Integrative And Molecular Studies Of Pain And Pain Control

疼痛和疼痛控制的综合和分子研究

• 批准号: 10691772
• 负责人: Andrew Mannes
• 金额: --
• 依托单位: CLINICAL CENTER
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10691772
• 关键词: Absence of pain sensation; Acute Pain; Address; Advanced Malignant Neoplasm; Affect; Afferent Neurons; Agonist; Analgesics; Anatomy; Animals; Autopsy; Axon; Basic Science; Behavior; Biological; Biopsy; Brain; Calcium; Calcium ion; Cell Nucleus; Cells; Central Nervous System; Cerebrospinal Fluid; Clinical; Clinical Research; Clinical Trials; Collaborations; Cooperative Research and Development Agreement; Copy Number Polymorphism; Coupled; Data; Dorsal; Dose; Exhibits; Fluorescence; Foundations; Gene Expression; Genes; Genetic; Genetic Variation; Goals; Health Sciences; Human; Immune; Immunofluorescence Immunologic; In Situ; In Situ Hybridization; In Vitro; Individual; Inflammation; Inflammatory; Injections; Injury; Intervention; Intractable Pain; Investigation; Ion Channel; Joints; Journals; Knowledge; Laboratories; Mechanics; Mediating; Messenger RNA; Methods; Modeling; Molecular; Molecular Biology; Morton' s Neuroma; Motor; National Center for Complementary and Integrative Health; Nerve; Nerve Endings; Nerve Fibers; Neuroma; Neuronal Plasticity; Neurons; Nociception; Nociceptors; Operative Surgical Procedures; Oregon; Organ Donor; Pain; Pain Research; Pain management; Paper; Pathologic; Pathology; Pathway interactions; Patients; Peptides; Peripheral; Peripheral Nerves; Peripheral Nervous System Diseases; Persistent pain; Phase I Clinical Trials; Phase III Clinical Trials; Physiological Processes; Population; Postoperative Pain; Postoperative Period; Pre-Clinical Model; Preparation; Process; Proprioception; Proteins; Protocols documentation; Publications; Publishing; RNA; Research; Resiniferatoxin; Route; Sampling; Sensory; Sensory Ganglia; Serious Adverse Event; Site; Skin; Spinal Cord; Spinal Ganglia; Stains; Stimulus; Surgical Oncology; Surgical incisions; Surgical wound; Synapses; System; TRPV1 gene; Techniques; Therapeutic; Therapeutic Agents; Thoracic Surgical Procedures; Time; Tissue Procurements; Tissues; Touch sensation; Toxicology; Transducers; Translational Research; Trigeminal Neuralgia; United States National Institutes of Health; Universities; Work; Writing; base; cancer pain; chronic pain; chronic painful condition; cytotoxicity; design; diabetic; experimental study; genomic locus; information processing; injured; lipidomics; nerve injury; neurotoxicity; neurotransmission; novel therapeutic intervention; osteoarthritis pain; overexpression; pain processing; pain sensation; pain sensitivity; pain signal; perineural; phase 1 study; pre-clinical; preclinical study; preservation; pressure; prevent; programs; receptor; small molecule; sodium ion; species difference; subcutaneous; tissue injury; transcriptome sequencing; transcriptomics; translational study; transmission process; wound healing

Summary Overview: This research program addresses basic molecular and physiological processes of nociceptive (pain-sensing) transmission in the peripheral and central nervous systems (CNS) and new ways to effectively control pain. The molecular research is performed using animal and in vitro cell-based models. We concentrate on primary afferent pain-sensing neurons located in dorsal root ganglion (DRG) that send axonal projections to skin and deep tissues and make connections within dorsal spinal cord, which is the first CNS site of synaptic information processing for pain. The mechanisms of transduction of pain stimuli are investigated through models of pathophysiological damage or using reductionist preparations such as primary DRG cultures or heterologous expression systems of ion channels or receptors. Our goals are (a) to understand the molecular and cell biological mechanisms of acute and chronic pain at the initial steps in the pain pathway, (b) to investigate mechanisms underlying human chronic pain disorders, (c) to explore neuronal plasticity and altered gene expression in persistent pain states, and (d) to use this knowledge to devise new treatments for pain. New Treatments for Pain: We address the new treatment goal through translational research coupled with human clinical trials to develop and introduce new molecular interventions for severe pain. Studies with the TRPV1 agonist resiniferatoxin (RTX) have resulted in a Phase I clinical trial for in patients with intractable pain from advanced cancer. RTX activates an influx of sodium and calcium ions and once bound to TRPV1, RTX props open the ion channel causing an intracellular calcium cytotoxicity. Depending on the route of administration RTX disables TRPV1 pain-sensing nerve endings or axons (i.e., the nerve fiber) or deletes the neuron entirely. RTX produces very