Botanical Modulation of AhR-ERÎ± by Crosstalk Inhibitors Promotes Estrogen (E2) Detoxification

串扰抑制剂对 AhR-ERα 的植物调节促进雌激素 (E2) 解毒

基本信息

批准号：
9977925
负责人：
Ryan Thomas Hitzman
金额：
$ 4.13万
依托单位：
UNIVERSITY OF ILLINOIS AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-16 至 2021-08-15
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9977925
关键词：
Agonist Alkaline Phosphatase Animals Antibodies Aryl Hydrocarbon Receptor Benign Binding Biological Assay Botanical dietary supplements Botanicals Breast Cancer Risk Factor CYP1A1 gene CYP1B1 gene Cancer Etiology Cell Culture Techniques Cell Line Cells Cessation of life Chemopreventive Agent Collaborations Complex Cytochrome P450 DNA Binding Down-Regulation Drug Metabolic Detoxication Endometrial Carcinoma Enzymes Epigenetic Process Epimedium Estradiol Estrogen Antagonists Estrogen Metabolism Estrogen Receptor alpha Estrogen Receptors Estrogen receptor positive Estrogens Exhibits Failure Fluorescence Polarization Fractionation Fright Gene Expression Genetic Transcription Goals Health HepG2 Hormone replacement therapy Human Humulus Immunohistochemistry In Vitro Knowledge Lead Luciferases MCF7 cell MG132 Measures Mediating Menopausal Symptom Methods Methylation Microsomes Modeling Mutation Outcome Pathway interactions Pharmacologic Substance Postmenopause Progesterone Property Proteasome Inhibitor Quantitative Reverse Transcriptase PCR Quinones Rattus Receptor Cell Reporter Reporting Research Response Elements Risk Safety Signal Transduction Standard Model Standardization Testing Tissue Stains Translating Up-Regulation Uterus Weight Woman Women&apos s Health Xenobiotics adduct aryl hydrocarbon receptor ligand carcinogenesis chemical carcinogenesis chromatin immunoprecipitation estrogenic genotoxicity hormone therapy horny goat weed in vivo inhibitor/antagonist insight malignant breast neoplasm novel chemoprevention

项目摘要

Botanical Modulation of AhR-ERα by Crosstalk Inhibitors Promotes Estrogen (E2) Detoxification Estrogen receptor (ER) positive breast cancer poses significant health risks for postmenopausal women, and is in part mediated by the estrogen (E2) metabolite, estradiol-3,4-quinone, which causes depurinating adducts and leads to mutations. P450 1B1 is the primary enzyme for the conversion of estradiol to the 4-hydroxylated product, which can result in a genotoxic 4-quinone, while P450 1A1, which is normally epigenetically inhibited by E2-activated estrogen receptor alpha (ERα), converts estradiol to a nongenotoxic 2-hydroxylated product. Activated aryl hydrocarbon receptor (AhR) induces degradation of ERα as well as the transcription of both CYP1A1 and CYP1B1, which are translated into P450 1A1 and 1B1 respectively. Activators and ligands of AhR (Crosstalk Inhibitors) have been shown to preferentially upregulate CYP1A1 and ultimately the nongenotoxic 2-hydroxylated estradiol metabolite (estrogen detoxification pathway), through downregulation of the epigenetic inhibition of CYP1A1. Women have increasingly turned to botanical dietary supplements (BDS), instead of traditional hormone replacement therapy (HRT), since the release of the findings that estrogen + progesterone increases breast cancer risk. Interestingly, icaritin, a bioactive compound from Epimedium sp., a botanical used for women's health purposes, has been shown to activate AhR. The hypothesis of this proposal is that some women's health botanicals contain antiestrogenic, AhR activating compounds which degrade ERα (Aim 1) and reverse epigenetic CYP1A1 inhibition to preferentially activate the estrogen detoxification pathway in an in vitro cell culture model (Aim 2) and an in vivo ovariectomized rat model (Aim 3). Inhibition of estrogen-dependent alkaline phosphatase activity in Ishikawa cells, and a failure to inhibit a fluorescent estradiol-ERα complex by botanical compounds in a fluorescence polarization enzyme assay will provide antiestrogenic compounds which do not act through ERα. These compounds will be subjected to in- cell western of MCF-7 (ER+) cells against an ERα antibody to find compounds which degrade ERα. Additional XRE-luciferase activity in HepG2 cells will indicate AhR activating compounds, while a decrease in methylation through a ChIP assay of DNMT and XRE, and qRT-PCR of CYP1A1/1B1 will show that the reversal of estradiol mediated epigenetic CYP1A1 inhibition leads to upregulation of the estrogen detoxification pathway by Crosstalk Inhibitors in vitro. This premise will be taken to an ovariectomized rat model involving E2 and botanical compounds before sacrificing and analyzing uterine weight, performing LC-MS analysis of the E2 metabolites, and quantifying ERα expression levels using immunohistochemistry. This research may reveal mechanisms by which some bioactive women's health BDS compounds exhibit antiestrogenic outcomes, and the importance of the E2 detoxification pathway in promoting wellness in an in vivo postmenopausal model.

