The assessment of estrogenic or anti-estrogenic activity of chemicals by the human stably transfected estrogen sensitive MELN cell line: Results of test performance and transferability

• Published in: Reproductive toxicology (Elmsford, N.Y.) Vol. 30; no. 1; pp. 60 - 72
• Main Authors: Witters, H., Freyberger, A., Smits, K., Vangenechten, C., Lofink, W., Weimer, M., Bremer, S., Ahr, P.H.-J., Berckmans, P.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01-08-2010
• Subjects: Agonist; Animal Testing Alternatives; Antagonist; Biological Assay - methods; Biological Assay - standards; Cell Culture Techniques; Cell Line, Tumor; Cell Survival - drug effects; Cytotoxicity; Dose-Response Relationship, Drug; Estrogen activity; Estrogen Antagonists - chemistry; Estrogen Antagonists - pharmacology; Estrogen Receptor alpha - agonists; Estrogen Receptor alpha - antagonists & inhibitors; Estrogen Receptor alpha - genetics; Estrogen Receptor beta - agonists; Estrogen Receptor beta - antagonists & inhibitors; Estrogen Receptor beta - genetics; hER transactivation assay; Humans; Luciferases - genetics; MELN cell line; Protein Binding; Reproducibility of Results; Transfection; Validation; Validation; Agonist; MELN cell line; hER transactivation assay; Estrogen activity; Cytotoxicity; Antagonist
• ISSN: 0890-6238; 1873-1708
• DOI: 10.1016/j.reprotox.2010.02.008

The need for development and validation of
in vitro hormone receptor transactivation assays as important alternative tools to study interactions with sex hormone receptors is outlined by international organisations, as such assays should be included in the OECD conceptual framework for the testing and assessment of endocrine active chemicals. Therefore as part of the European Union (EU)-sponsored 6th framework project ReProTect, the validation study with MELN cells, MCF-7 cells (ER+, estrogen receptor positive) which were stably transfected with the estrogen responsive gene ERE-βGlob-Luc-SVNeo was set up. Standard operating procedures including a prescreen assay for unknown chemicals, an ER-agonist assay and an ER-antagonist assay were developed at the Flemish Institute for Technological Research, Belgium, and successfully transferred to Bayer Schering Pharma AG, Germany. Test results were obtained for 16 chemicals, and it was demonstrated that the MELN assay is transferable, robust and reproducible which allowed to rank chemical compounds according to their strong to weak affinity for the estrogen-α receptor, or identify negative chemicals within the test range up to 10
−5
M. Besides the screening for agonism, we demonstrated the suitability of MELN cells to test for antagonistic activity, which is of added value compared to current validated assays. As the MELN assay successfully passed the first modules of the ECVAM validation procedure, it now should be considered for further steps including the definition of a prediction model and application domain to get it accepted as an alternative screening assay, contributing to the 3R's with a reduction of animal experiments.