Proliferation assays for estrogenicity testing with high predictive value for the in vivo uterotrophic effect

Non-Monotonic Dose Responses in Studies of Endocrine Disrupting Chemicals: Bisphenol a as a Case Study

“…It has also been suggested that NMDRCs are only relevant if they are observed for 'adverse' endpoints. Although this is not directly relevant to the current study, as in vitro endpoints are rarely acknowledged to represent adverse endpoints, there is certainly evidence that effects observed in in vitro studies are relevant to in vivo endpoints (see for example (Wang et al, 2012)), and there have been strong recommendations that results from in vitro studies be considered relevant to effects that are likely to be observed at other levels of biological organization (Schug et al, 2013). Furthermore, the development of Tox21, ToxCast, and other high throughput in vitro assays acknowledges that in vitro cell based assays and cell-free assays provide important tools that can and should inform chemical risk assessments (Knudsen et al, 2013;Mahadevan et al, 2011).…”

A 155-plex high-throughput in vitro coregulator binding assay for (anti-)estrogenicity testing evaluated with 23 reference compounds

“…these IC50 values are also in line with data reported by ICCVAM, i.e., 7.12×10 -7 M and 4.94×10 -9 M for tamoxifen and 4-hydroxytamoxifen, respectively, in the BG1luc eR transcriptional activation assay, demonstrating that the coregulator binding assay also is useful to test anti-estrogenic properties of compounds. Although tamoxifen and 4-hydroxytamoxifen are mainly reported to act as eR antagonists in breast and as eR agonists in uterus tissue (Shang and Brown, 2002), they are also able to inhibit the effect caused by ee2 in the uterotrophic assay and to induce breast cell proliferation in the E-screen (Fang et al, 2000;Yamasaki et al, 2003;Wang et al, 2012). the OMIY-bisphenol shows both agonistic and antagonistic effects in the uterotrophic assay, and when tested in proliferation assays it also behaves as an agonist and antagonist, demonstrating a biological effect profile nearly identical to tamoxifen (Wang et al, 2012).…”

“…Although tamoxifen and 4-hydroxytamoxifen are mainly reported to act as eR antagonists in breast and as eR agonists in uterus tissue (Shang and Brown, 2002), they are also able to inhibit the effect caused by ee2 in the uterotrophic assay and to induce breast cell proliferation in the E-screen (Fang et al, 2000;Yamasaki et al, 2003;Wang et al, 2012). the OMIY-bisphenol shows both agonistic and antagonistic effects in the uterotrophic assay, and when tested in proliferation assays it also behaves as an agonist and antagonist, demonstrating a biological effect profile nearly identical to tamoxifen (Wang et al, 2012). thus, transcriptional activation assays, cell proliferation assays, and the in vivo uterotrophic assay are capable of displaying both the eR agonistic and eR antagonistic properties of tamoxifen, 4-hydroxytamoxifen, and OMIY-bisphenol.…”

“…However, in several studies it was shown that t can induce cell proliferation in MCF-7/BOS cells (e-screen), and it has been demonstrated that this atypical response was mediated by activation of the ER. More specifically, the proliferative response induced by testosterone in the E-screen is partially due to its conversion into 17β-estradiol by aromatase (Wang et al, 2012), partially due to formation of other estrogenic metabolites (Wang et al, 2013), and also partially due to T, i.e., activation of ERα. These findings are in line with the observations in the present study, i.e., t is capable of activating ERα-LBD and induces subsequent binding of several coactivators.…”

“…Several reporter gene assays have been developed and applied as screening tools to determine the estrogenic/ anti-estrogenic activities of compounds, as they are cheap, fast, robust, and have been shown to produce relevant and reliable outcomes van der Burg et al, 2010;Plotan et al, 2012). Proliferation assays and low-density DNA microchipbased analysis of marker gene expression also have been shown to provide valuable tools for estrogenicity testing, and outcomes correlate well with the in vivo uterotrophic assay (Wang et al, 2012(Wang et al, , 2013, but these two assays are laborious and require 3-6 days. therefore, they are not ideal for the large-scale testing of chemicals with respect to initiatives such as ReACH.…”