Molecular Pharmacology of Estrogen Sulfotransferase

描述（由申请人提供）：雌激素磺基转移酶（EST）是一种II期代谢酶，可与类固醇激素雌激素偶联并失活。虽然人们对EST及其相关类固醇磺基转移酶的生物化学、结构生物学和药物遗传学等方面已经有了很多了解，但对EST及其相关类固醇磺基转移酶的分子药理学和体内功能还不完全了解。本提案的目的是验证EST是一种重要的肝外结合酶，调节组织雌激素敏感性，因此在局部雌激素代谢和稳态中起生理作用的假设。本研究的长期目标是了解EST活性的变化如何通过遗传（药物遗传学）或环境因素（药物-药物相互作用）在生理或治疗环境下改变组织雌激素敏感性。在我们的初步研究中，我们已经产生了一只EST敲除（KO）小鼠，将用于验证我们的假设。在本课题中，我们将追求以下具体目的：具体目的1：验证子宫和胎盘EST表达阻止胎儿雌激素毒性的假设。我们的初步发现支持了这一假设，雌性EST KO小鼠的生育能力降低。在缺乏EST的情况下，未代谢的雌激素可能对发育中的胎儿有毒或扰乱分娩过程。使用雌激素受体拮抗剂等药理学方法将应用于本研究。具体目的2：验证EST在维持间质细胞胆固醇稳态中起生理作用的假设。我们发现EST - KO小鼠间质细胞异常地积累中性脂质。我们将确定这些细胞中的异常脂质积累是否由清道夫受体（SRBI）介导，以及它是否依赖于雌激素受体的功能。生物化学、分子生物学和遗传学方法将被应用于解决这个问题。具体目的3：验证EST保护间质细胞免受雌激素诱导的类固醇生成抑制的假设，这种抑制发生在底物动员步骤或P450 17a-羟化酶步骤。具体目的4：验证EST以雄激素依赖的方式在男性生殖道附属结构中表达的假设。特异性目的3和4中的假设都得到了我们对EST KO小鼠的初步研究的支持。本研究将为了解雌激素磺基转移酶作为一种典型的II期类固醇磺基偶联酶的体内药理学和生理学提供分子基础。

DESCRIPTION (provided by applicant): Estrogen sulfotransferase (EST) is a phase II metabolic enzyme that sulfoconjugates and inactivates the steroid hormone estrogen. Although much has been learned on the biochemistry, structural biology and pharmacogenetics of EST and related steroid sulfotransferases, the molecular pharmacology and in vivo function of EST and other steroid sulfotransferases remain incompletely understood. The goal of this proposal is to test the hypothesis that EST is a significant extrahepatic conjugation enzyme that regulates tissue estrogen sensitivity, and as such plays a physiological role in local estrogen metabolism and homeostasis. The long-term objective of this study is to understand how variation in EST activity, through either genetic (pharmacogenetics) or environmental factors (drug-drug interaction), might alter tissue estrogen sensitivity under physiological or therapeutic settings. In our preliminary studies, we have generated an EST knockout (KO) mouse which will be used to test our hypothesis. In this proposal, we will pursue the following specific aims: Specific aim 1: To test the hypothesis that uterine and placental EST expression prevents fetal estrogen toxicity. This hypothesis is supported by our preliminary finding that female EST KO mice have reduced fertility. In the absence of EST, unmetabolized estrogen may become toxic to the developing fetus or disturb the parturition process. Pharmacological approaches such as the use of estrogen receptor antagonists will be applied to this investigation. Specific aim 2: To test the hypothesis that EST plays a physiological role in maintaining cholesterol homeostasis in Leydig cells. We have found that EST KO mouse Leydig cells abnormally accumulate neutral lipids. We will determine if abnormal lipid accumulation in these cells is mediated by the Scavenger Receptor, Class B, Type I (SRBI) and whether it is dependent on estrogen receptor function. Biochemical, molecular biological and genetic approaches will be applied to address this question. Specific aim 3: To test the hypothesis that EST protects Leydig cells from estrogen-induced inhibition of steroidogenesis and that such inhibition occurs either at the substrate mobilization step or at the P450 17a-hydroxylase step. Specific aim 4: To test the hypothesis that EST is expressed in the accessory structures of the male reproductive tract in an androgen-dependent manner. Both hypotheses in Specific aim 3 and 4 are supported by our preliminary study of the EST KO mice. The proposed studies will provide a molecular basis for understanding the in vivo pharmacology and physiology of estrogen sulfotransferase as a prototypical phase II steroid sulfoconjugation enzyme.