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Tsutomu Shimada - One of the best experts on this subject based on the ideXlab platform.
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comparative studies of in vitro inhibition of cytochrome p450 3a4 dependent testosterone 6β hydroxylation by roxithromycin and its metabolites Troleandomycin and erythromycin
Drug Metabolism and Disposition, 1998Co-Authors: Hiroshi Yamazaki, Tsutomu ShimadaAbstract:Roxithromycin has been shown to be a relatively weak inhibitor of cytochrome P450 (P450 or CYP)-dependent drug oxidations, compared with Troleandomycin. The potential for roxithromycin and its major metabolites found in human urine [namely the decladinosyl derivative (M1), O-dealkyl derivative (M2), and N-demethyl derivative (M3)] to inhibit testosterone 6beta-hydroxylation after metabolic activation by CYP3A4 was examined and compared with inhibition by Troleandomycin and erythromycin in vitro. Of roxithromycin and its studied metabolites, M3 was the most potent in inhibiting CYP3A4-dependent testosterone 6beta-hydroxylation by human liver microsomes and was activated to the inhibitory P450.Fe2+-metabolite complex to the greatest extent. Roxithromycin and its metabolites were N-demethylated by human liver microsomes, although the rates were slower than those measured with Troleandomycin and erythromycin as substrates. Recombinant human CYP3A4 in a baculovirus system coexpressing NADPH-P450 reductase was very active in catalyzing the N-demethylation of roxithromycin, M1, and M2, as well as Troleandomycin, erythromycin, and M3. The order for inhibition of CYP3A4-dependent testosterone 6beta-hydroxylation activities by these macrolide antibiotics in the recombinant CYP3A4 system was estimated to be Troleandomycin > erythromycin >/= M3 >/= M2 > M1 >/= roxithromycin. Erythromycin, roxithromycin, and its metabolites all failed to inhibit CYP1A2-dependent (R)-warfarin 7-hydroxylation and CYP2C9-dependent (S)-warfarin 7-hydroxylation but did inhibit CYP3A4-dependent (R)-warfarin 7-hydroxylation. These results suggest that roxithromycin itself is not as potent an inhibitor of CYP3A4 activities as are Troleandomycin and erythromycin, probably because of the slower metabolism of this compound to metabolites M1, M2, and M3 in humans.
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Formation in vitro of an inhibitory cytochrome P450·Fe2+-metabolite complex with roxithromycin and its decladinosyl, O-dealkyl and N-demethyl metabolites in rat liver microsomes
Xenobiotica; the fate of foreign compounds in biological systems, 1998Co-Authors: Hiroshi Yamazaki, Tsutomu ShimadaAbstract:1. Roxithromycin and its major metabolites found in rat and human urine, namely the decladinosyl derivative (M1), O-dealkyl derivative (M2) and N-demethyl derivative (M3), were incubated with rat liver microsomes and formation of an inhibitory cytochrome P450 (CYP)-metabolite complex and of formaldehyde (measurement of N-demethylation) were determined in vitro. Troleandomycin and erythromycin were also used for comparison. 2. Dexamethasone very significantly induced the microsomal N-demethylations of these macrolide antibiotics. The order of magnitude for the Vmax/Km ratio of N-demethylations by liver microsomes from dexamethasone-treated rats was Troleandomycin > erythromycin = M2 > roxithromycin > M3, M1. 3. Formation of an inhibitory P450 x Fe2+-metabolite complex was detected on incubation of these macrolide antibiotics with rat liver microsomes in the presence of an NADPH-generating system and the order of maximum complex formation was Troleandomycin > erythromycin > M2 > roxithromycin > M3 > M1. 4. Troleandomycin, erythromycin and M2 inhibited CYP3A-dependent testosterone 6beta-hydroxylation catalysed by liver microsomes from the dexamethasone-treated rat by 54, 33 and 23%, respectively, but roxithromycin, M3 and M1 were very weak by comparison. In the untreated rat, only testosterone 6beta-hydroxylation, but not testosterone 16alpha- and 2alpha-hydroxylation and androstenedione formation, activities were inhibited, indicating that inhibitory actions of these antibiotics are specific for CYP3A enzymes in liver microsomes. 5. These results support the view that formation of an inhibitory P450-metabolite complex is prerequisite for the inhibition of CYP3A-dependent substrate oxidations by rat liver microsomes and that M2 (and M3, to a lesser extent) may be the active metabolite that can form an inhibitory P450-metabolite complex by CYP3A enzyme(s).
