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Evaluation of a Multiplexed, Multispecies Nuclear Receptor Assay for Chemical Hazard Assessment
Citation:
Houck, K., A. Simha, A. Bone, J. Doering, S. Vliet, C. LaLone, A. Medvedev, AND S. Makarov. Evaluation of a Multiplexed, Multispecies Nuclear Receptor Assay for Chemical Hazard Assessment. TOXICOLOGY IN VITRO. Elsevier Science Ltd, New York, NY, 72:105016, (2021). https://doi.org/10.1016/j.tiv.2020.105016
Impact/Purpose:
Substantial progress has been made in developing approaches to identify xenobiotic chemicals that bind steroid hormone nuclear receptors with potential for adverse health effects through disruption of endocrine function. Of concern for human and environmental health are chemicals which activate estrogen receptor or inhibit androgen receptor pathways leading to adverse effects on individuals and populations. In addition, other nuclear receptors are recognized as having potential for modulation by xenobiotic chemicals affecting human and environmental health. Both in vivo and in vitro methods have been established and validated for identifying chemicals targeting these receptors. The in vitro assays are largely human cell line-based while the in vivo studies are primarily rodent-based and used as surrogates for understanding human health effects. However, environmental exposure scenarios encompass a diversity of other species with potential for adverse effects to varied populations. Developing assays and testing for effects for each species individually would be prohibitively expensive and time-consuming and thus new, more efficient approaches are needed. A new approach method, Sequence Alignment to Predict Across Species Susceptibility (SeqAPASS; seqapass.epa.gov/seqapass/), compares sequence similarity of receptors across hundreds of species as a way of evaluating conservation of protein targets and therefore predicting likelihood of chemical perturbations using only in silico methods. The underlying comparative genomics basis of this approach relies on gathering lines of evidence for conservation of the protein comparing a known sensitive or targeted species, in this case human, to all other species with sequence information available. The evaluation in SeqAPASS considers primary amino acid sequences, functional domains and critical amino acid residues involved in chemical protein interactions. The SeqAPASS tool is of obvious intuitive value for chemical prioritization purposes and screening level approaches, however it is desirable to develop testing strategies to fully understand how to apply the SeqAPASS output.
Description:
Sensitivity to potential endocrine disrupting chemicals in the environment varies across species and is influenced by sequence conservation of their nuclear receptor targets. Here, we evaluated a multiplexed, in vitro assay testing receptors relevant to endocrine and metabolic disruption from five species. The TRANS-FACTORIAL™ system of human nuclear receptors was modified to include additional species: mouse (Mus musculus), frog (Xenopus laevis), zebrafish (Danio rerio), chicken (Gallus gallus), and turtle (Chrysemys picta). Receptors regulating endocrine function and xenobiotic recognition were included, specifically: ERα, ERβ, AR, TRα, TRβ, PPARγ and PXR. The assay, ECOTOX-FACTORIAL™, was evaluated with 191 chemicals enriched with known receptor ligands. Hierarchical clustering of potency values demonstrated strong coherence of receptor families. Interspecies comparisons of responses within a receptor family showed moderate to high concordance for potencies under 50¿μM. PPARγ showed high concordance between mammalian species, 89%, but only 63% between mammalian and zebrafish. For chemicals with potencies below 1¿μM, concordances were 89–100% for all receptors except PXR. Concordance showed a strong positive relationship to ligand-binding domain sequence similarity and critical amino acid residues obtained by the Sequence Alignment to Predict Across Species Susceptibility (SeqAPASS) tool. In combination with SeqAPASS, ECOTOX-FACTORIAL may provide efficient screening of important receptors to identify species of high priority for effects monitoring.
URLs/Downloads:
DOI: Evaluation of a Multiplexed, Multispecies Nuclear Receptor Assay for Chemical Hazard Assessment
https://pubmed.ncbi.nlm.nih.gov/33049310/