The physiologic effects of estrogen action through the estrogen receptor (ER) are widespread, as this hormone exerts actions in both reproductive (e.g., uterus) and non-reproductive (e.g., bone, brain) tissues in both men and women. As such, the regulation of the activity of this ligand-activated transcription factor is highly relevant to the maintenance of normal physiology, and its disregulation can lead to various disease states, including one of the most deadly and well-recognized, breast cancer. As with many pathologies, early diagnosis provides the patient with much better chances of survival; however, once a diagnosis has been made, treatments that specifically target the molecular initiation of the disregulated cellular proliferation result in the greatest attenuation of tumor progression. As not all women have estrogen-receptor positive breast cancer, both the presence of ER and the functionality of the receptor at the start of treatment must be evaluated. With this information, ER positive breast cancers have conventionally been treated with either antagonist ligands for the receptor that shut off ER action or antagonists of the aromatase enzyme that produces the endogenous activating ligand estradiol. Unfortunately, within months to years, all of these tumors will eventually develop resistance to current treatments, making the discovery of novel methods of ER inhibition vital to continued success in control of breast cancer through targeted endocrine therapies. Herein, we first discuss a novel approach for the inhibition of the estrogen receptor, the use of small molecules to interrupt the protein-protein interaction between the ER and the primary coactivator in a cascade leading to the upregulation of many genes driving cellular proliferation. This "direct" inhibition will be contrasted with current methods whereby ER action is turned off via the "indirect" action of antagonist ligands. We have approached the identification and biological evaluation of these compounds from several different standpoints. In Chapter 2, we apply an assay capable of identifying these direct inhibitors of ER/coactivator binding that we have termed Coactivator Binding Inhibitors (CBIs). Through a collaboration with the Molecular Libraries Screening Center at Emory University, this assay was optimized and utilized in the screening of over 86,000 compounds in a search for potent compounds that work through this novel mechanism. Chapter 3 presents the work of various synthetic chemists in our laboratory who have made libraries of CBIs based on various structural scaffolds acquired by either directed design or high-throughput screening approaches. The structure-activity relationship of these compounds is explored through both "in vitro" and cell-based assays of ER action. Chapter 4 carries these studies further and examines the most potent compounds from Chapter 3 in more complete biological systems. These experiments provide information regarding CBI activity in a more complete model of breast cancer. The compounds are also evaluated on other nuclear hormone receptors and, most thoroughly, on the androgen receptor, the one nuclear hormone receptor known to bind coactivators through an interaction sequence comprising larger, hydrophobic amino acid side chains. Due to known difficulties with interrupting protein-protein interactions with small molecules, we attempted to tether both CBIs and ER ligands to a recruiting element that would bind a cellular chaperone protein that might provide a greater steric force more effective in preventing coactivator binding. These results will be discussed in Chapter 5. Finally, the determination of the ER status of a tumor is invaluable in choosing most effective treatment for a breast cancer patient. In Chapter 6 we study the protein product of a highly estrogen-regulated gene, carbonic anhydrase XII and examine the potential for this enzyme to act as an endogenous reporter of estrogen receptor action using imageable inhibitors of this protein. [The dissertation citations contained here are published with the permission of ProQuest LLC. Further reproduction is prohibited without permission. Copies of dissertations may be obtained by Telephone (800) 1-800-521-0600. Web page: http://www.proquest.com/en-US/products/dissertations/individuals.shtml.]