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Novel estrogen receptor ligands from plant genomics

来自植物基因组学的新型雌激素受体配体

基本信息

批准号：
8136646
负责人：
JOHN M. LITTLETON
金额：
$ 23.22万
依托单位：
NAPROGENIX, INC
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-01 至 2013-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8136646
关键词：
Agonist Area Binding Biological Assay Biological Factors Biotechnology Breast Cancer Cell Breast Cancer Treatment Businesses Cancer Cell Growth Cause of Death Cells Chemopreventive Agent Climate Data Development Drug Industry Estrogen Receptor alpha Estrogen Receptor beta Estrogen Receptors Fluorescence Fluorescence Microscopy Foundations Fractionation Genome Genomics Glycine max Green Fluorescent Proteins High Pressure Liquid Chromatography Hormone Responsive Human Image In Situ In Vitro Individual Intellectual Property Legal patent Libraries Licensing Ligands Link Malignant Neoplasms Mammalian Cell Marketing Methodology Mouse-ear Cress Mutagenesis Natural regeneration Nuclear Receptor Gene Nuclear Receptors Pharmaceutical Preparations Pharmacologic Substance Pharmacotherapy Phase Phenotype Plant Extracts Plant Genes Plant Roots Plantago Plants Population Process Production Reporter Genes Research Screening procedure Seedling Small Business Technology Transfer Research Source Soybeans Specific qualifier value Specificity Stream Switch Genes Technology Technology Transfer Testing Therapeutic Time Transgenes Transgenic Organisms United States National Institutes of Health Woman base cancer therapy combinatorial chemistry commercial application commercialization design drug discovery gain of function mutation high throughput screening in vivo interest malignant breast neoplasm mutant novel phase 1 study promoter protein expression public health relevance research and development success

项目摘要

DESCRIPTION (provided by applicant): Human nuclear receptor (HNR) ligands have potential value as pharmacotherapies or chemopreventive agents for many hormone-responsive cancers. The primary aim of this STTR is to transfer an "academic" plant genomics technology into drug discovery, targeting HNR ligands. Estrogen receptor (ER) ligands in plants will be used for proof of application, because these have clear commercial and therapeutic value in their own right. In phase1 transgenic Arabidopsis thaliana seedlings expressing a human ERalpha/GFP construct were used to screen extracts from a native plant library for ER ligands. 9 previously uninvestigated aquatic species were identified, including species with agonist or antagonist activity at ERs in mammalian cell-based screens. In phase 2, representatives of these species will be investigated further, using assay-guided HPLC fractionation to isolate active compounds, which will be tested on breast cancer cells in vitro and in vivo. However, the major aim in phase 2 is to apply the novel plant genomics technology to optimize the production of ER ligands in two plant species. In this technology, transgenic hairy root cultures of a plant species are generated, expressing either the ERalpha/GFP construct, or a novel ERbeta/GFP construct. These cultures are then subjected to activation tagging mutagenesis (ATM), which induces random gain-of-function mutations in the genome. Now, any individual mutant in which the production of ER ligands is increased, should be identifiable by bright green fluorescence. This rapidly screens the "genomic capability" of the species to generate ligands for either ERalpha or ERbeta. In phase 1, proof of concept for this approach was achieved in a mutant population of transgenic A thaliana seedlings expressing ERalpha/GFP, and in phase 2 we propose initially to apply this approach to hairy root cultures of Glycine max (soybean - known to contain high levels of ER ligands) and Cacalia plantaginea (a native plantain with high levels of ERalpha/beta agonist activity). Positive cultures will be characterized by their GFP expression, and on the activity of culture extracts in conventional screens for ER activity. Attempts will also be made to regenerate intact mutant plants with the same over-producing phenotype, and active fractions from mutant cultures or plants evaluated on breast cancer cells as above. The approach can be used with any HNR target, and the major aim is to show that the technology can be transferred to anti-cancer natural product discovery. The commercialization plan is aimed at licensing the technology, or partnering with major companies to apply the technology to targets specified by the partner. PUBLIC HEALTH RELEVANCE: The technology transfer in this STTR promises to rapidly evaluate plant species for appropriate therapeutic activity at human nuclear receptors. The example chosen is for plant compounds with activity at the two types of human estrogen receptors (ERalpha and beta) which have different therapeutic implications, particularly for breast cancer. This is the most common form of cancer in women and is the major cause of death for women under 60 in the USA. Many breast cancers are (initially at least) "hormone responsive" in that drugs which inhibit ERs markedly slow the growth of the cancer cells, and this is a very active research area in the pharmaceutical industry. Plants are a known rich source of ligands for the ER, but it has proven difficult to exploit this source because conventional plant drug discovery is often time-consuming and expensive. This technology promises to change this by applying genomics approaches to plant drug discovery in this area.