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Effects of roxithromycin, erythromycin and Troleandomycin on their N-demethylation by rat and human cytochrome P450 enzymes
Xenobiotica; the fate of foreign compounds in biological systems, 1996Co-Authors: Hiroshi Yamazaki, S. Hiroki, T. Urano, K. Inoue, Tsutomu ShimadaAbstract:1. The effects of treatment of rat with roxithromycin, erythromycin and Troleandomycin as well as other chemicals including typical cytochrome P450 inducers were examined in rat and human liver microsomes.2. Erythromycin and Troleandomycin but not roxithromycin caused slight increases in CYP3A1 levels and the N-demethylation of roxithromycin, erythromycin and Troleandomycin and oxidation of nifedipine in rat, but none of these chemicals induced significantly CYP2B1 levels or benzphetamine N-demethylation activities.3. Dexamethasone and pregnenolone 16α-carbonitrile induced CYP3A1 levels and N-demethylation of roxithromycin, erythromycin and Troleandomycin but not of benzphetamine, in rat liver microsomes. Treatment of rat with phenobarbital caused increases in both CYP2B1 and 3A1 levels and all of the N-demethylation activities examined. Phenytoin and metyrapone produced increases in contents of 2B1 and activities of benzphetamine N-demethylation as well as of roxithromycin, erythromycin and troleandomyci...
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Effects of erythromycin and roxithromycin on oxidation of testosterone and nifedipine catalyzed by CYP3A4 in human liver microsomes.
The Journal of toxicological sciences, 1996Co-Authors: Hiroshi Yamazaki, Tomoko Urano, Shigeru Hiroki, Tsutomu ShimadaAbstract:Roxithromycin and erythromycin were incubated with rat and human liver microsomal or reconstituted cytochrome P450 (P450 or CYP) monooxygenase systems in the presence of an NADPH-generating system, and the effects of these chemicals on testosterone 6 beta-hydroxylation and nifedipine oxidation activities were compared with those of typical CYP3A4 inhibitors including ketoconazole, Troleandomycin, and gestodene. Roxithromycin and erythromycin were found to be relatively weak inhibitors of testosterone 6 beta-hydroxylation and nifedipine oxidation activities by rat and human liver microsomes or by reconstituted systems containing CYP3A4/5. Formation of an inhibitory P450-metabolite complex was determined spectrally by incubating Troleandomycin with human liver microsomes; the extents of the complex formation were lesser in liver microsomes of humans than those of rats treated with dexamethasone. Erythromycin and roxithromycin were also activated slightly by rat liver microsomes to form P450.Fe(II)-metabolite complex, although these chemicals caused very little or undetectable levels, respectively, of spectral changes by human liver microsomes even when a human sample which contained relatively high levels of CYP3A4 was used. These results suggested that roxithromycin and erythromycin were relatively less potent to inhibit CYP3A4-catalytic activities in human liver microsomes, because of their low capabilities to form P450.Fe(II)-metabolite complex.
Evan D. Kharasch - One of the best experts on this subject based on the ideXlab platform.
