title: Targeting the Wnt pathway in zebrafish as a screening method to identify novel therapeutic compounds.
abstract: Activating mutations in the Wnt signaling pathway account for the initiation of greater than 90% of all colorectal cancers and this pathway has been implicated in numerous other diseases. Therefore, identifying small molecule inhibitors of this pathway is of critical importance towards identifying clinically relevant drugs. Numerous screens have been employed to identify therapeutic reagents, but none have made it to advanced clinical trials, suggesting that traditional screening methods are ineffective at identifying clinically relevant targets. Here, we describe a novel in vivo screen to identify small molecule inhibitors of the Wnt pathway. Specifically, treatment of zebrafish embryos with LiCl inhibits GSK3 kinase function, resulting in hyperactivation of the signaling pathway and an eyeless phenotype at 1 day post fertilization. Using the small molecule XAV939, a known inhibitor of Wnt signaling, we rescued the LiCl induced eyeless phenotype, confirming efficacy of the screen. We next tested our assay with 400 known small molecule kinase inhibitors, none of which should inhibit Wnt signaling below the level of GSK3 based on their known targets. Accordingly, none of these small molecules rescued the eyeless phenotype, which demonstrates the stringency of the assay. However, several of these small molecule kinase inhibitors did generate a non-Wnt phenotype in accordance with the kinase they targeted. Therefore, combining the efficacy, sensitivity, and stringency of this preliminary screen, this model will provide an alternative to the traditional in vitro screen, generating potentially clinical relevant drugs in a rapid and cost-effective way.
authors.affiliation: Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON Canada N1L 1A3.
authors.forename: Joshua K
authors.initials: JK
authors.lastname: Robertson
authors.forename: Kestral
authors.initials: K
authors.lastname: Danzmann
authors.forename: Sherise
authors.initials: S
authors.lastname: Charles
authors.forename: Katherine
authors.initials: K
authors.lastname: Blake
authors.forename: Annia
authors.initials: A
authors.lastname: Olivares
authors.forename: Solape
authors.initials: S
authors.lastname: Bamikole
authors.forename: Meghan
authors.initials: M
authors.lastname: Olson
authors.forename: Terence J
authors.initials: TJ
authors.lastname: Van  Raay
mesh_terms: D000818:Animals; D015179:Colorectal Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004625:Embryo, Nonmammalian / Q000187:drug effects; D047108:Embryonic Development / Q000187:drug effects / Q000235:genetics; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors; D006575:Heterocyclic Compounds, 3-Ring / Q000494:pharmacology; D018021:Lithium Chloride / Q000494:pharmacology; D010641:Phenotype; D060449:Wnt Signaling Pathway / Q000187:drug effects*; D015027:Zebrafish / Q000196:embryology / Q000378:metabolism*
chemical_list: D006575:Heterocyclic Compounds, 3-Ring; C544261:XAV939; D038362:Glycogen Synthase Kinase 3; D018021:Lithium Chloride
keywords: Cancer; LiCl; Wnt signaling; eyeless; small molecules; zebrafish