Development and validation of a higher throughput screening approach to genotoxicity testing using the GADD45a-GFP GreenScreen HC assay

Andrew W. Knight, Louise Birrell, Richard M. Walmsley

    Research output: Contribution to journalArticlepeer-review

    Abstract

    There is a pressing need to develop rapid yet accurate screening assays for the identification of genotoxic liability and for early hazard assessment in drug discovery. The GADD45a-GFP human cell-based genotoxicity assay (GreenScreen HC) has been reformatted to test 12 compounds per 96-well microplate in a higher throughput, automated screening mode and the protocol applied to the analysis of 1266 diverse, pharmacologically active compounds. Testing from a fixed starting concentration of 100 μM and over 3 serial dilutions, the hit rates for genotoxicity (7.3%) and cytotoxicity (33%) endpoints of the assay have been determined in a much wider chemical space than previously reported. The degree of interference from color, autofluorescence, and low solubility has also been assessed. The assay results have been compared to an in silico approach to genotoxicity assessment using Derek for Windows software. Where carcinogenicity data were available, GreenScreen HC demonstrated a higher specificity than in silico methods while identifying genotoxic species that were not highlighted for genotoxic liability in structure-activity relationship software. Higher throughput screening from a fixed, low concentration reduces sensitivity to less potent genotoxins, but the maintenance of the previously reported high specificity is essential in early hazard assessment where misclassification can lead to the needless rejection of potentially useful compounds in drug development. © 2009 Society for Biomolecular Sciences.
    Original languageEnglish
    Pages (from-to)16-30
    Number of pages14
    JournalJournal of Biomolecular Screening
    Volume14
    Issue number1
    DOIs
    Publication statusPublished - Jan 2009

    Keywords

    • GADD45a
    • Genotoxicity
    • Green fluorescent protein
    • GreenScreen HC
    • High-throughput screening
    • LOPAC

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