by omeprazole markedly increased M3 formation and the generation of a quinoneimine-cysteine conjugate (M6) identified as a downstream metabolite of M5. The nonselective P450 inhibitor 1-aminobenzotriazole reduced each of these metabolites (M1, M3, and M6). Collectively, these findings indicate that P450 3A activity is a key determinant of sunitinib N-dealkylation to the active metabolite M1, and P450 1A (and potentially 3A5) activity influences sunitinib bioactivation to the reactive quinoneimine metabolite. Accordingly, modulation of P450 activity due to genetic and/or nongenetic factors may impact the risk of sunitinib-associated toxicities.
| S-EPMC9131896 | biostudies-literature
Project description:In the present study, the validity of using a cocktail screening method in combination with a chemometrical data mining approach to evaluate metabolic activity and diversity of drug-metabolizing bacterial Cytochrome P450 (CYP) BM3 mutants was investigated. In addition, the concept of utilizing an in-house-developed library of CYP BM3 mutants as a unique biocatalytic synthetic tool to support medicinal chemistry was evaluated. Metabolic efficiency of the mutant library towards a selection of CYP model substrates, being amitriptyline (AMI), buspirone (BUS), coumarine (COU), dextromethorphan (DEX), diclofenac (DIC) and norethisterone (NET), was investigated. First, metabolic activity of a selection of CYP BM3 mutants was screened against AMI and BUS. Subsequently, for a single CYP BM3 mutant, the effect of co-administration of multiple drugs on the metabolic activity and diversity towards AMI and BUS was investigated. Finally, a cocktail of AMI, BUS, COU, DEX, DIC and NET was screened against the whole in-house CYP BM3 library. Different validated quantitative and qualitative (U)HPLC-MS/MS-based analytical methods were applied to screen for substrate depletion and targeted product formation, followed by a more in-depth screen for metabolic diversity. A chemometrical approach was used to mine all data to search for unique metabolic properties of the mutants and allow classification of the mutants. The latter would open the possibility of obtaining a more in-depth mechanistic understanding of the metabolites. The presented method is the first MS-based method to screen CYP BM3 mutant libraries for diversity in combination with a chemometrical approach to interpret results and visualize differences between the tested mutants.
| S-EPMC4723632 | biostudies-literature
Project description:Tacrolimus (Tac) is an effective remission inducer of refractory ulcerative colitis (UC). Gene polymorphisms result in interindividual variability in Tac pharmacokinetics. In this study, we aimed to examine the relationships between gene polymorphisms and the metabolism, pharmacokinetics, and therapeutic effects of Tac in patients with UC. Forty-five patients with moderate-to-severe refractory UC treated with Tac were retrospectively enrolled. Genotyping for cytochrome P450 (CYP) 3A4*1G, CYP3A5*3, CYP2C19*2, CYP2C19*3, nuclear receptor subfamily 1 group I member 2 (NR1I2)-25385C>T, ATP-binding cassette subfamily C member 2 (ABCC2)-24C>T, ABCC2 1249G>A, and ABCC2 3972C>T was performed. Concentration/dose (C/D) ratio, clinical therapeutic effects, and adverse events were evaluated. The C/D ratio of Tac in UC patients with the CYP3A4*1G allele was statistically lower than in those with the CYP3A4*1/*1 allele (P = 0.005) and significantly lower in patients with CYP3A5*3/*3 than in those with CYP3A5*1 (P < 0.001). Among patients with the CYP3A4*1G allele, the C/D ratio was significantly lower in patients with CYP3A5*1 than in those with CYP3A5*3/*3 (P = 0.001). Patients with the NR1I2-25385C/C genotype presented significantly more overall adverse events than those with the C/T or T/T genotype (P = 0.03). Although CYP3A4*1G and CYP3A5*3 polymorphisms were related to Tac pharmacokinetics, CYP3A5 presented a stronger effect than CYP3A4. The NR1I2-25385C/C genotype was related to the overall adverse events. The evaluation of these polymorphisms could be useful in the treatment of UC with Tac.
