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Probes to target the 3-way hotspot of IL1RacP to abolish aberrant interleukin inflammation

靶向 IL1RacP 3 路热点以消除异常白细胞介素炎症的探针

基本信息

批准号：
8944833
负责人：
Lance Allen Liotta
金额：
$ 59.52万
依托单位：
GEORGE MASON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-07-01 至 2018-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8944833
关键词：
Acute Affinity Agonist American Amino Acids Animal Model Arginine Arthritis Asthma Autoimmune Process Binding Biodistribution Biological Assay Biological Availability Cell Culture Techniques Cell Line Cell membrane Cell surface Chemistry Clinical Complex Cytokine Activation Degenerative polyarthritis Deuterium Disease Dose Drug Targeting Engineering Family Family member Hot Spot In Vitro Inflammation Inflammatory Inhibitory Concentration 50 Interleukin Therapy Interleukin-1 Interleukin-1 beta Interleukins Intra-Articular Injections Kineret Lead Ligand Binding Ligands Maps Mediating Methods Modeling Modification Molecular Monoclonal Antibodies Neonatal Obstructive Lung Diseases Peptide Hydrolases Peptides Pharmaceutical Preparations Play Positioning Attribute Protein Binding Protein Chemistry Proteins Psoriasis Publishing Receptor Signaling Recruitment Activity Role Signal Transduction Site Specificity Surface Surface Plasmon Resonance Tail Technology Testing Therapeutic Transmembrane Domain Traumatic Arthropathy Validation Work anakinra base crosslink cytokine design domain mapping in vitro Assay in vivo inhibitor/antagonist interleukin-1 receptor accessory protein member mouse model neoplastic new technology next generation novel osmotic minipump peptidomimetics prevent protein complex protein folding protein protein interaction public health relevance receptor small molecule

项目摘要

DESCRIPTION (provided by applicant): Aberrant interleukin family cytokine activation of inflammation characterizes a wide spectrum of autoimmune, inflammatory, neoplastic, and neonatal disorders, including arthritis, obstructive lung disease, psoriasis and asthma. We created a novel technology, protein painting, and used it to discover, and create, a novel peptide inhibitor, called Arg286p, that abolishes the necessary interaction of IL1-RAcP with the IL-1β-IL1Receptor complex. Arg286p constitutes a novel specific and potent approach to treat interleukin mediated inflammatory diseases. The binding interface contact points between interacting protein partners are important drug targets for the next generation of therapies that block such interactions. Employing a new class of small molecule affinity chemistry, protein painting has a positive hit specificity of 93% and yield up to ten fold higher compared to conventional cross linking or deuterium exchange methods. Using our technology to study the 3-way interaction of interleukin-1β (IL-1β, IL-1 receptor I (IL-1RI), and IL-1 receptor accessory protein (IL-1RAcP), we identified a highly conserved beta-loop Arg286 region of the IL-1RAcP protein that participates in a multivalent interaction with the ligand and the receptor. We validated this novel target to be necessary for IL1β-mediated signaling. We created a folded protein peptide Arg286p that corresponds to the Arg286 beta loop domain, and a monoclonal antibody that recognizes the IL-1RAcP Arg286 surface domain. Our Arg286 peptide compound abolished interleukin signaling in a cell culture model in a dose-dependent manner, compared to a control peptide lacking the arginine in the 286 position. The Arg286 peptide and the anti-Arg286 mAb also abolished formation of the IL-1β, IL-1RI, and IL-1RAcP 3-way complex, in vitro, in a dose-dependent manner. IL1-RAcP is an accessory protein that is recruited after the interleukin ligand binds to the receptor, and signaling requires all members of this three-way complex formation. Our new inhibitor Arg286p mimics a small key multivalent interaction region at the arginine 286 beta loop of IL1-RacP, thereby abolishing complex formation and preventing downstream inflammatory signaling. Since Arg286p acts downstream of receptor-ligand binding, it can massively amplify the potency for interleukin therapy even in the face of excess ligand, and may be superior to, or synergistic with, existing IL-1 therapies that competitively target the ligand. Under the Aims, we will create new structural modifications in this lead compound to optimize its affinity and stability for IL-1 and we will use it as a basis to create novel inhibitos specific for IL33 and IL36, for which clinical inhibitors do not exist. We will employ our protein painting technology in a novel iterative workflow to guide the design of peptide modulators (agonists and antagonists) that are either optimized in potency for IL-1 or specific for IL-33 and IL-36 in vitro, in three types of assays. The optimized version of our lead compound will be studied in a well-established animal model of osteoarthritis. Osteoarthritis causes suffering for 27 million Americans.

描述（由申请人提供）：炎症的异常白细胞介素家族细胞因子激活表征了广泛的自身免疫性、炎症性、肿瘤性和新生儿疾病，包括关节炎、阻塞性肺病、银屑病和哮喘。我们创造了一种新的技术，蛋白质绘画，并用它来发现和创造一种新的肽抑制剂，称为Arg 286 p，它消除了IL-1-RAcP与IL-1β-IL-1受体复合物的必要相互作用。Arg 286 p是治疗白细胞介素介导的炎症性疾病的一种新的特异性和有效的方法。相互作用蛋白质伴侣之间的结合界面接触点是阻断这种相互作用的下一代疗法的重要药物靶标。采用一类新的小分子亲和化学，蛋白质涂敷具有93%的阳性命中特异性，并且与常规交联或氘交换方法相比，产量高出10倍。利用我们的技术研究了白细胞介素1β（IL-1β）、IL-1受体I（IL-1 RI）和IL-1受体辅助因子（IL-1 RI）的3向相互作用，蛋白（IL-1 RAcP），我们确定了一个高度保守的β环Arg 286区的IL-1 RAcP蛋白，参与了多价相互作用的配体和受体。我们验证了这种新的靶点是IL 1 β介导的信号传导所必需的。我们创建了对应于Arg 286 β环结构域的折叠蛋白肽Arg 286 p和识别IL-1 RAcP Arg 286表面结构域的单克隆抗体。与在286位缺乏精氨酸的对照肽相比，我们的Arg 286肽化合物在细胞培养模型中以剂量依赖性方式消除了白细胞介素信号传导。Arg 286肽和抗Arg 286 mAb还以剂量依赖性方式在体外消除了IL-1β、IL-1 RI和IL-1 RAcP 3-way复合物的形成。IL 1-RAcP是一种辅助蛋白，在白细胞介素配体与受体结合后被招募，信号传导需要这种三向复合物形成的所有成员。我们的新抑制剂Arg 286 p模拟IL 1-RacP的精氨酸286 β环上的一个小的关键多价相互作用区域，从而消除复合物的形成并阻止下游炎症信号传导。由于Arg 286 p在受体-配体结合的下游起作用，即使面对过量的配体，它也可以大规模地放大白介素治疗的效力，并且可能上级竞争性靶向配体的现有IL-1治疗或与之协同。根据该目标，我们将在该先导化合物中进行新的结构修饰，以优化其对IL-1的亲和力和稳定性，并将其用作基础，以创建对IL 33和IL 36特异性的新型药物，其临床抑制剂不存在。我们将在一个新的迭代工作流程中使用我们的蛋白质绘制技术，以指导肽调节剂（激动剂和拮抗剂）的设计，这些肽调节剂在三种类型的测定中对IL-1的效力进行优化或对IL-33和IL-36具有特异性。我们的先导化合物的优化版本将在一个完善的骨关节炎动物模型中进行研究。骨关节炎导致2700万美国人遭受痛苦。