University of Oulu

Cytochrome P450 enzymes—in vitro, in vivo, and in silico studies

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Author: Turpeinen, Miia1,2
Organizations: 1University of Oulu, Faculty of Medicine, Department of Pharmacology and Toxicology
2University of Turku, Faculty of Medicine, Department of Pharmacology, Drug Development and Therapeutics
Format: ebook
Version: published version
Access: open
Online Access: PDF Full Text (PDF, 1 MB)
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Language: English
Published: 2006
Publish Date: 2006-10-10
Thesis type: Doctoral Dissertation
Defence Note: Academic dissertation to be presented, with the assent of the Faculty of Medicine of the University of Oulu, for public defence in the Auditorium of the Department of Pharmacology and Toxicology, on October 20th, 2006, at 12 noon
Reviewer: Professor Marja-Liisa Dahl
Doctor Amin Rostami-Hodjegan


Metabolism is a major determinant of the pharmacokinetic properties of most drugs and is often behind bioavailability problems, drug-drug interactions, and metabolic idiosyncrasies. Cytochrome P450 (CYP) enzymes are a superfamily of microsomal hemoproteins catalysing the metabolic reactions of several exogenous compounds. The majority of crucial steps within drug metabolism are in connection with CYP enzymes.

In the present study, in vivo, in vitro, and in silico approaches were applied and characterised to evaluate the effects of chemical entities on CYP-mediated metabolism. CYP2B6 was used as a target enzyme for these studies.

For evaluation of the CYP inhibition potential of new chemical entities, a novel in vitro test system utilising the n-in-one approach was developed. This method proved to be robust and applicable to screening purposes. Validation of the n-in-one assay was done by comparing its performance to commonly used in vitro techniques using six structurally diverse drugs. All assay types yield remarkably similar results with the majority of the CYP forms tested.

Several chemicals were screened in vitro and in silico in order to find potent and selective chemical inhibitors for CYP2B6. Ticlopidine, thioTEPA and 4-(4-chlorobenzylpyridine) were found to be highly effective inhibitors of CYP2B6. The selectivity of thioTEPA proved to be very high, whereas ticlopidine and 4-(4-chlorobenzylpyridine) also inhibited other CYPs. At a concentration level of 1 μM for ticlopidine and 0.1 μM for 4-(4-chlorobenzylpyridine), the inhibitory effect towards other CYPs was negligible.

Due to wide clinical use and relevance, clopidogrel and ticlopidine were selected for further in vivo interaction studies. Both clopidogrel and ticlopidine significantly inhibited the CYP2B6-catalysed bupropion hydroxylation and patients receiving either clopidogrel or ticlopidine are likely to need dose adjustments when treated with drugs primarily metabolised by CYP2B6. The effect of impaired kidney function on CYP2B6 activity and on bupropion pharmacokinetics was also explored. In patients with kidney disease, the bupropion AUC and Cmax were significantly higher and the apparent oral clearance of bupropion was notably lower compared to healthy controls.

The present results indicate that the in silico and in vitro methods used are helpful in predicting in vivo drug-drug interactions. The effective utilisation of these models in the early phases of drug discovery could therefore help to target the in vivo studies and to eliminate metabolically unfavourable drug candidates.

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Series: Acta Universitatis Ouluensis. D, Medica
ISSN-E: 1796-2234
ISBN: 951-42-8220-5
ISBN Print: 951-42-8219-1
Issue: 895
Copyright information: © University of Oulu, 2006. This publication is copyrighted. You may download, display and print it for your own personal use. Commercial use is prohibited.