RNA Targeted Screens of the Prion 5'UTR

朊病毒 5UTR 的 RNA 靶向筛选

基本信息

批准号：
7617517
负责人：
JACK T ROGERS
金额：
$ 17.59万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-09-30 至 2011-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7617517
关键词：
3&apos Untranslated Regions 5&apos Untranslated Regions Actins Alzheimer&apos s Disease Aminoglycosides Amyloid Amyloid beta-Protein Amyloid beta-Protein Precursor Amyotrophic Lateral Sclerosis Animal Model Animals Antibodies Appendix Base Pairing Binding Biological Assay Bovine Spongiform Encephalopathy Cell Line Cells Commit Condition DNA Data Disease Disease Progression Dose Drug Delivery Systems Elements Enzyme-Linked Immunosorbent Assay Ferritin Frequencies Gene Expression Genetic Translation H ferritin Hepatitis C Homeostasis Housing Human Infection Intervention Iron Iron-Regulatory Proteins Lead Left Letters Libraries Luciferases Macrolide Antibiotics Meat Mediating Messenger RNA Metalloproteins Modeling Molecular Bank Molecular Probes Mus Nerve Degeneration Neuroblastoma Neurodegenerative Disorders Neurons Oxidative Stress Parkinson Disease Pharmaceutical Chemistry Pharmaceutical Preparations PrPSc Proteins Prion Diseases Prions Production Proteins Public Health RNA Reagent Regulation Relative (related person)Reporter Reporting Resources Risk Running Scrapie Screening procedure Sequence Alignment Sheep Small RNA Source Specificity Synaptic Transmission Syndrome Testing Therapeutic Transcript Transfection Transferrin Transferrin Receptor Translations Universities Untranslated Regions Variant Viral Western Blotting alpha synuclein base beef design dopaminergic neuron experience high throughput screening in vivo infectious disease treatment inhibitor/antagonist neurotoxic neurotoxicity novel novel therapeutics prevent receptor relating to nervous system small molecule stem synuclein therapeutic target tissue culture wasting

项目摘要

DESCRIPTION (provided by applicant): We aim to set up a novel RNA based therapeutic strategy to screen and identify agents that slow down the spread of scrapies diseases to humans (Crueutzfeld-Jacob syndrome (CJD)) by limiting translation of the endogenous human cellular prion protein (PrPc). Limiting intracellular levels of endogenous PrPc should prevent this endogenous Cu/Zn metaloprotein from being a viable target for protein conversion by infectious PrP (PrPsc) from meat sources (i.e. Scapie from Sheep), and from beef (mad cow disease). Binding of small molecules to the 5' end of the PrPc transcript reduces its translational efficiency. This project is to set up the screening conditions to enable the use of the 5'untranslated region in front of the PrP (variant 2) transcript as a drug target so that we can identify novel and highly selective inhibitors that prevent intracellular prion (PrP) translation. We will use our prior expertise to high throughput screen the Alzheimer's APP 5'UTR, now to screen a chosen MLSCN drug library (HTS) and identify small molecule PrP UTR directed leads to be developed into effective prion translation inhibitors. Neurodegenerative diseases often occur due to the altered expression or aberrant folding of specific proteins, for example amyloid from APP in Alzheimer's disease, Prion protein in CJD and 1-synuclein in Parkinson's disease. However, our strategy is to prevent the translation of their key endogenous gene products in humans. Specificity of leads screened against the human PrP-vt2 5'UTR target will be assured after counter- screens with in-house stably transfected neural cell lines that each express the APP 5'UTR-luciferase and 1-syn 5'UTR-luciferase as key counter targets. This RNA based screening strategy will provide greater specificity to identify useful inhibitors of intracellular prion levels as a therapeutic strategy to offset this terrible neurodegenerative wasting disorder. Protein and DNA based approaches would be predicted to generate more off-target hits. PUBLIC HEALTH RELEVANCE: The data to be accumulated through this R21 molecular libraries screening cooperative mechanism will permit a high throughput transfection based screen of an important RNA target, the 5'untranslated region of the Parkinson's alpha synuclein (ASYN) transcript. This 5'UTR is an attractive therapeutic target for Parkinson's disease (PD), and newly identified ASYN 5'UTR specific leads may be developed by medicinal chemistry potentially to limit neurotoxic ASYN production in dopaminergic neurons. Use of 5'UTR specific MLSCN hits will probe the mechanism of translation of ASYN mRNA relevant to PD. Compounds directed to 5'UTRs of other mRNAs will probe mechanism of translation of the A2-amyloid precursor protein mRNA in Alzheimer's disease (SOD-1 mRNA in ALS, and PrP mRNA in Cruetzsfeld- Jacob Syndrome).

描述（由申请人提供）：我们旨在建立一种新型的基于RNA的治疗策略，以筛选和识别药物，以减慢刮擦疾病对人类的传播（Crueutzfeld-Jacob综合征（CJD）），通过限制内源性人类蜂窝状的翻译prion蛋白（PRPC）。限制内源性PRPC的细胞内水平应防止这种内源性Cu/Zn金属蛋白成为感染性PRP（PRPSC）的可行靶标（即来自绵羊的scapie）和牛肉（疯牛病）。小分子与PRPC转录本5'末端的结合降低了其翻译效率。该项目是为了建立筛查条件，以实现PRP（变体2）转录本前5'untranslated区域作为药物靶标的，以便我们可以识别出可预防细胞内prion（PRP）的新型且高度选择性的抑制剂。翻译。我们将利用先前的专业知识来高吞吐量屏幕Alzheimer的应用程序5'UTR，现在筛选选定的MLSCN药物库（HTS），并确定针对有效的prion型抑制剂的小分子PRP UTR引导。神经退行性疾病通常是由于特定蛋白的表达改变或异常折叠，例如阿尔茨海默氏病中APP的淀粉样蛋白的异常折叠，CJD中的prion蛋白和帕金森病中的1-核蛋白。但是，我们的策略是防止人类关键的内源基因产品的翻译。针对人PRP-VT2 5'UTR筛选的铅的特异性将在与内部稳定转染的神经细胞系进行反筛选后确保，每个神经细胞系都表达App 5'UTR-荧光酶和1-syn 5'utr-荧光酶关键计数器目标。这种基于RNA的筛查策略将提供更大的特异性，以确定细胞内prion水平的有用抑制剂作为一种治疗策略，以抵消这种可怕的神经退行性浪费障碍。将预测基于蛋白质和基于DNA的方法将产生更多的脱靶命中率。公共卫生相关性：通过此R21分子库筛选合作机制积累的数据将允许基于重要的RNA靶标的高通量转染屏幕，即帕金森氏症的5'untransalated区域，帕金森氏症的sns核蛋白（Asyn）转录。这5'UTR是帕金森氏病（PD）的有吸引力的治疗靶标，新近鉴定的ASYN 5'UTR特定铅可能是由药物化学可能开发的，可能会限制多巴胺能神经元中的神经毒性ASYN产生。使用5'UTR特异性MLSCN命中将探测与PD相关的ASYN mRNA翻译机理。针对其他mRNA的5'UTRS的化合物将探测阿尔茨海默氏病（ALS中的SOD-1 mRNA和cruetzsfeld- jacob综合征）中A2淀粉样蛋白前体蛋白mRNA翻译的机理。