Panely reportérových linií pro steroidní receptory určené k profilování chemických knihoven a vývoj selektivních ligandů pro estrogenové receptory alfa a beta

Sedlák, David

of the Ph.D. thesis Steroid hormone receptors represent a major target in the drug discovery. As ligand inducible transcription factors, their activity can be modulated by small lipophilic molecules. The first part of this work describes a preparation of two panels of potent, selective and robust luciferase reporter cell lines on the unified cellular background in U2OS osteosarcoma cell line. This system consists of two panels of stable luciferase reporter cell lines for estrogen receptor α (ERα), estrogen receptor β (ERβ), androgen receptor (AR), glucocorticoid receptor (GR), mineralocorticoid receptor (MR) and progesterone receptor (PR). The first panel of reporter cell lines relies on the expression of the chimeric steroid receptors created by the replacement of the N-terminal part of the steroid receptor molecule by Gal4 DNA binding domain (Gal4 DBD) binding to 9 copies of Gal4 upstream activation sequences (Gal4 UAS) in the promoter of the pGL4 luciferase reporter vector. In the second panel of reporter cell lines the activation of either synthetic promoter containing multiple hormone response elements or viral promoter derived from MMTV LTR is mediated by full- length exogenously expressed steroid receptors. We have extensively validated both panels using 28 well established ligands, carefully...

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