ReviewThe pivotal role of hepatocytes in drug discovery
Introduction
The liver is the body's largest internal organ, comprising approximately 2% of the total body mass. It is made up of many different cell types with distinct functions; the parenchymal cells (hepatocytes) constitute 65% of total liver cells, occupy over 80% of total liver volume [1] and are the primary site of metabolic drug transformation in the body. The majority of drugs are eliminated from the body predominantly by hepatic metabolism involving CYP-dependent oxidation (phase I metabolism) and most clinical DDIs occur via inhibition or induction of CYP enzymes.
Twenty years ago it was suggested that 80% of candidate drugs failed in development because of sub-optimal ADME [2]. Major liabilities such as metabolic instability and DDI potential have resulted in automated screening programs within drug discovery designed to optimise these parameters using in vitro tools [3]. As a result of this, in vitro drug metabolism and kinetic data have been used with increasing refinement to predict in vivo clearance and DDIs and this focus may account for purported improvements in late stage attrition statistics [4], [5]. For the in vitro study of drug metabolism in the liver, the most widely used tools are isolated hepatocytes, hepatic microsomes and heterologously expressed recombinant human CYPs. These have been incorporated into automated drug discovery methodologies in order to generate the data required for new molecular entity (NME) optimisation [6], [7], [8], [9], [10], [11], [12].
Availability of good quality, fresh liver tissue has limited human hepatocyte experiments but the progress in cryopreservation over the last decade has seen an explosion in the use of human hepatocytes within the pharmaceutical industry and academia allowing hepatocytes to become an ‘off-the-shelf’ reagent [13]. This review details the current and potential uses of hepatocytes in drug discovery from the perspective of the authors’ laboratory.
Section snippets
General use of isolated hepatocytes in drug discovery
In vivo hepatic metabolic clearance can be estimated successfully from the turnover or intrinsic clearance (CLint) derived from in vitro tissue preparations, such as hepatocytes and hepatic microsomes [14], [15]. Many laboratories use hepatic microsomes rather than hepatocytes to determine CLint[16], [17] since they have traditionally been viewed as a more flexible system with which to study oxidative biotransformations in terms of ease of preparation from many species, long-term storage and
Use of hepatocytes in the prediction of metabolic clearance
In the early 1990s in vitro systems (isolated hepatocytes, hepatic microsomes and liver slices) came into widespread use in the pharmaceutical industry. A general strategy for predicting in vivo clearance from in vitro data was presented by Houston [26] which illustrated how existing mathematical models for describing hepatic clearance (CLH), incorporating terms for liver blood flow (Q), CLint and blood binding (fub) [27], [28], [29], [30], could be adopted to facilitate IVIVE with CLint being
The use of hepatocyes to assess drug uptake
To date, there are two main pathways responsible for the uptake of endogenous and xenobiotic substrates into hepatocytes. The sodium-dependent route is catalyzed exclusively at the basolateral membrane by the sodium-taurocholate cotransporting polypeptide (NTCP). NTCP is primarily responsible for the uptake of conjugated bile salts and sulphated steroids but has a more limited role in the uptake of drugs [24]. The organic anion transporting polypeptides (OATPs) are a superfamily of enzymes that
Use of hepatocytes in the assessment of drug–drug interactions
Drug discovery departments in the pharmaceutical industry employ a battery of in vitro screens to assess the potential of new chemical entities to cause pharmacokinetic based DDIs via inhibition and induction of drug metabolising enzymes. Although metabolic DDIs have been reported for several enzyme families, it is recognised that inhibition and induction of CYP dependent metabolism are the most prevalent source of DDIs, which may lead to serious clinical consequences [89], [90].
Summary
Over the last 10–15 years, the use of hepatocytes for studying the routes and rates of drug metabolism has become relatively commonplace. The increase in popularity has been fuelled by the ability to optimise CYP-related biotransformation in Drug Discovery programmes, a desire to minimise affinity at the hERG channel (resulting in an increase in acidic phase II substrates) and the need to address phase II metabolism and hepatic transporter-related events.
In contrast, the application of
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