title: Cell-Based High-Throughput Screening for Aromatase Inhibitors in the Tox21 10K Library.
abstract: Multiple mechanisms exist for endocrine disruption; one nonreceptor-mediated mechanism is via effects on aromatase, an enzyme critical for maintaining the normal in vivo balance of androgens and estrogens. We adapted the AroER tri-screen 96-well assay to 1536-well format to identify potential aromatase inhibitors (AIs) in the U.S. Tox21 10K compound library. In this assay, screening with compound alone identifies estrogen receptor alpha (ERα) agonists, screening in the presence of testosterone (T) identifies AIs and/or ERα antagonists, and screening in the presence of 17β-estradiol (E2) identifies ERα antagonists. Screening the Tox-21 library in the presence of T resulted in finding 302 potential AIs. These compounds, along with 31 known AI actives and inactives, were rescreened using all 3 assay formats. Of the 333 compounds tested, 113 (34%; 63 actives, 50 marginal actives) were considered to be potential AIs independent of cytotoxicity and ER antagonism activity. Structure-activity analysis suggested the presence of both conventional (eg, 1, 2, 4, - triazole class) and novel AI structures. Due to their novel structures, 14 of the 63 potential AI actives, including both drugs and fungicides, were selected for confirmation in the biochemical tritiated water-release aromatase assay. Ten compounds were active in the assay; the remaining 4 were only active in high-throughput screen assay, but with low efficacy. To further characterize these 10 novel AIs, we investigated their binding characteristics. The AroER tri-screen, in high-throughput format, accurately and efficiently identified chemicals in a large and diverse chemical library that selectively interact with aromatase.
authors.affiliation: *Department of Cancer Biology, Beckman Research Institute of the City of Hope, Duarte, California 91010; schen@coh.org.
authors.forename: Shiuan
authors.initials: S
authors.lastname: Chen
authors.affiliation: Division of the National Toxicology Program, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709;
authors.forename: Jui-Hua
authors.initials: JH
authors.lastname: Hsieh
authors.affiliation: National Center for Advancing Translational Sciences, National Institutes of Health, Rockville, Maryland 20850; and.
authors.forename: Ruili
authors.initials: R
authors.lastname: Huang
authors.affiliation: National Center for Advancing Translational Sciences, National Institutes of Health, Rockville, Maryland 20850; and.
authors.forename: Srilatha
authors.initials: S
authors.lastname: Sakamuru
authors.affiliation: *Department of Cancer Biology, Beckman Research Institute of the City of Hope, Duarte, California 91010;
authors.forename: Li-Yu
authors.initials: LY
authors.lastname: Hsin
authors.affiliation: National Center for Advancing Translational Sciences, National Institutes of Health, Rockville, Maryland 20850; and.
authors.forename: Menghang
authors.initials: M
authors.lastname: Xia
authors.affiliation: Biostatistics and Computational Biology Branch, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709.
authors.forename: Keith R
authors.initials: KR
authors.lastname: Shockley
authors.affiliation: Division of the National Toxicology Program, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709;
authors.forename: Scott
authors.initials: S
authors.lastname: Auerbach
authors.affiliation: *Department of Cancer Biology, Beckman Research Institute of the City of Hope, Duarte, California 91010;
authors.forename: Noriko
authors.initials: N
authors.lastname: Kanaya
authors.affiliation: *Department of Cancer Biology, Beckman Research Institute of the City of Hope, Duarte, California 91010;
authors.forename: Hannah
authors.initials: H
authors.lastname: Lu
authors.affiliation: Division of the National Toxicology Program, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709;
authors.forename: Daniel
authors.initials: D
authors.lastname: Svoboda
authors.affiliation: Division of the National Toxicology Program, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709;
authors.forename: Kristine L
authors.initials: KL
authors.lastname: Witt
authors.affiliation: Division of the National Toxicology Program, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709;
authors.forename: B Alex
authors.initials: BA
authors.lastname: Merrick
authors.affiliation: Division of the National Toxicology Program, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709;
authors.forename: Christina T
authors.initials: CT
authors.lastname: Teng
authors.affiliation: Division of the National Toxicology Program, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709;
authors.forename: Raymond R
authors.initials: RR
authors.lastname: Tice
mesh_terms: D000818:Animals; D047072:Aromatase Inhibitors / Q000633:toxicity*; D004958:Estradiol / Q000494:pharmacology; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D061986:MCF-7 Cells / Q000187:drug effects; D054852:Small Molecule Libraries / Q000633:toxicity*; D013329:Structure-Activity Relationship; D013739:Testosterone / Q000494:pharmacology
chemical_list: D047072:Aromatase Inhibitors; D054852:Small Molecule Libraries; D013739:Testosterone; D004958:Estradiol
keywords: AroER tri-screen; Tox21 10K library; aromatase enzyme assay; environmental chemicals; quantitative high throughput screening