Genetic analysis of drug metabolizing phase-I enzymes CYP3A4 in Tibetan populations.

Science.gov (United States)

Liu, Lijun; Chang, Yu; Du, Shuli; Shi, Xugang; Yang, Hua; Kang, Longli; Jin, Tianbo; Yuan, Dongya; He, Yongjun

2017-06-01

The enzymatic activity of CYP3A4 results in broad interindividual variability in response to certain pharmacotherapies. The present study aimed to screen Tibetan volunteers for CYP3A4 genetic polymorphisms. Previous research has focussed on Han Chinese patients, while little is known about the genetic variation of CYP3A4 in the Tibetan populations. Here, we adopted DNA sequencing to investigate the promoter, exons and surrounding introns, and 3'-untranslated region of the CYP3A4 gene in 96 unrelated healthy Tibetan individuals.We identified 20 different CYP3A4 polymorphisms in the Tibetan population, including two novel variants (21824 A>G and 15580 G>C). In addition, we also determined the allele frequencies of CYP3A4*1A and CYP3A4*1H were 82.29% and 28.13%, respectively. CYP3A4*1P and *1G were relatively rare with frequencies of only 1.04% and 0.52%, respectively. Our results provide information on CYP3A4 polymorphisms in Tibetan individuals which may help to optimize pharmacotherapy effectiveness by providing personalized medicine to this ethnic group.

Microsomal cytochrome P450-3A4 (CYP3A4) nanobiosensor for the determination of 2,4-dichlorophenol-An endocrine disruptor compound

International Nuclear Information System (INIS)

Hendricks, Nicolette R.; Waryo, Tesfaye T.; Arotiba, Omotayo; Jahed, Nazeem; Baker, Priscilla G.L.; Iwuoha, Emmanuel I.

2009-01-01

Cytochrome P450-3A4 (CYP3A4) is a monooxygenase enzyme that plays a major role in the detoxification of bioactive compounds and hydrophobic xenobiotics (e.g. medicines, drugs, environmental pollutants, food supplements and steroids). Physiologically the monooxygenation reactions of this class II, microsomal, b-type heme enzyme, usually requires cytochrome P450 reductase, NADPH. A novel CYP3A4 biosensor system that essentially simplified the enzymatic redox processes by allowing electron transfer between the electrode and the enzyme redox centre to occur, without any need for the physiological redox partners, was developed for the detection of 2,4-dichlorophenol (2,4-DCP), a priority environmental pollutant and an endocrine disruptor. The biosensor, GC/Naf-Co(Sep) 3+ /CYP3A4/Naf, was constructed by encapsulating CYP3A4 in a Nafion-cobalt (III) sepulchrate (Naf-Co(Sep) 3+ ) composite film on a glassy carbon (GC) electrode. The responses of the biosensor to 2,4-dichlorophenol, erythromycin (CYP3A4 native substrate) and ketoconazole (CYP 3A4 natural inhibitor) were studied by cyclic and square wave voltammetric techniques. The detection limit (DL) of the biosensor for 2,4-dichlorophenol was 0.043 μg L -1 , which is by an order of magnitude lower than the EU limit (0.3 μg L -1 ) for any pesticide compound in ground water. The biosensor's DL is lower than the U.S. Environmental Protection Agency's drinking water equivalent level (DWEL) value for 2,4-DCP, which is 2 μg L -1

Microsomal cytochrome P450-3A4 (CYP3A4) nanobiosensor for the determination of 2,4-dichlorophenol-An endocrine disruptor compound

Energy Technology Data Exchange (ETDEWEB)

Hendricks, Nicolette R.; Waryo, Tesfaye T.; Arotiba, Omotayo; Jahed, Nazeem; Baker, Priscilla G.L. [SensorLab, Department of Chemistry, University of Western Cape, Moderddam Road, Bellville, Cape Town 7535 (South Africa); Iwuoha, Emmanuel I. [SensorLab, Department of Chemistry, University of Western Cape, Moderddam Road, Bellville, Cape Town 7535 (South Africa)], E-mail: eiwuoha@uwc.ac.za

2009-02-28

Cytochrome P450-3A4 (CYP3A4) is a monooxygenase enzyme that plays a major role in the detoxification of bioactive compounds and hydrophobic xenobiotics (e.g. medicines, drugs, environmental pollutants, food supplements and steroids). Physiologically the monooxygenation reactions of this class II, microsomal, b-type heme enzyme, usually requires cytochrome P450 reductase, NADPH. A novel CYP3A4 biosensor system that essentially simplified the enzymatic redox processes by allowing electron transfer between the electrode and the enzyme redox centre to occur, without any need for the physiological redox partners, was developed for the detection of 2,4-dichlorophenol (2,4-DCP), a priority environmental pollutant and an endocrine disruptor. The biosensor, GC/Naf-Co(Sep){sup 3+}/CYP3A4/Naf, was constructed by encapsulating CYP3A4 in a Nafion-cobalt (III) sepulchrate (Naf-Co(Sep){sup 3+}) composite film on a glassy carbon (GC) electrode. The responses of the biosensor to 2,4-dichlorophenol, erythromycin (CYP3A4 native substrate) and ketoconazole (CYP 3A4 natural inhibitor) were studied by cyclic and square wave voltammetric techniques. The detection limit (DL) of the biosensor for 2,4-dichlorophenol was 0.043 {mu}g L{sup -1}, which is by an order of magnitude lower than the EU limit (0.3 {mu}g L{sup -1}) for any pesticide compound in ground water. The biosensor's DL is lower than the U.S. Environmental Protection Agency's drinking water equivalent level (DWEL) value for 2,4-DCP, which is 2 {mu}g L{sup -1}.

Influence of genetic variants of CYP2D6, CYP2C9, CYP2C19 and CYP3A4 on antiepileptic drug metabolism in pediatric patients with refractory epilepsy.

Science.gov (United States)

López-García, Miguel A; Feria-Romero, Iris A; Serrano, Héctor; Rayo-Mares, Darío; Fagiolino, Pietro; Vázquez, Marta; Escamilla-Núñez, Consuelo; Grijalva, Israel; Escalante-Santiago, David; Orozco-Suarez, Sandra

2017-06-01

Identified the polymorphisms of CYP2D6, CYP2C9, CYP2C19 and CYP3A4, within a rigorously selected population of pediatric patients with drug-resistant epilepsy. The genomic DNA of 23 drug-resistant epilepsy patients and 7 patients with good responses were analyzed. Ten exons in these four genes were genotyped, and the drug concentrations in saliva and plasma were determined. The relevant SNPs with pharmacogenomics relations were CYP2D6*2 (rs16947) decreased your activity and CYP2D6*4 (rs1065852), CYP2C19*2 (rs4244285) and CYP3A4*1B (rs2740574) by association with poor metabolizer. The strongest risk factors were found in the AA genotype and allele of SNP rs3892097 from the CYP2D6 gene, followed by the alleles A and T of SNPs rs2740574 and rs2687116, respectively from CYP3A4. The most important concomitance was between homozygous genotype AA of rs3892097 and genotype AA of rs2740574 with 78.3% in drug-resistant epilepsy patients as compared to 14.3% in control patients. The results demonstrated the important role of the CYP 3A4*1B allelic variant as risk factor for developing drug resistance and CYP2D6, CYP2C19 SNPs and haplotypes may affect the response to antiepileptic drugs. Copyright © 2017. Published by Elsevier Urban & Partner Sp. z o.o.

Association between cytochrome CYP17A1, CYP3A4, and CYP3A43 polymorphisms and prostate cancer risk and aggressiveness in a Korean study population

OpenAIRE

Han, Jun Hyun; Lee, Yong Seong; Kim, Hae Jong; Lee, Shin Young; Myung, Soon Chul

2014-01-01

In this study, we evaluated genetic variants of the androgen metabolism genes CYP17A1, CYP3A4, and CYP3A43 to determine whether they play a role in the development of prostate cancer (PCa) in Korean men. The study population included 240 pathologically diagnosed cases of PCa and 223 age-matched controls. Among the 789 single-nucleotide polymorphism (SNP) database variants detected, 129 were reported in two Asian groups (Han Chinese and Japanese) in the HapMap database. Only 21 polymorphisms o...

CYP3A4 Mediates Oxidative Metabolism of the Synthetic Cannabinoid AKB-48.

Science.gov (United States)

Holm, Niels Bjerre; Nielsen, Line Marie; Linnet, Kristian

2015-09-01

Synthetic cannabinoid designer drugs have emerged as drugs of abuse during the last decade, and acute intoxication cases are documented in the scientific literature. Synthetic cannabinoids are extensively metabolized, but our knowledge of the involved enzymes is limited. Here, we investigated the metabolism of N-(1-adamantyl)-1-pentyl-1H-indazole-3-carboxamide (AKB-48), a compound identified in herbal blends from 2012 and onwards. We screened for metabolite formation using a panel of nine recombinant cytochrome P450 (CYP) enzymes (CYP1A2, 2B6, 2C8, 2C9, 2C18, 2C19, 2D6, 2E1, and 3A4) and compared the formed metabolites to human liver microsomal (HLM) incubations with specific inhibitors against CYP2D6, 2C19, and 3A4, respectively. The data reported here demonstrate CYP3A4 to be the major CYP enzyme responsible for the oxidative metabolism of AKB-48, preferentially performing the oxidation on the adamantyl moiety. Genetic polymorphisms are likely not important with regard to toxicity given the major involvement of CYP3A4. Adverse drug-drug interactions (DDIs) could potentially occur in cases with co-intake of strong CYP3A4 inhibitors, e.g., HIV antivirals and azole antifungal agents.

Dual-color fluorescence imaging to monitor CYP3A4 and CYP3A7 expression in human hepatic carcinoma HepG2 and HepaRG cells.

Directory of Open Access Journals (Sweden)

Saori Tsuji

Full Text Available Human adult hepatocytes expressing CYP3A4, a major cytochrome P450 enzyme, are required for cell-based assays to evaluate the potential risk of drug-drug interactions caused by transcriptional induction of P450 enzymes in early-phase drug discovery and development. However, CYP3A7 is preferentially expressed in premature hepatoblasts and major hepatic carcinoma cell lines. The human hepatocellular carcinoma cell line HepaRG possesses a high self-renewal capacity and can differentiate into hepatic cells similar to human adult hepatocytes in vitro. Transgenic HepaRG cells, in which the expression of fluorescent reporters is regulated by 35 kb regulatory elements of CYP3A4, have a distinct advantage over human hepatocytes isolated by collagenase perfusion, which are unstable in culture. Thus, we created transgenic HepaRG and HepG2 cells by replacing the protein-coding regions of human CYP3A4 and CYP3A7 with enhanced green fluorescent protein (EGFP and DsRed reporters, respectively, in a bacterial artificial chromosome vector that included whole regulatory elements. The intensity of DsRed fluorescence was initially high during the proliferation of transgenic HepaRG cells. However, most EGFP-positive cells were derived from those in which DsRed fluorescence was extinguished. Comparative analyses in these transgenic clones showed that changes in the total fluorescence intensity of EGFP reflected fold changes in the mRNA level of endogenous CYP3A4. Moreover, CYP3A4 induction was monitored by the increase in EGFP fluorescence. Thus, this assay provides a real-time evaluation system for quality assurance of hepatic differentiation into CYP3A4-expressing cells, unfavourable CYP3A4 induction, and fluorescence-activated cell sorting-mediated enrichment of CYP3A4-expressing hepatocytes based on the total fluorescence intensities of fluorescent reporters, without the need for many time-consuming steps.

A PXR reporter gene assay in a stable cell culture system: CYP3A4 and CYP2B6 induction by pesticides.

Science.gov (United States)

Lemaire, Géraldine; de Sousa, Georges; Rahmani, Roger

2004-12-15

A stable hepatoma cell line expressing the human pregnane X receptor (hPXR) and the cytochrome P4503A4 (CYP3A4) distal and proximal promoters plus the luciferase reporter gene was developed to assess the ability of several xenobiotic agents to induce CYP3A4 and CYP2B6. After selection for neomycin resistance, one clone, displaying high luciferase activity in response to rifampicin (RIF), was isolated and the stable expression of hPXR was confirmed by reverse transcription polymerase chain reaction (RT-PCR). Dose-response curves were generated by treating these cells with increasing concentrations of RIF, phenobarbital (PB), clotrimazole (CLOT) or 5beta-pregnane-3,20-dione (5beta-PREGN). The effective concentrations for half maximal response (EC50) were determined for each of these compounds. RIF was the most effective compound, with maximal luciferase activity induced at 10 microM. The agonist activities of PXR-specific inducers measured using our stable model were consistent with those measured in transient transfectants. The abilities of organochlorine (OC), organophosphate (OP) and pyrethroid pesticides (PY) to activate hPXR were also assessed and found to be consistent with the abilities of these compounds to induce CYP3A4 and CYP2B6 in primary culture of human hepatocytes. These results suggest that CYP3A4 and CYP2B6 regulation through PXR activation by persistent pesticides may have an impact on the metabolism of xenobiotic agents and endogenous steroid hormones. Our model provides a useful tool for studying hPXR activation and for identifying agents capable of inducing CYP3A4 and CYP2B6.

Assessment of human pregnane X receptor involvement in pesticide-mediated activation of CYP3A4 gene.

Science.gov (United States)

Matsubara, Tsutomu; Noracharttiyapot, Wachiraporn; Toriyabe, Takayoshi; Yoshinari, Kouichi; Nagata, Kiyoshi; Yamazoe, Yasushi

2007-05-01

Assessment of foreign chemical inducibility on CYP3A4 is necessary to optimize drug therapies. The properties of chemicals such as pesticides, however, are not well investigated. In the present study, properties of various pesticides on human CYP3A4 induction have been tested using HepG2-derived cells stably expressing the CYP3A4 promoter/enhancer (3-1-10 cells) and the human pregnane X receptor (hPXR)-small interfering RNA (siRNA) system. Among the examined pesticides, 13 pesticides were observed to activate the CYP3A4 gene. Surprisingly, pyributicarb was found to increase the CYP3A4 reporter activity at 0.1 to 1 microM more strongly than typical CYP3A4 inducer rifampicin. Expression of hPXR-siRNA clearly diminished the pyributicarb-stimulated CYP3A4 reporter activity in 3-1-10 cells and decreased the endogenous CYP3A4 mRNA levels in HepG2 cells. Pyributicarb caused enhancement of CYP3A4-derived reporter activity in mouse livers introduced with hPXR by adenovirus. These results indicate pyributicarb as a potent activator of CYP3A4 gene, suggesting the existence of pesticides leading to CYP3A4 induction in our environment.

Genomic variation in CYP3A4: type, frequencies and potential implications for pharmacogenetic understanding.

OpenAIRE

Creemer, O.

2012-01-01

The human cytochrome P450 3A subfamily metabolises endogenous substances and approximately half of all currently available drugs. There is marked inter-individual variation in hepatic expression of the major adult isoform, CYP3A4; the genetic component of this variability is estimated at 60-90% and, as yet, remains largely uncharacterised. Elucidation of genetic factors determining CYP3A4 activity would permit personalised dose-adjustment in therapies with CYP3A4 drug substrates. CYP3A4 genom...

Furocoumarins from grapefruit juice and their effect on human CYP 3A4 and CYP 1B1 isoenzymes.

Science.gov (United States)

Girennavar, Basavaraj; Poulose, Shibu M; Jayaprakasha, Guddadarangavvanahally K; Bhat, Narayan G; Patil, Bhimanagouda S

2006-04-15

Bioactive compounds present in grapefruit juice are known to increase the bioavailability of certain medications by acting as potent CYP 3A4 inhibitors. An efficient technique has been developed for isolation and purification of three furocoumarins. The isolated compounds have been tested for the inhibition of human CYP 1B1 isoform using specific substrates. Grapefruit juice was extracted with ethyl acetate (EtOAc) and the dried extract was loaded onto silica gel column chromatography. Further, column fractions were subjected to preparative HPLC to obtain three compounds. The purity of these compounds was analyzed by HPLC and structures were determined by NMR studies. The identified compounds, bergamottin, 6',7'-dihydroxybergamottin (DHB), and paradisin-A, were tested for their inhibitory effects on hydroxylase and O-dealkylase activities of human cytochrome P450 isoenzymes CYP 3A4 and CYP 1B1. Paradisin-A was found to be a potent CYP 3A4 inhibitor with an IC50 of 1.2 microM followed by DHB and bergamottin. All three compounds showed a substantial inhibitory effect on CYP 3A4 below 10 microM. Inhibitory effects on CYP 1B1 exhibited a greater variation due to the specificity of substrates. Paradisin A showed an IC50 of 3.56+/-0.12 microM for the ethoxy resorufin O-dealkylase (EROD) activity and 33.56+/-0.72 microM for the benzyloxy resorufin (BROD). DHB and bergamottin showed considerable variations for EROD and BROD activities with an IC50 of 7.17 microM and 13.86 microM, respectively.

Endosulfan-alpha Induces CYP26 and CYP3A4 by Activating the Pregnane X Receptor But Not the Constitutive Androstane Receptor

Science.gov (United States)

2006-01-01

CYP3A4 gene expression by organochlorine pesticides . Biochem Pharmacol 64:1513-1519. Dinham B (1993) The Pesticide Hazard. A Global Health and...Coumoul X, Diry M and Barouki R (2002) PXR-dependent induction of human CYP3A4 gene expression by organochlorine pesticides . Biochem Pharmacol 64:1513...system: CYP3A4 and CYP2B6 induction by pesticides . Biochem Pharmacol 68:2347-2358. 71 Nelson D (2003) Cytochrome P450 Homepage (http

CYP1A1, CYP3A5 and CYP3A7 polymorphisms and testicular cancer susceptibility.

Science.gov (United States)

Kristiansen, W; Haugen, T B; Witczak, O; Andersen, J M; Fosså, S D; Aschim, E L

2011-02-01

Testicular cancer (TC) incidence is increasing worldwide, but the aetiology remains largely unknown. An unbalanced level of oestrogens and androgens in utero is hypothesized to influence TC risk. Polymorphisms in genes encoding cytochrome P450 (CYP) enzymes involved in metabolism of reproductive hormones, such as CYP1A1, CYP3A5 and CYP3A7, may contribute to variability of an individual's susceptibility to TC. The aim of this case-control study was to investigate possible associations between different CYP genotypes and TC, as well as histological type of TC. The study comprised 652 TC cases and 199 controls of Norwegian Caucasian origin. Genotyping of the CYP1A1*2A (MspI), CYP1A1*2C (I462V), CYP1A1*4 (T461N), CYP3A5*3C (A6986G) and CYP3A7*2 (T409R) polymorphisms was performed using TaqMan allelic discrimination or sequencing. The CYP1A1*2A allele was associated with 44% reduced risk of TC with each polymorphic allele [odds ratio (OR) = 0.56, 95% confidence interval (CI) = 0.40-0.78, p(trend) = 0.001], whereas the CYP1A1*2C allele was associated with 56% reduced risk of TC with each polymorphic allele (OR = 0.44, 95% CI = 0.25-0.75, p(trend) = 0.003). The decreased risk per allele was significant for seminomas (OR = 0.46, 95% CI, 0.31-0.70, p(trend) < 0.001 and OR = 0.31, 95% CI = 0.14-0.66, p(trend) = 0.002, respectively), but only borderline significant for non-seminomas (OR = 0.65, 95% CI = 0.45-0.95, p(trend) = 0.027 and OR = 0.55, 95% CI = 0.30-1.01, p(trend) = 0.052, respectively). There were no statistically significant differences in the distribution of the CYP3A5*3C and CYP3A7*2 polymorphic alleles between TC cases and controls. This study suggests that polymorphisms in the CYP1A1 gene may contribute to variability of individual susceptibility to TC. © 2010 The Authors. International Journal of Andrology © 2010 European Academy of Andrology.

Interactions of endosulfan and methoxychlor involving CYP3A4 and CYP2B6 in human HepaRG cells.

Science.gov (United States)

Savary, Camille C; Jossé, Rozenn; Bruyère, Arnaud; Guillet, Fabrice; Robin, Marie-Anne; Guillouzo, André

2014-08-01

Humans are usually exposed to several pesticides simultaneously; consequently, combined actions between pesticides themselves or between pesticides and other chemicals need to be addressed in the risk assessment. Many pesticides are efficient activators of pregnane X receptor (PXR) and/or constitutive androstane receptor (CAR), two major nuclear receptors that are also activated by other substrates. In the present work, we searched for interactions between endosulfan and methoxychlor, two organochlorine pesticides whose major routes of metabolism involve CAR- and PXR-regulated CYP3A4 and CYP2B6, and whose mechanisms of action in humans remain poorly understood. For this purpose, HepaRG cells were treated with both pesticides separately or in mixture for 24 hours or 2 weeks at concentrations relevant to human exposure levels. In combination they exerted synergistic cytotoxic effects. Whatever the duration of treatment, both compounds increased CYP3A4 and CYP2B6 mRNA levels while differently affecting their corresponding activities. Endosulfan exerted a direct reversible inhibition of CYP3A4 activity that was confirmed in human liver microsomes. By contrast, methoxychlor induced this activity. The effects of the mixture on CYP3A4 activity were equal to the sum of those of each individual compound, suggesting an additive effect of each pesticide. Despite CYP2B6 activity being unchanged and increased with endosulfan and methoxychlor, respectively, no change was observed with their mixture, supporting an antagonistic effect. Altogether, our data suggest that CAR and PXR activators endosulfan and methoxychlor can interact together and with other exogenous substrates in human hepatocytes. Their effects on CYP3A4 and CYP2B6 activities could have important consequences if extrapolated to the in vivo situation. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

A search for new CYP3A4 variants as determinants of tacrolimus dose requirements in renal-transplanted patients.

Science.gov (United States)

Tavira, Beatriz; Coto, Eliecer; Diaz-Corte, Carmen; Alvarez, Victoria; López-Larrea, Carlos; Ortega, Francisco

2013-08-01

The CYP3A5*3 and CYP3A4*1B alleles have been related with tacrolimus (Tac) dose requirements. The rare CYP3A4*22 variant has also been associated with a significantly lower Tac dose. We genotyped the three single-nucleotide polymorphisms in 206 kidney-transplanted patients who received Tac as the primary immunosuppressor. CYP3A5*1 and CYP3A4*1B allele carriers received a significantly higher Tac dose (PCYP3A4*22 genotypes, either nominally or according to the CYP3A5 genotype (expressers vs. nonexpressers). Sequencing of CYP3A4 coding exons in a total of 15 patients revealed only one nonreported missense change (p.P227>T) in one patient. We concluded that CYP3A5*3 and CYP3A4*1B were the main determinants of the Tac dose-adjusted blood concentration in our cohort of renal-transplanted patients.

Hemodialysis does not alter in vitro hepatic CYP3A4 and CYP2D6 metabolic activity in uremic serum

Directory of Open Access Journals (Sweden)

Decker BS

2013-12-01

Full Text Available Brian S Decker,1,2 Kalisha D O'Neill,1,2 Mary A Chambers,1,2 James E Slaven,3 Zhangsheng Yu,3 David R Jones,2,4 Sharon M Moe1,21Division of Nephrology, 2Department of Medicine, 3Department of Biostatistics, 4Division of Clinical Pharmacology, Department of Medicine, School of Medicine, Indiana University, Indianapolis, IN, USAAbstract: There is a paucity of studies evaluating the change in liver metabolism in subjects receiving hemodialysis. The purpose of this study was to compare the effect of uremic toxins on hepatic cytochrome P450 (CYP3A4 and CYP2D6 metabolism before and after a 4-hour hemodialysis session. Midazolam and dextromethorphan were incubated with uremic serum collected from subjects before and after the 4-hour hemodialysis session. Analysis and quantification of the 1'-OH-midazolam and 4-OH-midazolam and dextrorphan metabolites were performed by high-pressure liquid chromatography/mass spectrometry. Statistical analysis using the Student's t-test (paired was used to compare the amount of metabolite formed. The mean amount of 1'-OH-midazolam, 4-OH-midazolam, and dextrorphan metabolites formed before and after hemodialysis did not significantly differ. There was no significant difference in CYP3A4 and CYP2D6 metabolic activity in uremic serum before and after hemodialysis.Keywords: hemodialysis, uremia, CYP3A4, CYP2D6, metabolism

[Toxic effect of trichloroethylene on liver cells with CYP3A4 gene defect].

Science.gov (United States)

Liao, R Y; Liu, S

2016-06-20

To investigate the toxic effect of trichloroethylene on liver cells with CYP3A4 gene defect. The normal human liver cells (L02 cells) and liver cells with CYP3A4 gene defect were exposed to trichloroethylene at different doses (0.0, 0.4, 0.8, 1.6, 3.2, and 6.4 mmol/L). CCK8 assay and RT-qPCR were used to measure cell viability and changes in the expression of apoptosis genes and oncogenes. After being exposed to trichloroethylene at doses of 1.6, 3.2, and 6.4 mmol/L, the liver cells with CYP3A4 gene defect showed significantly higher cell viability than L02 cells (0.91±0.06/0.89±0.05/0.85±0.07 vs 0.80±0.04/0.73±0.06/0.67±0.07, Ptrichloroethylene groups showed significant increases in the expression of the apoptosis genes caspase-3, caspase-8, and caspase-9 (PTrichloroethylene exposure has a less effect on the expression of apoptosis genes and oncogenes in liver cells with CYP3A4 gene defect than in normal human liver cells, suggesting that CYP3A4 gene defect reduces the inductive effect of trichloroethylene on apoptosis genes and oncogenes.

Three new shRNA expression vectors targeting the CYP3A4 coding sequence to inhibit its expression

Directory of Open Access Journals (Sweden)

Siyun Xu

2014-10-01

Full Text Available RNA interference (RNAi is useful for selective gene silencing. Cytochrome P450 3A4 (CYP3A4, which metabolizes approximately 50% of drugs in clinical use, plays an important role in drug metabolism. In this study, we aimed to develop a short hairpin RNA (shRNA to modulate CYP3A4 expression. Three new shRNAs (S1, S2 and S3 were designed to target the coding sequence (CDS of CYP3A4, cloned into a shRNA expression vector, and tested in different cells. The mixture of three shRNAs produced optimal reduction (55% in CYP3A4 CDS-luciferase activity in both CHL and HEK293 cells. Endogenous CYP3A4 expression in HepG2 cells was decreased about 50% at both mRNA and protein level after transfection of the mixture of three shRNAs. In contrast, CYP3A5 gene expression was not altered by the shRNAs, supporting the selectivity of CYP3A4 shRNAs. In addition, HepG2 cells transfected with CYP3A4 shRNAs were less sensitive to Ginkgolic acids, whose toxic metabolites are produced by CYP3A4. These results demonstrate that vector-based shRNAs could modulate CYP3A4 expression in cells through their actions on CYP3A4 CDS, and CYP3A4 shRNAs may be utilized to define the role of CYP3A4 in drug metabolism and toxicity.

An impact of CYP3A4 *1B polymorphism on rifampicin metabolism

Directory of Open Access Journals (Sweden)

H. O. Poludenko

2017-08-01

Full Text Available Until now, the enzyme systems responsible for biotransformation of the antituberculous drug rifampicin remain unknown. The aim of research was an investigation of the candidate enzymes involved in the biotransformation of rifampicin using the computer system PASS and an experimental study concerning the effect of the polymorphism of the biotransformation gene CYP3A4 *1B on the level of rifampicin in the blood of patients with pulmonary tuberculosis (РTB. The probability (Pa of certain pharmacological activity and the effect on putative enzyme systems of the human body of rifampicin has been calculated by the PASS method. Polymerase chain reaction revealed the polymorphism of the CYP3A4 *1B gene among healthy volunteers as well as patients with РTB. With a high degree of probability, according to PASS calculations, it was predicted that rifampicin undergo metabolism with the CYP3A4 enzyme - probability (Ra were 0.891. According to the genotype CYP3A4 *1B, 95.3% of the healthy donors carried a homozygous wild-type gene (i.e., had high enzymatic activity - AA genotype; the rest 4.7% - were carriers of the heterozygous AG genotype (moderate enzyme activity.The polymorphism of CYP3A4 *1B genotypes and alleles in the south-west of Ukraine was close to the results obtained in European countries. 91.4% and 8.6% of the patients with РTB had AA and AG genotype, correspondently. Thus, among the patients with РTB, the AG genotype was more often observed than among healthy volunteers. There was no significant difference in rifampicin concentration among РTB-patients concerning CYP3A4 * 1B polymorphism.

Metabolic Pathway of Icotinib In Vitro: The Differential Roles of CYP3A4, CYP3A5, and CYP1A2 on Potential Pharmacokinetic Drug-Drug Interaction.

Science.gov (United States)

Zhang, TianHong; Zhang, KeRong; Ma, Li; Li, Zheng; Wang, Juan; Zhang, YunXia; Lu, Chuang; Zhu, Mingshe; Zhuang, XiaoMei

2018-04-01

Icotinib is the first self-developed small molecule drug in China for targeted therapy of non-small cell lung cancer. To date, systematic studies on the pharmacokinetic drug-drug interaction of icotinib were limited. By identifying metabolite generated in human liver microsomes and revealing the contributions of major cytochromes P450 (CYPs) in the formation of major metabolites, the aim of the present work was to understand the mechanisms underlying pharmacokinetic and pharmacological variability in clinic. A liquid chromatography/UV/high-resolution mass spectrometer method was developed to characterize the icotinib metabolites. The formation of 6 major metabolites was studied in recombinant CYP isozymes and human liver microsomes with specific inhibitors to identify the CYPs responsible for icotinib metabolism. The metabolic pathways observed in vitro are consistent with those observed in human. Results demonstrated that the metabolites are predominantly catalyzed by CYP3A4 (77%∼87%), with a moderate contribution from CYP3A5 (5%∼15%) and CYP1A2 (3.7%∼7.5%). The contribution of CYP2C8, 2C9, 2C19, and 2D6 is insignificant. Based on our observations, to minimize drug-drug interaction risk in clinic, coprescription of icotinib with strong CYP3A inhibitors or inducers must be weighed. CYP1A2, a highly inducible enzyme in the smoking population, may also represent a determinant of pharmacokinetic and pharmacological variability of icotinib, especially in lung cancer patients with smoking history. Copyright © 2018 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

CYP2B6, CYP2D6, and CYP3A4 catalyze the primary oxidative metabolism of perhexiline enantiomers by human liver microsomes.

Science.gov (United States)

Davies, Benjamin J; Coller, Janet K; Somogyi, Andrew A; Milne, Robert W; Sallustio, Benedetta C

2007-01-01

The cytochrome P450 (P450)-mediated 4-monohydroxylations of the individual enantiomers of the racemic antianginal agent perhexiline (PHX) were investigated in human liver microsomes (HLMs) to identify stereoselective differences in metabolism and to determine the contribution of the polymorphic enzyme CYP2D6 and other P450s to the intrinsic clearance of each enantiomer. The cis-, trans1-, and trans2-4-monohydroxylation rates of (+)- and (-)-PHX by human liver microsomes from three extensive metabolizers (EMs), two intermediate metabolizers (IMs), and two poor metabolizers (PMs) of CYP2D6 were measured with a high-performance liquid chromatography assay. P450 isoform-specific inhibitors, monoclonal antibodies directed against P450 isoforms, and recombinantly expressed human P450 enzymes were used to define the P450 isoform profile of PHX 4-monohydroxylations. The total in vitro intrinsic clearance values (mean +/- S.D.) of (+)- and (-)-PHX were 1376 +/- 330 and 2475 +/- 321, 230 +/- 225 and 482 +/- 437, and 63.4 +/- 1.6 and 54.6 +/- 1.2 microl/min/mg for the EM, IM, and PM HLMs, respectively. CYP2D6 catalyzes the formation of cis-OH-(+)-PHX and trans1-OH-(+)-PHX from (+)-PHX and cis-OH-(-)-PHX from (-)-PHX with high affinity. CYP2B6 and CYP3A4 each catalyze the trans1- and trans2-4-monohydroxylation of both (+)- and (-)-PHX with low affinity. Both enantiomers of PHX are subject to significant polymorphic metabolism by CYP2D6, although this enzyme exhibits distinct stereoselectivity with respect to the conformation of metabolites and the rate at which they are formed. CYP2B6 and CYP3A4 are minor contributors to the intrinsic P450-mediated hepatic clearance of both enantiomers of PHX, except in CYP2D6 PMs.

Polychlorinated biphenyl (PCB) induction of CYP3A4 enzyme activity in healthy Faroese adults

DEFF Research Database (Denmark)

Petersen, Maria Skaalum; Halling, Jónrit; Damkier, Per

2007-01-01

The CYP3A4 enzyme is, along with other cytochrome P450 enzymes, involved in the metabolism of environmental pollutants and is highly inducible by these substances. A commercial polychlorinated biphenyl (PCB) mixture, 1,1,1,-trichloro-2-(o-chlorophenyl), 2-(p'-chlorophenyl)ethane (o,p'-DDT) and 1......,1,-dichloro-2,2-bis (p-chlorophenyl)ethene (p,p'-DDE) are known to induce CYP3A4 activity through activation of nuclear receptors, such as the pregnane X receptor. However, this induction of CYP3A4 has not yet been investigated in humans. Thus, the aim of the study was to determine the variability of the CYP3......A4 phenotype in regard to increased concentrations of PCBs and other persistent organohalogen pollutants (POPs) in healthy Faroese adults. In 310 randomly selected Faroese residents aged 18-60 years, the CYP3A4 activity was determined based on the urinary 6beta-hydroxycortisol/cortisol (6beta...