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role of cyp2b6 in stereoselective human methadone metabolism
Anesthesiology, 2008Co-Authors: Rheem A. Totah, Pamela Sheffels, Toni Roberts, Kenneth E. Thummel, Dale Whittington, Evan D. KharaschAbstract:Background: Metabolism and clearance of racemic methadone are stereoselective and highly variable, yet the mechanism remains largely unknown. Initial in vitro studies attributed methadone metabolism to cytochrome P4503A4 (CYP3A4). CYP3A4 was also assumed responsible for methadone clearance in vivo. Nevertheless, recent clinical data do not support a primary role for CYP3A4 and suggest that CYP2B6 may mediate methadone clearance. Expressed CYP2B6 and also CYP2C19 N-demethylate methadone in vitro. This investigation tested the hypothesis that CYPs 2B6, 3A4, and/or 2C19 are responsible for stereoselective methadone metabolism in human liver microsomes and in vivo. Methods: N-demethylation of racemic methadone and individual enantiomers by expressed CYPs 2B6, 2C19, and 3A4 was evaluated. Stereoselective microsomal methadone metabolism was quantified, compared with CYP 2B6 and 3A4 content, and probed using CYP isoform-selective inhibitors. A crossover clinical investigation (control, CYP2B6 and CYP3A4 induction by rifampin, CYP3A inhibition by Troleandomycin and grapefruit juice) evaluated stereoselective methadone disposition. Results: At clinical concentrations, methadone enantiomer N-demethylation by recombinant CYPs 2B6, 3A4, and 2C19 was S > R, S = R, and S R) occurred in livers expressing high levels of CYP2B6 compared with CYP3A4. Clopidogrel, Troleandomycin, and (+)-N-3-benzyl-nirvanol, selective inhibitors of CYPs 2B6, 3A4, and 2C19, respectively, inhibited microsomal methadone metabolism by 50-60%, 20-30%, and less than 10%. Only Inhibition by clopidogrel was stereoselective. Clinically, rifampin diminished both R- and S-methadone plasma concentrations, but Troleandomycin and grapefruit juice altered neither R- nor S-methadone concentrations. Plasma R/S-methadone ratios were increased by rifampin but unchanged by CYP3A Inhibition. Conclusions: These results suggest a significant role for CYP2B6, but not CYP3A, in stereoselective human methadone metabolism and disposition.
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Metabolism of alfentanil by cytochrome P4503A (CYP3A) enzymes
Drug metabolism and disposition: the biological fate of chemicals, 2004Co-Authors: Theresa Mariero Klees, Pamela Sheffels, Ola Dale, Evan D. KharaschAbstract:The synthetic opioid alfentanil is an analgesic and an in vivo probe for hepatic and first-pass CYP3A activity. Alfentanil is a particularly useful CYP3A probe because pupil diameter change is a surrogate for plasma concentrations, thereby affording noninvasive assessment of CYP3A. Alfentanil undergoes extensive CYP3A4 metabolism via two major pathways, forming noralfentanil and N-phenylpropionamide. This investigation evaluated alfentanil metabolism in vitro to noralfentanil and N-phenylpropionamide, by expressed CYP3A5 and CYP3A7 in addition to CYP3A4, with and without coexpressed or exogenous cytochrome b5. Effects of the CYP3A inhibitors Troleandomycin and ketoconazole were also determined. Rates of noralfentanil and N-phenylpropionamide formation by CYP3A4 and 3A5 in the absence of b5 were generally equivalent, although the metabolite formation ratio differed, whereas those by CYP3A7 were substantially less. CYP3A4 and 3A5 were equipotently inhibited by Troleandomycin, whereas ketoconazole was an order of magnitude more potent toward CYP3A4. Cytochrome b5 qualitatively and quantitatively altered alfentanil metabolism, with b5 coexpression having a greater effect than exogenous addition. Addition or coexpression of b5 markedly stimulated the formation of both metabolites and changed the formation of noralfentanil but not N-phenylpropionamide from apparent single-site to multisite Michaelis-Menten kinetics. These results demonstrate that alfentanil is a substrate for CYP3A5 in addition to CYP3A4, and the effects of the CYP3A inhibitors Troleandomycin and ketoconazole are CYP3A enzyme-selective. Alfentanil is one of the few CYP3A substrates that is metabolized in vitro as avidly by both CYP3A4 and 3A5. Polymorphic CYP3A5 expression may contribute to inter-individual variability in alfentanil metabolism.
