Microtiter Plate Assays for Inhibition of Human, Drug-Metabolizing Cytochromes P450

Crespi, Charles L.; Miller, Vaughn P.; Penman, Bruce W.

doi:10.1006/abio.1997.2145

Cited by 390 publications

(261 citation statements)

References 13 publications

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“…The formation of the fluorescent metabolites were monitored using the following excitation and emission wavelengths: CYP1A2, 405 and 460 nm; CYP2C9, 405 and 535 nm; CYP2C19, 405 and 460 nm; CYP2D6, 390 and 460 nm; and CYP3A4, 405 and 535 nm, respectively. The experiments were repeated six times for each enzyme, and IC 50 values for TS-011 on each enzyme were determined according to the method of Crespi et al (1997).…”

Section: Methodsmentioning

confidence: 99%

Beneficial Effects of a New 20-Hydroxyeicosatetraenoic Acid Synthesis Inhibitor, TS-011 [N-(3-Chloro-4-morpholin-4-yl) Phenyl-N′-hydroxyimido Formamide], on Hemorrhagic and Ischemic Stroke

Miyata

Seki²,

Tanaka³

et al. 2005

J Pharmacol Exp Ther

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The present study characterized the effects of TS-011 [N-(3-chloro-4-morpholin-4-yl) phenyl-NЈ-hydroxyimido formamide], a new selective inhibitor of the synthesis of 20-hydroxyeicosatetraenoic acid (20-HETE), on the metabolism of arachidonic acid by human and rat renal microsomes and the inhibitory effects of this compound on hepatic cytochrome P450 enzymes involved in drug metabolism. The effects of TS-011 on the fall in cerebral blood flow following subarachnoid hemorrhage (SAH) and in reducing infarct size in ischemic stroke models were also examined since 20-HETE may contribute to the development of cerebral vasospasm. TS-011 inhibited the synthesis of 20-HETE by human renal microsomes and recombinant CYP4A11 and 4F2, 4F3A, and 4F3B enzymes with IC 50 values around 10 to 50 nM. It had no effect on the activities of CYP1A, 2C9, 2C19, 2D6, or 3A4 enzymes. TS-011 inhibited the synthesis of 20-HETE by rat renal microsomes with an IC 50 of 9.19 nM, and it had no effect on epoxygenase activity at a concentration of 100 M. TS-011 (0.01-1 mg/kg i.v.) reversed the fall in cerebral blood flow and the increase in 20-HETE levels in the cerebrospinal fluid of rats after SAH. TS-011 also reduced the infarct volume by 35% following transient ischemic stroke and in intracerebral hemorrhage in rats. Injection of 20-HETE (8 or 12 mg/kg) into the carotid artery produced an infarct similar to that seen in the ischemic stroke model. These studies indicate that blockade of the synthesis of 20-HETE with TS-011 opposes cerebral vasospasm following SAH and reduces infarct size in ischemic models of stroke.Each year, 750,000 strokes occur in the United States. Approximately 80% of the strokes are ischemic, and 20% are hemorrhagic. Hemorrhagic stroke commonly occurs after rupture of aneurysms or head trauma. There is an initial period of cerebral ischemia lasting several hours due to a rise in cerebral spinal fluid (CSF) pressure and acute cerebral vasospasm. One-half of the patients that survive later develop delayed vasospasm. The mortality in the 1st month hovers around 50%. The majority of deaths (Ͼ60%) occur during the first 2 days and is associated with ischemic brain injury (Broderick et al., 1994). The factors that contribute to the acute fall in cerebral blood flow following subarachnoid hemorrhage (SAH) remain uncertain.Ischemic strokes are caused by blockage of cerebral arteries. In the ischemic neuronal tissue, there is depletion of

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Section: Methodsmentioning

confidence: 99%

Beneficial Effects of a New 20-Hydroxyeicosatetraenoic Acid Synthesis Inhibitor, TS-011 [N-(3-Chloro-4-morpholin-4-yl) Phenyl-N′-hydroxyimido Formamide], on Hemorrhagic and Ischemic Stroke

Miyata

Seki²,

Tanaka³

et al. 2005

J Pharmacol Exp Ther

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“…The assays of inhibition on human 2C9 enzyme activities were performed in a multiwell plate using CYP inhibition assay kit (GENTEST, Woburn, MA) as described previously (Crespi et al, 1997). Briefly, human CYP enzymes were obtained from baculovirus-infected insect cells.…”