Porcine foetal and neonatal CYP3A liver expression

Directory of Open Access Journals (Sweden)

Marie Louise Hiort Hermann

2011-05-01

Full Text Available Human cytochrome P450 3A7 (CYP3A7 and cytochrome P450 3A4 (CYP3A4 are hepatic metabolising enzymes which participates in the biotransformation of endo- and exogenous substances in foetuses and neonates respectively. These CYP3A enzymes display an inverse relationship: CYP3A7 is the dominant enzyme in the foetal liver, whereas the expression of CYP3A4 is low. After parturition there is a shift in the expression, thus CYP3A7 is down regulated, while the level of CYP3A4 gradually increases and becomes the dominant metabolising CYP3A enzyme in the adult. The minipig is increasingly being used as a model for humans in biomedical studies, because of its many similarities with the human physiology and anatomy. The aim of this study was to examine whether, as in humans, a shift is seen in the hepatic expression of a CYP3A7- like enzyme to cytochrome P450 3A29 (CYP3A29 (an orthologue to the human CYP3A4 in minipigs. This was elucidated by examining the hepatic mRNA expression of CYP3A7 and CYP3A29 in 39 foetuses and newborn Göttingen minipigs using quantitative real time polymerase chain reaction (qPCR. Furthermore the immunochemical level of CYP3A7-LE and CYP3A29 was measured in liver microsomes using western blotting. The expression of CYP3A29 was approximately 9- fold greater in neonates compared to foetuses, and a similar difference was reflected on the immunochemical level. It was not possible to detect a significant level of foetal CYP3A7 mRNA, but immunoblotting showed a visible difference depending on age. This study demonstrates an increase in the expression of CYP3A29, the CYP3A4 orthologue in perinatal minipigs as in humans, which suggests that the minipig could be a good model when testing for human foetal toxicity towards CYP3A4 substrates.

Genotype-phenotype associations for common CYP3A4 and CYP3A5 variants in the basal and induced metabolism of midazolam in European- and African-American men and women.

Science.gov (United States)

Floyd, Michael D; Gervasini, Guillermo; Masica, Andrew L; Mayo, Gail; George, Alfred L; Bhat, Kolari; Kim, Richard B; Wilkinson, Grant R

2003-10-01

CYP3A activity in adults varies between individuals and it has been suggested that this has a genetic basis, possibly related to variant alleles in CYP3A4 and CYP3A5 genes. Accordingly, genotype-phenotype associations were investigated under constitutive and induced conditions. Midazolam's systemic and oral clearances, and the erythromycin breath test (ERBT) were determined in 57 healthy subjects: 23 (11 men, 12 women) European- and 34 (14 men, 20 women) African-Americans. Studies were undertaken in the basal state and after 14-15 days pretreatment with rifampin. DNA was characterized for the common polymorphisms CYP3A4*1B, CYP3A5*3, CYP3A5*6 and CYP3A5*7 by direct sequencing, and for exon 21 and exon 26 variants of MDR1 by allele-specific, real-time polymerase chain reaction. In 95% of subjects, the basal systemic clearance of midazolam was unimodally distributed and variability was less than four-fold whereas, in 98% of the study population, oral clearance varied five-fold. No population or sex-related differences were apparent. Similar findings were observed with the ERBT. Rifampin pretreatment markedly increased the systemic (two-fold) and oral clearance (16-fold) of midazolam, and the ERBT (two-fold) but the variabilities were unchanged. No associations were noted between these phenotypic measures and any of the studied genotypes, except for oral clearance and its fold-increase after rifampin. These were related to the presence of CYP3A4*1B and the inversely linked CYP3A5*3 polymorphism, with the extent of induction being approximately 50% greater in CYP3A5*3 homozygotes compared to wild-type subjects. In most healthy subjects, variability in intestinal and hepatic CYP3A activity, using midazolam as an in-vivo probe, is modest and common polymorphisms in CYP3A4 and CYP3A5 do not appear to have important functional significance.

Time-dependent inhibition of CYP3A4 by gallic acid in human liver microsomes and recombinant systems.

Science.gov (United States)

Pu, Qiang-Hong; Shi, Liang; Yu, Chao

2015-03-01

1.Gallic acid is a main polyphenol in various fruits and plants. Inhibitory characteristics of gallic acid on CYP3A4 were still unclear. The objective of this work is hence to investigate inhibitory characteristics of gallic acid on CYP3A4 using testosterone as the probe substrate in human liver microsomes (HLMs) and recombinant CYP3A4 (rCYP3A4) systems. 2.Gallic acid caused concentration-dependent loss of CYP3A4 activity with IC50 values of 615.2 μM and 669.5 μM in HLM and rCYP3A4 systems, respectively. IC50-shift experiments showed that pre-incubation with gallic acid in the absence of NADPH contributed to 12- or 14-fold reduction of IC50 in HLM and rCYP3A4 systems, respectively, supporting a time-dependent inhibition. In HLM, time-dependent inactivation variables KI and Kinact were 485.8 μM and 0.05 min(-1), respectively. 3.Compared with the presence of NADPH, pre-incubation of gallic acid in the absence of NADPH markedly increased its inhibitory effects in HLM and rCYP3A4 systems. Those results indicate that CYP3A4 inactivation by gallic acid was independent on NADPH and was mainly mediated its oxidative products. 4.In conclusion, we showed that gallic acid weakly and time-dependently inactivated CYP3A4 via its oxidative products.

Enhancement of hepatic 4-hydroxylation of 25-hydroxyvitamin D3 through CYP3A4 induction in vitro and in vivo: implications for drug-induced osteomalacia.

Science.gov (United States)

Wang, Zhican; Lin, Yvonne S; Dickmann, Leslie J; Poulton, Emma-Jane; Eaton, David L; Lampe, Johanna W; Shen, Danny D; Davis, Connie L; Shuhart, Margaret C; Thummel, Kenneth E

2013-05-01

Long-term therapy with certain drugs, especially cytochrome P450 (P450; CYP)-inducing agents, confers an increased risk of osteomalacia that is attributed to vitamin D deficiency. Human CYP24A1, CYP3A4, and CYP27B1 catalyze the inactivation and activation of vitamin D and have been implicated in the adverse drug response. In this study, the inducibility of these enzymes and monohydroxylation of 25-hydroxyvitamin D3 (25OHD3) were evaluated after exposure to P450-inducing drugs. With human hepatocytes, treatment with phenobarbital, hyperforin, carbamazepine, and rifampin significantly increased the levels of CYP3A4, but not CYP24A1 or CYP27B1 mRNA. In addition, rifampin pretreatment resulted in an 8-fold increase in formation of the major metabolite of 25OHD3, 4β,25(OH)2D3. This inductive effect was blocked by the addition of 6',7'-dihydroxybergamottin, a selective CYP3A4 inhibitor. With human renal proximal tubular HK-2 cells, treatment with the same inducers did not alter CYP3A4, CYP24A1, or CYP27B1 expression. 24R,25(OH)2 D3 was the predominant monohydroxy metabolite produced from 25OHD3, but its formation was unaffected by the inducers. With healthy volunteers, the mean plasma concentration of 4β,25(OH)2D3 was increased 60% (p osteomalacia. Copyright © 2013 American Society for Bone and Mineral Research.

Porcine foetal and neonatal CYP3A liver expression

DEFF Research Database (Denmark)

Hermann-Bank, Marie Louise; Skaanild, Mette Tingleff

2011-01-01

enzyme in the foetal liver, whereas the expression of CYP3A4 is low. After parturition there is a shift in the expression, thus CYP3A7 is down regulated, while the level of CYP3A4 gradually increases and becomes the dominant metabolising CYP3A enzyme in the adult. The minipig is increasingly being used......3A4) in minipigs. This was elucidated by examining the hepatic mRNA expression of CYP3A7 and CYP3A29 in 39 foetuses and newborn Göttingen minipigs using quantitative real time polymerase chain reaction (qPCR). Furthermore the immunochemical level of CYP3A7-LE and CYP3A29 was measured in liver...

Systemic uptake of miconazole during vaginal suppository use and effect on CYP1A2 and CYP3A4 associated enzyme activities in women

DEFF Research Database (Denmark)

Kjærstad, Mia Birkhøj; Nielsen, Flemming; Nøhr-Jensen, Lene

2010-01-01

To investigate if the ordinary use of a vaginal suppository containing miconazole results in systemic absorption that is sufficient to affect the activities of CYP1A2 and CYP3A4, which are major drug- and steroid-metabolising enzymes.......To investigate if the ordinary use of a vaginal suppository containing miconazole results in systemic absorption that is sufficient to affect the activities of CYP1A2 and CYP3A4, which are major drug- and steroid-metabolising enzymes....

Effect of Garden Cress Seeds Powder and Its Alcoholic Extract on the Metabolic Activity of CYP2D6 and CYP3A4

Directory of Open Access Journals (Sweden)

Fahad I. Al-Jenoobi

2014-01-01

Full Text Available The powder and alcoholic extract of dried seeds of garden cress were investigated for their effect on metabolic activity of CYP2D6 and CYP3A4 enzymes. In vitro and clinical studies were conducted on human liver microsomes and healthy human subjects, respectively. Dextromethorphan was used as a common marker for measuring metabolic activity of CYP2D6 and CYP3A4 enzymes. In in vitro studies, microsomes were incubated with NADPH in presence and absence of different concentrations of seeds extract. Clinical investigations were performed in two phases. In phase I, six healthy female volunteers were administered a single dose of dextromethorphan and in phase II volunteers were treated with seeds powder for seven days and dextromethorphan was administered with last dose. The O-demethylated and N-demethylated metabolites of dextromethorphan were measured as dextrorphan (DOR and 3-methoxymorphinan (3-MM, respectively. Observations suggested that garden cress inhibits the formation of DOR and 3-MM metabolites. This inhibition of metabolite level was attributed to the inhibition of CYP2D6 and CYP3A4 activity. Garden cress decreases the level of DOR and 3-MM in urine and significantly increases the urinary metabolic ratio of DEX/DOR and DEX/3-MM. The findings suggested that garden cress seeds powder and ethanolic extract have the potential to interact with CYP2D6 and CYP3A4 substrates.

Characterization of the genetic variation present in CYP3A4 in three South African populations.

Science.gov (United States)

Drögemöller, Britt; Plummer, Marieth; Korkie, Lundi; Agenbag, Gloudi; Dunaiski, Anke; Niehaus, Dana; Koen, Liezl; Gebhardt, Stefan; Schneider, Nicol; Olckers, Antonel; Wright, Galen; Warnich, Louise

2013-01-01

The CYP3A4 enzyme is the most abundant human cytochrome P450 (CYP) and is regarded as the most important enzyme involved in drug metabolism. Inter-individual and inter-population variability in gene expression and enzyme activity are thought to be influenced, in part, by genetic variation. Although Southern African individuals have been shown to exhibit the highest levels of genetic diversity, they have been under-represented in pharmacogenetic research to date. Therefore, the aim of this study was to identify genetic variation within CYP3A4 in three South African population groups comprising of 29 Khoisan, 65 Xhosa and 65 Mixed Ancestry (MA) individuals. To identify known and novel CYP3A4 variants, 15 individuals were randomly selected from each of the population groups for bi-directional Sanger sequencing of ~600 bp of the 5'-upstream region and all thirteen exons including flanking intronic regions. Genetic variants detected were genotyped in the rest of the cohort. In total, 24 SNPs were detected, including CYP3A4(*)12, CYP3A4(*)15, and the reportedly functional CYP3A4(*)1B promoter polymorphism, as well as two novel non-synonymous variants. These putatively functional variants, p.R162W and p.Q200H, were present in two of the three populations and all three populations, respectively, and in silico analysis predicted that the former would damage the protein product. Furthermore, the three populations were shown to exhibit distinct genetic profiles. These results confirm that South African populations show unique patterns of variation in the genes encoding xenobiotic metabolizing enzymes. This research suggests that population-specific genetic profiles for CYP3A4 and other drug metabolizing genes would be essential to make full use of pharmacogenetics in Southern Africa. Further investigation is needed to determine if the identified genetic variants influence CYP3A4 metabolism phenotype in these populations.

Effect of Curcuma longa on CYP2D6- and CYP3A4-mediated metabolism of dextromethorphan in human liver microsomes and healthy human subjects.

Science.gov (United States)

Al-Jenoobi, Fahad Ibrahim; Al-Thukair, Areej A; Alam, Mohd Aftab; Abbas, Fawkeya A; Al-Mohizea, Abdullah M; Alkharfy, Khalid M; Al-Suwayeh, Saleh A

2015-03-01

Effect of Curcuma longa rhizome powder and its ethanolic extract on CYP2D6 and CYP3A4 metabolic activity was investigated in vitro using human liver microsomes and clinically in healthy human subjects. Dextromethorphan (DEX) was used as common probe for CYP2D6 and CYP3A4 enzymes. Metabolic activity of CYP2D6 and CYP3A4 was evaluated through in vitro study; where microsomes were incubated with NADPH in presence and absence of Curcuma extract. In clinical study phase-I, six healthy human subjects received a single dose (30 mg) of DEX syrup, and in phase-II DEX syrup was administered with Curcuma powder. The enzyme CYP2D6 and CYP3A4 mediated O- and N-demethylation of dextromethorphan into dextrorphan (DOR) and 3-methoxymorphinan (3-MM), respectively. Curcuma extract significantly inhibited the formation of DOR and 3-MM, in a dose-dependent and linear fashion. The 100 μg/ml dose of curcuma extract produced highest inhibition, which was about 70 % for DOR and 80 % for 3-MM. Curcuma significantly increases the urine metabolic ratio of DEX/DOR but the change in DEX/3-MM ratio was statistically insignificant. Present findings suggested that curcuma significantly inhibits the activity of CYP2D6 in in vitro as well as in vivo; which indicates that curcuma has potential to interact with CYP2D6 substrates.

Ibrutinib Dosing Strategies Based on Interaction Potential of CYP3A4 Perpetrators Using Physiologically Based Pharmacokinetic Modeling.

Science.gov (United States)

de Zwart, L; Snoeys, J; De Jong, J; Sukbuntherng, J; Mannaert, E; Monshouwer, M

2016-11-01

Based on ibrutinib pharmacokinetics and potential sensitivity towards CYP3A4-mediated drug-drug interactions (DDIs), a physiologically based pharmacokinetic approach was developed to mechanistically describe DDI with various CYP3A4 perpetrators in healthy men under fasting conditions. These models were verified using clinical data for ketoconazole (strong CYP3A4 inhibitor) and used to prospectively predict and confirm the inducing effect of rifampin (strong CYP3A4 inducer); DDIs with mild (fluvoxamine, azithromycin) and moderate inhibitors (diltiazem, voriconazole, clarithromycin, itraconazole, erythromycin), and moderate (efavirenz) and strong CYP3A4 inducers (carbamazepine), were also predicted. Ketoconazole increased ibrutinib area under the curve (AUC) by 24-fold, while rifampin decreased ibrutinib AUC by 10-fold; coadministration of ibrutinib with strong inhibitors or inducers should be avoided. The ibrutinib dose should be reduced to 140 mg (quarter of maximal prescribed dose) when coadministered with moderate CYP3A4 inhibitors so that exposures remain within observed ranges at therapeutic doses. Thus, dose recommendations for CYP3A4 perpetrator use during ibrutinib treatment were developed and approved for labeling. © 2016 American Society for Clinical Pharmacology and Therapeutics.

Characterization of the genetic variation present in CYP3A4 in three South African populations

Directory of Open Access Journals (Sweden)

Britt Ingrid Drögemöller

2013-02-01

Full Text Available TThe CYP3A4 enzyme is the most abundant human cytochrome P450 and is regarded as the most important enzyme involved in drug metabolism. Inter-individual and inter-population variability in gene expression and enzyme activity are thought to be influenced, in part, by genetic variation. Although Southern African individuals have been shown to exhibit the highest levels of genetic diversity, they have been under-represented in pharmacogenetic research to date. Therefore, the aim of this study was to identify genetic variation within CYP3A4 in three South African population groups comprising of 29 Khoisan, 65 Xhosa and 65 Mixed Ancestry individuals. To identify known and novel CYP3A4 variants, 15 individuals were randomly selected from each of the population groups for bi-directional Sanger sequencing of approximately 600 bp of the 5’-upstream region and all thirteen exons including flanking intronic regions. Genetic variants detected were genotyped in the rest of the cohort. In total, 24 SNPs were detected, including CYP3A4*12, CYP3A4*15, and the reportedly functional CYP3A4*1B promoter polymorphism, as well as two novel non-synonymous variants. These putatively functional variants, p.R162W and p.Q200H, were present in two of the three populations and all three populations, respectively, and in silico analysis predicted that the former would damage the protein product. Furthermore, the three populations were shown to exhibit distinct genetic profiles. These results confirm that South African populations show unique patterns of variation in the genes encoding xenobiotic metabolizing enzymes. This research suggests that population-specific genetic profiles for CYP3A4 and other drug metabolizing genes would be essential to make full use of pharmacogenetics in Southern Africa. Further investigation is needed to determine if the identified genetic variants influence CYP3A4 metabolism phenotype in these populations.

Indirubin, a component of Ban-Lan-Gen, activates CYP3A4 gene transcription through the human pregnane X receptor.

Science.gov (United States)

Kumagai, Takeshi; Aratsu, Yusuke; Sugawara, Ryosuke; Sasaki, Takamitsu; Miyairi, Shinichi; Nagata, Kiyoshi

2016-04-01

Ban-Lan-Gen is the common name for the dried roots of indigo plants, including Polygonum tinctorium, Isatis indigotica, Isatis tinctoria, and Strobilanthes cusia. Ban-Lan-Gen is frequently used as an anti-inflammatory and an anti-viral for the treatment of hepatitis, influenza, and various types of inflammation. One of the cytochrome P450 (CYP) enzymes, CYP3A4, is responsible for the metabolism of a wide variety of xenobiotics, including an estimated 60% of all clinically used drugs. In this study, we investigated the effect of Ban-Lan-Gen on the transcriptional activation of the CYP3A4 gene. Ban-Lan-Gen extract increased CYP3A4 gene reporter activity in a dose-dependent manner. Indirubin, one of the biologically active ingredients in the Ban-Lan-Gen, also dose-dependently increased CYP3A4 gene reporter activity. Expression of short hairpin RNA for the human pregnane X receptor (hPXR-shRNA) inhibited CYP3A4 gene reporter activity, and overexpression of human PXR increased indirubin- and rifampicin-induced CYP3A4 gene reporter activity. Furthermore, indirubin induced CYP3A4 mRNA expression in HepG2 cells. Taken together, these results indicate that indirubin, a component of Ban-Lan-Gen, activated CYP3A4 gene transcription through the activation of the human PXR. Copyright © 2016. Published by Elsevier Ltd.

In vitro study of modulatory effects of extracts of Strobilanthes Crispus on human cDNA-expressed cytochrome P450 2A6 (CYP2A6) and CYP3A4

Energy Technology Data Exchange (ETDEWEB)

Pan, Y.; Hsu, C.J.; Koh, R.I.; Ong, C.E.; Chieng, J.Y.

2016-07-01

Aim: Cytochrome P450 (CYP) 2A6 and CYP3A4 play important roles in biotransformation of endogenous substances as well as xenobiotics. Strobilanthes crispus (L.) Blume (S. crispus) has been found to have anti-cancer activities and this was suggested to be due to inhibition of enzymes involved in metabolic activation of procarcinogens. The purpose of this study was to look into the potential inhibitory effects of various extracts (aqueous, hexane, chloroform, ethyl acetate, and methanol) of S. crispus from leaf and stem on human cDNA-expressed CYP2A6 and CYP3A4 activities. Methods: The activity of CYP2A6 was examined via a fluorescence-based 7-hydroxylase coumarin assay. Meanwhile, high performance liquid chromatography (HPLC)-based testosterone 6β-hydroxylase assay was established to assess CYP3A4 activity. Results: It was shown that none of the extracts from both leaf and stem potently inhibited CYP2A6 and CYP3A4 activities with IC50values above 100μg/ml. Conclusion: The anticancer potency of S. crispus is unlikely due to the modulation of CYP2A6 and CYP3A4 activities, while other mechanisms might be involved and merits further investigation. On the other hand, potential drug-herb interactions occurring between CYP2A6 and CYP3A4 substrates and S. crispus preparations is relatively low, which requires further investigations via in vivo animal as well as clinical studies.

Cytochrome P450 CYP3A in marsupials: cloning and characterisation of the second identified CYP3A subfamily member, isoform 3A78 from koala (Phascolarctos cinereus).

Science.gov (United States)

El-Merhibi, Adaweyah; Ngo, Suong N T; Crittenden, Tamara A; Marchant, Ceilidh L; Stupans, Ieva; McKinnon, Ross A

2011-11-01

Cytochromes P450 (CYPs) are critically important in the oxidative metabolism of a diverse array of xenobiotics and endogenous substrates. Previously, we cloned and characterised the CYP2C, CYP4A, and CYP4B gene subfamilies from marsupials and demonstrated important species-differences in both activity and tissue expression of these CYP enzymes. Recently, we isolated the Eastern grey kangaroo CYP3A70. Here we have cloned and characterised the second identified member of marsupial CYP3A gene subfamily, CYP3A78 from the koala (Phascolarctos cinereus). In addition, we have examined the gender-differences in microsomal erythromycin N-demethylation activity (a CYP3A marker) and CYP3A protein expression across test marsupial species. Significant differences in hepatic erythromycin N-demethylation activity were observed between male and female koalas, with the activity detected in female koalas being 2.5-fold higher compared to that in male koalas (p<0.01). No gender-differences were observed in tammar wallaby or Eastern grey kangaroo. Immunoblot analysis utilising anti-human CYP3A4 antibody detected immunoreactive proteins in liver microsomes from all test male and female marsupials including the koala, tammar wallaby, and Eastern grey kangaroo, with no gender-differences detected across test marsupials. A 1610 bp koala hepatic CYP3A complete cDNA, designated CYP3A78, was cloned by reverse transcription-polymerase chain reaction approaches. It displays 64% nucleotide and 57% amino acid sequence identity to the Eastern grey kangaroo CYP3A70. The CYP3A78 cDNA encodes a protein of 515 amino acids, shares approximately 68% nucleotide and 56% amino acid sequence identity to human CYP3A4, and displays high sequence similarity to other published mammalian CYP3As from human, monkey, cow, pig, dog, rat, rabbit, mouse, hamster, and guinea pig. Collectively, this study provides primary molecular data regarding koala hepatic CYP3A78 gene and enables further functional analyses of CYP

In Vitro Evaluation of Reversible and Time-Dependent Inhibitory Effects of Kalanchoe crenata on CYP2C19 and CYP3A4 Activities.

Science.gov (United States)

Awortwe, Charles; Manda, Vamshi K; Avonto, Cristina; Khan, Shabana I; Khan, Ikhlas A; Walker, Larry A; Bouic, Patrick J; Rosenkranz, Bernd

2015-01-01

Kalanchoe crenata popularly known as "dog's liver" is used in most African countries for the treatment of chronic diseases such as diabetes, asthma and HIV/AIDS related infections. The evaluation of K. crenata for herb-drug interactions has not been reported. This study therefore aims to evaluate the risk of K. crenata for herb-drug interaction in vitro. Crude methanol and fractions of K. crenata were incubated and preincubated with recombinant human CYP2C19 and CYP3A4. Comparative studies were conducted in both human liver microsomes and recombinant human CYP to ascertain the inhibition profile of the crude extract and the various fractions. The cocktail approach of recombinant human CYPs was conducted to confirm the inhibition potential of the fractions in the presence of other CYPs. The results showed significant time-dependent inhibition of tested samples on CYP3A4 with crude methanol (39KC), fractions 45A, 45B and 45D given IC50 fold decrease of 3.29, 2.26, 1.91 and 1.49, respective. Time dependent kinetic assessment of 39KC and 45D showed KI and kinact values for 39KC as 1.77 µg/mL and 0.091 min(-1) while that of 45D were 6.45 µg/mL and 0.024 min(-1), respectively. Determination of kinact based on IC50 calculations yielded 0.015 and 0.04 min(-1) for 39KC and 45D, respectively. Cocktail approach exhibited fold decreases in IC50 for all test fractions on CYP3A4 within the ranges of 2.10 - 4.10. At least one phytoconstituent in the crude methanol extract of Kalanchoe crenata is a reversible and time-dependent inhibitor of CYP3A4.

Effect of Genetic Variability in the CYP4F2, CYP4F11, and CYP4F12 Genes on Liver mRNA Levels and Warfarin Response

Directory of Open Access Journals (Sweden)

J. E. Zhang

2017-05-01

Full Text Available Genetic polymorphisms in the gene encoding cytochrome P450 (CYP 4F2, a vitamin K oxidase, affect stable warfarin dose requirements and time to therapeutic INR. CYP4F2 is part of the CYP4F gene cluster, which is highly polymorphic and exhibits a high degree of linkage disequilibrium, making it difficult to define causal variants. Our objective was to examine the effect of genetic variability in the CYP4F gene cluster on expression of the individual CYP4F genes and warfarin response. mRNA levels of the CYP4F gene cluster were quantified in human liver samples (n = 149 obtained from a well-characterized liver bank and fine mapping of the CYP4F gene cluster encompassing CYP4F2, CYP4F11, and CYP4F12 was performed. Genome-wide association study (GWAS data from a prospective cohort of warfarin-treated patients (n = 711 was also analyzed for genetic variations across the CYP4F gene cluster. In addition, SNP-gene expression in human liver tissues and interactions between CYP4F genes were explored in silico using publicly available data repositories. We found that SNPs in CYP4F2, CYP4F11, and CYP4F12 were associated with mRNA expression in the CYP4F gene cluster. In particular, CYP4F2 rs2108622 was associated with increased CYP4F2 expression while CYP4F11 rs1060467 was associated with decreased CYP4F2 expression. Interestingly, these CYP4F2 and CYP4F11 SNPs showed similar effects with warfarin stable dose where CYP4F11 rs1060467 was associated with a reduction in daily warfarin dose requirement (∼1 mg/day, Pc = 0.017, an effect opposite to that previously reported with CYP4F2 (rs2108622. However, inclusion of either or both of these SNPs in a pharmacogenetic algorithm consisting of age, body mass index (BMI, gender, baseline clotting factor II level, CYP2C9∗2 rs1799853, CYP2C9∗3 rs1057910, and VKORC1 rs9923231 improved warfarin dose variability only by 0.5–0.7% with an improvement in dose prediction accuracy of ∼1–2%. Although there is complex

Effects of triazole fungicides on androgenic disruption and CYP3A4 enzyme activity.

Science.gov (United States)

Lv, Xuan; Pan, Liumeng; Wang, Jiaying; Lu, Liping; Yan, Weilin; Zhu, Yanye; Xu, Yiwen; Guo, Ming; Zhuang, Shulin

2017-03-01

Triazole fungicides are widely used as broad-spectrum fungicides, non-steroidal antiestrogens and for various industrial applications. Their residues have been frequently detected in multiple environmental and human matrices. The increasingly reported toxicity incidents have led triazole fungicides as emerging contaminants of environmental and public health concern. However, whether triazole fungicides behave as endocrine disruptors by directly mimicking environmental androgens/antiandrogens or exerting potential androgenic disruption indirectly through the inhibition of cytochrome P450 (CYP450) enzyme activity is yet an unresolved question. We herein evaluated five commonly used triazole fungicides including bitertanol, hexaconazole, penconazole, tebuconazole and uniconazole for the androgenic and anti-androgenic activity using two-hybrid recombinant human androgen receptor (AR) yeast bioassay and comparatively evaluated their effects on enzymatic activity of CYP3A4 by P450-Glo™ CYP3A4 bioassay. All five fungicides showed moderate anti-androgenic activity toward human AR with the IC 50 ranging from 9.34 μM to 79.85 μM. The anti-androgenic activity remained no significant change after the metabolism mediated by human liver microsomes. These fungicides significantly inhibited the activity of CYP3A4 at the environmental relevant concentrations and the potency ranks as tebuconazole > uniconazole > hexaconazole > penconazole > bitertanol with the corresponding IC 50 of 0.81 μM, 0.93 μM, 1.27 μM, 2.22 μM, and 2.74 μM, respectively. We found that their anti-androgenic activity and the inhibition potency toward CYP3A4 inhibition was significantly correlated (R 2 between 0.83 and 0.97, p pesticides and structurally similar chemicals should fully consider potential androgenic disrupting effects and the influences on the activity of CYP450s. Copyright © 2016 Elsevier Ltd. All rights reserved.

Epigenetic Determinants of CYP1A1 Induction by the Aryl Hydrocarbon Receptor Agonist 3,3',4,4',5-Pentachlorobiphenyl (PCB 126

Directory of Open Access Journals (Sweden)

Sabine U. Vorrink

2014-08-01

Full Text Available Many enzymes involved in xenobiotic metabolism, including cytochrome P450 (CYP 1A1, are regulated by the aryl hydrocarbon receptor (AhR. 3,3',4,4',5-Penta chlorobiphenyl (PCB 126 is a potent ligand for AhR and can thus induce the expression of CYP1A1. Interestingly, we observed that human carcinoma cell lines derived from different types of epithelial cells displayed divergent degrees of CYP1A1 induction after exposure to PCB 126. Since epigenetic mechanisms are known to be involved in cell type-specific gene expression, we sought to assess the epigenetic determinants of CYP1A1 induction in these carcinoma cell lines. In contrast to HepG2 hepatocarcinoma cells, HeLa cervical carcinoma cells showed significantly lower levels of CYP1A1 mRNA expression following PCB 126 exposure. Our results show that the two cell lines maintained differences in the chromatin architecture along the CYP1A1 promoter region. Furthermore, treatment with the epigenetic modifiers, trichostatin A (TSA and 5-aza-2'-deoxycytidine (5-Aza-dC, significantly increased the expression of CYP1A1 after PCB 126 treatment in HeLa cells. However, we did not observe apparent differences in methylation levels or specific location of CpG DNA methylation between the two cell lines in the analyzed CYP1A1 promoter region. Taken together, our findings suggest that the differences in CYP1A1 expression between HepG2 and HeLa cells are due to differences in the chromatin architecture of the CYP1A1 promoter and thus establish a role of epigenetic regulation in cell-specific CYP1A1 expression.

Cytochrome P450 CYP3A in marsupials: cloning and identification of the first CYP3A subfamily member, isoform 3A70 from Eastern gray kangaroo (Macropus giganteus).

Science.gov (United States)

El-Merhibi, Adaweyah; Ngo, Suong N T; Marchant, Ceilidh L; Height, Tamara A; Stupans, Ieva; McKinnon, Ross A

2012-09-15

Australian marsupials are unique fauna that have evolved and adapted to unique environments and thus it is likely that their detoxification systems differ considerably from those of well-studied eutherian mammals. Knowledge of these processes in marsupials is therefore vital to understanding the consequences of exposure to xenobiotics. Cytochromes P450 (CYPs) are critically important in the oxidative metabolism of a diverse array of both xenobiotics and endogenous substrates. In this study we have cloned and characterized CYP3A70, the first identified member of the CYP3A gene subfamily from Eastern gray kangaroo (Macropus giganteus). A 1665 base pair kangaroo hepatic CYP3A complete cDNA, designated CYP3A70, was cloned by reverse transcription-polymerase chain reaction approaches, which encodes a protein of 506 amino acids. The CYP3A70 cDNA shares approximately 71% nucleotide and 65% amino acid sequence homology to human CYP3A4 and displays high sequence similarity to other published mammalian CYP3As from human, monkey, cow, pig, dog, rat, rabbit, mouse, hamster, and guinea pig. Transfection of the CYP3A70 cDNAs into 293T cells resulted in stable cell lines expressing a CYP3A immuno-reactive protein that was recognized by a goat anti-human CYP3A4 polyclonal antibody. The anti-human CYP3A4 antibody also detected immunoreactive proteins in liver microsomes from all test marsupials, including the kangaroo, koala, wallaby, and wombat, with multiple CYP3A immunoreactive bands observed in kangaroo and wallaby tissues. Relatively, very low CYP catalytic activity was detected for the kangaroo CYP3A70 cDNA-expressed proteins (19.6 relative luminescent units/μg protein), which may be due to low protein expression levels. Collectively, this study provides primary molecular data regarding the Eastern kangaroo hepatic CYP3A70 gene and enables further functional analyses of CYP3A enzymes in marsupials. Copyright © 2012 Elsevier B.V. All rights reserved.

Evidences for CYP3A4 autoactivation in the desulfuration of dimethoate by the human liver.

Science.gov (United States)

Buratti, Franca M; Testai, Emanuela

2007-11-20

Dimethoate (DIM) is an organophosphorothionate (OPT) pesticide used worldwide as a systemic insecticide and acaricide. It is characterized by low-to-moderate acute mammalian toxicity; similarly to the other OPT pesticides, its mode of action is mediated by the inhibition of acetylcholinesterase (AChE), exerted by its toxic metabolite dimethoate-oxon or omethoate (OME), which is also used as a direct acting pesticide. Human hepatic DIM bioactivation to the toxic metabolite OME has been characterized by using c-DNA expressed human CYPs and human liver microsomes (HLM) also in the presence of CYP-specific chemical inhibitors, with a method based on AChE inhibition. The obtained kinetic parameters and AChE IC(50) have been compared with those previously obtained with other OPTs, indicating a lower efficiency in DIM desulfuration reaction and a lower potency in inhibiting AChE. Results showed that, similarly to the other OPTs tested so far, at low DIM concentration OME formation is mainly catalysed by CYP1A2, while the role of 3A4 is relevant at high DIM levels. Differently from the other OPTs, DIM desulfuration reaction showed an atypical kinetic profile, likely due to CYP3A4 autoactivation. The sigmoidicity degree of the activity curve increased with the level of CYP3A4 in HLM or disappeared in the presence of a CYP3A4 chemical inhibitor. This atypical kinetic behaviour can be considered one of the possible explanations for the recent findings that among patients hospitalized following OPT intoxication, DIM ingestion gave different symptoms and more severe poisoning (23.1% of fatal cases versus total) than chlorpyrifos (8% of deaths), which has a lower LD(50) value. Since DIM-poisoned patients poorly responded to pralidoxime, the possibility to use CYP3A4 inhibitors could be considered as a complementary treatment.