effective pain control in pre-clinical models. The central route involves administration into the cerebrospinal fluid around the spinal cord (intrathecal). To date, we have treated 19 patients with pain from advanced cancer. This Phase I study is provisionally complete although we plan to examine a higher dose tier. It and being readied for publication. Earlier we published studies of RTX injections around or directly into sensory ganglia which formed the basis of our protocol to treat localized chronic pain by periganglionic RTX injection. Other peripheral routes of RTX administration include injection into skin, joints, nerve bundles, or topically. Preclinical studies show that analgesia by these routes is long-lasting but reversible, since the peripheral nerve endings regrow. Human protocols for post-operative incision and neuroma pain indications were generated in collaboration with the Thoracic and Oncologic Surgery Branch, NCI and local podiatrists, respectively. The initial post-operative study is designed to evaluate preemptive treatment with RTX injected subcutaneously prior to incision. This protocol has passed scientific review and was evaluated by the FDA which asked for certain additional experiments which are pending. The protocol for treating Mortons neuroma will be by perineural injection just proximal to the neuroma. This protocol also has passed scientific review The IND has been prepared and will soon be submitted to the FDA. Early Translational Investigations: In this cycle we extended our systems approach for integrated RNA-Seq and lipidomics to humans through a our human intraoperative tissue biopsy protocol. We obtained samples from surgical wound margins over time. This longitudinal tissue procurement was completed, and we are presently analyzing the samples transcriptomically, anatomically, and lipidomically. During this cycle we have made a strong effort to incorporate direct in human studies of nociceptive molecular biology. This was prompted by our repeated observations of species differences in gene expression in dorsal root ganglion (DRG) and spinal cord. Most of the skin, nerve, DRG and spinal cord tissues are recovered from organ donors or obtained at autopsy of patients in NCI Laboratory of Pathology. These tissues are being analyzed intensively by whole tissue sequencing, single nucleus sequencing, in situ hybridization and immunofluorescence staining of proteins. In collaboration with the Dr. Ashok Kulkarnis lab, we have examined DRG from patients with painful diabetic peripheral neuropathy (DPN). The first paper describing these results was published in this cycle and a second one is about to be submitted. We observe a loss of a select subpopulation of nociceptors suggesting a dynamic interplay occurs between initial hyperexcitability and subsequent neurotoxicity. We emphasize the importance of enhanced focus on human studies in a recent publication in the Journal of Pain (PMID: 35504570). Basic Pain Mechanisms: Underlying the translational and clinical studies are investigations of neuronal function, behavior, and molecular biological mechanisms of pain transduction and wound healing. We systematically investigate molecular alterations at the first three steps in the pain pathway beginning with injured peripheral tissue, the dorsal root ganglion and the dorsal (sensory) spinal cord to obtain a comprehensive, quantitative foundational molecular understanding of nociceptive processes related to inflammation, surgical incision, and nerve injury. One of the main methods we used is called RNA-Seq with which we can sequence all of the mRNAs in a given tissue or cell population. Our work now integrates RNA-Seq as a component in most of our investigations. Another main method is multiplex fluorescence in situ and immunofluorescence anatomical studies of tissues in the nociceptive circuit. Using these techniques we now investigate humans with genetic variations that affect pain sensitivity. At present we are investigating a group of rare individuals with a copy number variant involving a specific gene locus. Individuals with three copies of this region exhibit profound decreases pain sensitivity. Using RNA-Seq we identified one gene in the locus of 25 that is an excellent candidate for mediating the analgesic action when overexpressed. The results are both compelling and informative and define a previously unidentified genetic mechanism for governing pain sensitivity. The RNA-Seq investigations provide new quantitative assessments of neuronal and glial genes as well as immune process related to nociceptive circuit function. Through this basic research we are obtaining a deeper understanding of mechanisms that trigger acute pain and sustain chronic pain and we are identifying molecular components to control pain.

