title: High-throughput screening normalized to biological response: application to antiviral drug discovery.
abstract: The process of conducting cell-based phenotypic screens can result in data sets from small libraries or portions of large libraries, making accurate hit picking from multiple data sets important for efficient drug discovery. Here, we describe a screen design and data analysis approach that allow for normalization not only between quadrants and plates but also between screens or batches in a robust, quantitative fashion, enabling hit selection from multiple data sets. We independently screened the MicroSource Spectrum and NCI Diversity Set II libraries using a cell-based phenotypic high-throughput screening (HTS) assay that uses an interferon-stimulated response element (ISRE)-driven luciferase-reporter assay to identify interferon (IFN) signal enhancers. Inclusion of a per-plate, per-quadrant IFN dose-response standard curve enabled conversion of ISRE activity to effective IFN concentrations. We identified 45 hits based on a combined z score ≥2.5 from the two libraries, and 25 of 35 available hits were validated in a compound concentration-response assay when tested using fresh compound. The results provide a basis for further analysis of chemical structure in relation to biological function. Together, the results establish an HTS method that can be extended to screening for any class of compounds that influence a quantifiable biological response for which a standard is available.
authors.affiliation: 1Drug Discovery Program, Pulmonary and Critical Care Medicine, Department of Medicine, Washington University School of Medicine, St. Louis, MO, USA.
authors.forename: Dhara A
authors.initials: DA
authors.lastname: Patel
authors.forename: Anand C
authors.initials: AC
authors.lastname: Patel
authors.forename: William C
authors.initials: WC
authors.lastname: Nolan
authors.forename: Guangming
authors.initials: G
authors.lastname: Huang
authors.forename: Arthur G
authors.initials: AG
authors.lastname: Romero
authors.forename: Nichole
authors.initials: N
authors.lastname: Charlton
authors.forename: Eugene
authors.initials: E
authors.lastname: Agapov
authors.forename: Yong
authors.initials: Y
authors.lastname: Zhang
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Holtzman
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology*; D055808:Drug Discovery / Q000379:methods; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D015870:Gene Expression; D005786:Gene Expression Regulation / Q000187:drug effects; D017930:Genes, Reporter; D057166:High-Throughput Screening Assays; D006801:Humans; D050835:Interferon Regulatory Factors / Q000378:metabolism; D015203:Reproducibility of Results; D020218:Response Elements; D054852:Small Molecule Libraries
chemical_list: D000998:Antiviral Agents; D050835:Interferon Regulatory Factors; D054852:Small Molecule Libraries
keywords: cell-based assay; interferon signal enhancer; phenotypic drug discovery; quantitative HTS; statin