The pivotal role of hepatocytes in drug discovery

Soars, Matthew G.; McGinnity, Dermot F.; Grime, Ken; Riley, Robert J.

doi:10.1016/j.cbi.2006.11.002

Cited by 154 publications

(117 citation statements)

References 103 publications

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“…The activity of members of the solute carrier and ATP-binding cassette families of transporter proteins may provide a mechanistic explanation for the limited success achieved for some drugs using standard (hepatic microsomal) in vitro experiments to predict pharmacokinetic aspects of clearance and drug-drug interaction (DDI) potential (Lam et al, 2006;Soars et al, 2007;Parker and Houston, 2008). Although various theoretical scenarios involving hepatic transporters can be proposed, specific examples with experimental confirmation of an unequivocal nature have been largely restricted to the statins (Lau et al, 2006;Shitara et al, 2006;Paine et al, 2008).…”

Section: Introductionmentioning

confidence: 99%

Comparative Use of Isolated Hepatocytes and Hepatic Microsomes for Cytochrome P450 Inhibition Studies: Transporter-Enzyme Interplay

Brown¹,

Wilby²,

Alder³

et al. 2010

Drug Metab Dispos

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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 K i 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 l/min/10 6 cells) properties.Inhibition studies were performed using two well established P450 probe substrates (theophylline and midazolam). Comparison of unbound K i 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 K i 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.

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Section: Introductionmentioning

confidence: 99%

Comparative Use of Isolated Hepatocytes and Hepatic Microsomes for Cytochrome P450 Inhibition Studies: Transporter-Enzyme Interplay

Brown¹,

Wilby²,

Alder³

et al. 2010

Drug Metab Dispos

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“…Although microsomes continue to be the first-line screening model for high throughput assays, the number of publications in the last 5 years citing metabolism in hepatocytes has increased by approximately 30% [13], partially due to commercial availability of the cryopreserved cells. Preparation of hepatocytes has been reported in detail by Soars et al [32], while Edwards et al [30] and Lake et al [33] have reported on the preparation of precision-cut human liver slices.…”

Section: Hepatocytes and Liver Slicesmentioning

confidence: 99%

The Conduct of Drug Metabolism Studies Considered Good Practice (II): In Vitro Experiments

Jia¹,

Liu²

2007

CDM

250

195

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In vitro drug metabolism studies, which are inexpensive and readily carried out, serve as an adequate screening mechanism to characterize drug metabolites, elucidate their pathways, and make suggestions for further in vivo testing. This publication is a sequel to part I in a series and aims at providing a general framework to guide designs and protocols of the in vitro drug metabolism studies considered good practice in an efficient manner such that it would help researchers avoid common pitfalls and misleading results. The in vitro models include hepatic and non-hepatic microsomes, cDNA-expressed recombinant human CYPs expressed in insect cells or human B lymphoblastoid, chemical P450 inhibitors, S9 fraction, hepatocytes and liver slices. Important conditions for conducting the in vitro drug metabolism studies using these models are stated, including relevant concentrations of enzymes, co-factors, inhibitors and test drugs; time of incubation and sampling in order to establish kinetics of reactions; appropriate control settings, buffer selection and method validation. Separate in vitro data should be logically integrated to explain results from animal and human studies and to provide insights into the nature and consequences of in vivo drug metabolism. This article offers technical information and data and addresses scientific rationales and practical skills related to in vitro evaluation of drug metabolism to meet regulatory requirements for drug development.

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“…In addition, since endogenous compounds such as bile salts rely on hepatic transporters to maintain normal hepatic physiology (e.g., bile flow), drugs that inhibit the function of key transporters may also cause important disturbances of endogenous substance elimination via the hepatobiliary system. The use of probe substrates and inhibitors to determine the relative importance of individual hepatic transporters has advanced the value of uptake data obtained using hepatocytes in drug discovery (Soars et al, 2007). However, as discussed by Wang et al (Wang et al, 2008), although the use of selective inhibitors of efflux transporters can provide useful mechanistic information on drug interactions involving efflux transporters, the potential cross-reaction of inhibitors with multiple transporters makes it difficult to discern the role of individual transporters in drug transport.…”

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confidence: 99%

Interaction of HIV protease inhibitors with OATP1B1, 1B3, and 2B1

et al. 2010

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The effects of human immunodeficiency virus (HIV) protease inhibitors (PI) on the accumulation of the fluorescent bile salt analogue cholyl-glycylamido-fluorescein (CGamF) were determined in organic anion transporting polypeptide (OATP)-1B1 and -1B3-expressing Chinese hamster ovary (CHO) cells. In addition, interaction studies in Caco-2 monolayers, known only to express the OATP2B1 isoform, were conducted using the established OATP substrate estrone 3-sulfate (E3S), since no CGamF accumulation was observed in Caco-2 monolayers. CGamF appeared an excellent substrate for the OATP1B subfamily, with net accumulation clearance values of 7.8 and 142 microl min(-1) mg(-1) protein in OATP1B1 and OATP1B3-transfected cells, respectively. K(i)-values reflecting inhibition of CGamF accumulation by HIV PI correlated well between OATP1B1 and OATP1B3-expressing cells. Lopinavir was the most potent inhibitor (K(i) = 0.5-1.4 microM) of OATP1B-mediated CGamF accumulation compared with atazanavir, darunavir, ritonavir, and saquinavir (K(i) between 1.4 and 3.3 microM). Inhibitory profiles towards OATP2B1-mediated E3S accumulation were different with only indinavir, saquinavir, and ritonavir showing substantial effects. In conclusion, OATP1B3 appears to be a major transport mechanism mediating sodium-independent CGamF accumulation in human liver, and CGamF could be used as a probe substrate for in vitro drug interaction studies. The remarkably potent inhibition of OATP1B1 by lopinavir may explain some clinically relevant drug interactions between lopinavir and OATP1B substrates such as fexofenadine.

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The pivotal role of hepatocytes in drug discovery

Cited by 154 publications

References 103 publications

Comparative Use of Isolated Hepatocytes and Hepatic Microsomes for Cytochrome P450 Inhibition Studies: Transporter-Enzyme Interplay

Comparative Use of Isolated Hepatocytes and Hepatic Microsomes for Cytochrome P450 Inhibition Studies: Transporter-Enzyme Interplay

The Conduct of Drug Metabolism Studies Considered Good Practice (II): In Vitro Experiments

Interaction of HIV protease inhibitors with OATP1B1, 1B3, and 2B1

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