title: A combined in vitro approach to improve the prediction of mitochondrial toxicants.
abstract: Drug induced mitochondrial dysfunction has been implicated in organ toxicity and the withdrawal of drugs or black box warnings limiting their use. The development of highly specific and sensitive in vitro assays in early drug development would assist in detecting compounds which affect mitochondrial function. Here we report the combination of two in vitro assays for the detection of drug induced mitochondrial toxicity. The first assay measures cytotoxicity after 24h incubation of test compound in either glucose or galactose conditioned media (Glu/Gal assay). Compounds with a greater than 3-fold toxicity in galactose media compared to glucose media imply mitochondrial toxicity. The second assay measures mitochondrial respiration, glycolysis and a reserve capacity with mechanistic responses observed within one hour following exposure to test compound. In order to assess these assays a total of 72 known drugs and chemicals were used. Dose-response data was normalised to 100× Cmax giving a specificity, sensitivity and accuracy of 100%, 81% and 92% respectively for this combined approach.
authors.affiliation: Cyprotex Discovery Ltd, BioHub at Alderley Park, Alderley Edge, Cheshire, SK10 4TG, UK.
authors.forename: Julie
authors.initials: J
authors.lastname: Eakins
authors.affiliation: Cyprotex Discovery Ltd, BioHub at Alderley Park, Alderley Edge, Cheshire, SK10 4TG, UK.
authors.forename: Caroline
authors.initials: C
authors.lastname: Bauch
authors.affiliation: Cyprotex Discovery Ltd, BioHub at Alderley Park, Alderley Edge, Cheshire, SK10 4TG, UK.
authors.forename: Heather
authors.initials: H
authors.lastname: Woodhouse
authors.affiliation: Cyprotex Discovery Ltd, BioHub at Alderley Park, Alderley Edge, Cheshire, SK10 4TG, UK.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Park
authors.affiliation: Cyprotex Discovery Ltd, BioHub at Alderley Park, Alderley Edge, Cheshire, SK10 4TG, UK.
authors.forename: Samantha
authors.initials: S
authors.lastname: Bevan
authors.affiliation: Cyprotex Discovery Ltd, BioHub at Alderley Park, Alderley Edge, Cheshire, SK10 4TG, UK.
authors.forename: Clive
authors.initials: C
authors.lastname: Dilworth
authors.affiliation: Cyprotex Discovery Ltd, BioHub at Alderley Park, Alderley Edge, Cheshire, SK10 4TG, UK. Electronic address: p.walker@cyprotex.com.
authors.forename: Paul
authors.initials: P
authors.lastname: Walker
mesh_terms: D001681:Biological Assay; D019069:Cell Respiration; D003470:Culture Media; D064420:Drug-Related Side Effects and Adverse Reactions; D005690:Galactose; D005947:Glucose; D006019:Glycolysis; D056945:Hep G2 Cells; D006801:Humans; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism
chemical_list: D003470:Culture Media; D005947:Glucose; D005690:Galactose
keywords: Extracellular flux assay; HepG2; In vitro; Mitochondria; Toxicity