概括 概述：该研究项目致力于解决外周和中枢神经系统（CNS）中伤害性（疼痛感知）传播的基本分子和生理过程以及有效控制疼痛的新方法。分子研究是使用动物和体外细胞模型进行的。我们专注于位于背根神经节 (DRG) 的初级传入疼痛感知神经元，这些神经元将轴突投射发送到皮肤和深层组织，并在背脊髓内建立连接，背脊髓是处理疼痛的突触信息的第一个中枢神经系统部位。通过病理生理学损伤模型或使用还原制剂（例如原代 DRG 培养物或离子通道或受体的异源表达系统）来研究疼痛刺激的转导机制。我们的目标是（a）在疼痛途径的最初步骤了解急性和慢性疼痛的分子和细胞生物学机制，（b）研究人类慢性疼痛疾病的机制，（c）探索神经元可塑性和改变的基因持续疼痛状态下的表达，以及（d）利用这些知识来设计新的疼痛治疗方法。 疼痛的新治疗方法：我们通过转化研究结合人体临床试验来实现新的治疗目标，以开发和引入针对严重疼痛的新分子干预措施。 TRPV1 激动剂树脂毒素 (RTX) 的研究已针对晚期癌症顽固性疼痛患者进行了 I 期临床试验。 RTX 激活钠离子和钙离子的流入，一旦与 TRPV1 结合，RTX 就会打开离子通道，导致细胞内钙细胞毒性。根据给药途径，RTX 会禁用 TRPV1 疼痛感应神经末梢或轴突（即神经纤维）或完全删除神经元。 RTX 在临床前模型中产生非常有效的疼痛控制。中枢途径涉及给药至脊髓周围的脑脊液（鞘内）。迄今为止，我们已经治疗了 19 名晚期癌症疼痛患者。尽管我们计划检查更高的剂量层，但这项第一期研究已暂时完成。它并准备出版。早些时候，我们发表了在感觉神经节周围或直接注射 RTX 的研究，这构成了我们通过神经节周围 RTX 注射治疗局部慢性疼痛的方案的基础。 RTX 给药的其他外周途径包括注射到皮肤、关节、神经束或局部。临床前研究表明，这些途径的镇痛作用是持久的，但可逆，因为周围神经末梢会再生。针对术后切口和神经瘤疼痛适应症的人体方案是分别与胸外科和肿瘤外科分部、NCI 和当地足病医生合作制定的。最初的术后研究旨在评估切口前皮下注射 RTX 的先发性治疗。该方案已通过科学审查，并由 FDA 进行评估，FDA 要求进行某些正在进行的额外实验。治疗莫顿神经瘤的方案是在神经瘤附近进行神经周围注射。 该方案也已通过科学审查，IND已准备就绪，即将提交给FDA。 早期转化研究：在这个周期中，我们通过人类术中组织活检方案将集成 RNA 测序和脂质组学的系统方法扩展到人类。随着时间的推移，我们从手术伤口边缘获取了样本。纵向组织采购已经完成，我们目前正在对样本进行转录组学、解剖学和脂质组学分析。 在这个周期中，我们做出了巨大的努力，将伤害性分子生物学的人类研究直接纳入其中。这是由于我们反复观察背根神经节（DRG）和脊髓基因表达的物种差异而得出的。 大多数皮肤、神经、背根神经节和脊髓组织是从器官捐献者身上回收的，或者是在 NCI 病理实验室对患者进行尸检时获得的。 通过全组织测序、单核测序、原位杂交和蛋白质免疫荧光染色对这些组织进行深入分析。我们与 Ashok Kulkarnis 博士实验室合作，检查了患有疼痛的糖尿病周围神经病变 (DPN) 患者的 DRG。 描述这些结果的第一篇论文已在本周期发表，第二篇论文即将提交。我们观察到特定伤害感受器亚群的丧失，表明最初的过度兴奋和随后的神经毒性之间存在动态相互作用。我们在《疼痛杂志》（PMID：35504570）最近发表的一篇文章中强调了加强对人类研究的关注的重要性。 基本疼痛机制：转化和临床研究的基础是对疼痛传导和伤口愈合的神经元功能、行为和分子生物学机制的研究。我们系统地研究疼痛通路前三个步骤的分子改变，从受伤的周围组织、背根神经节和背侧（感觉）脊髓开始，以获得与炎症、手术切口相关的伤害感受过程的全面、定量的基础分子理解。和神经损伤。我们使用的主要方法之一称为 RNA-Seq，通过它我们可以对给定组织或细胞群中的所有 mRNA 进行测序。我们现在的工作将 RNA-Seq 作为我们大多数研究的一个组成部分。另一种主要方法是对伤害感受回路中的组织进行多重荧光原位和免疫荧光解剖研究。 现在，我们利用这些技术来研究具有影响疼痛敏感性的基因变异的人类。目前，我们正在研究一组具有涉及特定基因位点的拷贝数变异的稀有个体。具有该区域三个副本的个体表现出疼痛敏感性的显着降低。使用 RNA-Seq，我们在 25 个基因座中鉴定出一个基因，该基因是过度表达时介导镇痛作用的绝佳候选基因。研究结果既令人信服又信息丰富，并定义了一种以前未知的控制疼痛敏感性的遗传机制。 RNA-Seq 研究为神经元和神经胶质基因以及与伤害感受回路功能相关的免疫过程提供了新的定量评估。通过这项基础研究，我们对引发急性疼痛和维持慢性疼痛的机制有了更深入的了解，并且我们正在确定控制疼痛的分子成分。