Developing chemical probes that target specific inflammasomes

开发针对特定炎症体的化学探针

• 批准号: 9101571
• 负责人: Tsan Sam Xiao
• 金额: $ 20.92万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9101571
• 关键词: Alzheimer' s Disease; Apoptosis; Atherosclerosis; Autoimmune Process; Base Sequence; Binding; Biochemical; Biological Assay; Bone Marrow; CASP1 gene; Caspase; Cell Death; Cell Line; Chemicals; Clinical; Collection; Development; Disease; Filament; Gout; Immune; Infection; Inflammatory; Interleukin-1 beta; Interleukin-18; Libraries; Ligand Binding Domain; Mediating; Methods; Molecular Conformation; Non-Insulin-Dependent Diabetes Mellitus; Physiological; Play; Proteins; Psoriasis; Receptor Signaling; Regulation; Reporting; Rest; Role; Signal Transduction; Specificity; Structural Models; Structure; Surface; Syndrome; Systemic Lupus Erythematosus; Testing; Therapeutic; United States National Institutes of Health; adapter protein; autoinflammatory; base; cytokine; drug development; ds-DNA; experience; in vitro Assay; inhibitor/antagonist; macrophage; marenostrin; meetings; novel; polymerization; programs; public health relevance; receptor; screening; sensor; small molecule; small molecule libraries; small molecule therapeutics; success; therapeutic development; tool; virtual

 DESCRIPTION (provided by applicant): The inflammasomes are crucial innate immune signaling platforms implicated in immune defense against infections and autoimmune/autoinflammatory disorders. Despite their important physiological and pathological functions, the critical need for inflammasome-specific chemical probes has not been met and none of the reported inflammasome inhibitors directly engage the inflammasome receptor/sensor proteins. This not only impedes the progress of mechanistic studies in the inflammasome field, but also hampers the development of specific and potent therapeutics for inflammatory disorders, such as systemic lupus erythematosus (SLE), psoriasis, cryopyrin-associated periodic syndromes (CAPS), gout, type 2 diabetes, Alzheimer's disease, and atherosclerosis. My lab has made major contributions to our understanding of the structure and function of the AIM2 and IFI16 inflammasomes: we established the structural basis of sequence-independent dsDNA recognition by AIM2 and IFI16, characterized the autoinhibition mechanism that regulates the AIM2 receptor, elucidated the signaling mechanisms mediated by the pyrin, CARD and TIR domains from the inflammasome receptors and signaling adapters. We will leverage our expertise and experience and test the hypothesis that chemical compounds that bind the inflammasome receptors/sensors or adapter at critical domain interfaces will stabilize different conformational states of their structures and thus modulate ther functions. To this end, we will explore a two-pronged approach to identify chemical probes specific for the AIM2 and NLRP3 inflammasomes. In aim 1, we will perform structure-based virtual screening for chemical probes that specifically bind AIM2, NLRP3 or ASC. We anticipate that small molecules that bind and stabilize the intramolecular domain-domain interfaces and the autoinhibited structures of AIM2 and NLRP3 may serve to suppress their activation. On the other hand, disruption of the ASC PYD oligomerization interface using chemical compounds that competitively inhibit the ASC filament formation may also inhibit inflammasome activation. In aim 2, we will carry out three independent in vitro assays that allow us to screen and validate inflammasome-specific chemical compounds that bind and regulate the ASC polymerization and speck formation. These assays include the thermofluor assay and the ASC polymerization assay using purified AIM2, NLRP3 and ASC proteins, and the ASC speck formation assay using a bone marrow-derived macrophage cell line stably expressing fluorescent cerulean-tagged ASC protein. We anticipate that chemical probes that specifically modulate the activities of either the AIM2 inflammasome, the NLRP3 inflammasome, or both through ASC, will be identified and validated. Importantly, the success of this project will fulfill unmet needs for hig quality and novel chemical tools to probe the function of specific inflammasomes, as well as to stimulate the development of specific and potent therapeutics against inflammatory disorders.

 描述（由适用提供）：炎症是针对感染和自身免疫/自身免疫性疾病的免疫防御中实施的至关重要的先天免疫信号平台。尽管它们具有重要的生理和病理功能，但尚未解决炎性特异性化学问题的关键需求，并且没有报道的炎性抑制剂直接吸引炎性体受体/传感器蛋白。 This not only impedes the progress of Mechanistic studies in the inflammasome field, but also hampers the development of specific and potential therapy for inflammatory disorders, such as systemic lupus erythematosus (SLE), psoriasis, cryopyrin-associated periodic syndromes (CAPS), get, type 2 diabetes, Alzheimer's disease, and atherosclerosis. My lab has made major contributions to our understanding of the structure and function of the AIM2 and IFI16 inflammasomes: we established the structural basis of sequence-independent dsDNA recognition by AIM2 and IFI16, characterised the autoinhibition mechanism that regulates the AIM2 receptor, elucidated the signaling mechanisms mediated by the pyrin, CARD and TIR domains from the inflammasome receptors and signaling adapters.我们将利用我们的专业知识和经验，并检验以下假设：在关键域界面上结合炎症体受体/传感器或适配器的化合物将稳定其结构的不同构造状态，从而调节功能。为此，我们将探索一种两种依据的方法，以识别针对AIM2和NLRP3炎症体特有的化学问题。在AIM 1中，我们将对特异性结合AIM2，NLRP3或ASC的化学问题进行基于结构的虚拟筛选。我们预计，结合和稳定分子内结构域域界面的小分子以及AIM2和NLRP3的自身抑制结构可能有助于抑制其激活。另一方面，使用竞争性抑制ASC细丝形成的化学化合物对ASC PYD寡聚界面的破坏也可能抑制炎性体的激活。在AIM 2中，我们将进行三个独立的体外测定法，使我们能够筛选和验证结合和调节ASC聚合和规格形成的炎性体特异性化合物。这些测定包括使用纯化的AIM2，NLRP3和ASC蛋白的热氟测定和ASC聚合测定，以及使用骨髓衍生的巨噬细胞系稳定表达荧光陶瓷标记的ASC蛋白的ASC规格形成测定。我们预计将发现和验证AIM2炎症体，NLRP3炎性体或两者兼而有之的化学问题。重要的是，该项目的成功将满足对高质量和新型化学工具的未满足需求，以探测特定炎症的功能，并刺激针对炎症性疾病的特定和潜在治疗的发展。