RNA Aptamers Selective for TPRV Channels

针对 TPRV 通道的选择性 RNA 适体

基本信息

批准号：
7656649
负责人：
ROGER Gordon O'NEIL
金额：
$ 18.75万
依托单位：
UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
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：
2008-08-01 至 2011-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7656649
关键词：
3-Dimensional Acidity Affinity Agonist Antibodies Behavior Binding Binding Sites Biological Assay Biological Availability Calcium Capsaicin Cells Chemicals Chili Pepper Classification Clinical Trials Cloning Dental caries Development Drug Delivery Systems Drug Kinetics Esthesia Event Evolution Exposure to Foundations Ganglia Goals Heating High temperature of physical object Hyperalgesia Image Imaging Techniques In Situ Inflammatory Ion Channel Kinetics Knockout Mice Libraries Ligands Mechanical Stress Modeling Modification Molecular Conformation Monoclonal Antibodies Mus Myxoid cyst Natural regeneration Nature Nerve Nerve Fibers Neurons Nociception Pain Pathway interactions Procedures Process Property Protein Binding Domain Protein Isoforms Proteins RNA Role Screening procedure Seminal Site Small RNA Specificity Stimulus Structure of trigeminal ganglion TRP channel TRPV channel TRPV1 gene Technology Testing Therapeutic Therapeutic Agents Time Trigeminal System Vanilloid Vertebral column Water afferent nerve aptamer base capsaicin receptor combinatorial chemistry drug candidate drug discovery in vivo interest member methyl group mouse model novel novel strategies novel therapeutics nuclease numb protein osmoreceptor pain behavior patch clamp pressure public health relevance spinal nerve posterior root success three dimensional structure tool

项目摘要

DESCRIPTION (provided by applicant): The overall goal of the project is to employ a novel, emerging combinatorial chemistry technology to identify and develop small RNA molecules that bind with high affinity and specificity to selected TRPV channel target sites as potential therapeutic agents for treatment of nociceptive behavior. Two isoforms of the vanilloid receptor subfamily of TRP channels, TRPV1 (the capsaicin receptor) and TRPV4 (an osmoreceptor), display polymodal gating behavior with sensitivity to numerous noxious stimuli. Both channels are widely distributed in small- to medium-sized neurons of the dorsal root and trigeminal ganglia. However, traditional drug discovery efforts to identify specific antagonist are relatively slow and have had limited success, although development efforts are intensifying. The recent discovery of small RNA molecules that fold into unique 3-D structures that bind with high affinity and specificity to protein binding domains, akin to monoclonal antibodies, has opened the door to development of a new, powerful, class of pharmacotherapeutic agent. The purpose of the present study is to generate selective RNA aptamers that bind with high affinity to TRPV1 and/or TRPV4 isoforms and act as antagonist for pharmacotherapeutic applications. Three specific aims are proposed to accomplish this goal: 1. To generate high affinity RNA aptamers with specificity for TRPV channel isoforms, TRPV1 and TRPV4. RNA aptamers will be identified from a large pool of random RNA molecules by systemic evolution, amplification, and enrichment of high affinity RNA ligands (aptamers). 2. To screen the high-affinity RNA aptamers for function as potential antagonist of the TRPV isoforms. High throughput, high-content, kinetic screening of identified RNA aptamers on calcium influx through TRPV1 or TRPV4 channels will be assessed to evaluate the potential of each identified aptamer as functional antagonist of TRPV1 and TRPV4. 3. To evaluate the potential function of the identified RNA aptamers as pharmacological tools/therapeutic agents. RNA aptamers identified from the kinetic screens will be evaluated as a pharmacotherapeutic tools using patch clamp analysis to directly assess aptamer effects on channel function. Promising aptamers will be further evaluated for therapeutic potential, in vivo, in established models of TRPV1- and TRPV4-dependent pain behavior. The studies will have far reaching impact both in providing a foundation for identification and development RNA aptamers as pharmacotherapeutic agents, and in providing new therapeutic tools for the specific treatment of nociceptive behavior, as well as for numerous other pathophysiological conditions associated with the TRPV ion channels. PUBLIC HEALTH RELEVANCE: The goal of the project is to use a novel, emerging, approach to generate small RNA molecules (RNA aptamers, 3-D folded) that bind with high affinity and specificity (10; 26) to target sites on specific channel proteins that are associated with sensing pain (TRPV channels) (40; 57; 61). RNA molecules that selectively block the function of the TRPV channels and, therefore, the pain sensation, would be specifically targeted. Generating such molecules would open a new platform for development of these, and other, compounds as potential pharmacological tools and therapeutic agents for the treatment of a broad range of painful conditions associated with noxious stimuli or inflammatory states (e.g., pressure, high temperature, nerve damage/tooth decay, hyperalgesia, and exposure to noxious chemicals) (12; 13; 17; 52; 78).