Polychlorinated biphenyl (PCB) induction of CYP3A4 enzyme activity in healthy Faroese adults

International Nuclear Information System (INIS)

Petersen, Maria Skaalum; Halling, Jonrit; Damkier, Per; Nielsen, Flemming; Grandjean, Philippe; Weihe, Pal; Brosen, Kim

2007-01-01

The CYP3A4 enzyme is, along with other cytochrome P450 enzymes, involved in the metabolism of environmental pollutants and is highly inducible by these substances. A commercial polychlorinated biphenyl (PCB) mixture, 1,1,1,-trichloro-2-(o-chlorophenyl), 2-(p'-chlorophenyl)ethane (o,p'-DDT) and 1,1,-dichloro-2,2-bis (p-chlorophenyl)ethene (p,p'-DDE) are known to induce CYP3A4 activity through activation of nuclear receptors, such as the pregnane X receptor. However, this induction of CYP3A4 has not yet been investigated in humans. Thus, the aim of the study was to determine the variability of the CYP3A4 phenotype in regard to increased concentrations of PCBs and other persistent organohalogen pollutants (POPs) in healthy Faroese adults. In 310 randomly selected Faroese residents aged 18-60 years, the CYP3A4 activity was determined based on the urinary 6β-hydroxycortisol/cortisol (6β-OHC/FC) ratio. POP exposures were assessed by measuring their concentrations in serum lipid. The results showed a unimodal distribution of the 6β-OHC/FC ratio with values ranging from 0.58 to 27.38. Women had a slightly higher 6β-OHC/FC ratio than men (p = 0.07). Confounder-adjusted multiple regression analysis showed significant associations between 6β-OHC/FC ratios and ΣPCB, PCB-TEQ and p,p'-DDE, o,p'-DDT and HCB, respectively, but the associations were statistically significant for men only

CAR/PXR provide directives for Cyp3a41 gene regulation differently from Cyp3a11.

Science.gov (United States)

Anakk, S; Kalsotra, A; Kikuta, Y; Huang, W; Zhang, J; Staudinger, J L; Moore, D D; Strobel, H W

2004-01-01

This study reports that Cyp3a41 gene contains 13 exons and is localized on the chromosome 5. CYP3A41 is a female-specific isoform that is predominantly expressed in the liver. Estrogen signaling is not responsible for its female specificity. CYP3A41 expression in kidney and brain is observed only in 50% of mice examined. PXR mediates dexamethasone-dependent suppression of CYP3A41. In contrast to CYP3A11, CYP3A41 expression is not induced by pregnenolone-16alpha-carbonitrile (PCN) in wild-type mice, but is significantly suppressed by PCN in PXR(-/-) mice. Phenobarbital and TCPOBOP induce CYP3A11 expression only in the presence of CAR, but have no effect on CYP3A41 expression. Immunoblot and erythromycin demethylase activity analysis reveal robust CYP3A induction after PCN treatment, which is poorly correlated to CYP3A41. These findings suggest a differential role for CAR/PXR in regulating individual CYP3A isoforms by previously characterized CYP3A inducers.

Mutation analysis of the human CYP3A4 gene 5' regulatory region: population screening using non-radioactive SSCP.

Science.gov (United States)

Hamzeiy, Hossein; Vahdati-Mashhadian, Nasser; Edwards, Helen J; Goldfarb, Peter S

2002-03-20

Human CYP3A4 is the major cytochrome P450 isoenzyme in adult human liver and is known to metabolise many xenobiotic and endogenous compounds. There is substantial inter-individual variation in the hepatic levels of CYP3A4. Although, polymorphic mutations have been reported in the 5' regulatory region of the CYP3A4 gene, those that have been investigated so far do not appear to have any effect on gene expression. To determine whether other mutations exist in this region of the gene, we have performed a new population screen on a panel of 101 human DNA samples. A 1140 bp section of the 5' proximal regulatory region of the CYP3A4 gene, containing numerous regulatory motifs, was amplified from genomic DNA as three overlapping segments. The 300 bp distal enhancer region at -7.9kb containing additional regulatory motifs was also amplified. Mutation analysis of the resulting PCR products was carried out using non-radioactive single strand conformation polymorphism (SSCP) and confirmatory sequencing of both DNA strands in those samples showing extra SSCP bands. In addition to detection of the previously reported CYP3A4*1B allele in nine subjects, three novel alleles were found: CYP3A4*1E (having a T-->A transversion at -369 in one subject), CYP3A4*1F (having a C-->G tranversion at -747 in 17 subjects) and CYP3A4*15B containing a nine-nucleotide insertion between -845 and -844 linked to an A-->G transition at -392 and a G-->A transition in exon 6 (position 485 in the cDNA) in one subject. All the novel alleles were heterozygous. No mutations were found in the upstream distal enhancer region. Our results clearly indicate that this rapid and simple SSCP approach can reveal mutant alleles in drug metabolising enzyme genes. Detection and determination of the frequency of novel alleles in CYP3A4 will assist investigation of the relationship between genotype, xenobiotic metabolism and toxicity in the CYP3A family of isoenzymes.

In vitro modulatory effects of Terminalia arjuna, arjunic acid, arjunetin and arjungenin on CYP3A4, CYP2D6 and CYP2C9 enzyme activity in human liver microsomes

Directory of Open Access Journals (Sweden)

Alice Varghese

2015-01-01

Full Text Available Terminalia arjuna is a tree having an extensive medicinal potential in cardiovascular disorders. Triterpenoids are mainly responsible for cardiovascular properties. Alcoholic and aqueous bark extracts of T. arjuna, arjunic acid, arjunetin and arjungenin were evaluated for their potential to inhibit CYP3A4, CYP2D6 and CYP2C9 enzymes in human liver microsomes. We have demonstrated that alcoholic and aqueous bark extract of T. arjuna showed potent inhibition of all three enzymes in human liver microsomes with IC50 values less than 50 μg/mL. Arjunic acid, arjunetin and arjungenin did not show significant inhibition of CYP enzymes in human liver microsomes. Enzyme kinetics studies suggested that the extracts of arjuna showed reversible non-competitive inhibition of all the three enzymes in human liver microsomes. Our findings suggest strongly that arjuna extracts significantly inhibit the activity of CYP3A4, CYP2D6 and CYP2C9 enzymes, which is likely to cause clinically significant drug–drug interactions mediated via inhibition of the major CYP isozymes.

Effect of Methamphetamine on Spectral Binding, Ligand Docking and Metabolism of Anti-HIV Drugs with CYP3A4

Science.gov (United States)

Ande, Anusha; Wang, Lei; Vaidya, Naveen K.; Li, Weihua; Kumar, Santosh; Kumar, Anil

2016-01-01

Cytochrome P450 3A4 (CYP3A4) is the major drug metabolic enzyme, and is involved in the metabolism of antiretroviral drugs, especially protease inhibitors (PIs). This study was undertaken to examine the effect of methamphetamine on the binding and metabolism of PIs with CYP3A4. We showed that methamphetamine exhibits a type I spectral change upon binding to CYP3A4 with δAmax and KD of 0.016±0.001 and 204±18 μM, respectively. Methamphetamine-CYP3A4 docking showed that methamphetamine binds to the heme of CYP3A4 in two modes, both leading to N-demethylation. We then studied the effect of methamphetamine binding on PIs with CYP3A4. Our results showed that methamphetamine alters spectral binding of nelfinavir but not the other type I PIs (lopinavir, atazanavir, tipranavir). The change in spectral binding for nelfinavir was observed at both δAmax (0.004±0.0003 vs. 0.0068±0.0001) and KD (1.42±0.36 vs.2.93±0.08 μM) levels. We further tested effect of methamphetamine on binding of 2 type II PIs; ritonavir and indinavir. Our results showed that methamphetamine alters the ritonavir binding to CYP3A4 by decreasing both the δAmax (0.0038±0.0003 vs. 0.0055±0.0003) and KD (0.043±0.0001 vs. 0.065±0.001 nM), while indinavir showed only reduced KD in presence of methamphetamine (0.086±0.01 vs. 0.174±0.03 nM). Furthermore, LC-MS/MS studies in high CYP3A4 human liver microsomes showed a decrease in the formation of hydroxy ritonavir in the presence of methamphetamine. Finally, CYP3A4 docking with lopinavir and ritonavir in the absence and presence of methamphetamine showed that methamphetamine alters the docking of ritonavir, which is consistent with the results obtained from spectral binding and metabolism studies. Overall, our results demonstrated differential effects of methamphetamine on the binding and metabolism of PIs with CYP3A4. These findings have clinical implication in terms of drug dose adjustment of antiretroviral medication, especially with ritonavir

Reductive metabolism of oxymatrine is catalyzed by microsomal CYP3A4

Directory of Open Access Journals (Sweden)

Liu W

2015-10-01

Full Text Available Wenqin Liu,1,2,* Jian Shi,1,2,* Lijun Zhu,2 Lingna Dong,1 Feifei Luo,2 Min Zhao,2 Ying Wang,2 Ming Hu,2,3 Linlin Lu,2 Zhongqiu Liu1,2 1Department of Pharmaceutics, School of Pharmaceutical Sciences, Southern Medical University, Guangzhou, Guangdong, People’s Republic of China; 2International Institute for Translational Chinese Medicine, Guangzhou University of Chinese Medicine, Guangzhou, Guangdong, People’s Republic of China; 3Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, University of Houston, Houston, TX, USA *These authors contributed equally to this work Abstract: Oxymatrine (OMT is a pharmacologically active primary quinolizidine alkaloid with various beneficial and toxic effects. It is confirmed that, after oral administration, OMT could be transformed to the more toxic metabolite matrine (MT, and this process may be through the reduction reaction, but the study on the characteristics of this transformation is limited. The aim of this study was to investigate the characteristics of this transformation of OMT in the human liver microsomes (HLMs and human intestinal microsomes (HIMs and the cytochrome P450 (CYP isoforms involved in this transformation. The current studies demonstrated that OMT could be metabolized to MT rapidly in HLMs and HIMs and CYP3A4 greatly contributed to this transformation. All HLMs, HIMs, and CYP3A4 isoform mediated reduction reaction followed typical biphasic kinetic model, and Km, Vmax, and CL were significant higher in HLMs than those in HIMs. Importantly, different oxygen contents could significantly affect the metabolism of OMT, and with the oxygen content decreased, the formation of metabolite was increased, suggesting this transformation was very likely a reduction reaction. Results of this in vitro study elucidated the metabolic pathways and characteristics of metabolism of OMT to MT and would provide a theoretical basis and guidance for the safe application of OMT

Modeling of drug-mediated CYP3A4 induction by using human iPS cell-derived enterocyte-like cells

International Nuclear Information System (INIS)

Negoro, Ryosuke; Takayama, Kazuo; Nagamoto, Yasuhito; Sakurai, Fuminori; Tachibana, Masashi; Mizuguchi, Hiroyuki

2016-01-01

Many drugs have potential to induce the expression of drug-metabolizing enzymes, particularly cytochrome P450 3A4 (CYP3A4), in small intestinal enterocytes. Therefore, a model that can accurately evaluate drug-mediated CYP3A4 induction is urgently needed. In this study, we overlaid Matrigel on the human induced pluripotent stem cells-derived enterocyte-like cells (hiPS-ELCs) to generate the mature hiPS-ELCs that could be applied to drug-mediated CYP3A4 induction test. By overlaying Matrigel in the maturation process of enterocyte-like cells, the gene expression levels of intestinal markers (VILLIN, sucrase-isomaltase, intestine-specific homeobox, caudal type homeobox 2, and intestinal fatty acid-binding protein) were enhanced suggesting that the enterocyte-like cells were maturated by Matrigel overlay. The percentage of VILLIN-positive cells in the hiPS-ELCs found to be approximately 55.6%. To examine the CYP3A4 induction potential, the hiPS-ELCs were treated with various drugs. Treatment with dexamethasone, phenobarbital, rifampicin, or 1α,25-dihydroxyvitamin D3 resulted in 5.8-fold, 13.4-fold, 9.8-fold, or 95.0-fold induction of CYP3A4 expression relative to that in the untreated controls, respectively. These results suggest that our hiPS-ELCs would be a useful model for CYP3A4 induction test. - Highlights: • The hiPS-ELCs were matured by Matrigel overlay. • The hiPS-ELCs expressed intestinal nuclear receptors, such as PXR, GR and VDR. • The hiPS-ELC is a useful model for the drug-mediated CYP3A4 induction test.

Modeling of drug-mediated CYP3A4 induction by using human iPS cell-derived enterocyte-like cells

Energy Technology Data Exchange (ETDEWEB)

Negoro, Ryosuke [Laboratory of Biochemistry and Molecular Biology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka 565-0871 (Japan); Takayama, Kazuo [Laboratory of Biochemistry and Molecular Biology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka 565-0871 (Japan); The Keihanshin Consortium for Fostering the Next Generation of Global Leaders in Research (K-CONNEX), Kyoto University, Kyoto 606-8302 (Japan); Laboratory of Hepatocyte Regulation, National Institute of Biomedical Innovation, Health and Nutrition, Osaka 567-0085 (Japan); Nagamoto, Yasuhito [Laboratory of Biochemistry and Molecular Biology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka 565-0871 (Japan); Laboratory of Hepatocyte Regulation, National Institute of Biomedical Innovation, Health and Nutrition, Osaka 567-0085 (Japan); Sakurai, Fuminori [Laboratory of Biochemistry and Molecular Biology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka 565-0871 (Japan); Laboratory of Regulatory Sciences for Oligonucleotide Therapeutics, Clinical Drug Development Project, Graduate School of Pharmaceutical Sciences, Osaka University Osaka 565-0871 (Japan); Tachibana, Masashi [Laboratory of Biochemistry and Molecular Biology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka 565-0871 (Japan); Mizuguchi, Hiroyuki, E-mail: mizuguch@phs.osaka-u.ac.jp [Laboratory of Biochemistry and Molecular Biology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka 565-0871 (Japan); Laboratory of Hepatocyte Regulation, National Institute of Biomedical Innovation, Health and Nutrition, Osaka 567-0085 (Japan); Global Center for Medical Engineering and Informatics, Osaka University, Osaka 565-0871 (Japan)

2016-04-15

Many drugs have potential to induce the expression of drug-metabolizing enzymes, particularly cytochrome P450 3A4 (CYP3A4), in small intestinal enterocytes. Therefore, a model that can accurately evaluate drug-mediated CYP3A4 induction is urgently needed. In this study, we overlaid Matrigel on the human induced pluripotent stem cells-derived enterocyte-like cells (hiPS-ELCs) to generate the mature hiPS-ELCs that could be applied to drug-mediated CYP3A4 induction test. By overlaying Matrigel in the maturation process of enterocyte-like cells, the gene expression levels of intestinal markers (VILLIN, sucrase-isomaltase, intestine-specific homeobox, caudal type homeobox 2, and intestinal fatty acid-binding protein) were enhanced suggesting that the enterocyte-like cells were maturated by Matrigel overlay. The percentage of VILLIN-positive cells in the hiPS-ELCs found to be approximately 55.6%. To examine the CYP3A4 induction potential, the hiPS-ELCs were treated with various drugs. Treatment with dexamethasone, phenobarbital, rifampicin, or 1α,25-dihydroxyvitamin D3 resulted in 5.8-fold, 13.4-fold, 9.8-fold, or 95.0-fold induction of CYP3A4 expression relative to that in the untreated controls, respectively. These results suggest that our hiPS-ELCs would be a useful model for CYP3A4 induction test. - Highlights: • The hiPS-ELCs were matured by Matrigel overlay. • The hiPS-ELCs expressed intestinal nuclear receptors, such as PXR, GR and VDR. • The hiPS-ELC is a useful model for the drug-mediated CYP3A4 induction test.

Contrasting exome constancy and regulatory region variation in the gene encoding CYP3A4: an examination of the extent and potential implications.

Science.gov (United States)

Creemer, Olivia J; Ansari-Pour, Naser; Ekong, Rosemary; Tarekegn, Ayele; Plaster, Christopher; Bains, Ripudaman K; Itan, Yuval; Bekele, Endashaw; Bradman, Neil

2016-06-01

CYP3A4 expression varies up to 100-fold among individuals, and, to date, genetic causes remain elusive. As a major drug-metabolizing enzyme, elucidation of such genetic causes would increase the potential for introducing personalized dose adjustment of therapies involving CYP3A4 drug substrates. The foetal CYP3A isoform, CYP3A7, is reported to be expressed in ∼10% of European adults and may thus contribute towards the metabolism of endogenous substances and CYP3A drug substrates. However, little is known about the distribution of the variant expressed in the adult. We resequenced the exons, flanking introns, regulatory elements and 3'UTR of CYP3A4 in five Ethiopian populations and incorporated data from the 1000 Genomes Project. Using bioinformatic analysis, we assessed likely consequences of observed CYP3A4 genomic variation. We also conducted the first extensive geographic survey of alleles associated with adult expression of CYP3A7 - that is, CYP3A7*1B and CYP3A7*1C. Ethiopia contained 60 CYP3A4 variants (26 novel) and more variants (>1%) than all non-African populations combined. No nonsynonymous mutation was found in the homozygous form or at more than 2.8% in any population. Seventy-nine per cent of haplotypes contained 3'UTR and/or regulatory region variation with striking pairwise population differentiation, highlighting the potential for interethnic variation in CYP3A4 expression. Conversely, coding region variation showed that significant interethnic variation is unlikely at the protein level. CYP3A7*1C was found at up to 17.5% in North African populations and in significant linkage disequilibrium with CYP3A5*3, indicating that adult expression of the foetal isoform is likely to be accompanied by reduced or null expression of CYP3A5.

Effect of a new functional CYP3A4 polymorphism on calcineurin inhibitors' dose requirements and trough blood levels in stable renal transplant patients.

Science.gov (United States)

Elens, Laure; van Schaik, Ron H; Panin, Nadtha; de Meyer, Martine; Wallemacq, Pierre; Lison, Dominique; Mourad, Michel; Haufroid, Vincent

2011-10-01

CYP3A4 is involved in the oxidative metabolism of many drugs and xenobiotics including the immunosuppressants tacrolimus (Tac) and cyclosporine (CsA). The objective of the study was to assess the potential influence of a new functional SNP in CYP3A4 on the pharmacokinetic parameters assessed by dose requirements and trough blood levels of both calcineurin inhibitors (CNI) in stable renal transplant patients. A total of 99 stable renal transplant patients receiving either Tac (n = 49) or CsA (n = 50) were genotyped for the CYP3A4 intron 6 C>T (rs35599367) and CYP3A5*3 SNPs. Trough blood levels ([Tac](0) or [CsA](0) in ng/ml), dose-adjusted [Tac](0) or [CsA](0) (ng/ml per mg/kg bodyweight) as well as doses (mg/kg bodyweight) required to achieve target concentrations were compared among patients according to allelic status for CYP3A4 and CYP3A5. Dose-adjusted concentrations were 2.0- and 1.6-fold higher in T-variant allele carriers for the CYP3A4 intron 6 C>T SNP compared with homozygous CC for Tac and CsA, respectively. When CYP3A4/CYP3A5 genotypes were combined, the difference was even more striking as the so-defined CYP3A poor metabolizer group presented dose-adjusted concentration 1.6- and 4.1-fold higher for Tac, and 1.5- and 2.2-fold higher for CsA than the intermediate metabolizer and extensive metabolizer groups, respectively. Multiple linear regression analysis revealed that, taken together, both CYP3A4 intron 6 and CYP3A5*3 SNPs explained more than 60 and 20% of the variability observed in dose-adjusted [Tac](0) and [CsA](0), respectively. The CYP3A4 intron 6 C>T polymorphism is associated with altered Tac and CsA metabolism. CYP3A4 intron 6 C>T along with CYP3A5*3 (especially for Tac) pharmacogenetic testing performed just before transplantation may help identifying patients at risk of CNI overexposure and contribute to limit CNI-related nephrotoxicity by refining the starting dose according to their genotype. Original submitted 5 May 2011; Revision

The mibefradil derivative NNC55-0396, a specific T-type calcium channel antagonist, exhibits less CYP3A4 inhibition than mibefradil.

Science.gov (United States)

Bui, Peter H; Quesada, Arnulfo; Handforth, Adrian; Hankinson, Oliver

2008-07-01

A novel mibefradil derivative, NNC55-0396, designed to be hydrolysis-resistant, was shown to be a selective T-type Ca(2+) channel inhibitor without L-type Ca(2+) channel efficacy. However, its effects on cytochromes P450 (P450s) have not previously been examined. We investigated the inhibitory effects of NNC55-0396 toward seven major recombinant human P450s--CYP3A4, CYP2D6, CYP1A2, CYP2C9, CYP2C8, CYPC19, and CYP2E1--and compared its effects with those of mibefradil and its hydrolyzed metabolite, Ro40-5966. Our results show that CYP3A4 and CYP2D6 are the two P450s most affected by mibefradil, Ro40-5966, and NNC55-0396. Mibefradil (IC(50) = 33 +/- 3 nM, K(i) = 23 +/- 0.5 nM) and Ro40-5966 (IC(50) = 30 +/- 7.8 nM, K(i) = 21 +/- 2.8 nM) have a 9- to 10-fold greater inhibitory activity toward recombinant CYP3A4 benzyloxy-4-trifluoromethylcoumarin-O-debenzylation activity than NNC55-0396 (IC(50) = 300 +/- 30 nM, K(i) = 210 +/- 6 nM). More dramatically, mibefradil (IC(50) = 566 +/- 71 nM, K(i) = 202 +/- 39 nM) shows 19-fold higher inhibition of CYP3A-associated testosterone 6beta-hydroxylase activity in human liver microsomes compared with NNC55-0396 (IC(50) = 11 +/- 1.1 microM, K(i) = 3.9 +/- 0.4 microM). Loss of testosterone 6beta-hydroxylase activity by recombinant CYP3A4 was shown to be time- and concentration-dependent with both compounds. However, NNC55-0396 (K(I) = 3.87 microM, K(inact) = 0.061/min) is a much less potent mechanism-based inhibitor than mibefradil (K(I) = 83 nM, K(inact) = 0.048/min). In contrast, NNC55-0396 (IC(50) = 29 +/- 1.2 nM, K(i) = 2.8 +/- 0.3 nM) and Ro40-5966 (IC(50) = 46 +/- 11 nM, K(i) = 4.5 +/- 0.02 nM) have a 3- to 4-fold greater inhibitory activity toward recombinant CYP2D6 than mibefradil (IC(50) = 129 +/- 21 nM, K(i) = 12.7 +/- 0.9 nM). Our results suggest that NNC55-0396 could be a more favorable T-type Ca(2+) antagonist than its parent compound, mibefradil, which was withdrawn from the market because of strong inhibition of CYP3A4.

PXR-dependent induction of human CYP3A4 gene expression by organochlorine pesticides.

Science.gov (United States)

Coumoul, Xavier; Diry, Monique; Barouki, Robert

2002-11-15

OCP are xenobiotics which display various toxic effects on animal and human health. One of their effects is to bind and activate estrogen receptor alpha (ERalpha). We have previously studied the down-regulation of induced CYP1A1 (cytochrome P450) expression by this class of molecules in mammary carcinoma cells and shown the importance of ERalpha in this process. However, an alternative mechanism was suggested by those experiments in hepatoma cells. In this study, we have performed Northern blot and transient transfection assays in various cell lines and shown that OCP activate human pregnane X receptor (PXR) and subsequent CYP3A4 mRNA expression. This effect is mediated by the distal xenobiotic responsive element modulator of the promoter. The induction of CYP3A4 by OCP was dose-dependent within the 1-10 microM range. The data suggest that chronic exposure to OCP could alter a major metabolite pathway in human liver and putatively modify the pharmacokinetics of drugs and pollutants.

A case report of a patient carrying CYP2C9*3/4 genotype with extremely low warfarin dose requirement.

Science.gov (United States)

Lee, Soo Youn; Nam, Myung Hyun; Kim, June Soo; Kim, Jong Won

2007-06-01

We report a case of intolerance to warfarin dosing due to impaired drug metabolism in a patient with CYP2C9*3/*4. A 73-yr-old woman with atrial fibrilation was taking warfarin. She attained a high prothrombin time international normalized ratio (INR) at the standard doses during the induction of anticoagulation and extremely low dose of warfarin (6.5 mg/week) was finally chosen to reach the target INR. Genotyping for CYP2C9 revealed that this patient had a genotype CYP2C9*3/*4. This is the first Korean compound heterozygote for CYP2C9*3 and *4. This case suggests the clinical usefulness of pharmacogenetic testing for individualized dosage adjustments of warfarin.

Association of CYP2B6, CYP3A5, and CYP2C19 genetic polymorphisms with sibutramine pharmacokinetics in healthy Korean subjects.

Science.gov (United States)

Kim, K A; Song, W K; Park, J Y

2009-11-01

We assessed the association of CYP2B6, CYP3A5, and CYP2C19 polymorphisms with sibutramine pharmacokinetics. Forty six healthy male subjects were enrolled, and their CYP2B6 (*4 and *6), CYP3A5 (*3), and CYP2C19 (*2, and *3) genotypes were analyzed. After a single 15-mg dose of sibutramine was administered, plasma concentrations of sibutramine and its metabolites, M1 and M2, were measured. CYP2B6 and CYP3A5 polymorphisms did not affect the pharmacokinetics of sibutramine and its metabolites. However, the CYP2C19 genotype substantially influenced plasma levels of sibutramine and its metabolites. The mean area under the curve (AUC) of sibutramine in CYP2C19 intermediate metabolizers (IMs; *1/*2 or *1/*3) and poor metabolizers (PMs; *2/*2, *2/*3)) was 18.5 and 252.2% higher, respectively, than the AUC in extensive metabolizers (EMs, *1/*1) (P sibutramine.

Coadministration of gemfibrozil and itraconazole has only a minor effect on the pharmacokinetics of the CYP2C9 and CYP3A4 substrate nateglinide

Science.gov (United States)

Niemi, Mikko; Backman, Janne T; Juntti-Patinen, Laura; Neuvonen, Mikko; Neuvonen, Pertti J

2005-01-01

Background and aims Gemfibrozil, and particularly its combination with itraconazole, greatly increases the area under the plasma concentration-time curve [AUC(0, ∞)] and response to the cytochrome P450 (CYP) 2C8 and 3A4 substrate repaglinide. In vitro, gemfibrozil is a more potent inhibitor of CYP2C9 than of CYP2C8. Our aim was to investigate the effects of the gemfibrozil-itraconazole combination on the pharmacokinetics and pharmacodynamics of another meglitinide analogue, nateglinide, which is metabolized by CYP2C9 and CYP3A4. Methods In a randomized crossover study with two phases, nine healthy subjects took 600 mg gemfibrozil and 100 mg itraconazole (first dose 200 mg) twice daily or placebo for 3 days. On day 3, they ingested a single 30-mg dose of nateglinide. Plasma nateglinide and blood glucose concentrations were measured for up to 12 h. Results During the gemfibrozil-itraconazole phase, the AUC(0, ∞) and Cmax of nateglinide were 47% (range 23–74%; P gemfibrozil and itraconazole had no effect on the formation of the M7 metabolite of nateglinide but impaired its elimination. The blood glucose response to nateglinide was not significantly changed by coadministration of gemfibrozil and itraconazole. Conclusions The combination of gemfibrozil and itraconazole has only a limited influence on the pharmacokinetics of nateglinide. This is in marked contrast to the substantial effect of this combination on the pharmacokinetics of repaglinide. The findings suggest that in vivo gemfibrozil, probably due to its metabolites, is a much more potent inhibitor of CYP2C8 than of CYP2C9. PMID:16042675

Pregnane X receptor-dependent induction of the CYP3A4 gene by o,p'-1,1,1,-trichloro-2,2-bis (p-chlorophenyl)ethane.

Science.gov (United States)

Medina-Díaz, Irma M; Arteaga-Illán, Georgina; de León, Mario Bermudez; Cisneros, Bulmaro; Sierra-Santoyo, Adolfo; Vega, Libia; Gonzalez, Frank J; Elizondo, Guillermo

2007-01-01

CYP3A4, the predominant cytochrome P450 (P450) expressed in human liver and intestine, contributes to the metabolism of approximately half the drugs in clinical use today. CYP3A4 catalyzes the 6beta-hydroxylation of a number of steroid hormones and is involved in the bioactivation of environmental procarcinogens. The expression of CYP3A4 is affected by several stimuli, including environmental factors such as insecticides and pesticides. The o,p'-1,1,1,-trichloro-2,2-bis (p-chlorophenyl)ethane (DDT) isomer of DDT comprises approximately 20% of technical grade DDT, which is an organochloride pesticide. We have recently shown that o,p'-DDT exposure increases CYP3A4 mRNA levels in HepG2 cells. To determine the mechanism by which o,p'-DDT induces CYP3A4 expression, transactivation and electrophoretic mobility shift assays were carried out, revealing that o,p'-DDT activates the CYP3A4 gene promoter through the pregnane X receptor (PXR). CYP3A4 gene promoter activation resulted in both an increase in CYP3A4 mRNA levels and an increase in the total CYP3A4 activity in HepG2 cells. We also observed induction of CYP3A4 and mouse Cyp3a11 mRNA in the intestine of CYP3A4-transgenic mice after exposure to 1 mg/kg o,p'-DDT. At higher doses, a decrease of CYP3A4 inducibility was observed together with an increase in levels of interleukin 6 mRNA, a proinflammatory cytokine that strongly represses CYP3A4 transcription. The present study indicates that regulation of other genes under PXR control may be altered by o,p'-DDT exposure.

Altered Protein Expression of Cardiac CYP2J and Hepatic CYP2C, CYP4A, and CYP4F in a Mouse Model of Type II Diabetes—A Link in the Onset and Development of Cardiovascular Disease?

Directory of Open Access Journals (Sweden)

Benoit Drolet

2017-10-01

Full Text Available Arachidonic acid can be metabolized by cytochrome P450 (CYP450 enzymes in a tissue- and cell-specific manner to generate vasoactive products such as epoxyeicosatrienoic acids (EETs-cardioprotective and hydroxyeicosatetraenoic acids (HETEs-cardiotoxic. Type II diabetes is a well-recognized risk factor for developing cardiovascular disease. A mouse model of Type II diabetes (C57BLKS/J-db/db was used. After sacrifice, livers and hearts were collected, washed, and snap frozen. Total proteins were extracted. Western blots were performed to assess cardiac CYP2J and hepatic CYP2C, CYP4A, and CYP4F protein expression, respectively. Significant decreases in relative protein expression of cardiac CYP2J and hepatic CYP2C were observed in Type II diabetes animals compared to controls (CYP2J: 0.80 ± 0.03 vs. 1.05 ± 0.06, n = 20, p < 0.001; (CYP2C: 1.56 ± 0.17 vs. 2.21 ± 0.19, n = 19, p < 0.01. In contrast, significant increases in relative protein expression of both hepatic CYP4A and CYP4F were noted in Type II diabetes mice compared to controls (CYP4A: 1.06 ± 0.09 vs. 0.18 ± 0.01, n = 19, p < 0.001; (CYP4F: 2.53 ± 0.22 vs. 1.10 ± 0.07, n = 19, p < 0.001. These alterations induced by Type II diabetes in the endogenous pathway (CYP450 of arachidonic acid metabolism may increase the risk for cardiovascular disease by disrupting the fine equilibrium between cardioprotective (CYP2J/CYP2C-generated and cardiotoxic (CYP4A/CYP4F-generated metabolites of arachidonic acid.

Role of CYP2B6 and CYP3A4 in the in vitro N-dechloroethylation of (R)- and (S)-ifosfamide in human liver microsomes.

Science.gov (United States)

Granvil, C P; Madan, A; Sharkawi, M; Parkinson, A; Wainer, I W

1999-04-01

The central nervous system toxicity of ifosfamide (IFF), a chiral antineoplastic agent, is thought to be dependent on its N-dechloroethylation by hepatic cytochrome P-450 (CYP) enzymes. The purpose of this study was to identify the human CYPs responsible for IFF-N-dechloroethylation and their corresponding regio- and enantioselectivities. IFF exists in two enantiomeric forms, (R) - and (S)-IFF, which can be dechloroethylated at either the N2 or N3 positions, producing the corresponding (R,S)-2-dechloroethyl-IFF [(R, S)-2-DCE-IFF] and (R,S)-3-dechloroethyl-IFF [(R,S)-3-DCE-IFF]. The results of the present study suggest that the production of (R)-2-DCE-IFF and (S)-3-DCE-IFF from (R)-IFF is catalyzed by different CYPs as is the production of (S)-2-DCE-IFF and (R)-3-DCE-IFF from (S)-IFF. In vitro studies with a bank of human liver microsomes revealed that the sample-to-sample variation in the production of (S)-3-DCE-IFF from (R)-IFF and (S)-2-DCE-IFF from (S)-IFF was highly correlated with the levels of (S)-mephenytoin N-demethylation (CYP2B6), whereas (R)-2-DCE-IFF production from (R)-IFF and (R)-3-DCE-IFF production from (S)-IFF were both correlated with the activity of testosterone 6beta-hydroxylation (CYP3A4/5). Experiments with cDNA-expressed P-450 and antibody and chemical inhibition studies supported the conclusion that the formation of (S)-3-DCE-IFF and (S)-2-DCE-IFF is catalyzed primarily by CYP2B6, whereas (R)-2-DCE-IFF and (R)-3-DCE-IFF are primarily the result of CYP3A4/5 activity.