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influence of hepatic and intestinal cytochrome p4503a activity on the acute disposition and effects of oral transmucosal fentanyl citrate
Anesthesiology, 2004Co-Authors: Evan D. Kharasch, Dale Whittington, Christine HofferAbstract:BACKGROUND Oral transmucosal fentanyl citrate (OTF) was developed to provide rapid analgesia and is specifically approved for treating breakthrough cancer pain. Fentanyl in OTF is absorbed across the oral mucosa, but a considerable portion is swallowed and absorbed enterally. Fentanyl metabolism is catalyzed by cytochrome P4503A4 (CYP3A). The role of intestinal or hepatic first-pass metabolism and CYP3A activity in OTF disposition is unknown. This investigation examined the influence of hepatic and intestinal CYP3A activity on the disposition and clinical effects of OTF. METHODS Healthy volunteers (n = 12) were studied in an Institutional Review Board-approved, randomized, balanced, four-way crossover. They received OTF (10 microg/kg) after hepatic/intestinal CYP3A induction by rifampin, hepatic/intestinal CYP3A inhibition by Troleandomycin, selective intestinal CYP3A inhibition by grapefruit juice, or nothing (control). Plasma fentanyl and norfentanyl concentrations were determined by mass spectrometry. Fentanyl effects were measured by dark-adapted pupil diameter and subjective self-assessments using visual analog scales. RESULTS : Peak plasma fentanyl concentrations, time to peak, and maximum pupil diameter change from baseline were unchanged after rifampin, Troleandomycin, and grapefruit juice. Fentanyl elimination, however, was significantly affected by CYP3A alterations. After control, rifampin, Troleandomycin and grapefruit juice, respectively, area under the curve of plasma fentanyl versus time was 5.9 +/- 3.7, 2.2 +/- 0.8,* 10.4 +/- 8.9,* and 5.8 +/- 3.3 h x ng/ml; norfentanyl/fentanyl plasma area under the curve ratios were 0.92 +/- 0.63, 3.2 +/- 1.8,* 0.08 +/- 0.14,* and 0.67 +/- 0.33 (*P < 0.05 versus control). DISCUSSION Peak fentanyl concentrations and clinical effects after OTF were minimally affected by altering both intestinal and hepatic CYP3A activity, whereas fentanyl metabolism, elimination, and duration of effects were significantly affected; selective intestinal CYP3A inhibition had minimal effects. This suggests that first-pass metabolism minimally influences OTF bioavailability. When treating breakthrough pain, with careful attention to maximal mucosal absorption and minimal swallowing, CYP3A variability and drug interactions are unlikely to affect the onset or magnitude of OTF analgesia; however, duration may be affected.
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simultaneous assessment of drug interactions with low and high extraction opioids application to parecoxib effects on the pharmacokinetics and pharmacodynamics of fentanyl and alfentanil
Anesthesiology, 2003Co-Authors: Andra E Ibrahim, Jennifer Feldman, Aziz Karim, Evan D. KharaschAbstract:BACKGROUND Parecoxib is a parenteral cyclooxygenase-2 (COX-2) inhibitor intended for perioperative analgesia. It is an inactive prodrug hydrolyzed in vivo to the active inhibitor valdecoxib, a substrate for hepatic cytochrome P450 3A4 (CYP3A4); hence, a potential exists for metabolic interactions with other CYP3A substrates. This study determined the effects of parecoxib on the pharmacokinetics and pharmacodynamics of the CYP3A substrates fentanyl and alfentanil compared with the CYP3A inhibitor Troleandomycin. Alfentanil is a low-extraction drug with a clearance that is highly susceptible to drug interactions; fentanyl is a high-extraction drug and, thus, is theoretically less vulnerable. We therefore also tested the hypothesis that the extraction ratio influences the consequence of altered hepatic metabolism of these opioids. METHODS After Institutional Review Board-approved, written, informed consent was obtained, 12 22- to 40-yr-old healthy volunteers were enrolled in the study. The protocol was a randomized, double-blinded, balanced, placebo-controlled, three-session (placebo, parecoxib, or Troleandomycin pretreatment) crossover. Subjects received both alfentanil (15 microg/kg) and fentanyl (5 microg/kg; 15-min intravenous infusion) 1 h after placebo, parecoxib (40 mg intravenously every 12 h), or Troleandomycin (every 6 h). Study sessions were separated by 7 or more days. Opioid concentrations in venous blood were determined by liquid chromatography-mass spectrometry. Pharmacokinetic parameters were determined by noncompartmental analysis. Opioid effects were determined by pupillometry, respiratory rate, and Visual Analog Scale scores. RESULTS There were no significant differences between the placebo and parecoxib treatments in alfentanil or fentanyl plasma concentration, maximum observed plasma concentration, area under the plasma time-concentration time curve, clearance, elimination half-life, or volume of distribution. However, disposition of alfentanil, and to a lesser extent fentanyl, was significantly altered by Troleandomycin. Clearances were reduced to 12% (0.64 +/- 0.25 ml. kg-1. min-1) and 61% (9.35 +/- 3.07) of control (5.53 +/- 2.16 and 15.3 +/- 5.0) for alfentanil and fentanyl (P < 0.001). Pupil diameter versus time curves were similar between placebo and parecoxib treatments but were significantly different after Troleandomycin. CONCLUSIONS Single-dose parecoxib does not alter fentanyl or alfentanil disposition or clinical effects and does not appear to cause significant CYP3A drug interactions. CYP3A inhibition decreases alfentanil clearance more than fentanyl clearance, confirming that the extraction ratio influences the consequence of altered hepatic drug metabolism. Modified cassette, or "cocktail," dosing is useful for assessing drug interactions in humans.
