title: The BlueScreen-384 assay as an indicator of genotoxic hazard potential in early-stage drug discovery.
abstract: High-throughput cell-based techniques that permit early detection of compound-induced genotoxic damage have recently become available. Methods based on induction of the GADD45a promoter are attractive because multiple intracellular mechanisms that detect genetic damage intersect at this checkpoint gene. Consequently, assays such as GreenScreen HC, which uses p53-competant human TK6 lymphoblastoid cells and a GADD45a-GFP reporter, have been developed. GreenScreen HC allows weekly testing of dozens of compounds using 96-well microplates, with high interassay consistency. BlueScreen HC is a recent advancement, coupling GADD45a to Gaussia luciferase, with several advantages over GADD45a-GFP including the potential for miniaturization. Here we describe implementation of a 384-well BlueScreen assay. For drug discovery programs carrying out iterative analogue synthesis around a chemical lead series, these assays permit assessment of compound genotoxic potential in parallel to, rather than subsequent to, determination of activity at a therapeutic target. We demonstrate comparability of BlueScreen-384 to GreenScreen HC and illustrate the use of BlueScreen-384 to explore the structure-activity relationship around a genotoxic lead molecule to identify nongenotoxic analogues. BlueScreen-384 can reduce the need for costly and time-consuming analogue testing in more traditional genotoxicity tests, such as the Ames test.
authors.affiliation: GlaxoSmithKline, Stevenage, Hertfordshire, UK.
authors.forename: Kate
authors.initials: K
authors.lastname: Simpson
authors.forename: Nicola
authors.initials: N
authors.lastname: Bevan
authors.forename: Paul
authors.initials: P
authors.lastname: Hastwell
authors.forename: Patrick
authors.initials: P
authors.lastname: Eidam
authors.forename: Poonam
authors.initials: P
authors.lastname: Shah
authors.forename: Elke
authors.initials: E
authors.lastname: Gogo
authors.forename: Steve
authors.initials: S
authors.lastname: Rees
authors.forename: Andrew
authors.initials: A
authors.lastname: Brown
mesh_terms: D001681:Biological Assay / Q000379:methods*; D055808:Drug Discovery; D017930:Genes, Reporter / Q000235:genetics; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism; D009152:Mutagenicity Tests; D009153:Mutagens / Q000633:toxicity*; D009154:Mutation / Q000235:genetics; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D015203:Reproducibility of Results; D013329:Structure-Activity Relationship; D013997:Time Factors; D000097642:GADD45 Proteins
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; D008156:Luciferases; D009153:Mutagens; D047428:Protein Kinase Inhibitors; D000097642:GADD45 Proteins