Influence of Donor and Recipient CYP3A4, CYP3A5, and ABCB1 Genotypes on Clinical Outcomes and Nephrotoxicity in Liver Transplant Recipients.

Science.gov (United States)

Debette-Gratien, Marilyne; Woillard, Jean-Baptiste; Picard, Nicolas; Sebagh, Mylène; Loustaud-Ratti, Véronique; Sautereau, Denis; Samuel, Didier; Marquet, Pierre

2016-10-01

This study investigated the influence of the CYP3A4*22, CYP3A5*3, and ABCB1 exons 12, 21, and 26 polymorphisms in donors and recipients on clinical outcomes and renal function in 170 liver transplant patients on cyclosporin A (CsA) or tacrolimus (Tac). Allelic discrimination assays were used for genotyping. Multivariate time-dependent Cox proportional hazard models, multiple linear regression using the generalized estimating equation and linear mixed-effect models were used for statistical analysis. Expression of CYP3A5 by either or both the donor and the recipient was significantly associated with lower Tac, but not CsA, dose-normalized trough levels. In the whole population, graft loss was only significantly associated with longer exposure to high calcineurin inhibitor (CNI) concentrations (hazard ratio, 6.93; 95% confidence interval, 2.13-22.55), P = 0.00129), whereas in the Tac subgroup, the risk of graft loss was significantly higher in recipient CYP3A5*1 expressers (hazard ratio, 3.39; 95% confidence interval, 1.52-7.58; P = 0.0028). Renal function was significantly associated with: (1) baseline modification of diet in renal disease (β = 0.51 ± 0.05; P < 0.0001); (2) duration of patient follow-up (per visit, β = -0.98 ± 0.22; P < 0.0001); and (3) CNI exposure (per quantile increase, β = -2.42 ± 0.59; P < 0.0001). No genetic factor was associated with patient survival, acute rejection, liver function test results, recurrence of viral or other initial liver disease, or renal function. This study confirms the effect of CYP3A5*3 on tacrolimus dose requirement in liver transplantation and shows unexpected associations between the type of, and exposure to, CNI and either chronic rejection or graft loss. None of the genetic polymorphisms studied had a noticeable impact on renal function degradation at 10 years.

Reduction in hepatic drug metabolizing CYP3A4 activities caused by P450 oxidoreductase mutations identified in patients with disordered steroid metabolism

International Nuclear Information System (INIS)

Flueck, Christa E.; Mullis, Primus E.; Pandey, Amit V.

2010-01-01

Research highlights: → Cytochrome P450 3A4 (CYP3A4), metabolizes 50% of drugs in clinical use and requires NADPH-P450 reductase (POR). → Mutations in human POR cause congenital adrenal hyperplasia from diminished activities of steroid metabolizing P450s. → We are reporting that mutations in POR may reduce CYP3A4 activity. → POR mutants Y181D, A457H, Y459H, V492E and R616X lost 99%, while A287P, C569Y and V608F lost 60-85% CYP3A4 activity. → Reduction of CYP3A4 activity may cause increased risk of drug toxicities/adverse drug reactions in patients with POR mutations. -- Abstract: Cytochrome P450 3A4 (CYP3A4), the major P450 present in human liver metabolizes approximately half the drugs in clinical use and requires electrons supplied from NADPH through NADPH-P450 reductase (POR, CPR). Mutations in human POR cause a rare form of congenital adrenal hyperplasia from diminished activities of steroid metabolizing P450s. In this study we examined the effect of mutations in POR on CYP3A4 activity. We used purified preparations of wild type and mutant human POR and in vitro reconstitution with purified CYP3A4 to perform kinetic studies. We are reporting that mutations in POR identified in patients with disordered steroidogenesis/Antley-Bixler syndrome (ABS) may reduce CYP3A4 activity, potentially affecting drug metabolism in individuals carrying mutant POR alleles. POR mutants Y181D, A457H, Y459H, V492E and R616X had more than 99% loss of CYP3A4 activity, while POR mutations A287P, C569Y and V608F lost 60-85% activity. Loss of CYP3A4 activity may result in increased risk of drug toxicities and adverse drug reactions in patients with POR mutations.

Cytochrome P450 induction by rifampicin in healthy subjects: determination using the Karolinska cocktail and the endogenous CYP3A4 marker 4beta-hydroxycholesterol.

Science.gov (United States)

Kanebratt, K P; Diczfalusy, U; Bäckström, T; Sparve, E; Bredberg, E; Böttiger, Y; Andersson, T B; Bertilsson, L

2008-11-01

The Karolinska cocktail, comprising caffeine, losartan, omeprazole, and quinine, was given before and after administration of rifampicin (20, 100, or 500 mg daily) to measure induction of cytochrome P450 (P450) enzymes. Rifampicin was given for 14 days to eight healthy subjects (all of whom possessed at least one wild-type CYP2C9 and one wild-type CYP2C19 gene) in each dose group. 4beta-hydroxycholesterol was assessed as an endogenous marker of CYP3A4 induction. A fourfold induction of CYP3A4 was seen at the highest dose by both quinine:3'-hydroxyquinine and 4beta-hydroxycholesterol measurements (P Karolinska cocktail and 4beta-hydroxycholesterol can be used for an initial screening of the induction properties of a drug candidate.

Contrasting exome constancy and regulatory region variation in the gene encoding CYP3A4: an examination of the extent and potential implications.

OpenAIRE

Creemer, O. J.; Ansari-Pour, N.; Ekong, R.; Tarekegn, A.; Plaster, C.; Bains, R. K.; Itan, Y.; Bekele, E.; Bradman, N.

2016-01-01

OBJECTIVE: CYP3A4 expression varies up to 100-fold among individuals, and, to date, genetic causes remain elusive. As a major drug-metabolizing enzyme, elucidation of such genetic causes would increase the potential for introducing personalized dose adjustment of therapies involving CYP3A4 drug substrates. The foetal CYP3A isoform, CYP3A7, is reported to be expressed in ∼10% of European adults and may thus contribute towards the metabolism of endogenous substances and CYP3A drug substrates. H...

Influencia de polimorfismos genéticos de CYP3A4/5 en la farmacocinética de levonorgestrel: estudio piloto

Directory of Open Access Journals (Sweden)

Iván Moreno

2012-12-01

Full Text Available Introducción. El levonorgestrel, un progestágeno sintético usado para endometriosis, dismenorrea y anticoncepción de emergencia, es rápida y completamente absorbido en el tubo digestivo. Su metabolismo es principalmente hepático, mediante las enzimas CYP3A4 y CYP3A5. Objetivo. El presente estudio tuvo como objetivo evaluar la asociación entre la farmacocinética de levonorgestrel y las variantes alélicas de CYP3A4*1B y CYP3A5*3. Materiales y métodos. En un grupo de 17 mujeres adultas sanas, que firmaron un consentimiento informado, se practicó genotipificación para CYP3A4*1B y CYP3A5*3 mediante PCR. Posteriormente, las voluntarias fueron sometidas a un estudio farmacocinético donde, luego de 12 horas de ayuno, recibieron una dosis de 0,75 mg de levonorgestrel. Se extrajeron muestras sanguíneas seriadas (0 a 24 horas y se determinaron las concentraciones de levonorgestrel mediante un método validado de UPLC-ms/ms, para luego obtener los parámetros farmacocinéticos. Todos los procedimientos consideraron los aspectos éticos de la Declaración de Helsinki y las buenas prácticas clínicas. Resultados. Las frecuencias genotípicas observadas para el grupo de estudio fueron 11,8 % para*1B/*1B; 5,8 % para *1/*1B, y 82,4 % para *1/*1 de CYP3A4*1B. Para CYP3A5*3, las frecuencias genotípicas fueron 70,5 % para *3/*3; 23,5 % para *1/*3, y 6,5 % para *1/*1. Se observa una interesante variabilidad entre las voluntarias que sugiere una relación con las variantes genéticas CYP3A, pero que no permite establecer una asociación estadísticamente significativa, presumiblemente debido albajo número de individuos homocigotos mutados de CYP3A4 y silvestres de CYP3A5. Conclusiones. Los polimorfismos genéticos podrían ser factores relevantes en la determinación de la variabilidad entre pacientes en las concentraciones plasmáticas de levonorgestrel, lo cual, sin embargo, no pudo ser establecido estadísticamente en este estudio. Por lo tanto

Monocrotophos Induces the Expression of Xenobiotic Metabolizing Cytochrome P450s (CYP2C8 and CYP3A4) and Neurotoxicity in Human Brain Cells.

Science.gov (United States)

Tripathi, Vinay Kumar; Kumar, Vivek; Pandey, Ankita; Vatsa, Pankhi; Dhasmana, Anupam; Singh, Rajat Pratap; Appikonda, Sri Hari Chandan; Hwang, Inho; Lohani, Mohtashim

2017-07-01

Expression of various cytochrome P450s (CYPs) in mammalian brain cells is well documented. However, such studies are hampered in neural/glial cells of human origin due to nonavailability of human brain cells. To address this issue, we investigated the expression and inducibility of CYP2C8 and CYP3A4 and their responsiveness against cyclophosphamide (CPA) and organophosphorus pesticide monocrotophos (MCP), a known developmental neurotoxicant in human neural (SH-SY5Y) and glial (U373-MG) cell lines. CPA induced significant expression of CYP2C8 and CYP3A4 in both types of cells in a time-dependent manner. Neural cell line exhibited relatively higher constitutive and inducible expression of CYPs than the glial cell line. MCP exposure alone could not induce the significant expression of CYPs, whereas the cells preexposed to CPA showed a significant response to MCP. Similar to the case of CPA induced expressions, neural cells were found to be more vulnerable than glial cells. Our data indicate differential expressions of CYPs in cultured human neural and glial cell lines. The findings were synchronized with protein ligand docking studies, which showed a significant modulatory capacity of MCP by strong interaction with CYP regulators-CAR and PXR. Similarly, the known CYP inducer CPA has also shown significant high docking scores with the two studied CYP regulators. We also observed a significant induction in reactive oxygen species (ROS), lipid peroxides (LPO), micronucleus (MN), chromosomal aberration (CA), and reduction in reduced glutathione (GSH) and catalase following the exposure of MCP. Moreover, the expressions of apoptotic markers such as caspase-3, caspase-9, Bax, and p53 were significantly upregulated, whereas the levels of antiapoptotic marker, Bcl2, was downregulated after the exposure of MCP in both cell lines. These findings confirm the involvement of ROS-mediated oxidative stress, which subsequently triggers apoptosis pathways in both human neural (SH-SY5Y

Genetic analysis of drug metabolizing phase-I enzymes CYP3A4 in ...

Indian Academy of Sciences (India)

LIJUN LIU1

3Key Laboratory of High Altitude Environment and Gene Related to Disease of Tibet Ministry of Education,. Xizang Minzu University ..... basic profile of CYP3A4 in the Tibetan population, and can be used to determine optimal dosage ... 19), and The Project of Young Teachers' Innovative Support. Programme in Universities ...

Stereoselective Inhibition of CYP2C19 and CYP3A4 by Fluoxetine and Its Metabolite: Implications for Risk Assessment of Multiple Time-Dependent Inhibitor Systems

Science.gov (United States)

Lutz, Justin D.; VandenBrink, Brooke M.; Babu, Katipudi N.; Nelson, Wendel L.; Kunze, Kent L.

2013-01-01

Recent guidance on drug-drug interaction (DDI) testing recommends evaluation of circulating metabolites. However, there is little consensus on how to quantitatively predict and/or assess the risk of in vivo DDIs by multiple time-dependent inhibitors (TDIs) including metabolites from in vitro data. Fluoxetine was chosen as the model drug to evaluate the role of TDI metabolites in DDI prediction because it is a TDI of both CYP3A4 and CYP2C19 with a circulating N-dealkylated inhibitory metabolite, norfluoxetine. In pooled human liver microsomes, both enantiomers of fluoxetine and norfluoxetine were TDIs of CYP2C19, (S)-norfluoxetine was the most potent inhibitor with time-dependent inhibition affinity constant (KI) of 7 μM, and apparent maximum time-dependent inhibition rate (kinact,app) of 0.059 min−1. Only (S)-fluoxetine and (R)-norfluoxetine were TDIs of CYP3A4, with (R)-norfluoxetine being the most potent (KI = 8 μM, and kinact,app = 0.011 min−1). Based on in-vitro-to-in-vivo predictions, (S)-norfluoxetine plays the most important role in in vivo CYP2C19 DDIs, whereas (R)-norfluoxetine is most important in CYP3A4 DDIs. Comparison of two multiple TDI prediction models demonstrated significant differences between them in in-vitro-to-in-vitro predictions but not in in-vitro-to-in-vivo predictions. Inclusion of all four inhibitors predicted an in vivo decrease in CYP2C19 (95%) and CYP3A4 (60–62%) activity. The results of this study suggest that adequate worst-case risk assessment for in vivo DDIs by multiple TDI systems can be achieved by incorporating time-dependent inhibition by both parent and metabolite via simple addition of the in vivo time-dependent inhibition rate/cytochrome P450 degradation rate constant (λ/kdeg) values, but quantitative DDI predictions will require a more thorough understanding of TDI mechanisms. PMID:23785064

Regulation of CYP3A4 by pregnane X receptor: The role of nuclear receptors competing for response element binding

Energy Technology Data Exchange (ETDEWEB)

Istrate, Monica A., E-mail: monicai@scripps.edu [Dr. Margarete Fischer-Bosch-Institute of Clinical Pharmacology, Stuttgart, Germany, and University of Tuebingen, Auerbachstr. 112, D-70376 Stuttgart (Germany); Nussler, Andreas K., E-mail: nuessler@uchir.me.tum.de [Department of Traumatology, Technical University Munich, Ismaningerstr. 22, 81675 Munich (Germany); Eichelbaum, Michel, E-mail: michel.eichelbaum@ikp-stuttgart.de [Dr. Margarete Fischer-Bosch-Institute of Clinical Pharmacology, Stuttgart, Germany, and University of Tuebingen, Auerbachstr. 112, D-70376 Stuttgart (Germany); Burk, Oliver, E-mail: oliver.burk@ikp-stuttgart.de [Dr. Margarete Fischer-Bosch-Institute of Clinical Pharmacology, Stuttgart, Germany, and University of Tuebingen, Auerbachstr. 112, D-70376 Stuttgart (Germany)

2010-03-19

Induction of the major drug metabolizing enzyme CYP3A4 by xenobiotics contributes to the pronounced interindividual variability of its expression and often results in clinically relevant drug-drug interactions. It is mainly mediated by PXR, which regulates CYP3A4 expression by binding to several specific elements in the 5' upstream regulatory region of the gene. Induction itself shows a marked interindividual variability, whose underlying determinants are only partly understood. In this study, we investigated the role of nuclear receptor binding to PXR response elements in CYP3A4, as a potential non-genetic mechanism contributing to interindividual variability of induction. By in vitro DNA binding experiments, we showed that several nuclear receptors bind efficiently to the proximal promoter ER6 and distal xenobiotic-responsive enhancer module DR3 motifs. TR{alpha}1, TR{beta}1, COUP-TFI, and COUP-TFII further demonstrated dose-dependent repression of PXR-mediated CYP3A4 enhancer/promoter reporter activity in transient transfection in the presence and absence of the PXR inducer rifampin, while VDR showed this effect only in the absence of treatment. By combining functional in vitro characterization with hepatic expression analysis, we predict that TR{alpha}1, TR{beta}1, COUP-TFI, and COUP-TFII show a strong potential for the repression of PXR-mediated activation of CYP3A4 in vivo. In summary, our results demonstrate that nuclear receptor binding to PXR response elements interferes with PXR-mediated expression and induction of CYP3A4 and thereby contributes to the interindividual variability of induction.

Tributyltin modulates 3,3',4,4',5-pentachlorobiphenyl (PCB-126)-induced hepatic CYP1A activity in channel catfish, Ictalurus punctatus.

Science.gov (United States)

Rice, C D; Roszell, L E

1998-10-09

Many harbor estuaries and their tributaries are contaminated with halogenated aromatic hydrocarbons (HAHs) and polycyclic aromatic hydrocarbons (PAHs). Planar congeners of these two classes initiate their toxic effects, including reproductive, developmental, and immunological dysfunction, primarily through the cytosolic arylhydrocabon receptor (Ahr). However, only rarely are aquatic environments contaminated with Ahr-binding contaminants alone. Instead, most are impacted by a variety of pollutants in mixture. Tributyltin (TBT), a common antifouling biocide, is also found in many harbor estuaries and their tributaries. Several reports indicate that TBT inhibits the cytochrome P-4501A system of fish, at least in vitro, and our recent studies with rodents indicate that TBT potentiates PCB-induced CYP1A. However, the effects of TBT on xenobiotic-induced CYP1A activity in aquatic organisms has been virtually unexplored. To this end, channel catfish, Ictalurus punctatus, were exposed to 3,3'4,4',5-pentachlorobiphenyl (PCB-126, PeCB), TBT, or both in combination, with corn oil (CO) serving as the carrier control. Immunoreactive CYP1A protein and ethoxyresorufin O-deethylase (EROD) activity were measured after (1) a single dose of 0.01, 0. 1, or 1 mg/kg of each or both in combination, and (2) 6 injections of 0.017, 1.7, or 17 microg/kg of each (or in combination) given every 3 d over a 16-d period to yield a cumulative dose of 0.01, 0.1, or 1 mg/kg. As expected, PeCB alone, but not TBT, greatly induced these two CYP1A parameters. Low and middle doses of TBT (0.01 and 0.1 mg/kg), but not the high dose, potentiated PeCB-induced activity at these same doses. This effect of TBT was even more pronounced in the repeated exposure study. Furthermore, EROD activity did not always reflect CYP1A protein induction; enzyme activity was inhibited by TBT at doses that potentiated protein induction (0.01 and 0.1 mg/kg). In summary, TBT potentiates PeCB-induced CYP1A in channel catfish at

Association of polymorphisms in CYP19A1 and CYP3A4 genes with lower urinary tract symptoms, prostate volume, uroflow and PSA in a population-based sample.

Science.gov (United States)

Berges, Richard; Gsur, Andrea; Feik, Elisabeth; Höfner, Klaus; Senge, Theodor; Pientka, Ludger; Baierl, Andreas; Michel, Martin C; Ponholzer, Anton; Madersbacher, Stephan

2011-04-01

The known importance of testosterone for the development of benign prostatic hyperplasia (BPH) prompted us to test the hypothesis whether polymorphisms of two genes (CYP19A1 and CYP3A4) involved in testosterone metabolism are associated with clinical BPH-parameters. A random sample of the population-based Herne lower urinary tract symptoms cohort was analysed. All these men underwent a detailed urological work-up. Two polymorphisms in the CYP19A1 gene [rs700518 in exon 4 (A57G); rs10046 at the 3'UTR(C268T)] and one in the 3'UTR of CYP3A4 [rs2740574 (A392G)] were determined by TaqMan assay from genomic DNA of peripheral blood. These polymorphisms were correlated to clinical and laboratory BPH-parameters. A total of 392 men (65.4 ± 7.0 years; 52-79 years) were analysed. Mean International Prostate Symptom Score (IPSS; 7.5), Q (max) (15.4 ml/s), prostate volume (31 ml) and prostate specific antigen (PSA) (1.8 ng/ml) indicated a typical elderly population. Both polymorphisms in the CYP19A1 gene were not correlated to age, IPSS, Q (max), prostate volume and post-void residual volume. Serum PSA was higher in men carrying the heterozygous rs10046 genotype (2.0 ± 0.1 ng/ml) than in those with the CC-genotype (1.7 ± 0.2 ng/ml, P = 0.012). Men carrying one a mutated allele of the CYP3A4 gene had smaller prostates (27.0 ± 2.0 vs. 32 ± 0.8 ml, P = 0.02) and lower PSA levels (1.6 ± 0.3 vs. 1.9 ± 0.1 ng/ml). The inconsistent associations observed herein and for other gene polymorphisms warrant further studies. In general, the data regarding the association of gene polymorphism to BPH-parameters suggest that this disease is caused by multiple rather than a single genetic variant. A rigorous patient selection based on anatomo-pathological and hormonal profile may possible reduce the number of confounders for future studies thus enabling a more detailed assessment of the association between genetic factors and BPH-parameters.

Gene structure of CYP3A4, an adult-specific form of cytochrome P450 in human livers, and its transcriptional control.

Science.gov (United States)

Hashimoto, H; Toide, K; Kitamura, R; Fujita, M; Tagawa, S; Itoh, S; Kamataki, T

1993-12-01

CYP3 A4 is the adult-specific form of cytochrome P450 in human livers [Komori, M., Nishio, K., Kitada, M., Shiramatsu, K., Muroya, K., Soma, M., Nagashima, K. & Kamataki, T. (1990) Biochemistry 29, 4430-4433]. The sequences of three genomic clones for CYP3A4 were analyzed for all exons, exon-intron junctions and the 5'-flanking region from the major transcription site to nucleotide position -1105, and compared with those of the CYP3A7 gene, a fetal-specific form of cytochrome P450 in humans. The results showed that the identity of 5'-flanking sequences between CYP3A4 and CYP3A7 genes was 91%, and that each 5'-flanking region had characteristic sequences termed as NFSE (P450NF-specific element) and HFLaSE (P450HFLa specific element), respectively. A basic transcription element (BTE) also lay in the 5'-flanking region of the CYP3A4 gene as seen in many CYP genes [Yanagida, A., Sogawa, K., Yasumoto, K. & Fujii-Kuriyama, Y. (1990) Mol. Cell. Biol. 10, 1470-1475]. The BTE binding factor (BTEB) was present in both adult and fetal human livers. To examine the transcriptional activity of the CYP3A4 gene, DNA fragments in the 5'-flanking region of the gene were inserted in front of the simian virus 40 promoter and the chloramphenicol acetyltransferase structural gene, and the constructs were transfected in HepG2 cells. The analysis of the chloramphenicol acetyltransferase activity indicated that (a) specific element(s) which could bind with a factor(s) in livers was present in the 5'-flanking region of the CYP3A4 gene to show the transcriptional activity.

Temporal kinetics and concentration-response relationships for induction of CYP1A, CYP2B, and CYP3A in primary cultures of beagle dog hepatocytes.

Science.gov (United States)

Graham, Richard A; Tyler, Lindsey O; Krol, Wojciech L; Silver, Ivin S; Webster, Lindsey O; Clark, Philip; Chen, Liangfu; Banks, Troy; LeCluyse, Edward L

2006-01-01

Compared to other species, little information is available on the xenobiotic-induced regulation of cytochrome P450 enzymes in the beagle dog. Dogs are widely used in the pharmaceutical industry for many study types, including those that will impact decisions on compound progression. The purpose of this study was (1) to determine the temporal kinetics of drug-induced changes in canine CYP1A, CYP2B, and CYP3A mRNA and enzymatic activity, and (2) to characterize concentration-response relationships for CYP1A2, CYP2B11, and CYP3A12 using primary cultures of canine hepatocytes treated with beta-naphthoflavone (BNF), phenobarbital (PB), and rifampin (RIF), respectively. CYP1A1 and CYP1A2 mRNA exhibited maximal expression (12,700-fold and 206-fold, respectively) after 36 h of treatment with BNF. PB treatment, but not RIF treatment, caused maximal induction of CYP2B11 mRNA (149-fold) after 48 h of treatment. CYP3A12 and CYP3A26 mRNA levels were increased maximally after 72 h of treatment with PB and RIF (CYP3A12, 35-fold and 18-fold, and CYP3A26, 72-fold and 22-fold with PB and RIF treatment, respectively). Concentration-response relationships for BNF induced 7-ethoxyresorufin O-dealkylation (EROD) (EC(50) = 7.8 +/- 4.2 microM), PB induced 7-benzyloxyresorufin O-dealkylation (BROD) (EC(50) = 123 +/- 30 microM), and PB and RIF induced testosterone 6beta-hydroxylation (EC(50) = 132 +/- 28 microM and 0.98 +/- 0.16 microM) resembled the relationship for human CYP induction compared to that of rodent. Interestingly, RIF had no effect on CYP2B11 expression, which represents a species difference overlooked in previous investigations. Overall, the induction of dog CYP1A, CYP2B, and CYP3A exhibits characteristics that are intermediate to those of rodent and human. (c) 2006 Wiley Periodicals, Inc.

Regulation of zebrafish CYP3A65 transcription by AHR2

International Nuclear Information System (INIS)

Chang, Chin-Teng; Chung, Hsin-Yu; Su, Hsiao-Ting; Tseng, Hua-Pin; Tzou, Wen-Shyong; Hu, Chin-Hwa

2013-01-01

CYP3A proteins are the most abundant CYPs in the liver and intestines, and they play a pivotal role in drug metabolism. In mammals, CYP3A genes are induced by various xenobiotics through processes mediated by PXR. We previously identified zebrafish CYP3A65 as a CYP3A ortholog that is constitutively expressed in gastrointestinal tissues, and is upregulated by treatment with dexamethasone, rifampicin or tetrachlorodibenzo-p-dioxin (TCDD). However, the underlying mechanism of TCDD-mediated CYP3A65 transcription is unclear. Here we generated two transgenic zebrafish, Tg(CYP3A65S:EGFP) and Tg(CYP3A65L:EGFP), which contain 2.1 and 5.4 kb 5′ flanking sequences, respectively, of the CYP3A65 gene upstream of EGFP. Both transgenic lines express EGFP in larval gastrointestinal tissues in a pattern similar to that of the endogenous CYP3A65 gene. Moreover, EGFP expression can be significantly induced by TCDD exposure during the larval stage. In addition, EGFP expression can be stimulated by kynurenine, a putative AHR ligand produced during tryptophan metabolism. AHRE elements in the upstream regulatory region of the CYP3A65 gene are indispensible for basal and TCDD-induced transcription. Furthermore, the AHR2 DNA and ligand-binding domains are required to mediate effective CYP3A65 transcription. AHRE sequences are present in the promoters of many teleost CYP3 genes, but not of mammalian CYP3 genes, suggesting that AHR/AHR2-mediated transcription is likely a common regulatory mechanism for teleost CYP3 genes. It may also reflect the different environments that terrestrial and aquatic organisms encounter. - Highlights: • Tg(CYP3A65:EGFP) and CYP3A65 exhibits identical expression pattern. • CYP3A65 can be significantly induced by TCDD or kynurenine. • The AHRE elements are required to mediate CYP3A65 transcription. • The AHR2 DNA and ligand-binding domains are required for CYP3A65 transcription. • AHRE elements are present in many teleost CYP3 genes, but not in

Regulation of zebrafish CYP3A65 transcription by AHR2

Energy Technology Data Exchange (ETDEWEB)

Chang, Chin-Teng; Chung, Hsin-Yu; Su, Hsiao-Ting; Tseng, Hua-Pin [Institute of Bioscience and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan (China); Tzou, Wen-Shyong [Institute of Bioscience and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan (China); Center of Excellence for Marine Bioenvironment and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan (China); Hu, Chin-Hwa, E-mail: chhu@mail.ntou.edu.tw [Institute of Bioscience and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan (China); Center of Excellence for Marine Bioenvironment and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan (China)

2013-07-15

CYP3A proteins are the most abundant CYPs in the liver and intestines, and they play a pivotal role in drug metabolism. In mammals, CYP3A genes are induced by various xenobiotics through processes mediated by PXR. We previously identified zebrafish CYP3A65 as a CYP3A ortholog that is constitutively expressed in gastrointestinal tissues, and is upregulated by treatment with dexamethasone, rifampicin or tetrachlorodibenzo-p-dioxin (TCDD). However, the underlying mechanism of TCDD-mediated CYP3A65 transcription is unclear. Here we generated two transgenic zebrafish, Tg(CYP3A65S:EGFP) and Tg(CYP3A65L:EGFP), which contain 2.1 and 5.4 kb 5′ flanking sequences, respectively, of the CYP3A65 gene upstream of EGFP. Both transgenic lines express EGFP in larval gastrointestinal tissues in a pattern similar to that of the endogenous CYP3A65 gene. Moreover, EGFP expression can be significantly induced by TCDD exposure during the larval stage. In addition, EGFP expression can be stimulated by kynurenine, a putative AHR ligand produced during tryptophan metabolism. AHRE elements in the upstream regulatory region of the CYP3A65 gene are indispensible for basal and TCDD-induced transcription. Furthermore, the AHR2 DNA and ligand-binding domains are required to mediate effective CYP3A65 transcription. AHRE sequences are present in the promoters of many teleost CYP3 genes, but not of mammalian CYP3 genes, suggesting that AHR/AHR2-mediated transcription is likely a common regulatory mechanism for teleost CYP3 genes. It may also reflect the different environments that terrestrial and aquatic organisms encounter. - Highlights: • Tg(CYP3A65:EGFP) and CYP3A65 exhibits identical expression pattern. • CYP3A65 can be significantly induced by TCDD or kynurenine. • The AHRE elements are required to mediate CYP3A65 transcription. • The AHR2 DNA and ligand-binding domains are required for CYP3A65 transcription. • AHRE elements are present in many teleost CYP3 genes, but not in

cDNA cloning and initial characterization of CYP3A43, a novel human cytochrome P450.

Science.gov (United States)

Domanski, T L; Finta, C; Halpert, J R; Zaphiropoulos, P G

2001-02-01

The RACE amplification technology was used on a novel CYP3A-like exon 1 sequence detected during the reverse transcriptase/polymerase chain reaction analysis of human CYP3A gene expression. This resulted in the identification of cDNAs encompassing the complete coding sequence of a new member of the CYP3A gene subfamily, CYP3A43. Interestingly, the majority of the cDNAs identified were characterized by alternative splicing events such as exon skipping and complete or partial intron inclusion. CYP3A43 expression was detected in liver, kidney, pancreas, and prostate. The amino acid sequence is 75% identical to that of CYP3A4 and CYP3A5 and 71% identical to CYP3A7. CYP3A43 differs from CYP3A4 at six amino acid residues, found within the putative substrate recognition sites of CYP3A4, that are known to be determinants of substrate selectivity. The N terminus of CYP3A43 was modified for efficient expression of the protein in Escherichia coli, and a 6X histidine tag was added at the C terminus to facilitate purification. CYP3A43 gave a reduced carbon monoxide difference spectra with an absorbance maximum at 450 nm. The level of heterologous expression was significantly lower than that observed for CYP3A4 and CYP3A5. Immunoblot analyses revealed that CYP3A43 comigrates with CYP3A4 in polyacrylamide gel electrophoresis but does separate from CYP3A5. Monooxygenase assays were performed under a variety of conditions, several of which yielded reproducible, albeit low, testosterone hydroxylase activity. The findings from this study demonstrate that there is a novel CYP3A member expressed in human tissues, although its relative contribution to drug metabolism has yet to be ascertained.

The effect of complementary and alternative medicines on CYP3A4-mediated metabolism of three different substrates : 7-benzyloxy-4-trifluoromethyl-coumarin, midazolam and docetaxel

NARCIS (Netherlands)

Mooiman, Kim D; Maas-Bakker, Roel F; Hendrikx, Jeroen J M A; Bank, Paul C D; Rosing, Hilde; Beijnen, Jos H; Schellens, Jan H M; Meijerman, Irma

OBJECTIVE: Concomitant use of complementary and alternative medicine (CAM) and anticancer drugs can affect the pharmacokinetics of anticancer drugs by inhibiting the metabolizing enzyme cytochrome P450 3A4 (CYP3A4) (EC 1.14.13.157). Several in vitro studies determined whether CAM can inhibit CYP3A4,

Overexpression of CYP3A4 in a COLO 205 Colon Cancer Stem Cell Model in vitro

International Nuclear Information System (INIS)

Olszewski, Ulrike; Liedauer, Richard; Ausch, Christoph; Thalhammer, Theresia; Hamilton, Gerhard

2011-01-01

Cancer stem cells (CSCs) seem to constitute a subpopulation of tumor cells that escape from chemotherapy and cause recurrent disease. Low proliferation rates, protection in a stem cell niche and overexpression of drug resistance proteins are considered to confer chemoresistance. We established an in vitro colon CSC-like model using the COLO 205 cell line, which revealed transiently increased expression of CD133 when transferred to serum-free stem cell culture medium. Assessment of global gene expression of COLO 205 cells under these conditions identified a set of upregulated genes including cytochrome P450 3A4 (CYP3A4) and aldehyde dehydrogenase 1A1 (ALDH1A1), as confirmed by real-time qPCR. ALDH1A1 is a CSC marker for certain tumor entities and confers resistance to cyclophosphamide. CYP3A4 is expressed in liver and colon and its overexpression seems particularly relevant in colon cancer, since it inactivates irinotecan and other xenobiotics, such as taxols and vinca alkaloids. In conclusion, this COLO 205 model provides evidence for CD133 induction concomitant with overexpression of CYP3A4, which, together with ATP-binding cassette, subfamily G, member 2 (ABCG2) and others, may have a role in chemoresistant colon CSCs and a negative impact on disease-free survival in colon cancer patients

CYP3A5 mRNA degradation by nonsense-mediated mRNA decay.

