Ligand Pharmacology of Estrogen Receptors

描述(申请人提供)：该项目的长期目标是阐明非基因组雌激素信号的分子机制，并创造新的配体，可以选择性地激活或阻止非基因组信号而不是基因组雌激素信号。雌激素基因组信号涉及核雌激素受体(ERA和ERB)的激活以及随后雌激素靶基因的转录调控。另一方面，雌激素的非基因组信号导致离子通道、酶和第二信使信号的快速激活。雌激素的许多非基因组效应发生在中脑，可能与绝经后女性面临的体温调节和情绪不稳定问题有关。这些反应的快速动力学和对转录/翻译抑制剂的不敏感性排除了核内ERs参与经典基因组机制的可能性；然而，非基因组信号的新的细胞机制尚不清楚。这项研究计划是围绕这样一种假设构建的，即一种新的雌激素膜相关G蛋白偶联受体(GPCR)至少介导了一些非基因组激素信号。支持这一假说的证据是，雌激素反应的GPCR迅速介导了下丘脑神经元中特定钾通道的抑制。此外，这种生理上相关的雌激素反应也是由一种新型的选择性雌激素受体调节剂(SERM)引起的，它与Era或ERB都没有结合亲和力。在特定目的1中，确定了一个孤立的GPCRa作为雌激素的候选受体，并建立了稳定表达该GPCRs的细胞系。在特定的目标2和3中，开发并进行了配体结合和配体激活分析，以建立和表征GPCRs的雌激素反应性。有了这些工具，新的SERM将被开发出来，激活(特定目标4)或阻止激活(特定目标5)雌激素反应GPCRs。本研究旨在确定快速、非基因组雌激素信号的分子机制和配体激活参数，从而为治疗更年期症状提供急需的安全疗法。

DESCRIPTION (provided by applicant): The long-term objectives of this project are to elucidate the molecular mechanism of non-genomic estrogen signaling and create novel ligands that can either activate or block non-genomic signaling selectively over genomic estrogen signaling. Estrogen genomic signaling involves activation of the nuclear estrogen receptors (ERa and ERB) and subsequent transcriptional regulation of estrogen target genes. On the other hand, nongenomic signaling by estrogen results in rapid activation of ion channels, enzymes, and second messenger signaling cascades. Many of these non-genomic effects of estrogen occur in the mid-brain and may be linked to the problems of thermal regulation and mood instability that post-menopausal women face. The rapid kinetics and insensitivity of these responses to transcription/translation inhibitors rules out the involvement of the nuclear ERs acting by the classical genomic mechanism; however, the novel cellular mechanism of nongenomic signaling is unclear. This research plan is constructed around the hypothesis that a novel membrane associated G protein-coupled receptor (GPCR) for estrogen mediates at least some of the non-genomic hormone signaling. Support for this hypothesis comes from evidence that an estrogen-responsive GPCR rapidly mediates the inhibition of a specific potassium channel in hypothalamic neurons. Moreover, this physiologically relevant estrogen response is also elicited by a novel selective estrogen receptor modulator (SERM) that has no binding affinity for either ERa or ERB. In specific aim 1, an orphan GPCR is identified as a candidate receptor for estrogen and a stable cell line expressing this GPCR is developed. In specific aims 2 and 3, ligand binding and ligand activation assays are developed and carried out to establish and characterize the estrogen responsiveness of the GPCR. With these tools in place, novel SERMs will be developed that either activate (specific aim 4) or block activation (specific aim 5) of the estrogen-responsive GPCR. This research aims to define the molecular mechanisms and ligand-activation parameters of rapid, non-genomic estrogen signaling which could lead to much needed safe therapeutics for treating the symptoms of menopause.