| S-EPMC8062093 | biostudies-literature
Project description:Background and aimLiver cirrhosis is associated with decreased hepatic cytochrome P4503A (CYP3A) activity but the pathogenesis of this phenomenon is not well elucidated. In this study, we examined if certain microRNAs (miRNA) are associated with decreased hepatic CYP3A activity in cirrhosis.MethodsHepatic CYP3A activity and miRNA microarray expression profiles were measured in cirrhotic (n=28) and normal (n=12) liver tissue. Hepatic CYP3A activity was measured via midazolam hydroxylation in human liver microsomes. Additionally, hepatic CYP3A4 protein concentration and the expression of CYP3A4 mRNA were measured. Analyses were conducted to identify miRNAs which were differentially expressed between two groups but also were significantly associated with lower hepatic CYP3A activity.ResultsHepatic CYP3A activity in cirrhotic livers was 1.7-fold lower than in the normal livers (0.28 ± 0.06 vs. 0.47 ± 0.07mL* min(-1)*mg protein(-1) (mean ± SEM), P=0.02). Six microRNAs (miR-155, miR-454, miR-582-5p, let-7f-1*, miR-181d, and miR-500) had >1.2-fold increase in cirrhotic livers and also had significant negative correlation with hepatic CYP3A activity (range of r = -0.44 to -0.52, P <0.05). Notably, miR-155, a known regulator of liver inflammation, had the highest fold increase in cirrhotic livers (2.2-fold, P=4.16E-08) and significantly correlated with hepatic CYP3A activity (r=-0.50, P=0.017). The relative expression (2(-ΔΔCt) mean ± SEM) of hepatic CYP3A4 mRNA was significantly higher in cirrhotic livers (21.76 ± 2.65 vs. 5.91 ± 1.29, P=2.04E-07) but their levels did not significantly correlate with hepatic CYP3A activity (r=-0.43, P=0.08).ConclusionThe strong association between certain miRNAs, notably miR-155, and lower hepatic CYP3A activity suggest that altered miRNA expression may regulate hepatic CYP3A activity.
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Project description:Physiological plasticity in a polluted system: A case of an uncultured bacterium and its cypome
| PRJEB38781 | ENA
Project description:Cytochrome P450 3A is the most important CYP subfamily in humans, and CYP3A4/CYP3A5 genetic variants contribute to inter-individual variability in drug metabolism. However, no information is available for bovine CYP3A (bCYP3A). Here we described bCYP3A missense single nucleotide variants (SNVs) and evaluated their functional effects. CYP3A28, CYP3A38 and CYP3A48 missense SNVs were identified in 300 bulls of Piedmontese breed through targeted sequencing. Wild-type and mutant bCYP3A cDNAs were cloned and expressed in V79 cells. CYP3A-dependent oxidative metabolism of testosterone (TST) and nifedipine (NIF) was assessed by LC-MS/MS. Finally, SNVs functional impact on TST hydroxylation was measured ex vivo in liver microsomes from individually genotyped animals. Thirteen missense SNVs were identified and validated. Five variants showed differences in CYP3A catalytic activity: three CYP3A28 SNVs reduced TST 6?-hydroxylation; one CYP3A38 variant increased TST 16?-hydroxylation, while a CYP3A48 SNV showed enhanced NIF oxidation. Individuals homozygous for rs384467435 SNV showed a reduced TST 6?-hydroxylation. Molecular modelling showed that most of SNVs were distal to CYP3A active site, suggesting indirect effects on the catalytic activity. Collectively, these findings demonstrate the importance of pharmacogenetics studies in veterinary species and suggest bCYP3A genotype variation might affect the fate of xenobiotics in food-producing species such as cattle.
| S-EPMC6927969 | biostudies-literature