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A gene expression biomarker for predictive toxicology to identify chemical modulators of NF-κB
Citation:
Korunes, K., J. Liu, R. Huang, M. Xia, K. Houck, AND C. Corton. A gene expression biomarker for predictive toxicology to identify chemical modulators of NF-κB. PLOS ONE . Public Library of Science, San Francisco, CA, 17(2):e0261854, (2022). https://doi.org/10.1371/journal.pone.0261854
Impact/Purpose:
Dysfunction of NF-κB plays a role in oncogenesis by inhibiting apoptosis, stimulating cell proliferation, and affecting inflammation and immunity in ways that create favorable environments for cancer (1). Misregulation of the NF-κB pathway is also linked to inflammatory diseases such as rheumatoid arthritis and asthma (2–4). Known environmental factors that are linked to NF-κB activation include cigarette smoke, nanoparticles, asbestos, and lead (6–9). A high-throughput screen for NF-κB chemical activators was carried out as part of the Environmental Protection Agency (EPA) ToxCast screening program (https://www.epa.gov/chemical-research/toxicity-forecasting) which encompasses ~700 HTS Tier 1 assays representing ~350 molecular targets that have been used to screen more than 3800 chemicals (10). Another HTS for NF-κB activators was carried out by the National Center for Advancing Translational Sciences (NCATS) as part of the Tox21 screening program (https://tripod.nih.gov/tox21/assays/), an ongoing effort to test the effects of over 10,000 compounds on nuclear receptors, stress response pathways, developmental pathways, and other cellular processes. These high-throughput screens pave the way for understanding the effects of environmentally relevant chemicals on NF-κB. In addition, a screen for NF-κB inhibitors was carried out on ~2800 clinically approved drugs and bioactive compounds from the NIH Chemical Genomics Center Pharmaceutical Collection (NPC) 21525397 in a NF-κB mediated beta-lactamase reporter gene assay (11) and demonstrated that many currently approved pharmaceuticals have previously unappreciated NF-κB signaling suppression activity. High-throughput transcriptomic (HTTr) technologies are being increasingly used to screen chemicals in human cell lines. In the EPA ToxCast screening program, HTTr strategies are now being used to replace the battery of individual ToxCast screening assays with targeted sequencing techniques (15) such as TempO-Seq (16). Compared to individual assays, HTTr technologies have the advantage of examining the effects of environmental chemicals on essentially all pathways operating in cell models, many of which are not examined by the current battery of ToxCast assays. A major challenge is how to interpret the gene expression profiles to identify the molecular targets of chemicals. A number of approaches have been used to interpret the HTTr profiles and these include pathway analysis and comparison to archived profiles of reference chemicals (17). Gene expression biomarkers have emerged as an alternative approach to accurately predict specific targets of chemicals. Biomarkers consist of sets of genes known or predicted to be regulated by a particular factor (18). The biomarker gene expression pattern is compared to gene expression profiles derived from human cells exposed to chemicals using a number of computational techniques that include correlation analysis (19). In the present study, we developed procedures for predicting NF-κB perturbation using HTTr data. We constructed a gene expression biomarker that accurately predicts NF-κB modulation after exposure to immunomodulatory factors and chemicals. We used the biomarker to screen a library of microarray profiles from cells treated with ~2600 organic chemicals to identify modulators of NF-κB. In addition, we carried out two HTS assays to identify NF-κB activators in large chemical sets of drugs and environmentally relevant chemicals and determined the overlap with those identified using the biomarker approach.
Description:
The nuclear factor-kappa B (NF-κB) is a transcription factor with important roles in inflammation, immune response, and oncogenesis. Dysregulation of NF-κB signaling is associated with inflammation and certain cancers. We developed a gene expression biomarker predictive of NF-κB modulation and used the biomarker to screen a large compendia of gene expression data. The biomarker consists of 108 genes responsive to tumor necrosis factor α in the absence but not the presence of IκB, an inhibitor of NF-κB. Using a set of 450 profiles from cells treated with immunomodulatory factors with known NF-κB activity, the balanced accuracy for prediction of NF-κB activation was > 90%. The biomarker was used to screen a microarray compendium consisting of 12,061 microarray comparisons from human cells exposed to 2,672 individual chemicals to identify chemicals that could cause toxic effects through NF-κB. There were 215 and 49 chemicals that were identified as putative or known NF-κB activators or suppressors, respectively. NF-κB activators were also identified using two high-throughput screening assays; 165 out of the ~3,800 chemicals (ToxCast assay) and 55 out of ~7,500 unique compounds (Tox21 assay) were identified as potential activators. A set of 32 chemicals not previously associated with NF-κB activation and which partially overlapped between the different screens were selected for validation in wild-type and NFKB1-null HeLa cells. Using RT-qPCR and targeted RNA-Seq, 31 of the 32 chemicals were confirmed to be NF-κB activators. These results comprehensively identify a set of chemicals that could cause toxic effects through NF-κB.
URLs/Downloads:
DOI: A gene expression biomarker for predictive toxicology to identify chemical modulators of NF-κB
https://pubmed.ncbi.nlm.nih.gov/35108274/