title: A High-Content Imaging Screen for Cellular Regulators of β-Catenin Protein Abundance.
abstract: Abnormal accumulation of β-catenin protein, a key transcriptional activator required for Wnt signaling, is the hallmark of many tumor types, including colon cancer. In normal cells, β-catenin protein level is tightly controlled by a multiprotein complex through the proteosome pathway. Mutations in the components of the β-catenin degradation complex, such as adenomatous polyposis coli (APC) and Axin, lead to β-catenin stabilization and the constitutive activation of target genes. Since the signal transduction of Wnt/β-catenin is mainly mediated by protein-protein interactions, this pathway has been particularly refractory to conventional target-based small-molecule screening. Here we designed a cellular high-content imaging assay to detect β-catenin protein through immunofluorescent staining in the SW480 colon cancer cell line, which has elevated β-catenin endogenously. We demonstrate that the assay is robust and specific to screen a focused biologically diverse chemical library set against known targets that play diverse cellular functions. We identified a number of hits that reduce β-catenin levels without causing cell death. These hits may serve as tools to understand the dynamics of β-catenin degradation. This study demonstrates that detecting cell-based β-catenin protein stability is a viable approach to identifying novel mechanisms of β-catenin regulation as well as small molecules of therapeutic potential.
authors.affiliation: R&D Target Sciences, GlaxoSmithKline, Upper Providence, Collegeville, PA, USA Xin.X.Zeng@gsk.com.
authors.forename: Xin
authors.initials: X
authors.lastname: Zeng
authors.affiliation: R&D Platform Technology Sciences, GlaxoSmithKine, Upper Providence, Collegeville, PA, USA.
authors.forename: Monica
authors.initials: M
authors.lastname: Montoute
authors.affiliation: R&D Platform Technology Sciences, GlaxoSmithKine, Upper Providence, Collegeville, PA, USA.
authors.forename: Tiger W
authors.initials: TW
authors.lastname: Bee
authors.affiliation: R&D Alternative Discovery and Development, GlaxoSmithKline, Upper Providence, Collegeville, PA, USA.
authors.forename: Hong
authors.initials: H
authors.lastname: Lin
authors.affiliation: R&D Platform Technology Sciences, GlaxoSmithKine, Upper Providence, Collegeville, PA, USA.
authors.forename: Lorena A
authors.initials: LA
authors.lastname: Kallal
authors.affiliation: R&D Platform Technology Sciences, GlaxoSmithKine, Upper Providence, Collegeville, PA, USA.
authors.forename: Yan
authors.initials: Y
authors.lastname: Liu
authors.affiliation: R&D Target Sciences, GlaxoSmithKline, Upper Providence, Collegeville, PA, USA.
authors.forename: Pankaj
authors.initials: P
authors.lastname: Agarwal
authors.affiliation: R&D Platform Technology Sciences, GlaxoSmithKine, Upper Providence, Collegeville, PA, USA.
authors.forename: Dayuan
authors.initials: D
authors.lastname: Wang
authors.affiliation: R&D Target Sciences, GlaxoSmithKline, Upper Providence, Collegeville, PA, USA.
authors.forename: Quinn
authors.initials: Q
authors.lastname: Lu
authors.affiliation: R&D Target Sciences, GlaxoSmithKline, Upper Providence, Collegeville, PA, USA.
authors.forename: Dwight
authors.initials: D
authors.lastname: Morrow
authors.affiliation: R&D Alternative Discovery and Development, GlaxoSmithKline, Upper Providence, Collegeville, PA, USA.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Pope
authors.affiliation: R&D Platform Technology Sciences, GlaxoSmithKine, Upper Providence, Collegeville, PA, USA.
authors.forename: Zining
authors.initials: Z
authors.lastname: Wu
mesh_terms: D011125:Adenomatous Polyposis Coli / Q000188:drug therapy; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D000918:Antibody Specificity; D000970:Antineoplastic Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D004354:Drug Screening Assays, Antitumor / Q000379:methods*; D057166:High-Throughput Screening Assays; D006801:Humans; D020128:Inhibitory Concentration 50; D054852:Small Molecule Libraries; D060449:Wnt Signaling Pathway / Q000187:drug effects*; D051176:beta Catenin / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; D000970:Antineoplastic Agents; D054852:Small Molecule Libraries; D051176:beta Catenin
keywords: Wnt; adenomatous polyposis coli (APC); colon cancer; high-content imaging; tankyrase (TNKS); β-catenin