Science.gov (United States)

Busi, Florent; Cresteil, Thierry

2005-09-01

The total CYP3A5 mRNA level is significantly greater in carriers of the CYP3A5*1 allele than in CYP3A5*3 homozygotes. Most of the CYP3A5*3 mRNA includes an intronic sequence (exon 3B) containing premature termination codons (PTCs) between exons 3 and 4. Two models were used to investigate the degradation of CYP3A5 mRNA: a CYP3A5 minigene consisting of CYP3A5 exons and introns 3 to 6 transfected into MCF7 cells, and the endogenous CYP3A5 gene expressed in HepG2 cells. The 3'-untranslated region g.31611C>T mutation has no effect on CYP3A5 mRNA decay. Splice variants containing exon 3B were more unstable than wild-type (wt) CYP3A5 mRNA. Cycloheximide prevents the recognition of PTCs by ribosomes: in transfected MCF7 and HepG2 cells, cycloheximide slowed down the degradation of exon 3B-containing splice variants, suggesting the participation of nonsense-mediated decay (NMD). When PTCs were removed from pseudoexon 3B or when UPF1 small interfering RNA was used to impair the NMD mechanism, the decay of the splice variant was reduced, confirming the involvement of NMD in the degradation of CYP3A5 splice variants. Induction could represent a source of variability for CYP3A5 expression and could modify the proportion of splice variants. The extent of CYP3A5 induction was investigated after exposure to barbiturates or steroids: CYP3A4 was markedly induced in a pediatric population compared with untreated neonates. However, no effect could be detected in either the total CYP3A5 RNA, the proportion of splice variant RNA, or the protein level. Therefore, in these carriers, induction is unlikely to switch on the phenotypic CYP3A5 expression in carriers of CYP3A5*3/*3.

CYP1A1 induction and CYP3A4 inhibition by the fungicide imazalil in the human intestinal Caco-2 cells-comparison with other conazole pesticides.

Science.gov (United States)

Sergent, Thérèse; Dupont, Isabelle; Jassogne, Coralie; Ribonnet, Laurence; van der Heiden, Edwige; Scippo, Marie-Louise; Muller, Marc; McAlister, Dan; Pussemier, Luc; Larondelle, Yvan; Schneider, Yves-Jacques

2009-02-10

Imazalil (IMA) is a widely used imidazole-antifungal pesticide and, therefore, a food contaminant. This compound is also used as a drug (enilconazole). As intestine is the first site of exposure to ingested drugs and pollutants, we have investigated the effects of IMA, at realistic intestinal concentrations, on xenobiotic-metabolizing enzymes and efflux pumps by using Caco-2 cells, as a validated in vitro model of the human intestinal absorptive epithelium. For comparison, other conazole fungicides, i.e. ketoconazole, propiconazole and tebuconazole, were also studied. IMA induced cytochrome P450 (CYP) 1A1 activity to the same extent as benzo(a)pyrene (B(a)P) or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), in a dose- and time-dependent manner. Cell-free aryl hydrocarbon receptor (AhR) binding assay and reporter gene assay suggested that IMA is not an AhR-ligand, implying that IMA-mediated induction should involve an AhR-independent pathway. Moreover, IMA strongly inhibited the CYP3A4 activity in 1,25-vitamin D(3)-induced Caco-2 cells. The other fungicides had weak or nil effects on CYP activities. Study of the apical efflux pump activities revealed that ketoconazole inhibited both P-glycoprotein (Pgp) and multidrug resistance-associated protein 2 (MRP-2) or breast cancer resistance protein (BCRP), whereas IMA and other fungicides did not. Our results imply that coingestion of IMA-contaminated food and CYP3A4- or CYP1A1-metabolizable drugs or chemicals could lead to drug bioavailability modulation or toxicological interactions, with possible adverse effects for human health.

CYP₃A₄ Mediates Oxidative Metabolism of the Synthetic Cannabinoid AKB-48

DEFF Research Database (Denmark)

Holm, Niels Bjerre; Nielsen, Line Marie; Linnet, Kristian

2015-01-01

the metabolism of N-(1-adamantyl)-1-pentyl-1H-indazole-3-carboxamide (AKB-48), a compound identified in herbal blends from 2012 and onwards. We screened for metabolite formation using a panel of nine recombinant cytochrome P450 (CYP) enzymes (CYP1A2, 2B6, 2C8, 2C9, 2C18, 2C19, 2D6, 2E1, and 3A4) and compared...

Inhibition of Cytochrome P450 (CYP3A4) Activity by Extracts from 57 Plants Used in Traditional Chinese Medicine (TCM)

Science.gov (United States)

Ashour, Mohamed L; Youssef, Fadia S; Gad, Haidy A; Wink, Michael

2017-01-01

Background: Herbal medicine is widely used all over the world for treating various health disorders. It is employed either alone or in combination with synthetic drugs or plants to be more effective. Objective: The assessment of the effect of both water and methanol extracts of 57 widely used plants from Traditional Chinese Medicine (TCM) against the main phase I metabolizing enzyme CYP3A4 in vitro for the first time. Materials and Methods: The inhibition of cytochrome P450 activity was evaluated using a luminescence assay. The principal component analysis (PCA) was used to correlate the inhibitory activity with the main secondary metabolites present in the plant extracts. Molecular modeling studies on CYP3A4 (PDB ID 4NY4) were carried out with 38 major compounds present in the most active plant extracts to validate the observed inhibitory effect. Results: Aqueous extracts of Acacia catechu, Andrographis paniculata, Arctium lappa, Areca catechu, Bupleurum marginatum, Chrysanthemum indicum, Dysosma versipellis, and Spatholobus suberectus inhibited CYP3A4 is more than 85% (at a dose of 100 μg/mL). The corresponding methanol extracts of A. catechu, A. paniculata, A. catechu, Mahonia bealei, and Sanguisorba officinalis inhibited the enzyme by more than 50%. Molecular modeling studies revealed that two polyphenols, namely hesperidin and rutin, revealed the highest fitting scores in the active sites of the CYP3A4 with binding energies equal to -74.09 and -71.34 kcal/mol, respectively. Conclusion: These results provide evidence that many TCM plants can inhibit CYP3A4, which might cause a potential interference with the metabolism of other concomitantly administered herbs or drugs. SUMMARY In this study, the inhibitory activity of the aqueous and methanol extracts of 57 widely used plants from Traditional Chinese Medicine (TCM) against the main phase I metabolizing enzyme CYP3A4 was tested in vitro for the first time.Aqueous extracts of Acacia catechu, Andrographis

CYP3A4 expression in breast cancer and its association with risk factors in Mexican women.

Science.gov (United States)

Floriano-Sanchez, Esau; Rodriguez, Noemi Cardenas; Bandala, Cindy; Coballase-Urrutia, Elvia; Lopez-Cruz, Jaime

2014-01-01

In Mexico, breast cancer (BCa) is the leading type of cancer in women. Cytochrome P450 (CYP450) is a superfamily of major oxidative enzymes that metabolize carcinogens and many antineoplastic drugs. In addition, these enzymes have influence on tumor development and tumor response to therapy. In this report, we analyzed the protein expression in patients with BCa and in healthy women. Links with some clinic-pathological characteristic were also assessed. Immunohistochemical analyses were conducted on 48 sets of human breast tumors and normal breast tissues enrolled in Hospital Militar de Especialidades de la Mujer y Neonatologia and Hospital Central Militar, respectively, during the time period from 2010 to 2011. Informed consent was obtained from all participants. Statistical analysis was performed using χ2 or Fisher exact tests to estimate associations and the Mann Whitney U test for comparison of group means. We found a significant CYP3A4 overexpression in BCa stroma and gland regions in comparison with healthy tissue. A significant association between protein expression with smoking, alcoholism and hormonal contraceptives use was also observed. Additionally, we observed estrogen receptor (ER) and progesterone receptor (PR) positive association in BCa. We suggest that CYP3A4 expression promotes BCa development and can be used in the prediction of tumor response to different treatments. One therapeutic approach may thus be to block CYP3A4 function.

Racial Differences in CYP3A4 Genotype and Survival Among Men Treated on Radiation Therapy Oncology Group (RTOG) 9202: A Phase III Randomized Trial

International Nuclear Information System (INIS)

Roach, Mack; Silvio, Michelle de; Rebbick, Timothy; Grignon, David; Rotman, Marvin; Wolkov, Harvey; Fisher, Barbara; Hanks, Gerald; Shipley, William U.; Pollack, Alan; Sandler, Howard; Watkins-Bruner, Deborah Ph.D.

2007-01-01

Purpose: Inherited genotypes may explain the inferior outcomes of African American (AA) men with prostate cancer. To understand how variation in CYP3A4 correlated with outcomes, a retrospective examination of the CYP3A4*1B genotype was performed on men treated with Radiation Therapy Oncology Group (RTOG) 92-02. Methods and Materials: From 1,514 cases, we evaluated 56 (28.4%) of 197 AA and 54 (4.3%) of 1,274 European American (EA) patients. All patients received goserelin and flutamide for 2 months before and during RT (STAD-RT) ± 24 months of goserelin (long-term androgen deprivation plus radiation [LTAD-RT]). Events studied included overall survival and biochemical progression using American Society for Therapeutic Radiology and Oncology consensus guidelines. Results: There were no differences in outcome in patients in with or without CYP3A4 data. There was an association between race and CYP3A4 polymorphisms with 75% of EAs having the Wild Type compared to only 25% of AA men (p <0.0001). There was no association between CYP3A4 classification or race and survival or progression. Conclusions: The samples analyzed support previously reported observations about the distribution of CYP3A4*1B genotype by race, but race was not associated with poorer outcome. However, patient numbers were limited, and selection bias cannot be completely ruled out

Cytochrome P450 CYP4DE1 and CYP6CW3v2 contribute to ethiprole resistance in Laodelphax striatellus (Fallén).

Science.gov (United States)

Elzaki, M E A; Zhang, W; Han, Z

2015-06-01

Laodelphax striatellus Fallén (Hemiptera: Delphacidae), a destructive pest of rice, has developed high resistance to multiple insecticides, threatening the success of pest management programmes. The present study investigated ethiprole resistance mechanisms in a field population that is highly resistant to ethiprole. That population was used to establish a laboratory population that was subjected to further selection to produce a resistant strain. Target genes were cloned and compared between the resistant and the susceptible strains, the role of detoxification enzymes was examined, and the relative expression levels of 71 detoxification enzyme genes were tested using quantitative real time (RT)-PCR. The laboratory selection enhanced the resistance from 107-fold to 180-fold. The Rdl-type target site mutation seldom occurred in the resistant strain and is unlikely to represent the major mechanism underlying the observed resistance. Of the three important detoxification enzymes, only P450 monooxygenase was found to be associated with ethiprole resistance. Moreover, two genes, CYP4DE1 and CYP6CW3v2, were found to be overexpressed in the resistant strain. Furthermore, gene-silencing via a double-stranded RNA feeding test was carried out, and the results showed that the mRNA levels of CYP4DE1 and CYP6CW3v2 were reduced in the resistant strain, whereas ethiprole susceptibility was increased. These results suggest that CYP4DE1 and CYP6CW3v2 play an important role in ethiprole resistance in L. striatellus. © 2015 The Royal Entomological Society.

Allosteric activation of midazolam CYP3A5 hydroxylase activity by icotinib - Enhancement by ketoconazole.

Science.gov (United States)

Zhuang, XiaoMei; Zhang, TianHong; Yue, SiJia; Wang, Juan; Luo, Huan; Zhang, YunXia; Li, Zheng; Che, JinJing; Yang, HaiYing; Li, Hua; Zhu, MingShe; Lu, Chuang

2016-12-01

Icotinib (ICO), a novel small molecule and a tyrosine kinase inhibitor, was developed and approved recently in China for non-small cell lung cancer. During screening for CYP inhibition potential in human liver microsomes (HLM), heterotropic activation toward CYP3A5 was revealed. Activation by icotinib was observed with CYP3A-mediated midazolam hydroxylase activity in HLM (∼40% over the baseline) or recombinant human CYP3A5 (rhCYP3A5) (∼70% over the baseline), but not in the other major CYPs including rhCYP3A4. When co-incubated with selective CYP3A4 inhibitor CYP3cide or monoclonal human CYP3A4 inhibitory antibody in HLM, the activation was extended to ∼60%, suggesting CYP3A5 might be the isozyme involved. Further, the relative activation was enhanced to ∼270% in rhCYP3A5 in the presence of ketoconazole. The activation was substrate and pathway dependent and observed only in the formation of 1'-OH-midazolam, and not 4-OH-midazolam, 6β-OH-testosterone, or oxidized nifedipine. The activation requires the presence of cytochrome b5 and it is only observed in the liver microsomes of dogs, monkeys, and humans, but not in rats and mice. Kinetic analyses of 1'-OH-midazolam formation showed that ICO increased the V max values in HLM and rhCYP3A5 with no significant changes in K m values. By adding CYP3cide with ICO to the incubation, the V max values increased 2-fold over the CYP3cide control. Addition of ketoconazole with ICO alone or ICO plus CYP3cide resulted in an increase in V max values and decrease in K m values compared to their controls. This phenomenon may be attributed to a new mechanism of CYP3A5 heterotropic activation, which warrants further investigation. Copyright © 2016 Elsevier Inc. All rights reserved.

Foetal and adult human CYP3A isoforms in the bioactivation of organophosphorothionate insecticides.

Science.gov (United States)

Buratti, Franca M; Leoni, Claudia; Testai, Emanuela

2006-12-15

In humans organophosphorothionate pesticides (OPT) prenatal exposure has been demonstrated. Since OPT-induced neurodevelopmental effects may be due to in situ bioactivation by foetal enzymes, the catalytic activity of the foetal CYP3A7 toward chlorpyrifos (CPF), parathion (PAR), malathion (MAL) and fenthion (FEN) has been assessed by using recombinant enzymes. A comparison with the adult isoforms CYP3A4 and CYP3A5 has been also carried out. CYP3A7 was able to produce significant levels of oxon or sulfoxide from the four OPTs in the range of tested concentrations (0.05-200 microM). When the efficiencies of CYP3A isoforms were compared, the ranking, expressed as CLi values, were: CPF=3A4>3A5>3A7; PAR=3A4>3A7>3A5; MAL=3A4>3A7>3A5; FEN (sulfoxide formation)=3A4>3A5>3A7. The CYP3A5 efficiency appeared to be more dependent on the single insecticide than its related isozyme CYP3A4. Our results indicate that the levels of toxic metabolite formed in situ by CYP3A7 from CPF, MAL and PAR but not from FEN have the chance to inhibit acetylcholinesterase, following prenatal exposure to OPTs. However, due to the smaller weight of foetal liver, the contribution to total OPT biotransformation is relatively low. On the other hand, our results clearly indicate that at low CPF concentrations, the formation of the non-toxic metabolites is highly favoured in the foetus.

Effect of ketoconazole-mediated CYP3A4 inhibition on clinical pharmacokinetics of panobinostat (LBH589), an orally active histone deacetylase inhibitor

NARCIS (Netherlands)

A.P. Hamberg (Paul); M.M. Woo (Margaret M.); L.C. Chen (Lin-Chi); J. Verweij (Jaap); M.G. Porro; L. Zhao (Ling); W. Li (Weili); D.A.J. van der Biessen (Diane); H.S. Sharma (Hari); T. Hengelage (Thomas); M.J.A. de Jonge (Maja)

2011-01-01

textabstractPurpose: Panobinostat is partly metabolized by CYP3A4 in vitro. This study evaluated the effect of a potent CYP3A inhibitor, ketoconazole, on the pharmacokinetics and safety of panobinostat. Methods: Patients received a single panobinostat oral dose on day 1, followed by 4 days wash-out

Coffee inhibition of CYP3A4 in vitro was not translated to a grapefruit-like pharmacokinetic interaction clinically.

Science.gov (United States)

Dresser, George K; Urquhart, Brad L; Proniuk, Julianne; Tieu, Alvin; Freeman, David J; Arnold, John Malcolm; Bailey, David G

2017-10-01

Grapefruit can augment oral medication bioavailability through irreversible (mechanism-based) inhibition of intestinal CYP3A4. Supplementary data from our recent coffee-drug interaction clinical study showed some subjects had higher area under the plasma drug concentration - time curve (AUC) and plasma peak drug concentration (Cmax) of the CYP3A4 probe felodipine compared to aqueous control. It was hypothesized that coffee might interact like grapefruit in responsive individuals. Beans from six geographical locations were consistently brewed into coffee that was separated chromatographically to a methanolic fraction for in vitro inhibition testing of CYP3A4 metabolism of felodipine at 1% coffee strength. The effect of simultaneous incubation and 10-min preincubation with coffee fractions determined whether coffee had direct and mechanism-based inhibitory activity. A subsequent five-way randomized balanced controlled crossover clinical study evaluated the clinical pharmacokinetic interaction with single-dose felodipine. Grapefruit juice, water, or three of the in vitro tested coffees were ingested at 300 mL alone 1 h before and then with felodipine. In vitro, all six coffees decreased felodipine metabolism for both simultaneous and preincubation exposure compared to corresponding control. Five coffees demonstrated mechanism-based inhibition. Grapefruit increased felodipine AUC 0-8 (25 vs. 13 ng.h/mL, P coffees caused no change in these parameters compared to water. Despite high in vitro potency of CYP3A4 inhibition, the coffees did not cause a clinical pharmacokinetic interaction possibly from insufficient amount of inhibitor(s) in coffee reaching intestinal CYP3A4 during the absorption phase of felodipine. The results of this study highlight the need for follow-up clinical testing when in vitro results indicate the possibility of an interaction. © 2017 The Authors. Pharmacology Research & Perspectives published by John Wiley & Sons Ltd, British

Development of Murine Cyp3a Knockout Chimeric Mice with Humanized Liver.

Science.gov (United States)

Kato, Kota; Ohbuchi, Masato; Hamamura, Satoko; Ohshita, Hiroki; Kazuki, Yasuhiro; Oshimura, Mitsuo; Sato, Koya; Nakada, Naoyuki; Kawamura, Akio; Usui, Takashi; Kamimura, Hidetaka; Tateno, Chise

2015-08-01

We developed murine CYP3A knockout ko chimeric mice with humanized liver expressing human P450S similar to those in humans and whose livers and small intestines do not express murine CYP3A this: approach may overcome effects of residual mouse metabolic enzymes like Cyp3a in conventional chimeric mice with humanized liver, such as PXB-mice [urokinase plasminogen activator/severe combined immunodeficiency (uPA/SCID) mice repopulated with over 70% human hepatocytes] to improve the prediction of drug metabolism and pharmacokinetics in humans. After human hepatocytes were transplanted into Cyp3a KO/uPA/SCID host mice, human albumin levels logarithmically increased until approximately 60 days after transplantation, findings similar to those in PXB-mice. Quantitative real-time-polymerase chain reaction analyses showed that hepatic human P450s, UGTs, SULTs, and transporters mRNA expression levels in Cyp3a KO chimeric mice were also similar to those in PXB-mice and confirmed the absence of Cyp3a11 mRNA expression in mouse liver and intestine. Findings for midazolam and triazolam metabolic activities in liver microsomes were comparable between Cyp3a KO chimeric mice and PXB-mice. In contrast, these activities in the intestine of Cyp3a KO chimeric mice were attenuated compared with PXB-mice. Owing to the knockout of murine Cyp3a, hepatic Cyp2b10 and 2c55 mRNA levels in Cyp3a KO/uPA/SCID mice (without hepatocyte transplants) were 8.4- and 61-fold upregulated compared with PXB-mice, respectively. However, human hepatocyte transplantation successfully restored Cyp2b10 level nearly fully and Cyp2c55 level partly (still 13-fold upregulated) compared with those in PXB-mice. Intestinal Cyp2b10 and 2c55 were also repressed by human hepatocyte transplantation in Cyp3a KO chimeric mice. Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.

Effect of botanical immunomodulators on human CYP3A4 inhibition: implications for concurrent use as adjuvants in cancer therapy.

Science.gov (United States)

Patil, Dada; Gautam, Manish; Gairola, Sunil; Jadhav, Suresh; Patwardhan, Bhushan

2014-03-01

Many botanical immunomodulators are used as adjuvants along with cancer chemotherapy. However, information on the impact of concurrent administration of such botanicals on pharmacokinetics of chemotherapy agents is inadequate. This study investigates inhibitory activities of 3 popular botanical adjuvants: ASPARAGUS RACEMOSU: (root aqueous extract; ARE), WITHANIA SOMNIFER: (root aqueous extract; WSE), and TINOSPORA CORDIFOLI: (stem aqueous extract, TCE) on human CYP3A4 isoenzyme, responsible for metabolism of several chemotherapy agents. . Testosterone 6-β hydroxylation was monitored using high-performance liquid chromatography as an indicator of CYP3A4 catalytic activities. Ketoconazole (positive control) and extracts were studied at their in vivo-relevant concentrations. TCE showed mild inhibition while no significant inhibitory activities were observed in WSE and ARE. TCE was further fractionated to obtain polar and nonpolar fractions. The nonpolar fraction showed significant CYP3A4 inhibition with IC50 13.06 ± 1.38 µg/mL. Major constituents of nonpolar fraction were identified using HPLC-DAD-MS profiling as berberine, jatrorrhizine, and palmatine, which showed IC50 values as 6.25 ± 0.30, 15.18 ± 1.59, and 15.53 ± 1.89 µg/mL, respectively. Our findings suggest that constituents of TCE extract especially protoberberine alkaloids have the potential to interact with cancer chemotherapy agents that are metabolized by CYP3A4 in vivo.

The far and distal enhancers in the CYP3A4 gene co-ordinate the proximal promoter in responding similarly to the pregnane X receptor but differentially to hepatocyte nuclear factor-4alpha.

Science.gov (United States)

Liu, Fu-Jun; Song, Xiulong; Yang, Dongfang; Deng, Ruitang; Yan, Bingfang

2008-01-01

CYP3A4 (cytochrome P450 3A4) is involved in the metabolism of more than 50% of drugs and other xenobiotics. The expression of CYP3A4 is induced by many structurally dissimilar compounds. The PXR (pregnane X receptor) is recognized as a key regulator for the induction, and the PXR-directed transactivation of the CYP3A4 gene is achieved through a co-ordinated mechanism of the distal module with the proximal promoter. Recently, a far module was found to support constitutive expression of CYP3A4. The far module, like the distal module, is structurally clustered by a PXR response element (F-ER6) and elements recognized by HNF-4alpha (hepatocyte nuclear receptor-4alpha). We hypothesized that the far module supports PXR transactivation of the CYP3A4 gene. Consistent with the hypothesis, fusion of the far module to the proximal promoter of CYP3A4 markedly increased rifampicin-induced reporter activity. The increase was synergistically enhanced when both the far and distal modules were fused to the proximal promoter. The increase, however, was significantly reduced when the F-ER6 was disrupted. Chromatin immunoprecipitation detected the presence of PXR in the far module. Interestingly, HNF-4alpha increased the activity of the distal-proximal fused promoter, but decreased the activity of the far-proximal fused promoter. Given the fact that induction of CYP3A4 represents an important detoxification mechanism, the functional redundancy and synergistic interaction in supporting PXR transactivation suggest that the far and distal modules ensure the induction of CYP3A4 during chemical insults. The difference in responding to HNF-4alpha suggests that the magnitude of the induction is under control through various transcriptional networks.

Oxidase uncoupling in heme monooxygenases: Human cytochrome P450 CYP3A4 in Nanodiscs

Energy Technology Data Exchange (ETDEWEB)

Grinkova, Yelena V.; Denisov, Ilia G.; McLean, Mark A. [Departments of Biochemistry and Chemistry, University of Illinois, 505 South Goodwin Avenue (United States); Sligar, Stephen G., E-mail: s-sligar@illinois.edu [Departments of Biochemistry and Chemistry, University of Illinois, 505 South Goodwin Avenue (United States)

2013-01-25

Highlights: ► Substantial reducing equivalents are lost in human P450 CYP3A4 via an oxidase channel. ► Substrate binding has a pronounced effect on uncoupling in cytochrome P450. ► Anionic phospholipids improve the overall coupling in CYP3A4 Nanodiscs. -- Abstract: The normal reaction mechanism of cytochrome P450 operates by utilizing two reducing equivalents to reduce atmospheric dioxygen, producing one molecule of water and an oxygenated product in an overall stoichiometry of 2 electrons:1 dioxygen:1 product. However, three alternate unproductive pathways exist where the intermediate iron–oxygen states in the catalytic cycle can yield reduced oxygen products without substrate metabolism. The first involves release of superoxide from the oxygenated intermediate while the second occurs after input of the second reducing equivalent. Superoxide rapidly dismutates and hence both processes produce hydrogen peroxide that can be cytotoxic to the organism. In both cases, the formation of hydrogen peroxide involves the same overall stoichiometry as oxygenases catalysis. The key step in the catalytic cycle of cytochrome P450 involves scission of the oxygen–oxygen bond of atmospheric dioxygen to produce a higher valent iron-oxo state termed “Compound I”. This intermediate initiates a radical reaction in the oxygenase pathway but also can uptake two additional reducing equivalents from reduced pyridine nucleotide (NADPH) and the flavoprotein reductase to produce a second molecule of water. This non-productive decay of Compound I thus yields an overall oxygen to NADPH ratio of 1:2 and does not produce hydrocarbon oxidation. This water uncoupling reaction provides one of a limited means to study the reactivity of the critical Compound I intermediate in P450 catalysis. We measured simultaneously the rates of NADPH and oxygen consumption as a function of substrate concentration during the steady-state hydroxylation of testosterone catalyzed by human P450 CYP3A4

CYP3A isoforms in Ewing's sarcoma tumours: an immunohistochemical study with clinical correlation.

Science.gov (United States)

Zia, Hamid; Murray, Graeme I; Vyhlidal, Carrie A; Leeder, J Steven; Anwar, Ahmed E; Bui, Marilyn M; Ahmed, Atif A

2015-04-01

Ewing's sarcoma is an aggressive malignancy of bone and soft tissue with high incidence of metastasis and resistance to chemotherapy. Cytochrome P450 (CYP) monooxygenases are a family of enzymes that are involved in the metabolism of exogenous and endogenous compounds, including anti-cancer drugs, and have been implicated in the aggressive behaviour of various malignancies. Tumour samples and clinical information including age, sex, tumour site, tumour size, clinical stage and survival were collected from 36 adult and paediatric patients with Ewing's sarcoma family tumours. Tissue microarrays slides were processed for immunohistochemical labelling for CYP3A4, CYP3A5 and CYP3A7 using liver sections as positive control. The intensity of staining was scored as negative, low or high expression and was analysed statistically for any association with patients' clinical information. Four cases were later excluded due to inadequate viable tissue. CYP3A4 staining was present in 26 (81%) cases with high expression noted in 13 (40%) of 32 cases. High expression was significantly associated with distant metastases (P Ewing's sarcoma tumours and high CYP3A4 expression may be associated with metastasis. Additional studies are needed to further investigate the role of CYP3A4 in the prognosis of these tumours. © 2015 The Authors. International Journal of Experimental Pathology © 2015 International Journal of Experimental Pathology.

Effects of strong CYP2D6 and 3A4 inhibitors, paroxetine and ketoconazole, on the pharmacokinetics and cardiovascular safety of tamsulosin

NARCIS (Netherlands)

Troost, Joachim; Tatami, Shinji; Tsuda, Yasuhiro; Mattheus, Michaela; Mehlburger, Ludwig; Wein, Martina; Michel, Martin C.

2011-01-01

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT: Tamsulosin metabolism involves both CYP2D6 and 3A4. However, data on potential drug-drug interactions between tamsulosin and inhibitors of CYP2D6 and 3A4 are limited and information on potential pharmacodynamic consequences of such pharmacokinetic

Alteration in the Expression of Cytochrome P450s (CYP1A1, CYP2E1, and CYP3A11 in the Liver of Mouse Induced by Microcystin-LR

Directory of Open Access Journals (Sweden)

Bangjun Zhang

2015-03-01

Full Text Available Microcystins (MCs are cyclic heptapeptide toxins and can accumulate in the liver. Cytochrome P450s (CYPs play an important role in the biotransformation of endogenous substances and xenobiotics in animals. It is unclear if the CYPs are affected by MCs exposure. The objective of this study was to evaluate the effects of microcystin-LR (MCLR on cytochrome P450 isozymes (CYP1A1, CYP2E1, and CYP3A11 at mRNA level, protein content, and enzyme activity in the liver of mice the received daily, intraperitoneally, 2, 4, and 8 µg/kg body weight of MCLR for seven days. The result showed that MCLR significantly decreased ethoxyresorufin-O-deethylase (EROD (CYP1A1 and erythromycin N-demthylase (ERND (CYP3A11 activities and increased aniline hydroxylase (ANH activity (CYP2E1 in the liver of mice during the period of exposure. Our findings suggest that MCLR exposure may disrupt the function of CYPs in liver, which may be partly attributed to the toxicity of MCLR in mice.

Molecular cloning and 3D model of first cytochrome P450 from CYP3A subfamily in saltwater crocodile (Crocodylus porosus).

Science.gov (United States)

Tabassum, Rabia

2017-10-18

Cytochrome P450s (CYPs) play critical role in oxidative metabolism of numerous xenobiotics and endogenous compounds. The first CYP3A subfamily member in saltwater crocodile has been cloned and modelled for three-dimensional (3D) structure. The full-length cDNA was obtained employing reverse transcription polymerase chain reaction (RT-PCR) strategy and rapid amplification of cDNA ends (RACE). The cDNA sequence of 1659 nucleotides includes 132 nucleotides from 5' untranslated region (UTR), an open reading frame of 1527 nucleotides encoding 509 amino acids designated as CYP3A163. The alignment of CYP3A163 sequence with CYP3A subfamily across the lineages exhibit the loss of 1 residue in birds and 7 residues in mammals in comparison to reptiles suggesting the adaptation processes during evolution. The amino acid identity of CYP3A163 with Alligator mississippiensis CYP3A77 and Homo sapiens CYP3A4 is 91% and 62% respectively. The 3D structure of CYP3A163 modelled using human CYP3A4 structure as a template with Phyre 2 software, represents high similarity with its functionally important motifs and catalytic domain. Both sequence and structure of CYP3A163 display the common and conserved features of CYP3A subfamily. Overall, this study provides primary molecular and structural data of CYP3A163 required to investigate the xenobiotic metabolism in saltwater crocodiles. Copyright © 2017 Elsevier Inc. All rights reserved.

Omeprazole and lansoprazole enantiomers induce CYP3A4 in human hepatocytes and cell lines via glucocorticoid receptor and pregnane X receptor axis.

Science.gov (United States)

Novotna, Aneta; Dvorak, Zdenek

2014-01-01

Benzimidazole drugs lansoprazole and omeprazole are used for treatment of various gastrointestinal pathologies. Both compounds cause drug-drug interactions because they activate aryl hydrocarbon receptor and induce CYP1A genes. In the current paper, we examined the effects of lansoprazole and omeprazole enantiomers on the expression of key drug-metabolizing enzyme CYP3A4 in human hepatocytes and human cancer cell lines. Lansoprazole enantiomers, but not omeprazole, were equipotent inducers of CYP3A4 mRNA in HepG2 cells. All forms (S-, R-, rac-) of lansoprazole and omeprazole induced CYP3A4 mRNA and protein in human hepatocytes. The quantitative profiles of CYP3A4 induction by individual forms of lansoprazole and omeprazole exerted enantiospecific patterns. Lansoprazole dose-dependently activated pregnane X receptor PXR in gene reporter assays, and slightly modulated rifampicin-inducible PXR activity, with similar potency for each enantiomer. Omeprazole dose-dependently activated PXR and inhibited rifampicin-inducible PXR activity. The effects of S-omeprazole were much stronger as compared to those of R-omeprazole. All forms of lansoprazole, but not omeprazole, slightly activated glucocorticoid receptor and augmented dexamethasone-induced GR transcriptional activity. Omeprazole and lansoprazole influenced basal and ligand inducible expression of tyrosine aminotransferase, a GR-target gene, in HepG2 cells and human hepatocytes. Overall, we demonstrate here that omeprazole and lansoprazole enantiomers induce CYP3A4 in HepG2 cells and human hepatocytes. The induction comprises differential interactions of omeprazole and lansoprazole with transcriptional regulators PXR and GR, and some of the effects were enantiospecific. The data presented here might be of toxicological and clinical importance, since the effects occurred in therapeutically relevant concentrations.

Interaksi Ekstrak Sambiloto (Andrographis paniculata (Burm.F.Ness dengan Glibenklamid terhadap Ekspresi Gen CYP3A4 pada Kultur Sel HepG2

Directory of Open Access Journals (Sweden)

Andzar Fikranus Shofa

2017-12-01

Full Text Available Diabetes melitus (DM merupakan salah satu penyakit metabolik kronis yang ditandai dengan hiperglikemia. Berbagai terapi dilakukan untuk mengatasi hiperglikemia, baik dengan menggunakan obat antidiabetes oral maupun tanaman herbal berkhasiat antidiabetes. Sambiloto (Andrographis paniculata (Burm.F.Nees merupakan salah satu herbal antidiabetes yang banyak dikonsumsi masyarakat Indonesia. Andrografolida merupakan zat aktif yang terdapat dalam sambiloto yang berperan sebagai agen antidiabetes. Namun, dalam banyak kasus kombinasi antara herbal dan obat sintesis menyebabkan interaksi jika digunakan pada waktu bersamaan. Penelitian ini bertujuan untuk memperoleh data interaksi ekstrak etanol herba sambiloto dengan glibenklamid terhadap ekspresi gen CYP3A4 pada kultur sel HepG2. Hasil penelitian ini menunjukkan bahwa terjadi penurunan ekspresi gen CYP3A4 pada pengujian sampel tunggal maupun kombinasi antara ekstrak etanol herba sambiloto dan glibenklamid dengan bertambahnya konsentrasi. Pada konsentrasi 50 dan 100 μg/mL sambiloto, glibenklamid, dan kombinasinya menyebabkan penurunan ekspresi gen CYP3A4 sekitar 0,82, 0,70; 0,89, 0,53; 0,84, 0,72. Berdasarkan hasil tersebut dapat disimpulkan terjadi penurunan ekspresi gen CYP3A4 dengan bertambahnya konsentrasi sampel.

