title: A robust high-content imaging approach for probing the mechanism of action and phenotypic outcomes of cell-cycle modulators.
abstract: High-content screening is increasingly used to elucidate changes in cellular biology arising from treatment with small molecules and biological probes. We describe a cell classifier for automated analysis of multiparametric data from immunofluorescence microscopy and characterize the phenotypes of 41 cell-cycle modulators, including several protein kinase inhibitors in preclinical and clinical development. This method produces a consistent assessment of treatment-induced phenotypes across experiments done by different biologists and highlights the prevalence of nonuniform and concentration-dependent cellular response to treatment. Contrasting cell phenotypes from high-content screening to kinase selectivity profiles from cell-free assays highlights the limited utility of enzyme potency ratios in understanding the mechanism of action for cell-cycle kinase inhibitors. Our cell-level approach for assessing phenotypic outcomes is reliable, reproducible and capable of supporting medium throughput analyses of a wide range of cellular perturbations.
authors.affiliation: Eli Lilly and Company, Lilly Corporate Center, Indianapolis, IN 46285, USA. sutherlandje@lilly.com
authors.forename: Jeffrey J
authors.initials: JJ
authors.lastname: Sutherland
authors.forename: Jonathan
authors.initials: J
authors.lastname: Low
authors.forename: Wayne
authors.initials: W
authors.lastname: Blosser
authors.forename: Michele
authors.initials: M
authors.lastname: Dowless
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Engler
authors.forename: Louis F
authors.initials: LF
authors.lastname: Stancato
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D002453:Cell Cycle / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects; D002477:Cells / Q000166:cytology* / Q000187:drug effects*; D003663:Decision Trees; D004305:Dose-Response Relationship, Drug; D045325:HCT116 Cells; D006801:Humans; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000378:metabolism; D010641:Phenotype; D047428:Protein Kinase Inhibitors / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D015203:Reproducibility of Results
chemical_list: D000970:Antineoplastic Agents; D047428:Protein Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases