Dataset Information

AroER tri-screen™ is a novel functional assay to estimate both estrogenic and estrogen precursor activity of chemicals or biological specimens.

ABSTRACT: The purpose of the study is to define AroER tri-screen's utility for identifying endocrine-disrupting chemicals (EDCs) that target aromatase and/or estrogen receptor (ER), and to measure the total estrogenic activity in biological specimens. ER-positive, aromatase-expressing MCF-7 breast cancer cells were stably transfected with an estrogen responsive element (ERE)-driven luciferase reporter plasmid to yield a new high-throughput screening platform-the AroER tri-screen. AroER tri-screen was capable of identifying estrogen precursors, such as cortodoxone, which function as estrogens through a two-step conversion process in aromatase-expressing tissue. Furthermore, the system proved useful for assessing EDC activity in biologically relevant samples. Estimating these activities is critical because natural estrogens and estrogenic EDCs are important factors in ER-positive breast cancer risk. As our research demonstrates, incorporating functionally active aromatase into the AroER tri-screen produces a powerful and unique tool to (1) identify new EDCs targeting aromatase and/or ER; (2) discover novel EDCs activated by aromatase; and (3) estimate overall estrogenic activities in biological samples as a potential intermediate risk factor for breast cancer.

SUBMITTER: Kanaya N

PROVIDER: S-EPMC4591046 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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AroER tri-screen™ is a novel functional assay to estimate both estrogenic and estrogen precursor activity of chemicals or biological specimens.

Kanaya Noriko N Nguyen Duc M DM Lu Hannah H Wang Yuan-Zhong YZ Hsin Li-Yu LY Petreas Myrto M Nelson David D Guo Weihong W Reynolds Peggy P Synold Tim T Chen Shiuan S

Breast cancer research and treatment 20150512 2

The purpose of the study is to define AroER tri-screen's utility for identifying endocrine-disrupting chemicals (EDCs) that target aromatase and/or estrogen receptor (ER), and to measure the total estrogenic activity in biological specimens. ER-positive, aromatase-expressing MCF-7 breast cancer cells were stably transfected with an estrogen responsive element (ERE)-driven luciferase reporter plasmid to yield a new high-throughput screening platform-the AroER tri-screen. AroER tri-screen was capa ...[more]

PMID: 25962693

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Project description:Estrogen receptor orthologues in molluscs may be targets for endocrine disruptors, although mechanistic evidence is lacking. Molluscs are reported to be highly susceptible to effects caused by very low concentrations of environmental estrogens which, if substantiated, would have a major impact on the risk assessment of many chemicals. The present paper describes the most thorough evaluation to-date of the susceptibility of Marisa cornuarietis ER and ERR gene transcription to modulation by vertebrate estrogens in vivo and in vitro. We investigated the effects of estradiol-17? and 4-tert-Octylphenol exposure on in vivo estrogen receptor (ER) and estrogen-related receptor (ERR) gene transcription in the reproductive and neural tissues of the gastropod snail M. cornuarietis over a 12-week period. There was no significant effect (p>0.05) of treatment on gene transcription levels between exposed and non-exposed snails. Absence of a direct interaction of estradiol-17? and 4-tert-Octylphenol with mollusc ER and ERR protein was also supported by in vitro studies in transfected HEK-293 cells. Additional in vitro studies with a selection of other potential ligands (including methyl-testosterone, 17?-ethinylestradiol, 4-hydroxytamoxifen, diethylstilbestrol, cyproterone acetate and ICI182780) showed no interaction when tested using this assay. In repeated in vitro tests, however, genistein (with mcER-like) and bisphenol-A (with mcERR) increased reporter gene expression at high concentrations only (>10(-6)M for Gen and >10(-5)M for BPA, respectively). Like vertebrate estrogen receptors, the mollusc ER protein bound to the consensus vertebrate estrogen-response element (ERE). Together, these data provide no substantial evidence that mcER-like and mcERR activation and transcript levels in tissues are modulated by the vertebrate estrogen estradiol-17? or 4-tert-Octylphenol in vivo, or that other ligands of vertebrate ERs and ERRs (with the possible exception of genistein and bisphenol A, respectively) would do otherwise.

Project description:A growing body of experimental evidence indicates that the in vitro effects of mixtures of estrogenic chemicals can be well predicted from the estrogenicity of their components by the concentration addition (CA) concept. However, some studies have observed small deviations from CA. Factors affecting the presence or observation of deviations could include: the type of chemical tested; number of mixture components; mixture design; and assay choice. We designed mixture experiments that address these factors, using mixtures with high numbers of components, chemicals from diverse chemical groups, assays with different in vitro endpoints and different mixture designs and ratios. Firstly, the effects of mixtures composed of up to 17 estrogenic chemicals were examined using estrogenicity assays with reporter-gene (ERLUX) and cell proliferation (ESCREEN) endpoints. Two mixture designs were used: 1) a 'balanced' design with components present in proportion to a common effect concentration (e.g. an EC(10)) and 2) a 'non-balanced' design with components in proportion to potential human tissue concentrations. Secondly, the individual and simultaneous ability of 16 potential modulator chemicals (each with minimal estrogenicity) to influence the assay outcome produced by a reference mixture of estrogenic chemicals was examined. Test chemicals included plasticizers, phthalates, metals, PCBs, phytoestrogens, PAHs, heterocyclic amines, antioxidants, UV filters, musks, PBDEs and parabens. In all the scenarios tested, the CA concept provided a good prediction of mixture effects. Modulation studies revealed that chemicals possessing minimal estrogenicity themselves could reduce (negatively modulate) the effect of a mixture of estrogenic chemicals. Whether the type of modulation we observed occurs in practice most likely depends on the chemical concentrations involved, and better information is required on likely human tissue concentrations of estrogens and of potential modulators. Successful prediction of the effects of diverse chemical combinations might be more likely if chemical profiling included consideration of effect modulation.

Dataset Information

AroER tri-screen™ is a novel functional assay to estimate both estrogenic and estrogen precursor activity of chemicals or biological specimens.

Publications

AroER tri-screen™ is a novel functional assay to estimate both estrogenic and estrogen precursor activity of chemicals or biological specimens.

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