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the role of cytochrome p450 3a4 in alfentanil clearance implications for interindividual variability in disposition and perioperative drug interactions
Anesthesiology, 1997Co-Authors: Evan D. Kharasch, Kenneth E. Thummel, Michael Russell, Douglas S Mautz, Kent L Kunze, Andrew T Bowdle, Kathy CoxAbstract:BACKGROUND There is considerable unexplained variability in alfentanil pharmacokinetics, particularly systemic clearance. Alfentanil is extensively metabolized in vivo, and thus systemic clearance depends on hepatic biotransformation. Cytochrome P450 3A4 was previously shown to be the predominant P450 isoform responsible for human liver microsomal alfentanil metabolism in vitro. This investigation tested the hypothesis that P450 3A4 is responsible for human alfentanil metabolism and clearance in vivo. METHODS Nine healthy male volunteers who provided institutionally approved written informed consent were studied in a three-way randomized crossover design. Each subject received alfentanil (20 micrograms/kg given intravenously) 30 min after midazolam (1 mg injected intravenously) on three occasions: control; high P450 3A4 activity (rifampin induction); and low P450 3A4 activity (selective inhibition by Troleandomycin). Midazolam is a validated selective in vivo probe for P450 3A4 activity. Venous blood was sampled for 24 h and plasma concentrations of midazolam and alfentanil and their primary metabolites 1'-hydroxymidazolam and noralfentanil were measured by gas chromatography-mass spectrometry. Pharmacokinetic parameters were determined by two-stage analysis using both noncompartmental and three-compartment models. RESULTS Plasma alfentanil concentration-time profiles depended significantly on P450 3A4 activity. Alfentanil noncompartmental clearance was 5.3 +/- 2.3, 14.6 +/- 3.8, and 1.1 +/- 0.5 ml.kg-1.min-1, and elimination half-life was 58 +/- 13, 35 +/- 7, and 630 +/- 374 min, respectively, in participants with normal (controls), high (rifampin), and low (Troleandomycin) P450 3A4 activity (means +/- SD; P < 0.05 compared with controls). Multicompartmental modeling suggested a time-dependent inhibition-resynthesis model for Troleandomycin effects on P450 3A4 activity, characterized as k10(t) = k10[1-phi e-alpha(t-tzero)], where k10(t) is the apparent time-dependent rate constant, k10 is the uninhibited rate constant, phi is the fraction of P450 3A4 inhibited, and alpha is the apparent P450 3A4 reactivation rate. Alfentanil clearance was calculated as V1 k10 for controls and men receiving rifampin, and as V1.average k10(t) for men receiving Troleandomycin. This clearance was 4.9 +/- 2.1, 13.2 +/- 3.6, and 1.5 +/- 0.8 ml.kg-1.min-1, respectively, in controls and in men receiving rifampin or Troleandomycin. There was a significant correlation (r = 0.97, P < 0.001) between alfentanil systemic clearance and P450 3A4 activity. CONCLUSIONS Modulation of P450 3A4 activity by rifampin and Troleandomycin significantly altered alfentanil clearance and disposition. These results strongly suggest that P450 3A4 is the major isoform of P450 responsible for clinical alfentanil metabolism and clearance. This observation, combined with the known population variability in P450 3A4 activity, provides a mechanistic explanation for the interindividual variability in alfentanil disposition. Furthermore, known susceptibility of human P450 3A4 activity to induction and inhibition provides a conceptual framework for understanding and predicting clinical alfentanil drug interactions. Finally, human liver microsomal alfentanil metabolism in vitro is confirmed as an excellent model for human alfentanil metabolism in vivo.