描述（由申请人提供）：该项目的总体目标是采用一种新颖的新兴组合化学技术来识别和开发与所选TRPV通道靶位点具有高亲和力和特异性结合的小RNA分子，作为潜在的治疗剂，以治疗伤害性行为。 TRP通道亚家族的两种同工型TRPV1（辣椒素受体）和TRPV4（一个osmoreceptor）显示了对众多有害刺激的敏感性的多峰传输行为。这两个通道都广泛分布在背根和三叉神经节的中小型神经元中。但是，尽管发展努力正在加剧，但传统的药物发现努力识别特定对手的速度相对较慢，并且取得了有限的成功。最近发现的小RNA分子折叠成具有高亲和力和对蛋白质结合结构域的特异性（类似于单克隆抗体）的独特的3D结构，为开发了一种新型，强大的药物治疗剂的开发打开了大门。本研究的目的是生成与TRPV1和/或TRPV4同工型高亲和力结合的选择性RNA适体，并充当药物治疗应用的拮抗剂。提出了三个特定的目标来实现这一目标：1。生成具有特异性的高亲和力RNA适体对TRPV通道同工型，TRPV1和TRPV4。通过全身进化，扩增和富集高亲和力RNA配体（Aptamers），将从大量的随机RNA分子中鉴定RNA适体。 2。筛选高亲和力的RNA适体，以作为TRPV同工型的潜在拮抗剂功能。将评估对通过TRPV1或TRPV4通道进行鉴定的RNA适体的高吞吐量，动力学筛选，以评估每个鉴定的适体的潜力，作为TRPV1和TRPV4的功能拮抗剂的潜力。 3。评估确定的RNA适体作为药理学工具/治疗剂的潜在功能。从动力学筛选中鉴定出的RNA适体将使用斑块夹分析评估作为药物治疗工具，以直接评估适合对通道功能的影响。在既定的TRPV1和TRPV4依赖性疼痛行为的既定模型中，将进一步评估有希望的适体的治疗潜力。这项研究将在为鉴定和开发RNA适体作为药物治疗剂的基础上以及为特异性治疗伤害性行为以及许多与TRPV离子通道相关的许多其他病理生理条件提供新的治疗工具方面具有很大的影响。公共卫生相关性：该项目的目的是使用一种新颖的，新兴的方法来产生与高亲和力和特异性（10; 26）结合的小RNA分子（RNA Aptamers，3-D折），以与特定通道蛋白上的靶位点结合，这些位点与感应疼痛（TRPV通道）相关的特定通道蛋白（40; 57; 57; 57; 61; 61; 61; 61; 61）。将有选择地阻断TRPV通道功能并因此将疼痛感觉的RNA分子被专门靶向。 Generating such molecules would open a new platform for development of these, and other, compounds as potential pharmacological tools and therapeutic agents for the treatment of a broad range of painful conditions associated with noxious stimuli or inflammatory states (e.g., pressure, high temperature, nerve damage/tooth decay, hyperalgesia, and exposure to noxious chemicals) (12; 13; 17; 52; 78).