Abstract
Thiabendazole (TBZ) and its major metabolite 5-hydroxythiabendazole (5OH-TBZ) were screened for potential time-dependent inhibition (TDI) against CYP1A2. Screen assays were carried out in the absence and presence of NADPH. TDI was observed with both compounds, with kinact and KI values of 0.08 and 0.02 min–1 and 1.4 and 63.3 μM for TBZ and 5OH-TBZ, respectively. Enzyme inactivation was time-, concentration-, and NADPH-dependent. Inactivation by TBZ was irreversible by dialysis and oxidation by potassium ferricyanide, and there was no protection by glutathione. 5OH-TBZ was a weak TDI of CYP1A2, and enzyme activity was recovered by dialysis. IC50 determination of TBZ and 5OH-TBZ showed both compounds to be potent inhibitors, with IC50 values of 0.83 and 13.05 μM, respectively. IC50 shift studies also demonstrated that TBZ was a TDI of CYP1A2. In silico methods identified the thiazole group as a TDI fragment and predicted it as the site of metabolism. The observation pointed to epoxidation of the thiazole and the benzyl rings of TBZ as possible routes of metabolism and mechanisms of TDI. Drug-drug interaction (DDI) simulation studies using SimCyp showed good predictions for competitive inhibition. However, predictions for mechanism-based inhibition (MBI)-based DDI were not in agreement with clinical observations. There was no TBZ accumulation upon chronic administration of the drug. The in vitro MBI findings might therefore not be capturing the in vivo situation in which the proposed bioactivation route is minor. This might be the case for TBZ in which, in vivo, UDP glucuronosyltransferases and sulfanotransferase metabolize and eliminate the 5OH-TBZ.
Footnotes
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This work was supported in part by the African Institute of Biomedical Science and Technology (Zimbabwe) [Grant DMPK-004]; International Programme in the Chemical Sciences (Sweden); European Union (AntiMal Project); and AstraZeneca (Sweden). R.T. is a recipient of the European Union AntiMal project Ph.D. scholarship for a joint project between African Institute of Biomedical Science and Technology and the University of Capetown.
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Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
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doi:10.1124/dmd.108.024604.
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ABBREVIATIONS: TBZ, thiabendazole; P450, cytochrome P450; CHC, 3-cyano-7-hydroxycoumarin; TDI, time-dependent inhibition/inhibitor; MBI, mechanism-based inhibition/inhibitor; CEC, 3-cyano-7-ethoxycoumarin; fm, fraction metabolized; 5OH-TBZ, 5-hydroxythiabendazole; AUC, area under the curve; ACN, acetonitrile.
- Received September 15, 2008.
- Accepted March 18, 2009.
- The American Society for Pharmacology and Experimental Therapeutics
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