M E Veronese - One of the best experts on this subject based on the ideXlab platform.
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in vitro proguanil activation to cycloguanil by human liver microsomes is mediated by cyp3a isoforms as well as by s mephenytoin hydroxylase
British Journal of Clinical Pharmacology, 1994Co-Authors: Donald J. Birkett, F.j. Gonzalez, John O Miners, Tommy B Andersson, D. Rees, M E VeroneseAbstract:1. The activation of proguanil to cycloguanil by human liver microsomes was studied to define the cytochrome P450 (CYP) isoforms involved in this reaction. 2. Apparent Km values for proguanil ranged from 35 microM to 183 microM with microsomes from four human livers. 3. There was a 6.3-fold range of activity with microsomes from seventeen human livers. Rates of proguanil activation correlated significantly with CYP3A activities (benzo[a]pyrene metabolism, caffeine 8-oxidation and omeprazole sulphone formation) and CYP3A immunoreactive content. There was also a highly significant correlation with rates of hydroxyomeprazole formation. Correlations with activities selective for CYP1A2, CYP2C9/10 and CYP2E1, and with immunoreactive CYP1A2 content were not significant. 4. Proguanil activation was inhibited by R,S-mephenytoin, Troleandomycin and by inhibitory anti-CYP3A antiserum and anti-CYP2C IgG and was activated by alpha-naphthoflavone. Inhibitors selective for CYP1A2, CYP2E1, CYP2A6 or CYP2C9/10 had little or no effect on proguanil activation. The extents of inhibition by R,S-mephenytoin, Troleandomycin and the two antibodies varied with the immunoreactive CYP3A content of the microsomes used. 5. It is concluded that proguanil activation to cycloguanil by human liver microsomes is mediated both by S-mephenytoin hydroxylase and isoforms of the CYP3A subfamily. This has implications for the use of proguanil as an in vivo probe for the S-mephenytoin poor metaboliser phenotype.
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In vitro proguanil activation to cycloguanil by human liver microsomes is mediated by CYP3A isoforms as well as by S‐mephenytoin hydroxylase.
British Journal of Clinical Pharmacology, 1994Co-Authors: Donald J. Birkett, F.j. Gonzalez, John O Miners, Tommy B Andersson, D. Rees, M E VeroneseAbstract:1. The activation of proguanil to cycloguanil by human liver microsomes was studied to define the cytochrome P450 (CYP) isoforms involved in this reaction. 2. Apparent Km values for proguanil ranged from 35 microM to 183 microM with microsomes from four human livers. 3. There was a 6.3-fold range of activity with microsomes from seventeen human livers. Rates of proguanil activation correlated significantly with CYP3A activities (benzo[a]pyrene metabolism, caffeine 8-oxidation and omeprazole sulphone formation) and CYP3A immunoreactive content. There was also a highly significant correlation with rates of hydroxyomeprazole formation. Correlations with activities selective for CYP1A2, CYP2C9/10 and CYP2E1, and with immunoreactive CYP1A2 content were not significant. 4. Proguanil activation was inhibited by R,S-mephenytoin, Troleandomycin and by inhibitory anti-CYP3A antiserum and anti-CYP2C IgG and was activated by alpha-naphthoflavone. Inhibitors selective for CYP1A2, CYP2E1, CYP2A6 or CYP2C9/10 had little or no effect on proguanil activation. The extents of inhibition by R,S-mephenytoin, Troleandomycin and the two antibodies varied with the immunoreactive CYP3A content of the microsomes used. 5. It is concluded that proguanil activation to cycloguanil by human liver microsomes is mediated both by S-mephenytoin hydroxylase and isoforms of the CYP3A subfamily. This has implications for the use of proguanil as an in vivo probe for the S-mephenytoin poor metaboliser phenotype.
