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Comparison of cell type specificities of stress pathway reporter assay ensemble response to environmental chemicals.
Citation:
Simmons, S. AND T. V. RAMABHADRAN. Comparison of cell type specificities of stress pathway reporter assay ensemble response to environmental chemicals. Presented at Society of Toxicology, Seattle, WA, March 16 - 20, 2008.
Impact/Purpose:
We will present the results comparing the fingerprints of sets of related compounds, such as metals and persistent chemicals, in human cell lines of different tissue origins to evaluate the utility of the ensemble approach as a predictor of toxicity that may also permit the inference of the mechanistic basis of toxicity.
Description:
The large number of environmental compounds that currently need characterization and prioritization for further toxicological study is a serious regulatory challenge facing the EPA. In addition to these agents comprising of pesticides, inerts, and high-production volume chemicals, new chemicals and novel materials of unknown toxicities such as nanoparticles are being introduced into the market each year. To meet this challenge, we are developing cell-based methods for the rapid, cost-efficient and high throughput assessment of toxicities of large numbers of environmental chemicals using in vitro reporter gene assays that measure the activation of key molecular stress pathways. The major molecular stress pathways are highly-conserved, well-characterized signaling pathways, triggered in response to environmental insult at doses significantly lower than those producing apical endpoints such as apoptosis. We have constructed and characterized an ensemble of reporter gene assays using promoter regions from canonical stress-responsive genes such as heme oxygenase I (HO) and heat shock protein 70 (hsp70) as well as multimerized synthetic elements that measure activation of stress-response factors such as Nuclear Factor kappa B (NFkB) and Nuclear Factor E2-related factor (Nrf2). The response of this assay ensemble is being used to obtain ‘toxicological fingerprints’ of compounds. To enable the delivery of reporters into cells of choice, these reporters were packaged into lentiviral vectors that allow for the rapid and efficient generation of stable reporter cell lines. We will present the results comparing the fingerprints of sets of related compounds, such as metals and persistent chemicals, in human cell lines of different tissue origins to evaluate the utility of the ensemble approach as a predictor of toxicity that may also permit the inference of the mechanistic basis of toxicity.