Genomic and Non-Genomic Actions of Estrogen Receptor Ligands

132 p.Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 2005.The physiological activities of estrogen are mediated by its binding to estrogen receptor proteins in target cells. In the nucleus, estrogen receptor alpha (ERalpha) and estrogen receptor beta (ERbeta) serve as ligand activated transcription factors and upon estrogen binding, they can stimulate or repress transcription of estrogen target genes. Classical estrogen receptor and other estrogen binding proteins at the cell membrane can also bind estrogen and rapidly activate intracellular signaling pathways in a ligand-sensitive and transcription independent manner. Here, we report on the characterization of two classes of ligands that should serve as molecular tools for the study of estrogen physiology: estrogen receptor subtype selective ligands and estrogen-dendrimer conjugates (EDCs). The activities of four novel estrogen receptor subtype selective ligands were characterized in this study. The activities of these ligands were examined at a range of promoter contexts where ER worked through a variety of mechanisms. All four ligands tested maintained both their biological activity and subtype-selectivity regardless of promoter context. As such, these ligands should be useful tools in studying the divergent biological roles of ERalpha and ERbeta. Estrogen-dendrimer conjugates are large, abiotic, non-degradable polyamidoamine (PAMAM) dendrimer macromolecules that are conjugated to multiple estrogen molecules through chemically robust linkages. Because of their charge and size, these estrogen-dendrimer conjugates (EDCs) remain outside the nucleus. They stimulate ERK, Shc and Src phosphorylation in MCF-7 breast cancer cells at low concentrations, yet they are very ineffective in stimulating transcription of endogenous estrogen target genes, being ca. 10,000-fold less potent than estradiol in genomic actions. In contrast to estradiol, EDC was not effective in stimulating breast cancer cell proliferation. Because these EDC ligands activate non-genomic activity at concentrations at which they do not alter the transcription of estrogen target genes, they should be useful in studying non-genomic pathways of estrogen action in a variety of target cells.U of I OnlyRestricted to the U of I community idenfinitely during batch ingest of legacy ETD