Hiroshi Yamazaki - One of the best experts on this subject based on the ideXlab platform.
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comparative studies of in vitro inhibition of cytochrome p450 3a4 dependent testosterone 6β hydroxylation by roxithromycin and its metabolites Troleandomycin and erythromycin
Drug Metabolism and Disposition, 1998Co-Authors: Hiroshi Yamazaki, Tsutomu ShimadaAbstract:Roxithromycin has been shown to be a relatively weak inhibitor of cytochrome P450 (P450 or CYP)-dependent drug oxidations, compared with Troleandomycin. The potential for roxithromycin and its major metabolites found in human urine [namely the decladinosyl derivative (M1), O-dealkyl derivative (M2), and N-demethyl derivative (M3)] to inhibit testosterone 6beta-hydroxylation after metabolic activation by CYP3A4 was examined and compared with inhibition by Troleandomycin and erythromycin in vitro. Of roxithromycin and its studied metabolites, M3 was the most potent in inhibiting CYP3A4-dependent testosterone 6beta-hydroxylation by human liver microsomes and was activated to the inhibitory P450.Fe2+-metabolite complex to the greatest extent. Roxithromycin and its metabolites were N-demethylated by human liver microsomes, although the rates were slower than those measured with Troleandomycin and erythromycin as substrates. Recombinant human CYP3A4 in a baculovirus system coexpressing NADPH-P450 reductase was very active in catalyzing the N-demethylation of roxithromycin, M1, and M2, as well as Troleandomycin, erythromycin, and M3. The order for inhibition of CYP3A4-dependent testosterone 6beta-hydroxylation activities by these macrolide antibiotics in the recombinant CYP3A4 system was estimated to be Troleandomycin > erythromycin >/= M3 >/= M2 > M1 >/= roxithromycin. Erythromycin, roxithromycin, and its metabolites all failed to inhibit CYP1A2-dependent (R)-warfarin 7-hydroxylation and CYP2C9-dependent (S)-warfarin 7-hydroxylation but did inhibit CYP3A4-dependent (R)-warfarin 7-hydroxylation. These results suggest that roxithromycin itself is not as potent an inhibitor of CYP3A4 activities as are Troleandomycin and erythromycin, probably because of the slower metabolism of this compound to metabolites M1, M2, and M3 in humans.
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Formation in vitro of an inhibitory cytochrome P450·Fe2+-metabolite complex with roxithromycin and its decladinosyl, O-dealkyl and N-demethyl metabolites in rat liver microsomes
Xenobiotica; the fate of foreign compounds in biological systems, 1998Co-Authors: Hiroshi Yamazaki, Tsutomu ShimadaAbstract:1. Roxithromycin and its major metabolites found in rat and human urine, namely the decladinosyl derivative (M1), O-dealkyl derivative (M2) and N-demethyl derivative (M3), were incubated with rat liver microsomes and formation of an inhibitory cytochrome P450 (CYP)-metabolite complex and of formaldehyde (measurement of N-demethylation) were determined in vitro. Troleandomycin and erythromycin were also used for comparison. 2. Dexamethasone very significantly induced the microsomal N-demethylations of these macrolide antibiotics. The order of magnitude for the Vmax/Km ratio of N-demethylations by liver microsomes from dexamethasone-treated rats was Troleandomycin > erythromycin = M2 > roxithromycin > M3, M1. 3. Formation of an inhibitory P450 x Fe2+-metabolite complex was detected on incubation of these macrolide antibiotics with rat liver microsomes in the presence of an NADPH-generating system and the order of maximum complex formation was Troleandomycin > erythromycin > M2 > roxithromycin > M3 > M1. 4. Troleandomycin, erythromycin and M2 inhibited CYP3A-dependent testosterone 6beta-hydroxylation catalysed by liver microsomes from the dexamethasone-treated rat by 54, 33 and 23%, respectively, but roxithromycin, M3 and M1 were very weak by comparison. In the untreated rat, only testosterone 6beta-hydroxylation, but not testosterone 16alpha- and 2alpha-hydroxylation and androstenedione formation, activities were inhibited, indicating that inhibitory actions of these antibiotics are specific for CYP3A enzymes in liver microsomes. 5. These results support the view that formation of an inhibitory P450-metabolite complex is prerequisite for the inhibition of CYP3A-dependent substrate oxidations by rat liver microsomes and that M2 (and M3, to a lesser extent) may be the active metabolite that can form an inhibitory P450-metabolite complex by CYP3A enzyme(s).
