Development of inlammasome inhibitors to be used as anti-inflammatory agents

开发用作抗炎剂的inlammasome抑制剂

• 批准号: 8403458
• 负责人: Shyam Biswal
• 金额: $ 4.05万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-21 至 2014-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8403458
• 关键词: Adrenal Cortex Hormones; Alzheimer' s Disease; Anti-Inflammatory Agents; Anti-inflammatory; Arthritis; Asbestos; Atherosclerosis; Attenuated; Bacteria; Biological Assay; Caspase-1; Cell Line; Cell-Free System; Cells; Chemicals; Cholesterol; Chronic; Cleaved cell; Collaborations; Complex; Data; Development; Diabetes Mellitus; Disease; Dissection; Dose; Flagellin; Fluorogenic Substrate; Future; Genomics; Goals; Gout; Human; Immune response; Incubated; Inflammation; Inflammatory; Inflammatory Response; Inhibitory Concentration 50; Interferons; Interleukin-18; Interleukin-6; Lead; Libraries; Ligands; Measures; Mediating; Mediator of activation protein; Molecular; Molecular Bank; Mus; NF-kappa B; Non-Insulin-Dependent Diabetes Mellitus; Pathway interactions; Pharmaceutical Preparations; Poly dA-dT; Pre-Clinical Model; Public Health; Pulmonary Fibrosis; Recombinants; Reporter; Role; Salmonella; Screening procedure; Silicon Dioxide; Silicosis; Stimulus; System; Testing; Therapeutic; United States National Institutes of Health; Uric Acid; Validation; Virus; arm; base; combat; cytokine; cytotoxic; follow-up; in vivo; inhibitor/antagonist; monocyte; novel; procaspase-1; programs; protein complex; response; small molecule; small molecule libraries

DESCRIPTION (provided by applicant): Inflammation is a central etiological component of numerous diseases, and anti-inflammatory drugs, such as corticosteroids, represent important therapies for combating a large number of diseases. However, many diseases do not adequately respond to current anti-inflammatory therapies. Thus new anti-inflammatory therapies are needed. The inflammasome is a recently identified arm of the innate immune response, which is becoming increasingly recognized as an important determinant of numerous inflammatory diseases. The inflammasome is a mult-protein complex that responds to numerous stimuli, including bacteria, viruses, silica, asbestos, cholesterol, and uric acid, to initiate a pro-inflammatory response that is mediated by the cytokines IL-1b, IL-18, and IL-33. Improper activation of the inflammasome has been implicated in multiple diseases, including gout, arthritis, diabetes, Alzheimer's, pulmonary fibrosis, and atherosclerosis. The role of the inflammasome in other disorders is still evolving, and inhibitors of this pathway will provide potential therapeutics for numerous diseases, and will provide mechanistic studies that will aid in dissection of this pathway. There are four inflammasome complexes identified thus far, and each inflammasome responds to a unique combination of molecular activators. However, a central component of all inflammasomes is its activation and secretion of the cytokines IL-1b, IL-18, and IL-33. Thus, secretion of IL-1b is a suitable marker for inflammasome activation, and identifying novel inhibitors of the inflammasome will provide potential therapies for a large number of inflammatory diseases. Secondary and tertiary studies will more directly measure activity of the inflammasome complex protein caspase-1. Specific Aim: To utilize a high throughput cell-based assay to identify small molecule inhibitors of the inflammasome. This proposal will focus on screening compounds to identify inhibitors of IL-1b, as a marker of inflammasome activity. We have developed a high throughput 1536-well assay using the AlphaLISA IL1b Kit (Perkin Elmer) to quantify IL-1b secretion from the human monocyte cell line THP-1. Our preliminary data demonstrates that we can readily detect IL-1b secretion from activated THP-1 cells and can quantify IC50 values of known inflammasome inhibitors. We have established a collaboration with the NIH Chemical Genomics Center (NCGC), and our chemical library will be derived from their Molecular Libraries Program. This library consists of an expansive set of small molecules totaling greater than 300,000 compounds. Drugs identified by this primary screen will be validated and characterized by secondary and tertiary screens to measure activity of caspase-1 and NF-kB, and secretion of other inflammasome-dependent and independent cytokines. Hit compounds will be validated in cells from mice deficient in various components of the inflammasome, and follow-up studies will examine efficacy in vivo. Successful completion of the proposed study will yield novel compounds that inhibit the inflammasome in cell-based systems.

描述（由申请人提供）：炎症是多种疾病的中心病因组成部分，抗炎药（例如皮质类固醇）代表了对抗大量疾病的重要疗法。但是，许多疾病对当前的抗炎疗法没有充分反应。因此，需要新的抗炎疗法。炎症体是最近确定的先天免疫反应的部门，该部门越来越被认为是多种炎症性疾病的重要决定因素。炎性体是一种多蛋白质复合物，对许多刺激有反应，包括细菌，病毒，二氧化硅，石棉，胆固醇和尿酸，以启动由细胞因子IL-1B，IL-18和IL-33介导的促炎反应。炎性体的激活不当与多种疾病有关，包括痛风，关节炎，糖尿病，阿尔茨海默氏症，肺纤维化和动脉粥样硬化。炎症体在其他疾病中的作用仍在发展，该途径的抑制剂将为许多疾病提供潜在的治疗疗法，并将提供机械研究，以帮助解剖这一途径。到目前为止，有四种炎性体复合物，每个炎症体对分子活化剂的独特组合做出反应。但是，所有炎症的一个核心成分是其激活和分泌细胞因子IL-1B，IL-18和IL-33。因此，IL-1B的分泌是炎症体激活的合适标记，鉴定炎性体的新型抑制剂将为大量炎症性疾病提供潜在的疗法。二级和三级研究将更直接地测量炎性体复合蛋白caspase-1的活性。具体目的：利用高吞吐细胞的测定法以鉴定炎症体的小分子抑制剂。该提案将着重于筛选化合物，以鉴定IL-1B的抑制剂，作为炎性体活动的标志。我们使用Alphalisa IL1B试剂盒（Perkin Elmer）开发了高吞吐量1536孔测定法，以量化人类单核细胞系THP-1的IL-1B分泌。我们的初步数据表明，我们可以很容易地检测到激活的THP-1细胞中的IL-1B分泌，并可以量化已知炎性体抑制剂的IC50值。我们已经与NIH化学基因组学中心（NCGC）建立了合作，我们的化学库将来自其分子图书馆计划。该库由一组膨胀的小分子组成，总分子大于300,000种化合物。该主要筛查确定的药物将得到验证和特征，并以次级和第三次筛选来测量caspase-1和NF-KB的活性，以及​​其他炎症体依赖性和独立细胞因子的分泌。 HIT化合物将在炎性体各种成分的小鼠的细胞中得到验证，随访研究将检查体内功效。成功完成拟议的研究将产生新的化合物，以抑制基于细胞的系统中的炎症体。