Development and Characterization of Novel SERMs

描述（由申请人提供）：本研究的目的是确定激素依赖性组织和癌症中雌激素激动和拮抗作用之间的分子和结构差异，并鉴定、开发和表征具有所需组织选择性雌激素或抗雌激素特性的新型化合物。因为这些作用由两种雌激素受体ER α和ER β中的一种或两种介导，所以了解受体-配体复合物的诱导构象变化和分析新受体配体的行为特征对于开发选择性雌激素受体调节剂（SERM）是必不可少的。因此，该项目的主要目标是将化合物在体外和体内的行为与结合到新型甾体和非甾体SERM的ER α和ER β的晶体结构相关联。这些信息将用于帮助修改或设计具有可预测的、改变的药理学性质的化合物。为了实现这些目标，我们将：1）确定人ER α和ER β蛋白质的详细晶体结构，包括LBD和高达全长受体的额外功能结构域，以及具有独特组织选择性和受体选择性性质的已知和新型SERMs。结晶将通过包含辅助调节肽和/或结晶分子伴侣（Fab和FN 3单体）来辅助，以稳定ER复合物。结构引导的诱变将用于定义功能结构域内和功能结构域之间的重要接触。该信息将用于修饰和/或设计具有改变的药理学性质的配体。2)在体外和肿瘤敏感性动物肿瘤模型中表征新型ER α和/或ER 0选择性配体，特别是发生自发性乳腺肿瘤的SCID小鼠异种移植物和C3（1）/SV 40 T抗原转基因小鼠。为了确定潜在的治疗效用，我们还将通过测量子宫营养反应、胆固醇水平和骨密度来研究候选配体在大鼠和小鼠中的体内药理学。将该行为与转染细胞中雌激素应答启动子的报告基因测定中两种ER的活性进行比较。ER亚型特异性相互作用的表征将有助于鉴定和/或产生对ER a和ER 3起不同作用并具有新的治疗性质的新化合物。详细的结构信息也将有助于揭示这种行为的分子基础。预期衍生自本研究的化合物可应用于治疗和/或预防激素敏感性癌症，特别是乳腺癌，并且还可用作激素替代疗法的药剂。

DESCRIPTION (provided by applicant): The objective of this investigation is to determine the molecular and structural distinctions between estrogen agonism and antagonism in hormone dependent tissues and cancers and to identify, develop and characterize novel compounds that have desired tissue-selective estrogenic or antiestrogenic properties. Because these effects are mediated by one or both of the two estrogen receptors, ERa and ER(J, knowledge of the induced conformational changes of receptor-ligand complexes and analysis of behavior profiles for novel receptor ligands is essential for developing selective estrogen receptor modulators (SERMs). Therefore, a major goal of this project is to correlate compound behavior in vitro and in vivo with the crystallographic structures of ERa and ER(3 bound to novel steroidal and nonsteroidal SERMs. This information will be used to help modify or design compounds with predictable, altered pharmacological properties. To accomplish these goals we will: 1) Determine the detailed crystallographic structures of the human ERa and ER(5 proteins, including the LBDs and additional functional domains up to full-length receptor, in combination with known and novel SERMs with unique tissue-selective and receptor-selective properties. Crystallizations will be assisted by the inclusion of coregulator peptides and/or crystallization chaperones (Fab and FN3 monobody) to stabilize ER complexes. Structure guided mutagenesis will be used to define important contacts within and between functional domains. This information will be used to modify and/or design ligands with altered pharmacological properties. 2) Characterize novel ERa and/or ER0- selective ligands in in vitro and in hormone-sensitive animal tumor models, especially SCID mouse xenografts and C3(1)/SV40 T-antigen transgenic mice that develop spontaneous mammary tumors. To determine potential therapeutic utility, we will also study the in vivo pharmacology of candidate ligands in rats and mice by measuring uterotrophic response, cholesterol levels, and bone density. This behavior will be compared with the activities of both ERs in reporter assays of estrogen responsive promoters in transfected cells. The characterization of ER subtype-specific interactions will facilitate the identification and/or creation of new compounds that act differently on ERa and ER3 and possess novel therapeutic properties. Detailed structure information will also help reveal the molecular basis for such behavior. It is anticipated that compounds derived from this study may have application in the treatment and/or prevention of hormone sensitive cancers, especially breast cancer, and may also have utility as agents for hormone replacement therapy.