High-content screen using zebrafish (Danio rerio) embryos identifies a novel kinase activator and inhibitor.
High-content screen using zebrafish (Danio rerio) embryos identifies a novel kinase activator and inhibitor.
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DOI：
10.1016/j.bmcl.2017.02.068
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发表时间：
2017-05-01
影响因子：
2.7
通讯作者：
Hazlehurst LA
中科院分区：
文献类型：
--
作者：
Geldenhuys WJ;Bergeron SA;Mullins JE;Aljammal R;Gaasch BL;Chen WC;Yun J;Hazlehurst LA
在这份报告中，我们利用斑马鱼(Danio Rerio)胚胎在表型高含量筛选(HCS)中识别抗癌药物发现计划中的新线索。我们最初使用含有内质网(ER)酶的黄素腺苷二核苷酸(FAD)和内质网氧化还原酶(ERO1)抑制剂EN460来验证我们的HCS模型。在胚胎发育早期，10μM剂量的EN460对胚胎有明显的致死作用，表现出剂量效应效应。HCS活动利用一个小型文库发现了一种很有前途的先导化合物，一种名为萘基苯甲酸衍生物的化合物1，它对斑马鱼胚胎的脊索和肌肉发育具有显著的剂量和时间依赖效应。通过对369个激酶成员的筛选，我们发现化合物1是PIM3K抑制因子(IC_(50)=4.078μM)和DAPK_1激动剂/激活剂(EC_(50)=39.525μM)。据我们所知，这是第一个小分子激活DAPK1激酶的例子。我们提供了一个假设的模型，当化合物1与DAPK1虚拟对接时，增加了ATP结合域中的磷酸转移。我们的数据表明，可观察到的表型变化可以用于未来的斑马鱼筛选，以识别通过类似的分子信号通路发挥作用的化合物。
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 10 μM 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 = 4.078 μM) and surprisingly a DAPK1 kinase agonist/activator (EC50 = 39.525 μM). 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.