Abstract
Accurate assignment of the concentration of victim drug/inhibitor available at the enzyme active site, both in vivo and within an in vitro incubation, is an essential requirement in rationalizing and predicting drug-drug interactions. Inhibitor accumulation within the liver, whether as a result of active transport processes or intracellular binding, may best be accounted for using hepatocytes rather than hepatic microsomes to estimate in vitro inhibitory potency. The aims of this study were to compare Ki values determined in rat liver microsomes and freshly isolated rat hepatocytes of four cytochrome P450 (P450) inhibitors (clarithromycin, enoxacin, nelfinavir, and saquinavir) with known hepatic transporter involvement and a range of uptake (cell/medium concentration ratios 20–3000) and clearance (10–1200 μl/min/106 cells) properties. Inhibition studies were performed using two well established P450 probe substrates (theophylline and midazolam). Comparison of unbound Ki values showed marked differences between the two in vitro systems for inhibition of metabolism. In two cases (clarithromycin and enoxacin, both low-clearance drugs), inhibitory potency in hepatocytes markedly exceeded that in microsomes (10- to 20-fold), and this result was consistent with their high cell/medium concentration ratios. For nelfinavir and saquinavir (high-clearance, extensively metabolized drugs), the opposite trend was seen in the Ki values: despite very high cell/medium concentration ratios, stronger inhibition was evident within microsomal preparations. Hence, the consequences of hepatic accumulation resulting from uptake transporters vary according to the clearance of the inhibitor. This study demonstrates that transporter-enzyme interplay can result in differences in inhibitory potency between microsomes and hepatocytes and hence drug-drug interaction predictions that are not always intuitive.
Footnotes
This work was supported in part by a consortium of pharmaceutical companies (GlaxoSmithKline, Lilly, Novartis, Pfizer, and Servier) within the Centre for Applied Pharmacokinetic Research at the University of Manchester. A.J.W. was supported by a Biotechnology and Biological Sciences Research Council Collaborative Award in Science and Engineering studentship with Roche Products Ltd.
Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
doi:10.1124/dmd.110.035824.
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ABBREVIATIONS:
- DDI
- drug-drug interaction
- P450
- cytochrome P450
- CLAR
- clarithromycin
- ENX
- enoxacin
- NFV
- nelfinavir
- SQV
- saquinavir
- YKpu
- true hepatocyte/medium unbound drug concentration ratio reflecting purely distribution processes
- Kpu, app
- apparent hepatocyte/medium unbound drug concentration ratio reflecting both elimination and distribution processes
- 1,3-DMU
- 1,3-dimethyluric acid.
- Received August 9, 2010.
- Accepted September 16, 2010.
- Copyright © 2010 by The American Society for Pharmacology and Experimental Therapeutics
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