串扰抑制剂对AHR-ERα的植物调制促进雌激素（E2）解毒雌激素受体（ER）阳性乳腺癌对绝经后妇女具有重大健康风险，并且部分由雌激素（E2）代谢产物介导的雌二醇-3,4-夸酮，导致加合物脱落并导致突变。 P450 1B1是将雌二醇转化为4-羟化的主要酶产物，可能导致遗传毒性4夸酮，而P450 1A1通常在表观遗传上抑制通过E2激活的雌激素受体α（ERα），将雌二醇转化为非核2-羟基化产物。活化的芳基烃受体（AHR）诱导ERα的降解以及两者的转录 CYP1A1和CYP1B1分别转化为P450 1A1和1B1。激活剂和配体 AHR（串扰抑制剂）已被证明优先上调CYP1A1，最终是非核2-羟化雌二醇代谢物（雌激素解毒途径），通过下调 CYP1A1的表观遗传学抑制。妇女已经增加了植物饮食补充剂（BDS），自从雌激素 +的发现释放以来，而不是传统的骑马替代疗法（HRT）孕酮会增加乳腺癌的风险。有趣的是，iCaritin，一种来自Epimedium sp。的生物活性化合物，A 用于妇女健康目的的植物性已被证明可以激活AHR。这个假设提议是一些女性健康植物体含有抗雌激素，AHR激活化合物，这些化合物，这些化合物降解ERα（AIM 1）和反向表观遗传CYP1A1抑制优先激活雌激素体外细胞培养模型（AIM 2）和体内卵巢切除大鼠模型（AIM 3）中的解毒途径。抑制雌激素依赖性的酒精磷酸酶活性在石川细胞中，并且未能抑制A 荧光雌激素化合物在荧光极化酶测定中通过荧光雌二醇-erα复合物将提供不通过ERα起作用的抗雌激素化合物。这些化合物将受到内在 MCF-7（ER+）细胞的细胞西部针对ERα抗体，以找到降解ERα的化合物。额外的 HEPG2细胞中的XRE-环酶活性将表明AHR激活化合物，而甲基化降低通过DNMT和XRE的芯片测定，以及CYP1A1/1B1的QRT-PCR将表明逆转雌二醇介导的表观遗传学CYP1A1抑制导致雌激素解毒途径的上调通过体外串扰抑制剂。此前提将被带到涉及E2和植物化合物在牺牲和分析子宫重量之前，对E2进行LC-MS分析代谢物，并使用免疫组织化学来量化ERα表达水平。这项研究可能揭示了某些生物活性女性健康BDS化合物暴露于抗雌激素结果的机制，并 E2解毒途径在绝经后模型中促进健康方面的重要性。