Citation:

Wei, Z., S. Sakamuru, L. Zhang, J. Zhao, R. Huang, N. Kleinstreuer, Y. Chen, Y. Shu, T. Knudsen, AND M. Xia. Identification and profiling of environmental chemicals that inhibit the TGFβ/SMAD signaling pathway. CHEMICAL RESEARCH IN TOXICOLOGY. American Chemical Society, Washington, DC, 32(12):2433-2444, (2019). https://doi.org/10.1021/acs.chemrestox.9b00228

Impact/Purpose:

Current prenatal toxicity testing guidelines monitor the potential action of toxicants at many levels of normal development and assess disruption of overall processes of developmental, reproductive, and teratogenic effects of pharmaceuticals, food additives, and pesticides.67−69 Faster, inexpensive high-throughput in vitro assays, such as the SBE-bla assay, could be used as a quantitative high-throughput (qHTS) screen to identify chemicals for developmental toxicity assessment.

Description:

The transforming growth factor beta (TGFβ) superfamily of secreted signaling molecules and their cognate receptors regulate cell fate and behaviors relevant to many developmental and disease processes. Disruption of TGFβ signaling during embryonic development can, for example, affect morphogenesis and differentiation through complex pathways that may be SMAD (Small Mothers Against Decapentaplegic) -dependent or SMAD-independent. In the present study, the SMAD Binding Element (SBE)-beta lactamase (bla) HEK 293T cell line, which specifically responds to the activation of the SMAD2/3/4 complex, was used in a quantitative high-throughput screening (qHTS) assay to identify potential TGFβ disruptors in the Tox21 10K compound library. From the primary screening, we identified several kinase inhibitors, organometallic compounds, and dithiocarbamates (DTCs) that inhibited TGFβ1-induced SMAD signaling of reporter gene activation independent of cytotoxicity. Counter-screen of SBE antagonists on human embryonic neural stem cells demonstrated cytotoxicity, providing additional evidence to further test these compounds for developmental toxicity. We profiled the inhibitory patterns of putative SBE antagonists toward other developmental signaling pathways, including wingless-related integration site (WNT), retinoic acid α receptor (RAR) and sonic hedgehog (SHH). These results show that the SBE-bla assay is an effective qHTS approach for identifying chemicals that disrupt TGFβ/SMAD signaling.

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