Sex difference in induction of hepatic CYP2B and CYP3A subfamily enzymes by nicardipine and nifedipine in rats

International Nuclear Information System (INIS)

Konno, Yoshihiro; Sekimoto, Masashi; Nemoto, Kiyomitsu; Degawa, Masakuni

2004-01-01

Male and female of F344 rats were treated per os with nicardipine (Nic) and nifedipine (Nif), and changes in the levels of mRNA and protein of hepatic cytochrome P450 (P450) enzymes, CYP2B1, CYP2B2, CYP3A1, CYP3A2, CYP3A9, and CYP3A18 were examined. Furthermore, hepatic microsomal activities for pentoxyresorufin O-dealkylation (PROD) and nifedipine oxidation, which are mainly mediated by CYP2B and CYP3A subfamily enzymes, respectively, were measured. Analyses of RT-PCR and Western blotting revealed that Nic and Nif induced predominantly CYP3A and CYP2B enzymes, respectively. As for the gene activation of CYP2B enzymes, especially CYP2B1, Nif showed high capacity in both sexes of rats, whereas Nic did a definite capacity in the males but little in the females. Gene activations of CYP3A1, CYP3A2, and CYP3A18 by Nic occurred in both sexes of rats, although that of CYP3A9 did only in the male rats. Although gene activations of CYP3A1 and CYP3A2 by Nif were observed in both sexes of rats, a slight activation of the CYP3A9 gene occurred only in female rats, and the CYP3A18 gene activation, in neither male nor female rats. Thus, changes in levels of the mRNA or protein of CYP2B and CYP3A enzymes, especially CYP2B1 and CYP3A2, were closely correlated with those in hepatic PROD and nifedipine oxidation activities, respectively. The present findings demonstrate for the first time the sex difference in the Nic- and Nif-mediated induction of hepatic P450 enzymes in rats and further indicate that Nic and Nif show different specificities and sex dependencies in the induction of hepatic P450 enzymes

Polymorphisms in CYP1A1 and CYP3A5 Genes Contribute to the Variability in Granisetron Clearance and Exposure in Pregnant Women with Nausea and Vomiting.

Science.gov (United States)

Bustos, Martha L; Zhao, Yang; Chen, Huijun; Caritis, Steve N; Venkataramanan, Raman

2016-12-01

Nausea and vomiting affect up to 90% of pregnant women. Granisetron is a potent and highly selective serotonin receptor antagonist and is an effective antiemetic. Findings from a prior study in pregnant women demonstrated a large interindividual variability in granisetron exposure. Granisetron is primarily metabolized by the cytochrome P450 (CYP) enzymes CYP1A1 and CYP3A and is likely a substrate of the ABCB1 transporter. Single-nucleotide polymorphisms (SNPs) in CYP3A, CYP1A1, and ABCB1 can alter drug metabolism. This study evaluated the influence of polymorphisms in CYP3A4, CYP3A5, CYP1A1, and ABCB1 on the pharmacokinetic properties of granisetron in pregnant women. The study enrolled 16 pregnant women (gestational age of 12-19 wks). All patients had nausea and vomiting and were treated with granisetron 1 mg. Granisetron plasma concentrations were determined using liquid chromatography tandem-mass spectrometry. The patients' genotype was determined using TaqMan SNP Genotyping Assays. The Hardy-Weinberg equilibrium was assessed by comparing observed and expected genotype frequencies, using the exact test. Intravenous granisetron clearance was used as the dependent variable for analysis of associations. Of 16 patients, 25% were homozygous for the allele variant CYP3A5*3 and had a significantly lower granisetron clearance and increased area under the plasma concentration-versus-time curve (AUC) compared with nonhomozygous patients. Approximately one-third of patients (n=5) were carriers for the allele variant CYP1A1*2A and had a significantly higher granisetron clearance and decreased AUC. We did not find significant differences in the AUC or clearance for any SNPs in CYP3A4 and ABCB1 genes. Polymorphisms in CYP3A5 and CYP1A1 account for some of the variability in systemic clearance and exposure of granisetron in pregnant women. © 2016 Pharmacotherapy Publications, Inc.

Modeling Chemical Interaction Profiles: I. Spectral Data-Activity Relationship and Structure-Activity Relationship Models for Inhibitors and Non-inhibitors of Cytochrome P450 CYP3A4 and CYP2D6 Isozymes

Directory of Open Access Journals (Sweden)

Richard D. Beger

2012-03-01

Full Text Available An interagency collaboration was established to model chemical interactions that may cause adverse health effects when an exposure to a mixture of chemicals occurs. Many of these chemicals—drugs, pesticides, and environmental pollutants—interact at the level of metabolic biotransformations mediated by cytochrome P450 (CYP enzymes. In the present work, spectral data-activity relationship (SDAR and structure-activity relationship (SAR approaches were used to develop machine-learning classifiers of inhibitors and non-inhibitors of the CYP3A4 and CYP2D6 isozymes. The models were built upon 602 reference pharmaceutical compounds whose interactions have been deduced from clinical data, and 100 additional chemicals that were used to evaluate model performance in an external validation (EV test. SDAR is an innovative modeling approach that relies on discriminant analysis applied to binned nuclear magnetic resonance (NMR spectral descriptors. In the present work, both 1D 13C and 1D 15N-NMR spectra were used together in a novel implementation of the SDAR technique. It was found that increasing the binning size of 1D 13C-NMR and 15N-NMR spectra caused an increase in the tenfold cross-validation (CV performance in terms of both the rate of correct classification and sensitivity. The results of SDAR modeling were verified using SAR. For SAR modeling, a decision forest approach involving from 6 to 17 Mold2 descriptors in a tree was used. Average rates of correct classification of SDAR and SAR models in a hundred CV tests were 60% and 61% for CYP3A4, and 62% and 70% for CYP2D6, respectively. The rates of correct classification of SDAR and SAR models in the EV test were 73% and 86% for CYP3A4, and 76% and 90% for CYP2D6, respectively. Thus, both SDAR and SAR methods demonstrated a comparable performance in modeling a large set of structurally diverse data. Based on unique NMR structural descriptors, the new SDAR modeling method complements the existing SAR

Comparative study of polymorphism frequencies of the CYP2D6, CYP3A5, CYP2C8 and IL-10 genes in Mexican and Spanish women with breast cancer.

Science.gov (United States)

Alcazar-González, Gregorio Antonio; Calderón-Garcidueñas, Ana Laura; Garza-Rodríguez, María Lourdes; Rubio-Hernández, Gabriela; Escorza-Treviño, Sergio; Olano-Martin, Estibaliz; Cerda-Flores, Ricardo Martín; Castruita-Avila, Ana Lilia; González-Guerrero, Juan Francisco; le Brun, Stéphane; Simon-Buela, Laureano; Barrera-Saldaña, Hugo Alberto

2013-10-01

Pharmacogenetic studies in breast cancer (BC) may predict the efficacy of tamoxifen and the toxicity of paclitaxel and capecitabine. We determined the frequency of polymorphisms in the CYP2D6 gene associated with activation of tamoxifen, and those of the genes CYP2C8, CYP3A5 and DPYD associated with toxicity of paclitaxel and capecitabine. We also included a IL-10 gene polymorphism associated with advanced tumor stage at diagnosis. Genomic DNAs from 241 BC patients from northeast Mexico were genotyped using DNA microarray technology. For tamoxifen processing, CYP2D6 genotyping predicted that 90.8% of patients were normal metabolizers, 4.2% ultrarapid, 2.1% intermediate and 2.9% poor metabolizers. For paclitaxel and the CYP2C8 gene, 75.3% were normal, 23.4% intermediate and 1.3% poor metabolizers. Regarding the DPYD gene, only one patient was a poor metabolizer. For the IL-10 gene, 47.1% were poor metabolizers. These results contribute valuable information towards personalizing BC chemotherapy in Mexican women.

First Analysis of the Association Between CYP3A4/5, ABCB1 Genetic Polymorphisms and Oxcarbazepine Metabolism and Transport in Chinese Epileptic Patients with Oxcarbazepine Monotherapy and Bitherapy.

Science.gov (United States)

Wang, Ping; Yin, Tao; Ma, Hong-ying; Liu, Dan-Qi; Sheng, Yangh-ao; Zhou, Bo-Ting

2015-01-01

Oxcarbazepine (OXC) is widely used in anti-epileptic treatment. Cytochrome P450 3A4 (CYP3A4), cytochrome P450 3A5(CYP3A5), and ATP-binding cassette sub-family B member 1 (ABCB1) are potential genes involved in OXC metabolisms and transport in vivo. This study aims to examine the genetic effects of CYP3A4, CYP3A5, and ABCB1 on OXC metabolism and transport in Chinese epileptic patients using OXC as monotherapy and bitherapy with lamotrigine (LTG), levetiracetam (LEV), or valproic acid (VPA). Sixty-six Chinese epileptic patients were recruited from Xiangya Hospital Central South University, of whom 40 patients were receiving OXC monotherapy, 11 patients were placed in the OXC bitherapy group combined with one enzyme-inducing anti-epileptic drugs (LTG or LEV), and 15 patients were placed in the OXC bitherapy group combined with VPA. Oxcarbazepine and its main metabolite 10-hydrocarbazepine (MHD) plasma concentrations were measured using high performance liquid chromatography (HPLC)-UV method. In addition, eight single nucleotide polymorphisms (SNPs) in CYP3A4, CYP3A5, ABCB1 gene were genotyped by polymerase chain reaction-improved multiple ligase detection reaction (PCR-iMLDR). In the OXC+VPA group, ABCB1 rs2032582 and rs2032582-rs10234411-rs1045642 TAG haplotype were associated with MHD and MHD+OXC plasma concentration before permutation test. In OXC monotherapy and OXC+ LTG/LEV groups, no significant association between genetic polymorphisms in CYP3A4/5, ABCB1 gene and OXC plasma concentration parameters were observed. CYP3A4/5 and ABCB1 genetic variants might not take part in the metabolism and transport of MHD and OXC among epileptic patients using OXC monotherapy and bitherapy in combination with LEV, LTG or VPA.

Isolation of CYP3A5P cDNA from human liver: a reflection of a novel cytochrome P-450 pseudogene.

Science.gov (United States)

Schuetz, J D; Guzelian, P S

1995-03-14

We have isolated, from a human liver cDNA library, a 1627 bp CYP3A5 cDNA variant (CYP3A5P) that contains several large insertions, deletions, and in-frame termination codons. By comparison with the genomic structure of other CYP3A genes, the major insertions in CYP3A5P cDNA demarcate the inferred sites of several CYP3A5 exons. The segments inserted in CYP3A5P have no homology with splice donor acceptor sites. It is unlikely that CYP3A5P cDNA represents an artifact of the cloning procedures since Southern blot analysis of human genomic DNA disclosed that CYP3A5P cDNA hybridized with a DNA fragment distinct from fragments that hybridized with either CYP3A5, CYP3A3 or CYP3A4. Moreover, analysis of adult human liver RNA on Northern blots hybridized with a CYP3A5P cDNA fragment revealed the presence of an mRNA with the predicted size of CYP3A5P. We conclude that CYP3A5P cDNA was derived from a separate gene, CYP3A5P, most likely a pseudogene evolved from CYP3A5.

Increased expression of CYP4Z1 promotes tumor angiogenesis and growth in human breast cancer

International Nuclear Information System (INIS)

Yu, Wei; Chai, Hongyan; Li, Ying; Zhao, Haixia; Xie, Xianfei; Zheng, Hao; Wang, Chenlong; Wang, Xue; Yang, Guifang; Cai, Xiaojun; Falck, John R.; Yang, Jing

2012-01-01

Cytochrome P450 (CYP) 4Z1, a novel CYP4 family member, is over-expressed in human mammary carcinoma and associated with high-grade tumors and poor prognosis. However, the precise role of CYP4Z1 in tumor progression is unknown. Here, we demonstrate that CYP4Z1 overexpression promotes tumor angiogenesis and growth in breast cancer. Stable expression of CYP4Z1 in T47D and BT-474 human breast cancer cells significantly increased mRNA expression and production of vascular endothelial growth factor (VEGF)-A, and decreased mRNA levels and secretion of tissue inhibitor of metalloproteinase-2 (TIMP-2), without affecting cell proliferation and anchorage-independent cell growth in vitro. Notably, the conditioned medium from CYP4Z1-expressing cells enhanced proliferation, migration and tube formation of human umbilical vein endothelial cells, and promoted angiogenesis in the zebrafish embryo and chorioallantoic membrane of the chick embryo. In addition, there were lower levels of myristic acid and lauric acid, and higher contents of 20-hydroxyeicosatetraenoic acid (20-HETE) in CYP4Z1-expressing T47D cells compared with vector control. CYP4Z1 overexpression significantly increased tumor weight and microvessel density by 2.6-fold and 1.9-fold in human tumor xenograft models, respectively. Moreover, CYP4Z1 transfection increased the phosphorylation of ERK1/2 and PI3K/Akt, while PI3K or ERK inhibitors and siRNA silencing reversed CYP4Z1-mediated changes in VEGF-A and TIMP-2 expression. Conversely, HET0016, an inhibitor of the CYP4 family, potently inhibited the tumor-induced angiogenesis with associated changes in the intracellular levels of myristic acid, lauric acid and 20-HETE. Collectively, these data suggest that increased CYP4Z1 expression promotes tumor angiogenesis and growth in breast cancer partly via PI3K/Akt and ERK1/2 activation. -- Highlights: ► CYP4Z1 overexpression promotes human breast cancer growth and angiogenesis. ► The pro-angiogenic effects of CYP4Z1 have

Increased expression of CYP4Z1 promotes tumor angiogenesis and growth in human breast cancer

Energy Technology Data Exchange (ETDEWEB)

Yu, Wei [Department of Pharmacology, School of Medicine, Wuhan University, Wuhan 430071 (China); Chai, Hongyan [Center for Gene Diagnosis, Zhongnan Hospital, Wuhan University, Wuhan 430071 (China); Li, Ying; Zhao, Haixia; Xie, Xianfei; Zheng, Hao; Wang, Chenlong; Wang, Xue [Department of Pharmacology, School of Medicine, Wuhan University, Wuhan 430071 (China); Yang, Guifang [Department of Pathology, Zhongnan Hospital, Wuhan University, Wuhan 430071 (China); Cai, Xiaojun [Department of Ophthalmology, Zhongnan Hospital, Wuhan University, Wuhan 430071 (China); Falck, John R. [Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, TX 75390 (United States); Yang, Jing, E-mail: yangjingliu@yahoo.com.cn [Department of Pharmacology, School of Medicine, Wuhan University, Wuhan 430071 (China); Research Center of Food and Drug Evaluation, Wuhan University, Wuhan 430071 (China)

2012-10-01

Cytochrome P450 (CYP) 4Z1, a novel CYP4 family member, is over-expressed in human mammary carcinoma and associated with high-grade tumors and poor prognosis. However, the precise role of CYP4Z1 in tumor progression is unknown. Here, we demonstrate that CYP4Z1 overexpression promotes tumor angiogenesis and growth in breast cancer. Stable expression of CYP4Z1 in T47D and BT-474 human breast cancer cells significantly increased mRNA expression and production of vascular endothelial growth factor (VEGF)-A, and decreased mRNA levels and secretion of tissue inhibitor of metalloproteinase-2 (TIMP-2), without affecting cell proliferation and anchorage-independent cell growth in vitro. Notably, the conditioned medium from CYP4Z1-expressing cells enhanced proliferation, migration and tube formation of human umbilical vein endothelial cells, and promoted angiogenesis in the zebrafish embryo and chorioallantoic membrane of the chick embryo. In addition, there were lower levels of myristic acid and lauric acid, and higher contents of 20-hydroxyeicosatetraenoic acid (20-HETE) in CYP4Z1-expressing T47D cells compared with vector control. CYP4Z1 overexpression significantly increased tumor weight and microvessel density by 2.6-fold and 1.9-fold in human tumor xenograft models, respectively. Moreover, CYP4Z1 transfection increased the phosphorylation of ERK1/2 and PI3K/Akt, while PI3K or ERK inhibitors and siRNA silencing reversed CYP4Z1-mediated changes in VEGF-A and TIMP-2 expression. Conversely, HET0016, an inhibitor of the CYP4 family, potently inhibited the tumor-induced angiogenesis with associated changes in the intracellular levels of myristic acid, lauric acid and 20-HETE. Collectively, these data suggest that increased CYP4Z1 expression promotes tumor angiogenesis and growth in breast cancer partly via PI3K/Akt and ERK1/2 activation. -- Highlights: ► CYP4Z1 overexpression promotes human breast cancer growth and angiogenesis. ► The pro-angiogenic effects of CYP4Z1 have

MDR1 haplotypes conferring an increased expression of intestinal CYP3A4 rather than MDR1 in female living-donor liver transplant patients.

Science.gov (United States)

Hosohata, Keiko; Masuda, Satohiro; Yonezawa, Atsushi; Katsura, Toshiya; Oike, Fumitaka; Ogura, Yasuhiro; Takada, Yasutsugu; Egawa, Hiroto; Uemoto, Shinji; Inui, Ken-Ichi

2009-07-01

This study investigated whether haplotypes in the multidrug resistance 1 (MDR1) gene had effects on mRNA expression levels of MDR1 and cytochrome P450 (CYP) 3A4, and on the pharmacokinetics of tacrolimus in living-donor liver transplant (LDLT) patients, considering the gender difference. Haplotype analysis of MDR1 with G2677T/A and C3435T was performed in 63 de novo Japanese LDLT patients (17 to 55 years; 44.4% women). The expression levels of MDR1 and CYP3A4 mRNAs in jejunal biopsy specimens were quantified by real-time PCR. Intestinal CYP3A4 mRNA expression levels (amol/microg total RNA) showed significantly higher values in women carrying the 2677TT-3435TT haplotype (median, 10.7; range, 5.92-15.2) than those with 2677GG-3435CC (3.03; range 1.38-4.68) and 2677GT-3435CT (median, 4.31; range, 0.07-9.42) (P = 0.022), but not in men (P = 0.81). However, MDR1 haplotype did not influence mRNA expression levels of MDR1 nor the concentration/dose ratio [(ng/mL)/(mg/day)] of oral tacrolimus for the postoperative 7 days, irrespective of gender. MDR1 haplotype may have a minor association with the tacrolimus pharmacokinetics after LDLT, but could be a good predictor of the inter-individual variation of intestinal expression of CYP3A4 in women.

CYP3A5 Polymorphism In Serbian Paediatric Epileptic Patients On Carbamazepine Treatment

Directory of Open Access Journals (Sweden)

Milovanovic Dragana Dragas

2015-06-01

Full Text Available Carbamazepine exhibits significant inter-individual variability in its efficacy and safety, which leads to unpredictable therapy outcomes for the majority of patients. Although its complex biotransformation depends on CYP3A5 activity, evidence of association between carbamazepine treatment outcomes and CYP3A5 functional variations remains inconclusive. The aim of the present study was to investigate the distribution of two of the functionally important CYP3A5 variants *2 and *3 as well as their effects on carbamazepine dose requirements, plasma concentrations and clearance in a Serbian population. The study involved 40 paediatric epileptic patients on steady-state carbamazepine treatment. Genotyping was conducted using the PCR-RFLP method, and carbamazepine plasma concentrations were determined using the HPLC method. CYP3A5*2 and *3 polymorphisms were found at frequencies of 0.0% and 97.5%, respectively, which corresponds well to previously published data for Caucasians. No differences in CYP3A5*3 allele frequencies were detected among epileptic patients in comparison to healthy volunteers within similar ethnic populations (p>0.08, indicating that CYP3A5 polymorphism does not represent a risk factor for epilepsy development. There was an observed tendency towards lower dosage requirements (mean±SD: 15.06±4.45 mg/kg vs. 18.74±5.55 mg/kg; p=0.26, higher plasma concentrations (mean±SD: 0.45±0.13 mg/kg vs. 0.38±0.03 mg/kg; p=0.47 and lower clearance (mean±SD: 0.14±0.05 mg/kg vs. 0.15±0.01 mg/kg; p=0.79 of carbamazepine in homozygous carriers of CYP3A5*3/*3 compared to heterozygous CYP3A5*1A/*3 Serbians. Because these genotype groups did not differ significantly in terms of their carbamazepine pharmacokinetics parameters, the proposed effects of CYP3A5*3 on carbamazepine metabolism could not be confirmed.

Impact of CYP2C8*3 polymorphism on in vitro metabolism of imatinib to N-desmethyl imatinib.

Science.gov (United States)

Khan, Muhammad Suleman; Barratt, Daniel T; Somogyi, Andrew A

2016-01-01

1. Imatinib is metabolized to N-desmethyl imatinib by CYPs 3A4 and 2C8. The effect of CYP2C8*3 genotype on N-desmethyl imatinib formation was unknown. 2. We examined imatinib N-demethylation in human liver microsomes (HLMs) genotyped for CYP2C8*3, in CYP2C8*3/*3 pooled HLMs and in recombinant CYP2C8 and CYP3A4 enzymes. Effects of CYP-selective inhibitors on N-demethylation were also determined. 3. A single-enzyme Michaelis-Menten model with autoinhibition best fitted CYP2C8*1/*1 HLM (n = 5) and recombinant CYP2C8 kinetic data (median ± SD Ki = 139 ± 61 µM and 149 µM, respectively). Recombinant CYP3A4 showed two-site enzyme kinetics with no autoinhibition. Three of four CYP2C8*1/*3 HLMs showed single-enzyme kinetics with no autoinhibition. Binding affinity was higher in CYP2C8*1/*3 than CYP2C8*1/*1 HLM (median ± SD Km = 6 ± 2 versus 11 ± 2 µM, P=0.04). CYP2C8*3/*3 (pooled HLM) also showed high binding affinity (Km = 4 µM) and single-enzyme weak autoinhibition (Ki = 449 µM) kinetics. CYP2C8 inhibitors reduced HLM N-demethylation by 47-75%, compared to 0-30% for CYP3A4 inhibitors. 4. In conclusion, CYP2C8*3 is a gain-of-function polymorphism for imatinib N-demethylation, which appears to be mainly mediated by CYP2C8 and not CYP3A4 in vitro in HLM.

A comparative pharmacokinetic study in healthy volunteers of the effect of carbamazepine and oxcarbazepine on cyp3a4

DEFF Research Database (Denmark)

Andreasen, Astrid-Helene; Brøsen, Kim; Damkier, Per

2007-01-01

PURPOSE: Carbamazepine (CBZ) and oxcarbazepine (OXCZ) are well-known inducers of drug metabolism via CYP3A4. Indirect interaction studies and clinical experience suggest that CBZ has a stronger potential in this regard than OXCZ. However this has never been subject to a direct comparative study. We...

Characterization of midazolam metabolism in locusts: The role of CYP3A4-like enzyme in the formation of 1'-OH and 4-OH midazolam

DEFF Research Database (Denmark)

Olsen, Line Rørbæk; Gabel-Jensen, Charlotte; Wubshet, Sileshi Gizachew

2016-01-01

) were in the same range as reported in humans (in locusts: 7-23 and 33-85 µM for the formation of the 1'-OH and 4-OH metabolites, respectively). 3. The formation of hydroxylated metabolites could successfully be inhibited by co-administration of ketoconazole, a known CYP3A4/5 inhibitor. 4. Besides phase...

Lentiviral transgenic microRNA-based shRNA suppressed mouse cytochromosome P450 3A (CYP3A expression in a dose-dependent and inheritable manner.

Directory of Open Access Journals (Sweden)

Yong Wang

Full Text Available Cytochomosome P450 enzymes (CYP are heme-containing monooxygenases responsible for oxidative metabolism of many exogenous and endogenous compounds including drugs. The species difference of CYP limits the extent to which data obtained from animals can be translated to humans in pharmacodynamics or pharmacokinetics studies. Transgenic expression of human CYP in animals lacking or with largely reduced endogenous CYP counterparts is recognized as an ideal strategy to correct CYP species difference. CYP3A is the most abundant CYP subfamily both in human and mammals. In this study, we designed a microRNA-based shRNA (miR-shRNA simultaneously targeting four members of mouse CYP3A subfamily (CYP3A11, CYP3A16, CYP3A41 and CYP3A44, and transgenic mice expressing the designed miR-shRNA were generated by lentiviral transgenesis. Results showed that the CYP3A expression level in transgenic mice was markedly reduced compared to that in wild type or unrelated miR-shRNA transgenic mice, and was inversely correlated to the miR-shRNA expression level. The CYP3A expression levels in transgenic offspring of different generations were also remarkably lower compared to those of controls, and moreover the inhibition rate of CYP3A expression remained comparable over generations. The ratio of the targeted CYP3A transcriptional levels was comparable between knockdown and control mice of the same gender as detected by RT-PCR DGGE analysis. These data suggested that transgenic miR-shRNA suppressed CYP3A expression in a dose-dependent and inheritable manner, and transcriptional levels of the targeted CYP3As were suppressed to a similar extent. The observed knockdown efficacy was further confirmed by enzymatic activity analysis, and data showed that CYP3A activities in transgenic mice were markedly reduced compared to those in wild-type or unrelated miR-shRNA transgenic controls (1.11±0.71 vs 5.85±1.74, 5.9±2.4; P<0.01. This work laid down a foundation to further knock

Characterization CYP1A2, CYP2C9, CYP2C19 and CYP2D6 polymorphisms using HRMA in Psychiatry patients with schizophrenia and bipolar disease for personalized medicine.

Science.gov (United States)

Yenilmez, Ebru Dundar; Tamam, Lut; Karaytug, Onur; Tuli, Abdullah

2018-06-19

The interindividual genetic variations in drug metabolizing enzymes effects the impact and toxicity in plenty of drugs. The CYP1A2, CYP2C9, CYP2C19 and CYP2D6 gene polymorphisms characterized using high resolution melting analysis (HRMA) in follow-up patients in psychiatry clinic as a preliminary preparation for personalized medicine. Genotyping of CYP1A2*1F, CYP2C9 *2, *3, CYP2C19 *2, *3 and *17 and CYP2D6 *3, *4 was conducted in 101 patients using HRMA. Genotype and allele frequencies of the CYP variants were found to be in equilibrium with the Hardy-Weinberg equation. The frequency of the CYP1A2*1F allele in schizophrenia and bipolar disease was 0.694 and 0.255, respectively. The CYP2C9 allele frequencies were 0.087 (CYP2C9*2), and 0.549 (CYP2C9*3) for bipolar; 0.278 (CYP2C9*2) and 0.648 (CYP2C9*3) in schizophrenias. The CYP2C19*2 and *17 allele frequencies was 0.111 and 0.185 in schizophrenia and variant *2 was 0.117 and variant *17 was 0.255 in bipolar group. The frequency of the CYP2D6*3 allele was 0.027 in schizophrenias. The frequencies for the CYP2D6*4 variant was 0.092 and 0.096 in schizophrenia and bipolar groups, respectively. The knowledge in pharmacogenomics and also the developments in molecular genetics are growing rapidly. In the future this can be expected to provide new methodologies in the prediction of the activity in drug metabolizing enzymes. The HRMA is a rapid and useful technique to identify the genotypes for drug dosage adjustment before therapy in psychiatry patients. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Up-regulatation of CYP3A expression through pregnent X receptor by praeruptorin D isolated from Peucedanum praeruptorum Dunn.

Science.gov (United States)

Huang, Ling; Huang, Min; Li, Yu-Hua; Li, Rui-Ming; Zeng, Yu; Kuang, Shao-Yi; Zhang, Li; Wang, Yi-Tao; Bi, Hui-Chang

2013-07-09

Qianhu, the dried roots of Peucedanum praeruptorum DUNN (Umbelliferae), is a well-known traditional Chinese medicinal herb which was officially listed in the Chinese Pharmacopoeia. Praeruptorin D (PD) is one of the major active constituents of Peucedanum praeruptorum Dunn (Qianhu). The Pregnane X receptor (PXR) is an orphan nuclear receptor and plays a pivotal role in the activation of human cytochrome P450 3A4 (CYP3A4) gene. The purpose of this study was to investigate the effect of PD on the PXR-mediated transactivation of CYP3A4, and thus to predict potential herb-drug interactions between PD, Qianhu, and the other co-administered drugs that metabolized by CYP3A4. The effect of PD on the Cyp3a11, mPXR mRNA expression in mice primary hepatocytes was measured using real-time PCR. The gene expression, protein expression, and catalytic activity of CYP3A4 in the LS174T cells after transfected with PXR expression plasmids were determined by real-time PCR, Western blot analysis, and LC-MS/MS based CYP3A4 substrate assay. The results revealed that the level of Cyp3a11 gene expression in mice primary hepatocytes was significantly increased by PD, but PD cannot induce the mPXR gene expression. On the other hand, CYP3A4 mRNA, protein expression and functional activity in PXR-over-expression LS174T cells were significantly increased by PD through PXR-mediated pathway; conversely, no significant change was found in the untransfected cells. These findings suggest that PD can significantly up-regulate CYP3A4 expression and activity via the PXR-mediated pathway and this should be taken into consideration to predict any potential herb-drug interactions when PD and Peucedanum praeruptorum Dunn are co-administered with other drugs. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Metabolismus des tabakspezifischen Nitrosamins N'-Nitrosonornicotin in Gewebeschnitten von Mäusen, Ratten und Menschen sowie Möglichkeiten der Hemmung durch Nicotin, Cotinin und Phenethylisothiocyanat

OpenAIRE

Lassnack, Bettina

2005-01-01

Das Ziel dieser Arbeit war es, speziesspezifische Unterschiede in der Metabolisierung des tabakspezifischen Nitrosamins N'-Nitrosonornicotin (NNN) in den Organen Lunge und Leber von Mäusen, Ratten und Mensch zu untersuchen. Gewebeschnitte von Lunge und Leber der Maus, Ratte und des Menschen wurden unter identischen Versuchsbedingungen mit [5-3H]-NNN sechs Stunden inkubiert. Es kamen je 8 Konzentrationen von 0,001 bis 1,2 µM zum Einsatz. Bei Maus und Ratte wurden pro Konzentration die Schnitte...

Effects of strong CYP2D6 and 3A4 inhibitors, paroxetine and ketoconazole, on the pharmacokinetics and cardiovascular safety of tamsulosin

Science.gov (United States)

Troost, Joachim; Tatami, Shinji; Tsuda, Yasuhiro; Mattheus, Michaela; Mehlburger, Ludwig; Wein, Martina; Michel, Martin C

2011-01-01

AIM To determine the effect of the strong CYP2D6 inhibitor paroxetine and strong CYP3A4 inhibitor ketoconazole on the pharmacokinetics and safety (orthostatic challenge) of tamsulosin. METHODS Two open-label, randomized, two-way crossover studies were conducted in healthy male volunteers (extensive CYP2D6 metabolizers). RESULTS Co-administration of multiple oral doses of 20 mg paroxetine once daily with a single oral dose of the 0.4 mg tamsulosin HCl capsule increased the adjusted geometric mean (gMean) values of Cmax and AUC(0,∞) of tamsulosin by factors of 1.34 (90% CI 1.21, 1.49) and 1.64 (90% CI 1.44, 1.85), respectively, and increased the terminal half-life (t1/2) of tamsulosin HCl from 11.4 h to 15.3 h. Co-administration of multiple oral doses of 400 mg ketoconazole once dailywith a single oral dose of the 0.4 mg tamsulosin increased the gMean values of Cmax and AUC(0,∞) of tamsulosin by a factor of 2.20 (90% CI 1.96, 2.45) and 2.80 (90% CI 2.56, 3.07), respectively. The terminal half-life was slightly increased from 10.5 h to 11.8 h. These pharmacokinetic changes were not accompanied by clinically significant alterations of haemodynamic responses during orthostatic stress testing. CONCLUSION The exposure to tamsulosin is increased upon co-administration of strong CYP2D6 inhibitors and even more so of strong 3A4 inhibitors, but neither PK alteration was accompanied by clinically significant haemodynamic changes during orthostatic stress testing. PMID:21496064

CYP4F18-Deficient Neutrophils Exhibit Increased Chemotaxis to Complement Component C5a

Directory of Open Access Journals (Sweden)

Rachel Vaivoda

2015-01-01

Full Text Available CYP4Fs were first identified as enzymes that catalyze hydroxylation of leukotriene B4 (LTB4. CYP4F18 has an unusual expression in neutrophils and was predicted to play a role in regulating LTB4-dependent inflammation. We compared chemotaxis of wild-type and Cyp4f18 knockout neutrophils using an in vitro assay. There was no significant difference in the chemotactic response to LTB4, but the response to complement component C5a increased 1.9–2.25-fold in knockout cells compared to wild-type (P < 0.01. This increase was still observed when neutrophils were treated with inhibitors of eicosanoid synthesis. There were no changes in expression of other CYP4 enzymes in knockout neutrophils that might compensate for loss of CYP4F18 or lead to differences in activity. A mouse model of dextran sodium sulfate colitis was used to investigate the consequences of increased C5a-dependent chemotaxis in vivo, but there was no significant difference in weight loss, disease activity, or colonic tissue myeloperoxidase between wild-type and Cyp4f18 knockout mice. This study demonstrates the limitations of inferring CYP4F function based on an ability to use LTB4 as a substrate, points to expanding roles for CYP4F enzymes in immune regulation, and underscores the in vivo challenges of CYP knockout studies.