Section: Cyp 2c9 Inhibition Assaymentioning

confidence: 99%

Pharmacokinetic Drug Interaction between Carvedilol and Ticlopidine in Rats

Choi¹

2010

Biomolecules and Therapeutics

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-This study was designed to investigate the effects of ticlopidine on the pharmacokinetics of carvedilol after oral or intravenous administration of carvedilol in rats. Carvedilol was administered orally (3 mg/kg) or intravenously (1 mg/kg) without or with oral administration of ticlopidine (4, 12 mg/kg) to rats. The effects of ticlopidine on P-glycoprotein (P-gp) and cytochrome P450 (CYP) 2C9 activity were also evaluated. Ticlopidine inhibited CYP2C9 activity in a concentration-dependent manner with 50% inhibition concentration (IC 50 ) of 25.2 μM. In addition, ticlopidine could not significantly enhance the cellular accumulation of rhodamine 123 in MCF-7/ADR cells overexpressing P-gp. Compared with the control group (given carvedilol alone), the area under the plasma concentration-time curve (AUC) was significantly (12 mg/kg, p＜0.05) increased by 14-41%, and the peak concentration (C max ) was significantly (12 mg/kg, p＜0.05) increased by 10.7-73.3% in the presence of ticlopidine after oral administration of carvedilol. Consequently, the relative bioavailability (R.B.) of carvedilol was increased by 1.14-to 1.41-fold and the absolute bioavailability (A.B.) of carvedilol in the presence of ticlopidine was increased by 36.2-38.5%. Compared to the i.v. control, ticlopidine could not significantly change the pharmacokinetic parameters of i.v. administered carvedilol. The enhanced oral bioavailability of carvedilol may result from inhibition of CYP2C9-mediated metabolism rather than P-gpmediated efflux of carvedilol in the intestinal and/or in liver and renal eliminatin of carvedilol by ticlopidine.

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“…The various aqueous herbal product extracts were then evaluated for their ability to inhibit CYP3A4-mediated metabolism of a reference substrate using an in vitro fluorimetric microtiter plate assay adapted and modified Budzinski et al (2000) from that originally reported by Crespi et al (1997) and later described by GENTEST Corporation (GENTEST, 1998a). Briefly, assays were performed in clear-bottom, opaque-welled microtiter plates (96-well, Corning Costar, model no.…”

Section: Cyp3a4 Inhibitionmentioning

confidence: 99%

The Interaction of Selected Phytochemicals, HIV Drugs, and Commercial-Source Herbal Teas and Capsules with Human Cytochrome P450 3A4 and P-glycoprotein

Budzinski

Foster

Trudeau

et al. 2008

Pharmaceutical Biology

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As the popularity of natural health product (NHP) use increases, unfortunately, so does the frequency of cases reporting suspected adverse interactions. Adverse reactions associated with concomitant NHP-therapeutic use may result from competing interactions at the level of the key xenobiotic-biotransforming phase I enzyme cytochrome P450 3A4 (CYP3A4) or the membrane-bound ATP-dependent protein pump P-glycoprotein (P-gp). In this study, selected NHPs of interest to the HIV+ community were assessed for their ability to affect these two important mechanisms of drug disposition. Briefly, commercial-source teas and powdered extracts in capsule formulations were examined for their ability to inhibit CYP3A4-mediated metabolism of the coadministered reference substrate dibenzyl-fluorescein, and their ability to stimulate P-gp ATPase activity. Among the herbal capsules, it was found that aqueous extracts of two different goldenseal (Hydrastis canadensis L.) products were the most inhibitory of CYP3A4-mediated metabolism among all NHPs tested (IC 50 : 3.03 and 3.23 mg/mL). Goldenseal and milk thistle [Silybum marianum (L.) Gaertn.] teas were found to stimulate P-gp ATPase to a greater degree than the reference positive control verapamil (20 µM). As well, 70% ethanol extracts of one goldenseal product (designated NRP 121) and aqueous extracts of another (designated NRP 17) had the highest relative P-gp ATPase activity overall. Milk thistle and goldenseal products were further analyzed * Dedicated to Professor John Thor Arnason of the University of Ottawa, Department of Biology, on the occasion of his sixtieth birthday.

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Microtiter Plate Assays for Inhibition of Human, Drug-Metabolizing Cytochromes P450

Cited by 390 publications

References 13 publications

Beneficial Effects of a New 20-Hydroxyeicosatetraenoic Acid Synthesis Inhibitor, TS-011 [N-(3-Chloro-4-morpholin-4-yl) Phenyl-N′-hydroxyimido Formamide], on Hemorrhagic and Ischemic Stroke

Beneficial Effects of a New 20-Hydroxyeicosatetraenoic Acid Synthesis Inhibitor, TS-011 [N-(3-Chloro-4-morpholin-4-yl) Phenyl-N′-hydroxyimido Formamide], on Hemorrhagic and Ischemic Stroke

Pharmacokinetic Drug Interaction between Carvedilol and Ticlopidine in Rats

The Interaction of Selected Phytochemicals, HIV Drugs, and Commercial-Source Herbal Teas and Capsules with Human Cytochrome P450 3A4 and P-glycoprotein

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