The Acute Extracellular Flux (XF) Assay to Assess Compound Effects on Mitochondrial Function

• Published in: Journal of biomolecular screening Vol. 20; no. 3; pp. 422 - 429
• Main Authors: Wang, Ruolan, Novick, Steven J., Mangum, James B., Queen, Kennedy, Ferrick, David A., Rogers, George W., Stimmel, Julie B.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.03.2015
• Subjects: acute; Automation; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical - methods; Hep G2 Cells; High-Throughput Screening Assays; Humans; Mitochondria - drug effects; Mitochondria - metabolism; mitochondrial dysfunction; Reproducibility of Results; screening assay; Small Molecule Libraries; toxicities; acute; toxicities; mitochondrial dysfunction; screening assay
• ISSN: 2472-5552; 2472-5560; 1552-454X
• DOI: 10.1177/1087057114557621

Numerous investigations have linked mitochondrial dysfunction to adverse health outcomes and drug-induced toxicity. The pharmaceutical industry is challenged with identifying mitochondrial liabilities earlier in drug development and thereby reducing late-stage attrition. Consequently, there is a demand for reliable, higher-throughput screening methods for assessing the impact of drug candidates on mitochondrial function. The extracellular flux (XF) assay described here is a plate-based method in which galactose-conditioned HepG2 cells were acutely exposed to test compounds, then real-time changes in the oxygen consumption rate and extracellular acidification rate were simultaneously measured using a Seahorse Bioscience XF-96 analyzer. The acute XF assay was validated using marketed drugs known to modulate mitochondrial function, and data analysis was automated using a spline curve fitting model developed at GlaxoSmithKline. We demonstrate that the acute XF assay is a robust, sensitive screening platform for evaluating drug-induced effects on mitochondrial activity in whole cells.