A Case of 3,4-Dimethoxyamphetamine (3,4-DMA) and 3,4-Methylenedioxymethamphetamine (MDMA) Toxicity with Possible Metabolic Interaction.

Science.gov (United States)

Darracq, Michael A; Thornton, Stephen L; Minns, Alicia B; Gerona, Roy R

2016-01-01

We present a case of "ecstasy" ingestion revealing 3,4-methylenedioxymethamphetamine (MDMA) and 3,4-dimethoxyamphetamine (3,4-DMA) and absence of cytochrome P450 (CYP)-2D6 MDMA metabolites. A 19-year-old presented following a seizure. Initial vital signs were normal. Laboratories were normal with the exception of sodium 127 mEq/L and urine drugs of abuse screen positive for amphetamines. Twelve hours later, serum sodium was 114 mEq/L and a second seizure occurred. After receiving hypertonic saline (3%), the patient had improvement in mental status and admitted to taking "ecstasy" at a rave prior to her initial presentation. Liquid chromatography-time-of-flight mass spectrometry (LC-TOF/MS) of serum and urine revealed MDMA, 3,4-DMA, and the CYP-2B6 MDMA metabolites 3,4-methylendioxyamphetamine (MDA) and 4-hydroxy-3-methoxyamphetamine (HMA). The CYP2D6 metabolites of MDMA, 3,4-dihydromethamphetamine (HHMA) and 4-hydroxy-3-methoxymethamphetamine (HMMA), were detected at very low levels. This case highlights the polypharmacy which may exist among users of psychoactive illicit substances and demonstrates that concurrent use of MDMA and 3,4-DMA may predispose patients to severe toxicity. Toxicologists and other healthcare providers should be aware of this potential toxicity.

Genotyping and phenotyping of CYP2D6 and CYP3A isoenzymes in patients with alcohol use disorder: correlation with haloperidol plasma concentration.

Science.gov (United States)

Sychev, Dmitry A; Zastrozhin, Mikhail S; Miroshnichenko, Igor I; Baymeeva, Natalia V; Smirnov, Valery V; Grishina, Elena A; Ryzhikova, Kristina A; Mirzaev, Karin B; Markov, Dmitry D; Skryabin, Valentin Y; Snalina, Nataliya E; Nosikova, Polina G; Savchenko, Ludmila M; Bryun, Evgeny A

2017-09-26

Haloperidol is used for the treatment of alcohol use disorders in patients with signs of alcohol-related psychosis. Haloperidol therapy poses a high risk of adverse drug reactions (ADR). Contradictory data, which include the effects of genetic polymorphisms in genes encoding the elements of haloperidol biotransformation system on haloperidol metabolism rate and plasma drug concentration ratio, are described in patients with different genotypes. The primary objective of this study was to investigate the effects of CYP2D6 and CYP3A5 genetic polymorphisms on haloperidol equilibrium concentration in patients with alcohol use disorder. The study included 69 male patients with alcohol use disorder. Genotyping was performed using the allele-specific real-time PCR. CYP2D6 and CYP3A were phenotyped with HPLC-MS using the concentration of endogenous substrate of the enzyme and its urinary metabolites [6-hydroxy-1,2,3,4-tetrahydro-β-carboline(6-HO-THBC) to pinoline ratio for CYP2D6 and 6-β-hydroxycortisol to cortisol ratio for CYP3A]. The equilibrium plasma concentration was determined using LC-MS-MS. Results indicated that both C/D indexes and equilibrium concentration levels depend on CYP2D6 genetic polymorphism, but only in patients receiving haloperidol intramuscular injections [0.26 (0.09; 0.48) vs. 0.54 (0.44; 0.74), p=0.037]. The study demonstrates that CYP2D6 genetic polymorphism (1846G>A) can affect haloperidol concentration levels in patients with alcohol use disorder.

Characterization of the genetic variation present in CYP3A4 in three South African populations

OpenAIRE

Drögemöller, Britt; Plummer, Marieth; Korkie, Lundi; Agenbag, Gloudi; Dunaiski, Anke; Niehaus, Dana; Koen, Liezl; Gebhardt, Stefan; Schneider, Nicol; Olckers, Antonel; Wright, Galen; Warnich, Louise

2013-01-01

TThe CYP3A4 enzyme is the most abundant human cytochrome P450 and is regarded as the most important enzyme involved in drug metabolism. Inter-individual and inter-population variability in gene expression and enzyme activity are thought to be influenced, in part, by genetic variation. Although Southern African individuals have been shown to exhibit the highest levels of genetic diversity, they have been under-represented in pharmacogenetic research to date. Therefore, the aim of this study wa...

Influence of CYP3A5 genetic variation on everolimus maintenance dosing after cardiac transplantation.

Science.gov (United States)

Lesche, Dorothea; Sigurdardottir, Vilborg; Setoud, Raschid; Englberger, Lars; Fiedler, Georg M; Largiadèr, Carlo R; Mohacsi, Paul; Sistonen, Johanna

2015-12-01

Everolimus (ERL) has become an alternative to calcineurin inhibitors (CNIs) due to its renal-sparing properties, especially in heart transplant (HTx) recipients with kidney dysfunction. However, ERL dosing is challenging due to its narrow therapeutic window combined with high interindividual pharmacokinetic variability. Our aim was to evaluate the effect of clinical and genetic factors on ERL dosing in a pilot cohort of 37 HTx recipients. Variants in CYP3A5, CYP3A4, CYP2C8, POR, NR1I2, and ABCB1 were genotyped, and clinical data were retrieved from patient charts. While ERL trough concentration (C0 ) was within the targeted range for most patients, over 30-fold variability in the dose-adjusted ERL C0 was observed. Regression analysis revealed a significant effect of the non-functional CYP3A5*3 variant on the dose-adjusted ERL C0 (p = 0.031). ERL dose requirement was 0.02 mg/kg/d higher in patients with CYP3A5*1/*3 genotype compared to patients with CYP3A5*3/*3 to reach the targeted C0 (p = 0.041). ERL therapy substantially improved estimated glomerular filtration rate (28.6 ± 6.6 mL/min/1.73 m(2)) in patients with baseline kidney dysfunction. Everolimus pharmacokinetics in HTx recipients is highly variable. Our preliminary data on patients on a CNI-free therapy regimen suggest that CYP3A5 genetic variation may contribute to this variability. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

CYP1A2 and NAT2 phenotyping and 3-aminobiphenyl and 4-aminobiphenyl hemoglobin adduct levels in smokers and non-smokers

International Nuclear Information System (INIS)

Sarkar, Mohamadi; Stabbert, Regina; Kinser, Robin D.; Oey, Jan; Rustemeier, Klaus; Holt, Klaus von; Schepers, Georg; Walk, Roger A.; Roethig, Hans J.

2006-01-01

Some aromatic amines are considered to be putative bladder carcinogens. Hemoglobin (Hb) adducts of 3-aminobiphenyl (3-ABP) and 4-aminobiphenyl (4-ABP) have been used as biomarkers of exposure to aromatic amines from cigarette smoke. One of the goals of this study was to determine intra- and inter-individual variability in 3-ABP and 4-ABP Hb adducts and to explore the predictability of ABP Hb adduct levels based on caffeine phenotyping. The study was conducted in adult smokers (S, n = 65) and non-smokers (NS, n 65). The subjects were phenotyped for CYP1A2 and NAT2 using urinary caffeine metabolites. Blood samples were collected twice within 6 weeks and adducts measured by GC/MS. The levels of 4-ABP Hb adducts were significantly (p < 0.0001) greater in S (34.5 ± 21.06 pg/g Hb) compared to NS (6.3 ± 3.02 pg/g Hb). The levels of 3-ABP Hb adducts were below the limit of quantification (BLOQ) in most (82%) of the NS and about 10-fold lower in S (3.6 ± 3.29 pg/g Hb) compared to 4-ABP Hb adducts. No differences were observed in the adduct levels between weeks 1 and 6 in the smokers, suggesting that a single sample would be adequate to monitor cigarette smoke exposure. The regression model developed with CYP1A2, NAT2 phenotype and number of cigarettes smoked (NCIG) accounted for 47% of the variability in 3-ABP adducts, whereas 32% variability in 4-ABP adducts was accounted by CYP1A2 and NCIG. The ratio of 4-ABP Hb adducts in adult S:NS was ∼ 5:1, whereas 3-ABP Hb adducts levels were BLOQ in some S, exhibited large interindividual variability (∼ 91% compared to 57% for 4-ABP Hb) and poor dose response relationship. Therefore, 4-ABP Hb adduct levels may be a more useful biomarker of aminobiphenyl exposure from cigarette smoke

Model based on GRID-derived descriptors for estimating CYP3A4 enzyme stability of potential drug candidates

Science.gov (United States)

Crivori, Patrizia; Zamora, Ismael; Speed, Bill; Orrenius, Christian; Poggesi, Italo

2004-03-01

A number of computational approaches are being proposed for an early optimization of ADME (absorption, distribution, metabolism and excretion) properties to increase the success rate in drug discovery. The present study describes the development of an in silico model able to estimate, from the three-dimensional structure of a molecule, the stability of a compound with respect to the human cytochrome P450 (CYP) 3A4 enzyme activity. Stability data were obtained by measuring the amount of unchanged compound remaining after a standardized incubation with human cDNA-expressed CYP3A4. The computational method transforms the three-dimensional molecular interaction fields (MIFs) generated from the molecular structure into descriptors (VolSurf and Almond procedures). The descriptors were correlated to the experimental metabolic stability classes by a partial least squares discriminant procedure. The model was trained using a set of 1800 compounds from the Pharmacia collection and was validated using two test sets: the first one including 825 compounds from the Pharmacia collection and the second one consisting of 20 known drugs. This model correctly predicted 75% of the first and 85% of the second test set and showed a precision above 86% to correctly select metabolically stable compounds. The model appears a valuable tool in the design of virtual libraries to bias the selection toward more stable compounds. Abbreviations: ADME - absorption, distribution, metabolism and excretion; CYP - cytochrome P450; MIFs - molecular interaction fields; HTS - high throughput screening; DDI - drug-drug interactions; 3D - three-dimensional; PCA - principal components analysis; CPCA - consensus principal components analysis; PLS - partial least squares; PLSD - partial least squares discriminant; GRIND - grid independent descriptors; GRID - software originally created and developed by Professor Peter Goodford.

Polymorphism of CYP3A4 and ABCB1 genes increase the risk of neuropathy in breast cancer patients treated with paclitaxel and docetaxel

Directory of Open Access Journals (Sweden)

Kus T

2016-08-01

Full Text Available Tulay Kus,1 Gokmen Aktas,1 Mehmet Emin Kalender,1 Abdullah Tuncay Demiryurek,2 Mustafa Ulasli,1 Serdar Oztuzcu,3 Alper Sevinc,1 Seval Kul,4 Celaletdin Camci1 1Department of Internal Medicine, Division of Medical Oncology, University of Gaziantep, Gaziantep Oncology Hospital, Gaziantep, Turkey; 2Department of Medical Pharmacology, 3Department of Medical Biology, Faculty of Medicine, University of Gaziantep, Gaziantep, Turkey; 4Department of Biostatistics, Faculty of Medicine, University of Gaziantep, Gaziantep, Turkey Background: Interindividual variability of pharmacogenetics may account for unpredictable neurotoxicities of taxanes. Methods: From March 2011 to June 2015, female patients with operable breast cancer who had received docetaxel- or paclitaxel-containing adjuvant chemotherapy were included in this study. All patients were treated with single-agent paclitaxel intravenously (IV 175 mg/m2 every 3 weeks for four cycles, or IV 80 mg/m2 weekly for 12 cycles, and IV 100 mg/m2 docetaxel for four cycles as adjuvant treatment. We evaluated the relationship between neurotoxicity of taxanes and single-nucleotide polymorphisms of ABCB1, CYP3A4, ERCC1, ERCC2, FGFR4, TP53, ERBB2, and CYP2C8 genes. Taxane-induced neurotoxicity during the treatment was evaluated according to the National Cancer Institute Common Toxicity Criteria version 4.03 prior to each cycle. Chi-squared tests were used to compare the two groups, and multivariate binary logistic regression models were used for determining possible risk factors of neuropathy. Results: Pharmacogenetic analysis was performed in 219 females. ABCB1 3435 TT genotype had significantly higher risk for grade ≥2 neurotoxicity (odds ratio [OR]: 2.759, 95% confidence interval [CI]: 1.172–6.493, P: 0.017 compared to TC and CC genotype, and also CYP3A4 392 AA and AG genotype had significantly higher risk for grade ≥2 neurotoxicity (OR: 2.259, 95% CI: 1.033–4.941, P: 0.038 compared to GG genotype. For

Fentanyl Enhances Hepatotoxicity of Paclitaxel via Inhibition of CYP3A4 and ABCB1 Transport Activity in Mice

Science.gov (United States)

Pan, Jia-Hao; Bi, Bing-Tian; Feng, Kun-Yao; Huang, Wan; Zeng, Wei-An

2015-01-01

Fentanyl, a potent opioid analgesic that is used to treat cancer pain, is commonly administered with paclitaxel in advanced tumors. However, the effect of fentanyl on the hepatotoxicity of paclitaxel and its potential mechanism of action is not well studied. The purpose of this study was to investigate the effect of fentanyl on the hepatotoxicity of paclitaxel and its potential mechanisms of action. Pharmacokinetic parameters of paclitaxel were tested using reversed phase high-performance liquid chromatography (RP-HPLC). Aspartate transaminase (AST), alanine aminotransferase (ALT), and mouse liver histopathology were examined. Moreover, the cytotoxicity of anti-carcinogens was examined using 1-(4, 5-dimethylthiazol-2-yl)-3,5-diphenylformazan (MTT), and the intracellular accumulation of doxorubicin and rhodamine 123 was detected by flow cytometry. Furthermore, the expression of ABCB1 and the activity of ABCB1 ATPase and CYP3A4 were also examined. In this study, the co-administration of fentanyl and paclitaxel prolonged the half-life (t1/2) of paclitaxel from 1.455 hours to 2.344 hours and decreased the clearance (CL) from 10.997 ml/h to 7.014 ml/h in mice. Fentanyl significantly increased the levels of ALT in mice to 88.2 U/L, which is more than 2-fold higher than the level detected in the control group, and it increased the histological damage in mouse livers. Furthermore, fentanyl enhanced the cytotoxicity of anti-carcinogens that are ABCB1 substrates and increased the accumulation of doxorubicin and rhodamine 123. Additionally, fentanyl stimulated ABCB1 ATPase activity and inhibited CYP3A4 activity in the liver microsomes of mice. Our study indicates that the obvious hepatotoxicity during this co-administration was due to the inhibition of CYP3A4 activity and ABCB1 transport activity. These findings suggested that the accumulation-induced hepatotoxicity of paclitaxel when it is combined with fentanyl should be avoided. PMID:26633878

Role of genetic polymorphisms of CYP1A1, CYP3A5, CYP2C9, CYP2D6, and PON1 in the modulation of DNA damage in workers occupationally exposed to organophosphate pesticides

International Nuclear Information System (INIS)

Singh, Satyender; Kumar, Vivek; Vashisht, Kapil; Singh, Priyanka; Banerjee, Basu Dev; Rautela, Rajender Singh; Grover, Shyam Sunder; Rawat, Devendra Singh; Pasha, Syed Tazeen; Jain, Sudhir Kumar; Rai, Arvind

2011-01-01

Organophosphate pesticides (OPs) are primarily metabolized by several xenobiotic metabolizing enzymes (XMEs). Very few studies have explored genetic polymorphisms of XMEs and their association with DNA damage in pesticide-exposed workers. The present study was designed to determine the role of genetic polymorphisms of CYP1A1, CYP3A5, CYP2C9, CYP2D6, and PON1 in the modulation of DNA damage in workers occupationally exposed to OPs. We examined 284 subjects including 150 workers occupationally exposed to OPs and 134 normal healthy controls. The DNA damage was evaluated using the alkaline comet assay and genotyping was done using PCR–RFLP. The results revealed that the PONase activity toward paraoxonase and AChE activity was found significantly lowered in workers as compared to control subjects (p < 0.001). Workers showed significantly higher DNA damage compared to control subjects (14.37 ± 2.15 vs. 6.24 ± 1.37 tail% DNA, p < 0.001). Further, the workers with CYP2D6*3 PM and PON1 (QQ and MM) genotypes were found to have significantly higher DNA damage when compared to other genotypes (p < 0.05). In addition, significant increase in DNA damage was also observed in workers with concomitant presence of certain CYP2D6 and PON1 (Q192R and L55M) genotypes which need further extensive studies. In conclusion, the results indicate that the PON1 and CYP2D6 genotypes can modulate DNA damage elicited by some OPs possibly through gene-environment interactions. -- Highlights: ► Role of CYP1A1, CYP3A5, CYP2C, CYP2D6 and PON1 genotypes on DNA damage. ► Workers exposed to some OPs demonstrated increased DNA damage. ► CYP2D6 *3 PM and PON1 (Q192R and L55M) genotypes are associated with DNA damage. ► Concomitant presence of certain CYP2D6 and PON1 genotypes can increase DNA damage.

Amine-free melanin-concentrating hormone receptor 1 antagonists: Novel 1-(1H-benzimidazol-6-yl)pyridin-2(1H)-one derivatives and design to avoid CYP3A4 time-dependent inhibition.

Science.gov (United States)

Igawa, Hideyuki; Takahashi, Masashi; Shirasaki, Mikio; Kakegawa, Keiko; Kina, Asato; Ikoma, Minoru; Aida, Jumpei; Yasuma, Tsuneo; Okuda, Shoki; Kawata, Yayoi; Noguchi, Toshihiro; Yamamoto, Syunsuke; Fujioka, Yasushi; Kundu, Mrinalkanti; Khamrai, Uttam; Nakayama, Masaharu; Nagisa, Yasutaka; Kasai, Shizuo; Maekawa, Tsuyoshi

2016-06-01

Melanin-concentrating hormone (MCH) is an attractive target for antiobesity agents, and numerous drug discovery programs are dedicated to finding small-molecule MCH receptor 1 (MCHR1) antagonists. We recently reported novel pyridine-2(1H)-ones as aliphatic amine-free MCHR1 antagonists that structurally featured an imidazo[1,2-a]pyridine-based bicyclic motif. To investigate imidazopyridine variants with lower basicity and less potential to inhibit cytochrome P450 3A4 (CYP3A4), we designed pyridine-2(1H)-ones bearing various less basic bicyclic motifs. Among these, a lead compound 6a bearing a 1H-benzimidazole motif showed comparable binding affinity to MCHR1 to the corresponding imidazopyridine derivative 1. Optimization of 6a afforded a series of potent thiophene derivatives (6q-u); however, most of these were found to cause time-dependent inhibition (TDI) of CYP3A4. As bioactivation of thiophenes to form sulfoxide or epoxide species was considered to be a major cause of CYP3A4 TDI, we introduced electron withdrawing groups on the thiophene and found that a CF3 group on the ring or a Cl adjacent to the sulfur atom helped prevent CYP3A4 TDI. Consequently, 4-[(5-chlorothiophen-2-yl)methoxy]-1-(2-cyclopropyl-1-methyl-1H-benzimidazol-6-yl)pyridin-2(1H)-one (6s) was identified as a potent MCHR1 antagonist without the risk of CYP3A4 TDI, which exhibited a promising safety profile including low CYP3A4 inhibition and exerted significant antiobesity effects in diet-induced obese F344 rats. Copyright © 2016 Elsevier Ltd. All rights reserved.

CYP3A5 polymorphisms in renal transplant recipients: influence on tacrolimus treatment

Directory of Open Access Journals (Sweden)

Chen L

2018-03-01

Full Text Available Lucy Chen,1 G V Ramesh Prasad2 1Kidney Transplant Program, St Michael’s Hospital, Toronto, ON, Canada; 2Division of Nephrology, St Michael’s Hospital, Toronto, ON, Canada Abstract: Tacrolimus is a commonly used immunosuppressant after kidney transplantation. It has a narrow therapeutic range and demonstrates wide interindividual variability in pharmacokinetics, leading to potential underimmunosuppression or toxicity. Genetic polymorphism in CYP3A5 enzyme expression contributes to differences in tacrolimus bioavailability between individuals. Individuals carrying one or more copies of the wild-type allele *1 express CYP3A5, which increases tacrolimus clearance. CYP3A5 expressers require 1.5 to 2-fold higher tacrolimus doses compared to usual dosing to achieve therapeutic blood concentrations. Individuals with homozygous *3/*3 genotype are CYP3A5 nonexpressers. CYP3A5 nonexpression is the most frequent phenotype in most ethnic populations, except blacks. Differences between CYP3A5 genotypes in tacrolimus disposition have not translated into differences in clinical outcomes, such as acute rejection and graft survival. Therefore, although genotype-based dosing may improve achievement of therapeutic drug concentrations with empiric dosing, its role in clinical practice is unclear. CYP3A5 genotype may predict differences in absorption of extended-release and immediate-release oral formulations of tacrolimus. Two studies found that CYP3A5 expressers require higher doses of tacrolimus in the extended-release formulation compared to immediate release. CYP3A5 genotype plays a role in determining the impact of interacting drugs, such as fluconazole, on tacrolimus pharmacokinetics. Evidence conflicts regarding the impact of CYP3A5 genotype on risk of nephrotoxicity associated with tacrolimus. Further study is required. Keywords: calcineurin inhibitor, graft, pharmacogenomics, kidney, genotype

Patientenakzeptanz und Verträglichkeit von Risedronat zur Osteoporosetherapie - Ergebnisse einer Phase IV-Studie

Directory of Open Access Journals (Sweden)

Pusch HH

2001-01-01

Full Text Available Anwendungsbeobachtungen (Phase IV-Studien haben nach den erfolgreich absolvierten klinischen Prüfungen (Phase III den Zweck, die Verträglichkeit, Sicherheit, Compliance und Akzeptanz eines Präparates in der täglichen praktischen Medizin zu erfassen und auszuwerten. Eine AWB ist prospektiv aufgebaut. Die 1121 in dieser Studie erfaßten Patienten stellen ein solides Fundament für die statistische Aufarbeitung der erhobenen Daten dar. Die Ergebnisse zu den Themen Patientendaten, Diagnosestellung, Risikofaktoren, Frakturen und Mobilität zeigen deutlich, daß hier ein spezielles Patientengut vorliegt: Der Patient mit Osteoporose mit den typischen alterskorrelierten Begleiterkrankungen. Die Osteoporose verläuft, so lange keine Frakturen eintreten, praktisch schmerzfrei - ein Grund, warum diese Erkrankung so oft und so lange ignoriert wird. Begleitende Beschwerden gehen nach der derzeitigen Lehrmeinung stets auf degenerative oder entzündlich-rheumatische Prozesse am Bewegungsapparat zurück. Aus diesem Blickwinkel bieten die Ergebnisse dieser Studie eine Überraschung: Die im Bereich der Wirbelsäule beobachtete und statistisch hochsignifikante Schmerzlinderung. Sie läßt sich jedoch mit der Hemmung des Knochenabbaues durch Risedronat alleine nicht befriedigend erklären. Möglicherweise liegen hier andere, bisher unbekannte pharmakologische Effekte dieser Substanz vor. Zusätzliche Hinweise ergeben sich daraus, daß beim Mb. Paget ähnliche Wirkungen auf die Begleitschmerzen bei Bisphosphonatbehandlung berichtet wurden. Die analgetische Wirkung von Calcitonin ist bekannt, sie beruht auf einer zentralnervösen Wirkung, ebenso wie auf der Hemmung der Freisetzung von freien Kalzium-Ionen, die als Schmerzmediator angesehen werden, aus dem Knochen. Vielleicht liegen hier vergleichbare Effekte vor, die in den Phase II und III Studien nicht zutage treten konnten. Diese überraschenden Ergebnisse sollten Anlaß für weitere Forschungen sein. Die

Effect of CYP2C9, VKORC1, CYP4F2 and GGCX genetic variants on warfarin maintenance dose and explicating a new pharmacogenetic algorithm in South Indian population.

Science.gov (United States)

Krishna Kumar, Dhakchinamoorthi; Shewade, Deepak Gopal; Loriot, Marie-Anne; Beaune, Philippe; Balachander, Jayaraman; Sai Chandran, B V; Adithan, Chandrasekaran

2014-01-01

To determine the influence of genetic polymorphisms on warfarin maintenance dose and to explicate an algorithm using the pharmacogenetic and clinical factors to determine the maintenance and/or starting dose of warfarin in South Indian patients receiving warfarin therapy. Patients receiving stabilized warfarin therapy (n=257) were included in the study. Single nucleotide polymorphisms (SNPs) of CYP2C9 (rs1799853 and rs1057910), VKORC1 (rs9923231, rs7196161, rs2884737, rs9934438, rs8050894, rs2359612 and rs7294), CYP4F2 (rs2108622) and GGCX (rs11676382) were genotyped by the quantitative real time-PCR method. The mean daily maintenance dose of warfarin was found to be 4.7 ± 2.1 mg/day. Patients with the CYP2C9*1/*2, *1/*3 and *2/*3 variant genotypes required a 51.0 (2.8 mg), 60.9 (2.3 mg) and 62.2 % (2.2 mg) lower daily maintenance dose of warfarin, respectively, than those patients with the CYP2C9*1/*1 wild-type genotype (5.2 mg) (pmaintenance dose. Genetic polymorphisms of CYP2C9, VKORC1, CYP4F2 and GGCX are important predictive factors of warfarin maintenance dose, and the developed algorithm will be useful to predict the required maintenance and/or starting warfarin dose in South Indian populations.

Role of genetic polymorphisms of CYP1A1, CYP3A5, CYP2C9, CYP2D6, and PON1 in the modulation of DNA damage in workers occupationally exposed to organophosphate pesticides

Energy Technology Data Exchange (ETDEWEB)

Singh, Satyender [Division of Biochemistry and Biotechnology, National Centre for Disease Control 22, Sham Nath Marg, Delhi-110054 (India); Kumar, Vivek [Environmental Biochemistry and Molecular Biology laboratory, Department of Biochemistry, University College of Medical Sciences and GTB Hospital, University of Delhi, Dilshad Garden, Delhi-110095 (India); Vashisht, Kapil; Singh, Priyanka [Division of Biochemistry and Biotechnology, National Centre for Disease Control 22, Sham Nath Marg, Delhi-110054 (India); Banerjee, Basu Dev, E-mail: banerjeebd@hotmail.com [Environmental Biochemistry and Molecular Biology laboratory, Department of Biochemistry, University College of Medical Sciences and GTB Hospital, University of Delhi, Dilshad Garden, Delhi-110095 (India); Rautela, Rajender Singh; Grover, Shyam Sunder; Rawat, Devendra Singh; Pasha, Syed Tazeen [Division of Biochemistry and Biotechnology, National Centre for Disease Control 22, Sham Nath Marg, Delhi-110054 (India); Jain, Sudhir Kumar [Centre for Epidemiology and Parasitic Diseases, National Centre for Disease Control 22, Sham Nath Marg, Delhi-110054 (India); Rai, Arvind [Division of Biochemistry and Biotechnology, National Centre for Disease Control 22, Sham Nath Marg, Delhi-110054 (India)

2011-11-15

Organophosphate pesticides (OPs) are primarily metabolized by several xenobiotic metabolizing enzymes (XMEs). Very few studies have explored genetic polymorphisms of XMEs and their association with DNA damage in pesticide-exposed workers. The present study was designed to determine the role of genetic polymorphisms of CYP1A1, CYP3A5, CYP2C9, CYP2D6, and PON1 in the modulation of DNA damage in workers occupationally exposed to OPs. We examined 284 subjects including 150 workers occupationally exposed to OPs and 134 normal healthy controls. The DNA damage was evaluated using the alkaline comet assay and genotyping was done using PCR-RFLP. The results revealed that the PONase activity toward paraoxonase and AChE activity was found significantly lowered in workers as compared to control subjects (p < 0.001). Workers showed significantly higher DNA damage compared to control subjects (14.37 {+-} 2.15 vs. 6.24 {+-} 1.37 tail% DNA, p < 0.001). Further, the workers with CYP2D6*3 PM and PON1 (QQ and MM) genotypes were found to have significantly higher DNA damage when compared to other genotypes (p < 0.05). In addition, significant increase in DNA damage was also observed in workers with concomitant presence of certain CYP2D6 and PON1 (Q192R and L55M) genotypes which need further extensive studies. In conclusion, the results indicate that the PON1 and CYP2D6 genotypes can modulate DNA damage elicited by some OPs possibly through gene-environment interactions. -- Highlights: Black-Right-Pointing-Pointer Role of CYP1A1, CYP3A5, CYP2C, CYP2D6 and PON1 genotypes on DNA damage. Black-Right-Pointing-Pointer Workers exposed to some OPs demonstrated increased DNA damage. Black-Right-Pointing-Pointer CYP2D6 *3 PM and PON1 (Q192R and L55M) genotypes are associated with DNA damage. Black-Right-Pointing-Pointer Concomitant presence of certain CYP2D6 and PON1 genotypes can increase DNA damage.

Investigations for assessing the health hazards of dichloromethane exposure. Final report; Untersuchungen zur Abschaetzung des gesundheitlichen Risikos infolge Dichlormethanexposition. Schlussbericht

Energy Technology Data Exchange (ETDEWEB)

Pankow, D. [Halle-Wittenberg Univ., Halle (Germany). Inst. fuer Pharmakologie und Toxikologie

1994-04-01

The oxidative CYP2E1-catalyzed dechlorination of dichloremethane (DCM) is a saturable pathway with high affinity and low capacity. In general, due to increasing DCM concentration in the inhaled air COHb values up to 12% are expected, but higher COHb levels were found according to some case reports. It was the aim of this study to characterize the DCM-derived COHb formation and to test effects of DCM on the cardiovascular system using animal models. The results show that a high CYP2E1 activity correlates with enhanced COHb formation. The COHb formation is inhibited and the DCM level in the blood enhanced after simultaneous exposure to DCM and solvents or drugs known as substrates of CYP2E1. Therefore, the risk assessment of DCM exposure must comprimise a possible additional uptake of a substrate or inducer of CYP2E1. Existing PBPK models proved to be an insufficient description in cases of an accidental scenario, a short exposure to very high concentration of DCM. The moderate CO hypoxia following DCM exposure is determined not only by the carboxyhemoglobinemia but also by an inhibition of cytochrome c oxidase activity. Cardiovascular dysfunctions, mainly arrhythmias, are detectable after exposure to relatively high DCM concentration as well as under pathophysiological conditions which per se influenced the cardiovascular system. (orig.) [Deutsch] Die zu CO fuehrende oxydative, CYP2E1-katalysierte Dechlorierung von Dichlormethan (DCM) verfuegt ueber eine hohe Affinitaet bei begrenzter Kapazitaet, es handelt sich um einen saettigbaren Stoffwechselweg. Im allgemeinen werden trotz steigender DCM-Konzentration in der eingeatmeten Luft Werte von 10-12% COHb nicht ueberschritten. Vergiftungsfaelle mit weitaus hoeheren COHb-Spiegeln wurden aber beschrieben. Ziel der tierexperimentellen Untersuchungen war es, die DCM-bedingte COHb-Bildung naeher zu charakterisieren und die Wirkung von DCM auf kardiovaskulaere Funktionen zu pruefen. Die Resultate zeigen, dass es bei

Relative contributions of the major human CYP450 to the metabolism of icotinib and its implication in prediction of drug-drug interaction between icotinib and CYP3A4 inhibitors/inducers using physiologically based pharmacokinetic modeling.

Science.gov (United States)

Chen, Jia; Liu, Dongyang; Zheng, Xin; Zhao, Qian; Jiang, Ji; Hu, Pei

2015-06-01

Icotinib is an anticancer drug, but relative contributions of CYP450 have not been identified. This study was carried out to identify the contribution percentage of CYP450 to icotinib and use the results to develop a physiologically based pharmacokinetic (PBPK) model, which can help to predict drug-drug interaction (DDI). Human liver microsome (HLM) and supersome using relative activity factor (RAF) were employed to determine the relative contributions of the major human P450 to the net hepatic metabolism of icotinib. These values were introduced to develop a PBPK model using SimCYP. The model was validated by the observed data in a Phase I clinical trial in Chinese healthy subjects. Finally, the model was used to simulate the DDI with ketoconazole or rifampin. Final contribution of CYP450 isoforms determined by HLM showed that CYP3A4 provided major contributions to the metabolism of icotinib. The percentage contributions of the P450 to the net hepatic metabolism of icotinib were determined by HLM inhibition assay and RAF. The AUC ratio under concomitant use of ketoconazole and rifampin was 3.22 and 0.55, respectively. Percentage of contribution of CYP450 to icotinib metabolism was calculated by RAF. The model has been proven to fit the observed data and is used in predicting icotinib-ketoconazole/rifampin interaction.