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Effects of roxithromycin, erythromycin and Troleandomycin on their N-demethylation by rat and human cytochrome P450 enzymes
Xenobiotica; the fate of foreign compounds in biological systems, 1996Co-Authors: Hiroshi Yamazaki, S. Hiroki, T. Urano, K. Inoue, Tsutomu ShimadaAbstract:1. The effects of treatment of rat with roxithromycin, erythromycin and Troleandomycin as well as other chemicals including typical cytochrome P450 inducers were examined in rat and human liver microsomes.2. Erythromycin and Troleandomycin but not roxithromycin caused slight increases in CYP3A1 levels and the N-demethylation of roxithromycin, erythromycin and Troleandomycin and oxidation of nifedipine in rat, but none of these chemicals induced significantly CYP2B1 levels or benzphetamine N-demethylation activities.3. Dexamethasone and pregnenolone 16α-carbonitrile induced CYP3A1 levels and N-demethylation of roxithromycin, erythromycin and Troleandomycin but not of benzphetamine, in rat liver microsomes. Treatment of rat with phenobarbital caused increases in both CYP2B1 and 3A1 levels and all of the N-demethylation activities examined. Phenytoin and metyrapone produced increases in contents of 2B1 and activities of benzphetamine N-demethylation as well as of roxithromycin, erythromycin and troleandomyci...
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Effects of erythromycin and roxithromycin on oxidation of testosterone and nifedipine catalyzed by CYP3A4 in human liver microsomes.
The Journal of toxicological sciences, 1996Co-Authors: Hiroshi Yamazaki, Tomoko Urano, Shigeru Hiroki, Tsutomu ShimadaAbstract:Roxithromycin and erythromycin were incubated with rat and human liver microsomal or reconstituted cytochrome P450 (P450 or CYP) monooxygenase systems in the presence of an NADPH-generating system, and the effects of these chemicals on testosterone 6 beta-hydroxylation and nifedipine oxidation activities were compared with those of typical CYP3A4 inhibitors including ketoconazole, Troleandomycin, and gestodene. Roxithromycin and erythromycin were found to be relatively weak inhibitors of testosterone 6 beta-hydroxylation and nifedipine oxidation activities by rat and human liver microsomes or by reconstituted systems containing CYP3A4/5. Formation of an inhibitory P450-metabolite complex was determined spectrally by incubating Troleandomycin with human liver microsomes; the extents of the complex formation were lesser in liver microsomes of humans than those of rats treated with dexamethasone. Erythromycin and roxithromycin were also activated slightly by rat liver microsomes to form P450.Fe(II)-metabolite complex, although these chemicals caused very little or undetectable levels, respectively, of spectral changes by human liver microsomes even when a human sample which contained relatively high levels of CYP3A4 was used. These results suggested that roxithromycin and erythromycin were relatively less potent to inhibit CYP3A4-catalytic activities in human liver microsomes, because of their low capabilities to form P450.Fe(II)-metabolite complex.
Karl-friedrich Sewing - One of the best experts on this subject based on the ideXlab platform.
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Identification of drugs inhibiting the in vitro metabolism of tacrolimus by human liver microsomes
British journal of clinical pharmacology, 1996Co-Authors: Uwe Christians, Gudrun Schmidt, Augustinus Bader, Alfons Lampen, Schottmann R, A Linck, Karl-friedrich SewingAbstract:1. Tacrolimus, an immunosuppressive macrolide, is metabolized by enzymes of the cytochrome P450 3A subfamily. In this study, 34 drugs were tested for their interactions with tacrolimus metabolism by human liver microsomes. 2. Fifteen drugs which inhibit the in vitro metabolism of tacrolimus were identified: bromocriptine, corticosterone, dexamethasone, ergotamine, erythromycin, ethinyloestradiol, josamycin, ketoconazole, miconazole, midazolam, nifedipine, omeprazole, tamoxifen, Troleandomycin and verapamil.
