Identification and Profiling of Environmental Chemicals That Inhibit the TGFβ/SMAD Signaling Pathway

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 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. Counterscreen of SBE antagonists on human embryonic
neural stem cells demonstrated cytotoxicity, providing additional
evidence to support evaluation of 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). The profiling results from SBE-bla assay identify chemicals that disrupt TGFβ/SMAD signaling
as part of an integrated qHTS approach for prioritizing putative developmental
toxicants.