The influence of CYP3A5 polymorphisms on haloperidol treatment in patients with alcohol addiction

Directory of Open Access Journals (Sweden)

Zastrozhin MS

2017-12-01

Full Text Available Mikhail Sergeevich Zastrozhin,1,2 Elena Anatolievna Grishina,1 Kristina Anatolievna Ryzhikova,1 Valery Valerievich Smirnov,3 Ludmila Mikhailovna Savchenko,1 Evgeny Alekseevich Bryun,1,2 Dmitry Alekseevich Sychev1 1Russian Medical Academy of Continuous Professional Education, Ministry of Health of the Russian Federation, 2Moscow Research and Practical Centre on Addictions, Moscow Department of Healthcare, 3National Research Center Institute of Immunology, Federal Medical-Biological Agency, Moscow, Russia Background: Isoenzymes CYP2D6 and CYP3A4, the activity of which varies widely, are involved in metabolism of haloperidol and may influence its profile of efficacy and safety.Objective: The primary aim of this study was to estimate the relationship between CYP3A5 gene polymorphism, activity of the CYP3A isoenzyme, and the risk of development of adverse drug reactions by haloperidol in patients with alcohol abuse.Methods: Sixty-six male alcohol-addicted patients participated in the study. The safety of haloperidol was evaluated by Udvalg for Kliniske Undersogelser Side Effect Rating Scale (UKU and Simpson–Angus Scale for extrapyramidal symptoms (SAS. The activity of CYP3A was evaluated by determining the concentrations of an endogenous substrate of this isoenzyme (cortisol and its urinary metabolite (6-beta-hydroxycortisol, 6-B-HC. Genotyping of CYP3A5*3 was performed by real-time polymerase chain reaction with allele-specific hybridization.Results: The frequency of A-allele occurrence in Russian population was very poor (2.27%. CYP3A5*3 polymorphism had no influence on safety profile indicators of haloperidol (UKU scale: p=0.55, SAS scale: p=0.64. In addition, there was no statistical significant difference between the values of indexes of the metabolic ratio (6-B-HC/cortisol in groups with different genotypes of CYP3A5*3: GG 5.00 (3.36; 6.39 vs AG 5.26 (2.10; 6.78 (p=0.902.Conclusion: The frequency of A-allele occurrence of CYP3A5*3 in Russian

Global pharmacogenomics: distribution of CYP3A5 polymorphisms and phenotypes in the Brazilian population.

Directory of Open Access Journals (Sweden)

Guilherme Suarez-Kurtz

Full Text Available The influence of self-reported "race/color", geographical origin and genetic ancestry on the distribution of three functional CYP3A5 polymorphisms, their imputed haplotypes and inferred phenotypes was examined in 909 healthy, adult Brazilians, self-identified as White, Brown or Black ("race/color" categories of the Brazilian census. The cohort was genotyped for CYP3A5*3 (rs776746, CYP3A5*6 (rs10264272 and CYP3A5*7 (rs41303343, CYP3A5 haplotypes were imputed and CYP3A5 metabolizer phenotypes were inferred according to the number of defective CYP3A5 alleles. Estimates of the individual proportions of Amerindian, African and European ancestry were available for the entire cohort. Multinomial log-linear regression models were applied to infer the statistical association between the distribution of CYP3A5 alleles, haplotypes and phenotypes (response variables, and self-reported Color, geographical region and ancestry (explanatory variables. We found that Color per se or in combination with geographical region associates significantly with the distribution of CYP3A5 variant alleles and CYP3A5 metabolizer phenotypes, whereas geographical region per se influences the frequency distribution of CYP3A5 variant alleles. The odds of having the default CYP3A5*3 allele and the poor metabolizer phenotype increases continuously with the increase of European ancestry and decrease of African ancestry. The opposite trend is observed in relation to CYP3A5*6, CYP3A5*7, the default CYP3A5*1 allele, and both the extensive and intermediate phenotypes. No significant effect of Amerindian ancestry on the distribution of CYP3A5 alleles or phenotypes was observed. In conclusion, this study strongly supports the notion that the intrinsic heterogeneity of the Brazilian population must be acknowledged in the design and interpretation of pharmacogenomic studies, and dealt with as a continuous variable, rather than proportioned in arbitrary categories that do not capture the

Effects of CYP3A5, CYP2C19, and CYP2B6 on the clinical efficacy and adverse outcomes of sibutramine therapy: a crucial role for the CYP2B6*6 allele.

Science.gov (United States)

Hwang, In Cheol; Park, Ji Young; Ahn, Hong Yup; Kim, Kyoung Kon; Suh, Heuy Sun; Ko, Ki Dong; Kim, Kyoung-Ah

2014-01-20

Various cytochrome P450 isoforms modulate sibutramine activity and influence sibutramine plasma levels and pharmacokinetics. However, there are no available data to demonstrate the association of these polymorphisms with the clinical outcomes of sibutramine administration. This study was a sub-investigation of a 12-week, double-blind, placebo-controlled trial examining the additive effect of orlistat on sibutramine. The final analysis was restricted to 101 women who had fulfilled the protocol. We evaluated the effects of genetic polymorphisms of CYP3A5, CYP2C19 and CYP2B6 on the % weight loss and the occurrence of adverse events. The change of pulse rate from baseline value was affected by both CYP2B6 and CYP3A5 genetic polymorphisms (Psibutramine treatment. Copyright © 2013 Elsevier B.V. All rights reserved.

Roles of Human CYP2A6 and Monkey CYP2A24 and 2A26 Cytochrome P450 Enzymes in the Oxidation of 2,5,2',5'-Tetrachlorobiphenyl.

Science.gov (United States)

Shimada, Tsutomu; Kakimoto, Kensaku; Takenaka, Shigeo; Koga, Nobuyuki; Uehara, Shotaro; Murayama, Norie; Yamazaki, Hiroshi; Kim, Donghak; Guengerich, F Peter; Komori, Masayuki

2016-12-01

2,5,2',5'-Tetrachlorobiphenyl (TCB) induced type I binding spectra with cytochrome P450 (P450) 2A6 and 2A13, with K s values of 9.4 and 0.51 µM, respectively. However, CYP2A6 oxidized 2,5,2',5'-TCB to form 4-hydroxylated products at a much higher rate (∼1.0 minute -1 ) than CYP2A13 (∼0.02 minute -1 ) based on analysis by liquid chromatography-tandem mass spectrometry. Formation of 4-hydroxy-2,5,2',5'-TCB by CYP2A6 was greater than that of 3-hydroxy-2,5,2',5'-TCB and three other hydroxylated products. Several human P450 enzymes, including CYP1A1, 1A2, 1B1, 2B6, 2D6, 2E1, 2C9, and 3A4, did not show any detectable activities in oxidizing 2,5,2',5'-TCB. Cynomolgus monkey CYP2A24, which shows 95% amino acid identity to human CYP2A6, catalyzed 4-hydroxylation of 2,5,2',5'-TCB at a higher rate (∼0.3 minute -1 ) than CYP2A26 (93% identity to CYP2A6, ∼0.13 minute -1 ) and CYP2A23 (94% identity to CYP2A13, ∼0.008 minute -1 ). None of these human and monkey CYP2A enzymes were catalytically active in oxidizing other TCB congeners, such as 2,4,3',4'-, 3,4,3',4'-, and 3,5,3',5'-TCB. Molecular docking analysis suggested that there are different orientations of interaction of 2,5,2',5'-TCB with the active sites (over the heme) of human and monkey CYP2A enzymes, and that ligand interaction energies (U values) of bound protein-ligand complexes show structural relationships of interaction of TCBs and other ligands with active sites of CYP2A enzymes. Catalytic differences in human and monkey CYP2A enzymes in the oxidation of 2,5,2',5'-TCB are suggested to be due to amino acid changes at substrate recognition sites, i.e., V110L, I209S, I300F, V365M, S369G, and R372H, based on the comparison of primary sequences. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

CYP3A5*3 and ABCB1 61A>G Significantly Influence Dose-adjusted Trough Blood Tacrolimus Concentrations in the First Three Months Post-Kidney Transplantation.

Science.gov (United States)

Hu, Rong; Barratt, Daniel T; Coller, Janet K; Sallustio, Benedetta C; Somogyi, Andrew A

2018-03-30

Tacrolimus (TAC) is a first-line immunosuppressant used to prevent organ rejection after kidney transplantation. There is large inter-individual variability in its pharmacokinetics. Single nucleotide polymorphisms (SNPs) in genes encoding TAC metabolizing enzymes cytochromes P450 3A4/5 (CYP3A4/5), P-glycoprotein efflux transporter (ABCB1), their expression regulator pregnane X receptor (NR1I2) and CYP3A co-factor cytochrome P450 reductase (POR) have been studied for their effects on tacrolimus disposition. However, except for CYP3A5*3, controversies remain about their roles in predicting dose-adjusted trough blood TAC concentrations (C 0 /D). This study aimed to investigate the effects of ABCB1 (61A>G, 1199G>A, 1236C>T, 2677G>T and 3435C>T), CYP3A4*22, CYP3A5*3, NR1I2 (8055C>T, 63396C>T and -25385C>T) and POR*28 SNPs on TAC C 0 /D. In total, 165 kidney transplant recipients were included in this study. SNPs were genotyped by probe-based real-time polymerase chain reaction. Associations between log-transformed whole blood TAC C 0 /D (measured at 1 and 3 months post-transplant) and genotypes/haplotypes were assessed by linear mixed effects analysis, controlling for age, sex and haematocrit. It was observed that CYP3A5 expressors (*1/*1 + *1/*3) (p = 5.5 × 10 -16 ) and ABCB1 61G allele carriers (p = 0.001) had lower log-transformed TAC C 0 /D (56% and 26% lower geometric mean TAC C 0 /D, respectively) and accounted for approximately 30% and 4%, respectively, of log-transformed TAC C 0 /D variability in the first 3 months post-transplant. In conclusion, CYP3A5*3 is a major, and ABCB1 61A>G is a novel, although minor, genetic factor affecting TAC C 0 /D in kidney transplant recipients. © 2018 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).

Aspartame Administration and Insulin Treatment Altered Brain Levels of CYP2E1 and CYP3A2 in Streptozotocin-Induced Diabetic Rats.

Science.gov (United States)

Nosti-Palacios, Rosario; Gómez-Garduño, Josefina; Molina-Ortiz, Dora; Calzada-León, Raúl; Dorado-González, Víctor Manuel; Vences-Mejía, Araceli

2014-07-01

This study demonstrates that aspartame consumption and insulin treatment in a juvenile diabetic rat model leads to increase in cytochrome P450 (CYP) 2E1 and CYP3A2 isozymes in brain. Diabetes mellitus was induced in postweaned 21-day-old Wistar male rat by streptozotocin. Animals were randomly assigned to one of the following groups: untreated control, diabetic (D), D-insulin, D-aspartame, or the D-insulin + aspartame-treated group. Brain and liver tissue samples were used to analyze the activity of CYP2E1 and CYP3A2 and protein levels. Our results indicate that combined treatment with insulin and aspartame in juvenile diabetic rats significantly induced CYP2E1 in the cerebrum and cerebellum without modifying it in the liver, while CYP3A2 protein activity increased both in the brain and in the liver. The induction of CYP2E1 in the brain could have important in situ toxicological effects, given that this CYP isoform is capable of bioactivating various toxic substances. Additionally, CYP3A2 induction in the liver and brain could be considered a decisive factor in the variation of drug response and toxicity. © The Author(s) 2014.

Cloning and expression of SgCYP450-4 from Siraitia grosvenorii

Directory of Open Access Journals (Sweden)

Dongping Tu

2016-10-01

Full Text Available CYP450 plays an essential role in the development and growth of the fruits of Siraitia grosvenorii. However, little is known about the SgCYP450-4 gene in S. grosvenorii. Here, based on transcriptome data, a full-length cDNA sequence of SgCYP450-4 was cloned by reverse transcriptase-polymerase chain reaction (RT-PCR and rapid-amplification of cDNA ends (RACE strategies. SgCYP450-4 is 1677 bp in length (GenBank accession No. AEM42985.1 and contains a complete open reading frame (ORF of 1422 bp. The deduced protein was composed of 473 amino acids, the molecular weight is 54.01 kDa, the theoretical isoelectric point (PI is 8.8, and the protein was predicted to possess cytochrome P450 domains. SgCYP450-4 gene was highly expressed in root, diploid fruit and fruit treated with hormone and pollination. At 10 days after treatment with pollination and hormones, the expression of SgCYP450-4 had the highest level and then decreased over time, which was consistent with the development of fruits of S. Grosvenorii. Hormonal treatment could significantly induce the expression of SgCYP450-4. These results provide a reference for regulation of fruit development and the use of parthenocarpy to generate seedless fruit, and provide a scientific basis for the production of growth regulator application agents.

Effect of CYP3A perpetrators on ibrutinib exposure in healthy participants.

Science.gov (United States)

de Jong, Jan; Skee, Donna; Murphy, Joe; Sukbuntherng, Juthamas; Hellemans, Peter; Smit, Johan; de Vries, Ronald; Jiao, Juhui James; Snoeys, Jan; Mannaert, Erik

2015-08-01

Ibrutinib (PCI-32765), a potent covalent inhibitor of Bruton's tyrosine kinase, has shown efficacy against a variety of B-cell malignancies. Given the prominent role of CYP3A in ibrutinib metabolism, effect of coadministration of CYP3A perpetrators with ibrutinib was evaluated in healthy adults. Ibrutinib (120 mg [Study 1, fasted], 560 mg [studies 2 (fasted), and 3 (nonfasted)]) was given alone and with ketoconazole [Study 1; 400 mg q.d.], rifampin [Study 2; 600 mg q.d.], and grapefruit juice [GFJ, Study 3]. Lower doses of ibrutinib were used together with CYP3A inhibitors [Study 1: 40 mg; Study 3: 140 mg], as safety precaution. Under fasted condition, ketoconazole increased ibrutinib dose-normalized (DN) exposure [DN-AUClast: 24-fold; DN-C max: 29-fold], rifampin decreased ibrutinib exposure [C max: 13-fold; AUClast: 10-fold]. Under nonfasted condition, GFJ caused a moderate increase [DN-C max: 3.5-fold; DN-AUC: 2.2-fold], most likely through inhibition of intestinal CYP3A. Half-life was not affected by CYP perpetrators indicating the interaction was mainly on first-pass extraction. All treatments were well-tolerated.

Effects of CYP2C9*1/*3 genotype on the pharmacokinetics of flurbiprofen in Korean subjects.

Science.gov (United States)

Lee, Yun-Jeong; Byeon, Ji-Yeong; Kim, Young-Hoon; Kim, Se-Hyung; Choi, Chang-Ik; Bae, Jung-Woo; Sohn, Uy-Dong; Jang, Choon-Gon; Lee, Jeongmi; Lee, Seok-Yong

2015-06-01

The aim of this study was to investigate the impact of CYP2C9*1/*3 genotype on the pharmacokinetics of flurbiprofen and its metabolite. The CYP2C9 genotypes were determined with the use of polymerase chain reaction and restriction fragment and DNA sequencing analysis in 358 healthy Koreans. Among them, twenty individuals with CYP2C9*1/*1 (n = 12) or CYP2C9*1/*3 (n = 8) genotypes received a single 40 mg oral dose of flurbiprofen. The plasma concentrations of flurbiprofen and its metabolite, 4'-hydroxyflurbiprofen were measured by HPLC. AUCinf of flurbiprofen was significantly higher and its clearance was significantly lower in the CYP2C9*1/*3 individuals than in those with CYP2C9*1/*1. The AUC ratio of 4'-hydroxyflurbiprofen to flurbiprofen was significantly lower in the CYP2C9*1/*3 individuals than in those with CYP2C9*1/*1. These results indicate that the individuals carrying of CYP2C9*3 have significant reduction in flurbiprofen metabolism. The clinical use of this information may allow for more efficient personalized pharmacotherapy.

Genetic polymorphisms in MDR1 and CYP3A4 genes in Asians and the influence of MDR1 haplotypes on cyclosporin disposition in heart transplant recipients.

Science.gov (United States)

Chowbay, Balram; Cumaraswamy, Sivathasan; Cheung, Yin Bun; Zhou, Qingyu; Lee, Edmund J D

2003-02-01

Intestinal cytochrome P450 3A4 (CYP3A4) and P-glycoprotein (P-gp) both play a vital role in the metabolism of oral cyclosporine (CsA). We investigated the genetic polymorphisms in CYP3A4(promoter region and exons 5, 7 and 9) and MDR1 (exons 12, 21 and 26) genes and the impact of these polymorphisms on the pharmacokinetics of oral CsA in stable heart transplant patients (n = 14). CYP3A4 polymorphisms were rare in the Asian population and transplant patients. Haplotype analysis revealed 12 haplotypes in the Chinese, eight in the Malays and 10 in the Indians. T-T-T was the most common haplotype in all ethnic groups. The frequency of the homozygous mutant genotype at all three loci (TT-TT-TT) was highest in the Indians (31%) compared to 19% and 15% in the Chinese and Malays, respectively. In heart transplant patients, CsA exposure (AUC(0-4 h), AUC(0-12 h) and C(max)) was high in patients with the T-T-T haplotypes compared to those with C-G-C haplotypes. These findings suggest that haplotypes rather than genotypes influence CsA disposition in transplant patients.

Level of CYP4G19 Expression Is Associated with Pyrethroid Resistance in Blattella germanica

Directory of Open Access Journals (Sweden)

Guang-zhou Guo

2010-01-01

Full Text Available German cockroaches have become a large problem in the Shenzhen area because of their pesticide resistance, especially to pyrethroid. A pyrethroid called “Jia Chong Qing” to prevent pests for a long time were found to be resistant to “Jia Chong Qing” with resistance index of 3.88 measured using RT-PCR and immunohistochemistry analysis showed that both CYP4G19 mRNA and CYP4G19 protein expression levels in the wild strain were substantially higher than that of a sensitive strain. dsRNA segments derived from the target gene CYP4G19 were prepared using in vitro transcription and were microinjected into abdomens of the wild strain. Two to eight days after injection, the result showed that CYP4G19 mRNA expressions were significantly reduced in the groups injected with dsRNAs.

Impact of the CYP2C8 *3 polymorphism on the drug-drug interaction between gemfibrozil and pioglitazone.

Science.gov (United States)

Aquilante, Christina L; Kosmiski, Lisa A; Bourne, David W A; Bushman, Lane R; Daily, Elizabeth B; Hammond, Kyle P; Hopley, Charles W; Kadam, Rajendra S; Kanack, Alexander T; Kompella, Uday B; Le, Merry; Predhomme, Julie A; Rower, Joseph E; Sidhom, Maha S

2013-01-01

The objective of this study was to determine the extent to which the CYP2C8*3 allele influences pharmacokinetic variability in the drug-drug interaction between gemfibrozil (CYP2C8 inhibitor) and pioglitazone (CYP2C8 substrate). In this randomized, two phase crossover study, 30 healthy Caucasian subjects were enrolled based on CYP2C8*3 genotype (n = 15, CYP2C8*1/*1; n = 15, CYP2C8*3 carriers). Subjects received a single 15 mg dose of pioglitazone or gemfibrozil 600 mg every 12 h for 4 days with a single 15 mg dose of pioglitazone administered on the morning of day 3. A 48 h pharmacokinetic study followed each pioglitazone dose and the study phases were separated by a 14 day washout period. Gemfibrozil significantly increased mean pioglitazone AUC(0,∞) by 4.3-fold (P gemfibrozil administration was significantly influenced by CYP2C8 genotype. Specifically, CYP2C8*3 carriers had a 5.2-fold mean increase in pioglitazone AUC(0,∞) compared with a 3.3-fold mean increase in CYP2C8*1 homozygotes (P = 0.02). CYP2C8*3 is associated with decreased pioglitazone plasma exposure in vivo and significantly influences the pharmacokinetic magnitude of the gemfibrozil-pioglitazone drug-drug interaction. Additional studies are needed to evaluate the impact of CYP2C8 genetics on the pharmacokinetics of other CYP2C8-mediated drug-drug interactions. © 2012 The Authors. British Journal of Clinical Pharmacology © 2012 The British Pharmacological Society.

Impact of the CYP2C8 *3 polymorphism on the drug–drug interaction between gemfibrozil and pioglitazone

Science.gov (United States)

Aquilante, Christina L; Kosmiski, Lisa A; Bourne, David W A; Bushman, Lane R; Daily, Elizabeth B; Hammond, Kyle P; Hopley, Charles W; Kadam, Rajendra S; Kanack, Alexander T; Kompella, Uday B; Le, Merry; Predhomme, Julie A; Rower, Joseph E; Sidhom, Maha S

2013-01-01

AIM The objective of this study was to determine the extent to which the CYP2C8*3 allele influences pharmacokinetic variability in the drug–drug interaction between gemfibrozil (CYP2C8 inhibitor) and pioglitazone (CYP2C8 substrate). METHODS In this randomized, two phase crossover study, 30 healthy Caucasian subjects were enrolled based on CYP2C8*3 genotype (n = 15, CYP2C8*1/*1; n = 15, CYP2C8*3 carriers). Subjects received a single 15 mg dose of pioglitazone or gemfibrozil 600 mg every 12 h for 4 days with a single 15 mg dose of pioglitazone administered on the morning of day 3. A 48 h pharmacokinetic study followed each pioglitazone dose and the study phases were separated by a 14 day washout period. RESULTS Gemfibrozil significantly increased mean pioglitazone AUC(0,∞) by 4.3-fold (P gemfibrozil administration was significantly influenced by CYP2C8 genotype. Specifically, CYP2C8*3 carriers had a 5.2-fold mean increase in pioglitazone AUC(0,∞) compared with a 3.3-fold mean increase in CYP2C8*1 homozygotes (P= 0.02). CONCLUSION CYP2C8*3 is associated with decreased pioglitazone plasma exposure in vivo and significantly influences the pharmacokinetic magnitude of the gemfibrozil–pioglitazone drug-drug interaction. Additional studies are needed to evaluate the impact of CYP2C8 genetics on the pharmacokinetics of other CYP2C8-mediated drug–drug interactions. PMID:22625877

CHRNA3 and CYP3A5*3 genotype, lung function and chronic obstructive pulmonary disease in the general population

DEFF Research Database (Denmark)

Kaur-Knudsen, Diljit; Bojesen, Stig E; Nordestgaard, Børge G

2014-01-01

OBJECTIVE: Genetic variations are most likely an additional risk factor besides tobacco smoking per se for the risk of chronic obstructive pulmonary disease (COPD). In this study, we compared genetic variants influencing the effect of smoking on COPD, that is, the effect of the well-known splicin.......1-2.2) for GOLD 3-4. This association could not be found in never-smokers. No association was found for CYP3A5*3. CONCLUSION: The CHRNA3 genotype is associated with decreased lung function and risk of COPD among ever-smokers, whereas this was not the case for CYP3A5*3....... were genotyped. Information on spirometry, hospital admissions and smoking behaviour was recorded. Endpoints were lung function and COPD. RESULTS: For CHRNA3, the percentage of forced expiratory volume in 1 s (FEV1%) predicted was 89.3, 90.6 and 92.4% in homozygous, heterozygous and noncarrier ever-smokers...... (P-trendsmokers was 1.5 [95...

Association of CYP3A polymorphisms with the pharmacokinetics of cyclosporine A in early post-renal transplant recipients in China

Institute of Scientific and Technical Information of China (English)

Xiang-guang MENG; Ji-ye YIN; Xiang-dong PENG; Qi PEI; Wei ZHANG; Guo WANG; Meng HE; Min LIU; Jing-ke YANG; Hong-hao ZHOU; Cheng-xian GUO; Guo-qing FENG; Ying-chun ZHAO; Bo-ting ZHOU; Jian-le HAN; Xin CHEN; Yong SHI; Hong-yao SHI

2012-01-01

Aim: To evaluate retrospectively the association of cytochrome P450 3A (CYP3A) and ATP-binding cassette sub-family B member 1(ABCB1) gene polymorphisms with the pharmacokinetics of cyclosporine A (CsA) in Chinese renal transplant patients.Methods: One hundred and twenty-six renal transplant patients were recruited.Blood samples were collected,and corresponding clinical indices were recorded on the seventh day after the procedure.The patients were genotyped for CYP3A4*1G,CYP3A5*3C,ABCB1 1236 C＞T,ABCB1 2677 G＞T/A,and ABCB1 3435 C＞T polymorphisms.Whole blood trough concentrations of CsA at time zero (Co)were measured before the drug administration.A multiple regression model was developed to analyze the effects of genetic factors on the CsA dose-adjusted Co (Co/dose) based on several clinical indices.Results: The CYP3A5*3C polymorphism influenced the Co and Co/dose of CsA,which were significantly higher in patients with the GG genotype than in patients with the AA or GA genotypes.No significant differences were detected for other SNPs (CYP3A4*1G,ABCB1 1236 C＞T,ABCB1 2677 G＞T/A,and ABCB1 3435 C＞T).In a univariate analysis using Pearson's correlation test,age,hemoglobin,blood urea nitrogen and blood creatinine levels were significantly correlated with the log-transformed CsA Co/dose.In the multiple regression model,CYP3A5*3C,age,hemoglobin and blood creatinine level were associated with the log-transformed CsA Co/dose.Conclusion: CYP3A5*3C correlates with the Co/dose of CsA on the seventh day after renal transplantation.The allele is a putative indicator for the optimal CsA dosage in the early phase of renal transplantation in the Chinese population.

Efficacy of piroxicam for postoperative pain after lower third molar surgery associated with CYP2C8*3 and CYP2C9

Directory of Open Access Journals (Sweden)

Calvo AM

2017-07-01

Full Text Available Adriana Maria Calvo, Paulo Zupelari-Gonçalves, Thiago José Dionísio, Daniel Thomas Brozoski, Flávio Augusto Faria, Carlos Ferreira Santos Department of Biological Sciences, Bauru School of Dentistry, University of São Paulo, São Paulo, Brazil Objective: Nonsteroidal anti-inflammatory drugs (NSAIDs are metabolized by the cytochrome P450 enzymes (CYPs, predominantly CYP2C8 and CYP2C9. The aim of this study was to evaluate the possible association of polymorphisms in the CYP2C8*3 and CYP2C9 genes with the clinical efficacy of oral piroxicam (20 mg daily for 4 days after lower third molar surgeries with regard to postoperative pain, swelling, trismus, adverse reactions, need for rescue medication and the volunteer’s overall satisfaction. Materials and methods: For this purpose, 102 volunteers were genotyped for CYP2C8*3 and CYP2C9 polymorphisms. Briefly, genomic DNA was isolated from saliva collected from volunteers subjected to invasive lower third molar surgeries, and the preoperative, intraoperative and postoperative parameters were collected and analyzed. Results: An equal amount of piroxicam sufficiently managed postoperative pain and inflammatory symptoms, with visual analog pain scores typically <40 mm for all genotypes investigated. Furthermore, only two out of 102 volunteers heterozygous for CYP2C8*3 and CYP2C9*3 reported adverse side effects. Conclusion: In general, slow metabolizers of piroxicam, who were volunteers with mutant alleles, were indifferent from normal metabolizers with the wild-type alleles and therefore did not require specialized piroxicam doses to manage postoperative pain and inflammation. Keywords: piroxicam, lower third molar surgery, P450, CYP2C8, CYP2C9, pharmacogenetics 

High allele frequency of CYP2C9*3 (rs1057910) in a Negrito's subtribe population in Malaysia; Aboriginal people of Jahai.

Science.gov (United States)

Rosdi, Rasmaizatul Akma; Mohd Yusoff, Narazah; Ismail, Rusli; Soo Choon, Tan; Saleem, Mohamed; Musa, Nurfadhlina; Yusoff, Surini

2016-09-01

CYP2C9 gene polymorphisms modulate inter-individual variations in the human body's responses to various endogenous and exogenous drug substrates. To date, little is known about the CYP2C9 gene polymorphisms among the aboriginal populations of the world, including those in Malaysia. To characterise and compare the CYP2C9 polymorphisms (CYP2C9*2, CYP2C9*3, CYP2C9*4 and CYP2C9*5) between one of Malaysia's aboriginal populations, Jahai, with the national major ethnic, Malay. To also compare the allele frequencies from these two populations with available data of other aboriginal populations around the world. The extracted DNA of 155 Jahais and 183 Malays was genotyped for CYP2C9 polymorphisms using a nested multiplex allele-specific polymerase chain reaction technique. The results were confirmed by DNA direct sequencing. Genotyping results revealed that CYP2C9*2, CYP2C9*4 and CYP2C9*5 were absent in Jahais, while only the latter two were absent in Malays. The CYP2C9*3 allelic frequency in Jahais was 36.2%, making them the most frequent carriers of the allele thus far reported in any ethnic group from Southeast Asia. The high frequency of CYP2C9*3 and the absence of CYP2C9*2 in Jahais suggest that genetic drift may be occurring in this ethnic group. This is the first study to determine the CYP2C9 polymorphisms in an aboriginal population in Malaysia.

A Novel Polymorphism in the Promoter of the CYP4A11 Gene Is Associated with Susceptibility to Coronary Artery Disease

Science.gov (United States)

Sirotina, Svetlana; Ponomarenko, Irina; Kharchenko, Alexander; Bykanova, Marina; Bocharova, Anna; Vagaytseva, Kseniya; Solodilova, Maria

2018-01-01

Enzymes CYP4A11 and CYP4F2 are involved in biosynthesis of vasoactive 20-hydroxyeicosatetraenoic acid and may contribute to pathogenesis of coronary artery disease (CAD). We investigated whether polymorphisms of the CYP4A11 and CYP4F2 genes are associated with the risk of CAD in Russian population. DNA samples from 1323 unrelated subjects (637 angiographically confirmed CAD patients and 686 age- and sex-matched healthy individuals) were genotyped for polymorphisms rs3890011, rs9332978, and rs9333029 of CYP4A11 and rs3093098 and rs1558139 of CYP4F2 by using the Mass-ARRAY 4 system. SNPs rs3890011 and rs9332978 of CYP4A11 were associated with increased risk of CAD in women: OR = 1.26, 95% CI: 1.02–1.57, P = 0.004, and Q = 0.01 and OR = 1.45, 95% CI: 1.13–1.87, P = 0.004, and Q = 0.01, respectively. Haplotype G-C-A of CYP4A11 was associated with increased risk of CAD (adjusted OR = 1.41, 95% CI: 1.12–1.78, and P = 0.0036). Epistatic interactions were found between rs9332978 of CYP4A11 and rs1558139 of CYP4F2 (P interaction = 0.025). In silico analysis allowed identifying that SNP rs9332978 is located at a binding site for multiple transcription factors; many of them are known to regulate the pathways involved in the pathogenesis of CAD. This is the first study in Europeans that reported association between polymorphism rs9332978 of CYP4A11 and susceptibility to coronary artery disease. PMID:29484037

QSAR development and profiling of 72,524 REACH substances for PXR activation and CYP3A4 induction

DEFF Research Database (Denmark)

Abildgaard Rosenberg, Sine; Xia, M.; Huang, R.

2017-01-01

,524 substances pre-registered under the EU chemicals regulation, REACH, and the models could predict 52.5% to 71.9% of the substances within their respective applicability domains. These predictions can, for example, be used for priority setting and in weight-of-evidence assessments of chemicals. Statistical...... analyses of the experimental drug dataset and the QSAR-predicted set of REACH substances were performed to identify similarities and differences in frequencies of overlapping positive results for PXR binding, PXR activation and CYP3A4 induction between the two datasets....

The enhanced atorvastatin hepatotoxicity in diabetic rats was partly attributed to the upregulated hepatic Cyp3a and SLCO1B1

Science.gov (United States)

Shu, Nan; Hu, Mengyue; Ling, Zhaoli; Liu, Peihua; Wang, Fan; Xu, Ping; Zhong, Zeyu; Sun, Binbin; Zhang, Mian; Li, Feng; Xie, Qiushi; Liu, Xiaodong; Liu, Li

2016-01-01

Liver injury is a common adverse effect of atorvastatin. This study aimed to investigate atorvastatin-induced hepatotoxicity in diabetic rats induced by high-fat diet combined with streptozotocin. The results showed that 40 mg/kg atorvastatin was lethal to diabetic rats, whose mean survival time was 6.2 days. Severe liver injury also occurred in diabetic rats treated with 10 mg/kg and 20 mg/kg atorvastatin. The in vitro results indicated that atorvastatin cytotoxicity in hepatocytes of diabetic rats was more severe than normal and high-fat diet feeding rats. Expressions and activities of hepatic Cyp3a and SLCO1B1 were increased in diabetic rats, which were highly correlated with hepatotoxicity. Antioxidants (glutathione and N-Acetylcysteine), Cyp3a inhibitor ketoconazole and SLCO1B1 inhibitor gemfibrozil suppressed cytotoxicity and ROS formation in primary hepatocytes of diabetic rats. In HepG2 cells, up-regulations of CYP3A4 and SLCO1B1 potentiated hepatotoxicity and ROS generation, whereas knockdowns of CYP3A4 and SLCO1B1 as well as CYP3A4/SLCO1B1 inhibitions showed the opposite effects. Phenobarbital pretreatment was used to induce hepatic Cyp3a and SLCO1B1 in rats. Phenobarbital aggravated atorvastatin-induced hepatotoxicity, while decreased plasma exposure of atorvastatin. All these findings demonstrated that the upregulations of hepatic Cyp3a and SLCO1B1 in diabetic rats potentiated atorvastatin-induced hepatotoxicity via increasing ROS formation. PMID:27624558

A study on CYP1A inhibitory action of E-2-(4'-methoxybenzylidene)-1-benzosuberone and some related chalcones and cyclic chalcone analogues