High-content screen using zebrafish (Danio rerio) embryos identifies a novel kinase activator and inhibitor.
In this report we utilized zebrafish (Danio rerio) embryos in a phenotypical high-content screen (HCS) to identify novel leads in a cancer drug discovery program. We initially validated our HCS model using the flavin adenosine dinucleotide (FAD) containing endoplasmic reticulum (ER) enzyme, endoplasmic reticulum oxidoreductase (ERO1) inhibitor EN460. EN460 showed a dose response effect on the embryos with a dose of 10μM being significantly lethal during early embryonic development. The HCS campaign which employed a small library identified a promising lead compound, a naphthyl-benzoic acid derivative coined compound 1 which had significant dosage and temporally dependent effects on notochord and muscle development in zebrafish embryos. Screening a 369 kinase member panel we show that compound 1 is a PIM3 kinase inhibitor (IC50=4.078μM) and surprisingly a DAPK1 kinase agonist/activator (EC50=39.525μM). To our knowledge this is the first example of a small molecule activating DAPK1 kinase. We provide a putative model for increased phosphate transfer in the ATP binding domain when compound 1 is virtually docked with DAPK1. Our data indicate that observable phenotypical changes can be used in future zebrafish screens to identify compounds acting via similar molecular signaling pathways.
Department of Pharmaceutical Sciences, School of Pharmacy, West Virginia University, Morgantown, WV 26506, United States. Electronic address: werner.geldenhuys@hsc.wvu.edu.
Werner J
WJ
Geldenhuys
Department of Biology, Eberly College of Arts and Sciences, West Virginia University, Morgantown, WV 26506, United States.
Sadie A
SA
Bergeron
Department of Biology, Eberly College of Arts and Sciences, West Virginia University, Morgantown, WV 26506, United States.
Jackie E
JE
Mullins
Department of Pharmaceutical Sciences, School of Pharmacy, West Virginia University, Morgantown, WV 26506, United States.
Rowaa
R
Aljammal
Department of Pharmaceutical Sciences, School of Pharmacy, West Virginia University, Morgantown, WV 26506, United States.
Briah L
BL
Gaasch
Department of Pharmaceutical Sciences, School of Pharmacy, West Virginia University, Morgantown, WV 26506, United States.
Wei-Chi
WC
Chen
Department of Integrative Medical Sciences, College of Medicine, Northeast Ohio Medical University, Rootstown, OH 44272, United States.
June
J
Yun
Department of Pharmaceutical Sciences, School of Pharmacy, West Virginia University, Morgantown, WV 26506, United States.
Lori A
LA
Hazlehurst
D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology; D019817:Benzoic Acid / Q000737:chemistry / Q000494:pharmacology; D064550:Death-Associated Protein Kinases / Q000378:metabolism; D055808:Drug Discovery / Q000379:methods*; D004354:Drug Screening Assays, Antitumor / Q000379:methods; D004625:Embryo, Nonmammalian / Q000187:drug effects* / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D020536:Enzyme Activators / Q000737:chemistry* / Q000494:pharmacology*; D009369:Neoplasms / Q000188:drug therapy / Q000201:enzymology; D047428:Protein Kinase Inhibitors / Q000737:chemistry* / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D015027:Zebrafish / Q000196:embryology*; D029961:Zebrafish Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism
D000970:Antineoplastic Agents; D020536:Enzyme Activators; D047428:Protein Kinase Inhibitors; D029961:Zebrafish Proteins; D019817:Benzoic Acid; D064550:Death-Associated Protein Kinases; D017346:Protein Serine-Threonine Kinases
Cancer; Compound library; Kinase; Notochord; Phenotypic screen; Somites; Zebrafish