title: Quantification of Low-Level Drug Effects Using Real-Time, in vitro Measurement of Oxygen Consumption Rate.
abstract: There is a general need to detect toxic effects of drugs during preclinical screening. We propose that increased sensitivity of xenobiotics toxicity combined with improved in vitro physiological recapitulation will more accurately assess potentially toxic perturbations of cellular biochemistry that are near in vivo pharmacological exposure levels. Importantly, measurement of such cytopathologies avoids activating mechanisms mediating toxicity at suprapharmacologic levels not relevant to in vivo effects. We present a sensitive method to measure changes in oxygen consumption rate (OCR), a well-established parameter reflecting a potential hazard, in response to exposure to pharmacologic levels of drugs using a flow culture system and state of the art oxygen sensing system. We tested metformin and acetaminophen on rat liver slices to illustrate the method. The features of the method include continuous and very stable measurement of OCR over the course of 48 h in liver slices in a continuous flow chamber with the ability to resolve changes as small as 0.3%/h. Kinetic modeling of metformin inhibition of OCR over a wide range of concentrations revealed both a slow and fast mechanism, where the fast mechanism activated only at concentrations above 0.6 mM. For both drugs, small amounts of inhibition were reversible, but higher decrements were irreversible. Overall the study highlights the advantages of measuring low-level toxicity so as to avoid the common extrapolations made about drug toxicity based on effects of drugs tested at suprapharmacologic levels.
authors.affiliation: *Department of Medicine, University of Washington, Seattle, Washington, 98195;
authors.forename: Adam
authors.initials: A
authors.lastname: Neal
authors.affiliation: *Department of Medicine, University of Washington, Seattle, Washington, 98195;
authors.forename: Austin M
authors.initials: AM
authors.lastname: Rountree
authors.affiliation: Center for Commercialization, University of Washington, Seattle, Washington 98195;
authors.forename: Craig W
authors.initials: CW
authors.lastname: Philips
authors.affiliation: Department of Environmental and Occupational Health Sciences, University of Washington, Seattle, Washington 98105;
authors.forename: Terrance J
authors.initials: TJ
authors.lastname: Kavanagh
authors.affiliation: Department of Drug Safety and Metabolism, AstraZeneca, Cambridge Science Park, Milton Road, Milton, Cambridge CB4 0FZ, UK; and.
authors.forename: Dominic P
authors.initials: DP
authors.lastname: Williams
authors.affiliation: Department of Drug Safety and Metabolism, AstraZeneca, Cambridge Science Park, Milton Road, Milton, Cambridge CB4 0FZ, UK; and.
authors.forename: Peter
authors.initials: P
authors.lastname: Newham
authors.affiliation: EnTox Sciences, LLC, Mercer Island, Washington 98040.
authors.forename: Gamal
authors.initials: G
authors.lastname: Khalil
authors.affiliation: EnTox Sciences, LLC, Mercer Island, Washington 98040.
authors.forename: Daniel L
authors.initials: DL
authors.lastname: Cook
authors.affiliation: *Department of Medicine, University of Washington, Seattle, Washington, 98195; EnTox Sciences, LLC, Mercer Island, Washington 98040. isweet@u.washington.edu.
authors.forename: Ian R
authors.initials: IR
authors.lastname: Sweet
mesh_terms: D000082:Acetaminophen / Q000633:toxicity*; D000818:Animals; D056486:Chemical and Drug Induced Liver Injury / Q000209:etiology* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D066298:In Vitro Techniques; D007700:Kinetics; D008099:Liver / Q000187:drug effects* / Q000378:metabolism; D008687:Metformin / Q000633:toxicity*; D008954:Models, Biological; D010101:Oxygen Consumption / Q000187:drug effects*; D017207:Rats, Sprague-Dawley; D020127:Recovery of Function; D018570:Risk Assessment; D018675:Toxicity Tests / Q000379:methods*
chemical_list: D000082:Acetaminophen; D008687:Metformin
keywords: acetaminophen; drug toxicity; liver slices; mathematical modeling; metformin; oxygen consumption rate