Abstract
Hepatic clearance (CLH) prediction is a critical parameter to estimate human dose. However, CLH underpredictions are common, especially for slowly metabolized drugs, and may be attributable to drug properties that pose challenges for conventional in vitro absorption, distribution, metabolism, and elimination (ADME) assays, resulting in nonvalid data, which prevents in vitro to in vivo extrapolation and CLH predictions. Other processes, including hepatocyte and biliary distribution via transporters, can also play significant roles in CLH. Recent advances in understanding the interplay of metabolism and drug transport for clearance processes have aided in developing the extended clearance model. In this study, we demonstrate proof of concept of a novel two-step assay enabling the measurement of multiple kinetic parameters from a single experiment in plated human primary hepatocytes with and without transporter and cytochrome P450 inhibitors—the hepatocyte uptake and loss assay (HUpLA). HUpLA accurately predicted the CLH of eight of the nine drugs (within twofold of the observed CLH). Distribution clearances were within threefold of observed literature values in standard uptake and efflux assays. In comparison, the conventional suspension hepatocyte stability assay poorly predicted the CLH. The CLH of only two drugs was predicted within twofold of the observed CLH. Therefore, HUpLA is advantageous by enabling the measurement of enzymatic and transport processes concurrently within the same system, alleviating the need for applying scaling factors independently. The use of primary human hepatocytes enables physiologically relevant exploration of transporter-enzyme interplay. Most importantly, HUpLA shows promise as a sensitive measure for low-turnover drugs. Further evaluation across different drug characteristics is needed to demonstrate method robustness.
SIGNIFICANCE STATEMENT The hepatocyte uptake and loss assay involves measuring four commonly derived in vitro hepatic clearance endpoints. Since endpoints are generated within a single test system, it blunts experimental error originating from assays otherwise conducted independently. A key advantage is the concept of removing drug-containing media following intracellular drug loading, enabling the measurement of drug reappearance rate in media as well as the measurement of loss of total drug in the test system unencumbered by background quantities of drug in media otherwise present in a conventional assay.
Footnotes
- Received April 25, 2024.
- Accepted July 15, 2024.
This manuscript was sponsored and funded by AbbVie.
All authors are (or were) employees of AbbVie and may own AbbVie stock. AbbVie contributed to the design and participated in the collection, analysis, and interpretation of data and in writing, reviewing, and approving the final abstract, which contains no proprietary AbbVie data. Mei Feng is a former AbbVie employee and has no conflict of interest.
↵1Current affiliation: Biopharmaceutical Innovation and Optimization Center, The University of Kansas, Lawrence, Kansas.
↵This article has supplemental material available at dmd.aspetjournals.org.
- Copyright © 2024 by The American Society for Pharmacology and Experimental Therapeutics