描述(由申请人提供)：人类核受体(HNR)配体具有潜在的价值，可作为许多激素反应癌症的药物治疗或化学预防药物。这项研究的主要目的是将一种“学术”植物基因组学技术转化为药物发现，以HNR配体为靶标。植物中的雌激素受体(ER)配体将被用作应用的证据，因为这些配体本身具有明显的商业和治疗价值。在阶段1中，表达人ERpha/GFP结构的转基因拟南芥幼苗被用来筛选从天然植物文库中提取的ER配体。鉴定了9种以前未被调查的水生物种，包括在哺乳动物细胞筛选的ERs上具有激动剂或拮抗剂活性的物种。在第二阶段，将进一步研究这些物种的代表，使用检测引导的高效液相分离来分离活性化合物，这将在体外和体内的乳腺癌细胞上进行测试。然而，第二阶段的主要目标是应用新的植物基因组学技术来优化两个植物物种的ER配体的生产。在这项技术中，产生了植物物种的转基因毛状根培养物，表达ERpha/GFP结构或新的ERbeta/GFP结构。然后，这些培养物受到激活标签突变(ATM)的影响，这会在基因组中诱导随机的功能获得突变。现在，任何产生ER配体增加的个体突变，都应该可以通过明亮的绿色荧光来识别。这可以快速筛选出该物种产生ERpha或ERbeta配体的“基因组能力”。在第一阶段，这种方法的概念在表达ERpha/GFP的转基因拟南车草苗的突变群体中得到了验证，在第二阶段，我们最初建议将这种方法应用于大豆(已知含有高水平的ER配体)和Cacalia ants agina(一种具有高水平ERα/β激动剂活性的本土车前草)的毛状根培养。阳性培养的特征是其绿色荧光蛋白的表达，以及在传统的ER活性筛查中培养物提取物的活性。还将尝试再生具有相同过度生产表型的完整突变植物，以及来自突变培养物或如上所述在乳腺癌细胞上评估的植物的活性部分。该方法可以用于任何HNR靶点，主要目的是表明该技术可以转移到抗癌天然产物的发现上。商业化计划旨在许可该技术，或与大公司合作，将该技术应用于合作伙伴指定的目标。公共卫生相关性：这项技术转让承诺快速评估植物物种在人类核受体上的适当治疗活性。所选择的例子是对两种类型的人类雌激素受体(ERα和B受体)具有活性的植物化合物，这两种受体具有不同的治疗意义，特别是对乳腺癌。这是女性最常见的癌症形式，也是美国60岁以下女性死亡的主要原因。许多乳腺癌(至少在最初)是“激素反应”的，因为抑制ER的药物显著减缓了癌细胞的生长，这是制药业非常活跃的研究领域。植物是已知的内质网配体的丰富来源，但事实证明很难开发这一来源，因为传统的植物药物发现通常既耗时又昂贵。这项技术有望通过将基因组学方法应用于这一领域的植物药物发现来改变这一点。