A Forward Chemical Screen in Zebrafish Identifies a Retinoic Acid Derivative with Receptor Specificity

Abstract

BackgroundRetinoids regulate key developmental pathways throughout life, and have potential uses for differentiation therapy. It should be possible to identify novel retinoids by coupling new chemical reactions with screens using the zebrafish embryonic model.Principal FindingsWe synthesized novel retinoid analogues and derivatives by amide coupling, obtaining 80–92% yields. A small library of these compounds was screened for bioactivity in living zebrafish embryos. We found that several structurally related compounds significantly affect development. Distinct phenotypes are generated depending on time of exposure, and we characterize one compound (BT10) that produces specific cardiovascular defects when added 1 day post fertilization. When compared to retinoic acid (ATRA), BT10 shows similar but not identical changes in the expression pattern of embryonic genes that are known targets of the retinoid pathway. Reporter assays determined that BT10 interacts with all three RAR receptor sub-types, but has no activity for RXR receptors, at all concentrations tested.ConclusionsOur screen has identified a novel retinoid with specificity for retinoid receptors. This lead compound may be useful for manipulating components of retinoid signaling networks, and may be further derivatized for enhanced activity.

Chemical screening in zebrafish for novel biological and therapeutic discovery

“…Less frequently conducted are nonspecific screens that score any morphological change observed. These have been carried out to determine compound bioactivity in broad terms; the readouts being any observable perturbation of development(Das, McCartin, Liu, Peterson, & Evans, 2010; Jung et al, 2005; Khersonsky et al, 2003; Moon et al, 2002; Peterson, Link, Dowling, & Schreiber, 2000; Sachidanandan, Yeh, Peterson, & Peterson, 2008; Spring, Krishnan, Blackwell, & Schreiber, 2002; Sternson, Louca, Wong, & Schreiber, 2001; Torregroza, Evans, & Das, 2009; Wong, Sternson, Louca, Hong, & Schreiber, 2004). Often, in-house synthetic libraries containing one specific pharmacophore are screened on wild-type zebrafish embryos.…”

“…An example of such a study is a zebrafish screen carried out with novel retinoid analogues that discovered a novel retinoid with retinoid receptor specificity. This lead compound is useful for probing the biology of the retinoic acid signaling pathways (Das et al, 2010). A second aspect of a synthetic library is that compounds can be designed with tags, allowing for target ID through protein pull-down.…”

15 years of zebrafish chemical screening

“…The same assay used to conduct the initial zebrafish chemical screen can also be used to quickly make conclusions about the effects of structural modifications on chemical activity. There are now many examples of SAR studies using zebrafish assays [10–12, 14, 35, 36, 48, 52–54]. In vivo SAR has advantages over in vitro SAR.…”

Zebrafish—As an integrative model for twenty‐first century toxicity testing

“…On one hand, the amenability of the system to screening has enabled fast and efficient assessment of large chemical libraries and the identification of compounds with specific activity on key biological pathways (Peal et al, 2010;Taylor et al, 2010). For example, when screened in zebrafish embryos, compounds designed to modulate transforming growth factor beta (TGFb) (Torregroza et al, 2009) and mimic retinoic acids (Das et al, 2010) elicited a variety of defects similarly seen in mammalian models. Subsequent analyses confirmed inhibition of TGFb-regulated pathways and interaction of retinoic acid receptors (RARs).…”