Structure, function and engineering of immune cytokine receptor signaling

免疫细胞因子受体信号传导的结构、功能和工程

基本信息

批准号：
9297185
负责人：
Kenan Christopher GARCIA
金额：
$ 39.25万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-04-01 至 2018-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9297185
关键词：
Address Affinity Antibodies Architecture Autoimmunity Award Binding Biological Biology Biophysics Cells Chemistry Chemosensitization Complement Complex Cryoelectron Microscopy Crystallography Cytokine Receptors Cytokine Signaling Development Diabetes Mellitus Dimerization Disease model Electron Microscopy Engineering Evolution Exhibits Experimental Autoimmune Encephalomyelitis Extracellular Structure Gap Junctions Gene Expression Geometry Goals Grant Homeostasis Human IL6ST gene Image Immune Immune System Diseases Immunotherapeutic agent Immunotherapy In Vitro Instruction Interferons Interleukin 2 Receptor Interleukin 2 Receptor Gamma Interleukin 6 Receptor Interleukin-1 Interleukin-13 Interleukin-15 Interleukin-17 Interleukin-2 Interleukin-4 Interleukin-6 Janus kinase Knowledge Left Length Ligand Binding Ligands Link Malignant Neoplasms Mediating Molecular Molecular Conformation Molecular Profiling Outcome Paint Pharmaceutical Preparations Play Privatization Property Protein Engineering Proto-Oncogene Proteins c-akt Receptor Signaling Regulatory T-Lymphocyte Reporting Resolution Role STAT protein Signal Pathway Signal Transduction Specificity Stat5 protein Structural Protein Structure Structure-Activity Relationship System T-Lymphocyte Subsets Techniques Testing Therapeutic Time Toxic effect Translating Variant X-Ray Crystallography base cell type combinatorial conformer cytokine cytokine receptor gp130 design dimer extracellular functional outcomes graft vs host disease human disease immune system function immunoregulation improved in vivo in vivo Model insight interleukin-15 receptor interleukin-23 interleukin-6 receptor alpha leukocyte proliferation melanoma network dysfunction novel pleiotropism preference public health relevance receptor receptor binding reconstitution reconstruction structural biology

项目摘要

DESCRIPTION (provided by applicant): Immunoregulatory cytokines engage transmembrane signaling receptors in order to mediate a wide range of functions including leukocyte proliferation, differentiation, and expansion. Most immunoregulatory cytokines possess both redundant and distinct activities that are critical to normal immune homeostasis, but this functional pleiotropy presents a major problem for the effective, targeted use of these cytokines as drugs. Cytokine pleiotropy is a consequence of a small number of shared receptors, such as common gamma chain and gp130, engaging many different cytokines, which then activate overlapping intracellular signaling pathways through a limited set of JAK and STAT proteins. During the prior term of this award, we gained an appreciation for the extracellular structural architectures of a spectrum of different cytokine complexes with shared receptors, including those of IL-1, IL-2, IL-4, IL-6, IL-13, IL-17, IL-23, and IFN, which exhibited an astonishing diversity of heterodimeric signaling geometries. In this renewal application, we focus our studies on the pleiotropic cytokines IL-2 and IL-15, to ask how extracellular structures of the receptor-cytokine complexes influence transmembrane signaling, intracellular activation of JAK and STAT, and subsequent in vivo function. We wish to determine if the binding chemistry and geometry of the IL-2 and IL-15 receptor complex subunits plays a role in modulating signaling specificity, and whether "tuning" signaling through structure-based cytokine engineering of receptor interactions is a viable means of developing novel immunotherapeutics with enhanced efficacy, cell subset preferences, and reduced toxicity. The overall goals of this highly collaborative proposal are: Aim 1- to determine the biophysical basis for the functional redundancy and specificity exhibited by two "natural surrogate" γc cytokines, IL-2 and IL-15, that act through shared signaling receptors (IL-2Rß and γc) but private alpha-receptors; Aim 2- to utilize structure-based protein engineering to attempt to create IL- 2 variants with diverse signaling properties and T cell subset preferences, that may be more effective immunotherapeutics as assessed by collaborators in a variety of in vivo disease models; and Aim 3- to determine the mechanistic basis of antibody potentiation of IL-2 activity towards distinct T cell subsets, as well as discover new potentiating antibodies that could remodel the conformation of wild-type IL-2 and alter its biological activity. Finally, in Aim 4, we continue to pursue structural information on how cytokine receptor intracellular segments engage Janus Kinase (JAK) molecules, by reconstituting an entire full-length cytokine receptor transmembrane complex, bound to both cytokine and intracellular JAK for imaging by crystallography and electron microscopy. In this fashion, by combining structure (X-ray crystallography, Electron Microscopy, and NMR), protein engineering, signaling, and in vivo studies, we propose to obtain a complete molecular snapshot of shared cytokine receptor signaling from the initial engagement of ligand through the activation of intracellular signaling cascades.

描述（由申请人提供）：免疫调节细胞因子与跨膜信号传导受体结合，以介导多种功能，包括白细胞增殖、分化和扩增。大多数免疫调节细胞因子具有对正常免疫稳态至关重要的冗余和独特的活性。功能多效性对这些细胞因子作为药物的有效性和针对性使用提出了一个主要问题。细胞因子多效性是少数共享的结果。受体，例如常见的伽马链和 gp130，与许多不同的细胞因子结合，然后通过一组有限的 JAK 和 STAT 蛋白激活重叠的细胞内信号传导途径。一系列具有共享受体的不同细胞因子复合物，包括 IL-1、IL-2、IL-4、IL-6、IL-13、IL-17、IL-23 和 IFN 的复合物，表现出惊人的多样性在这个更新应用中，我们将研究重点放在多效性细胞因子 IL-2 和 IL-15 上，以探究受体-细胞因子复合物的细胞外结构如何影响跨膜信号传导、JAK 和 STAT 的细胞内激活以及随后的影响。我们希望确定 IL-2 和 IL-15 受体复合物亚基的结合化学和几何结构是否在调节信号传导特异性中发挥作用，以及是否通过“调节”信号传导。基于结构的细胞因子受体相互作用工程是开发新型免疫疗法的可行方法，该疗法具有增强的功效、细胞亚群偏好和降低的毒性。这一高度协作提案的总体目标是：目标 1-确定功能冗余的生物物理基础。两种“天然替代”γc 细胞因子 IL-2 和 IL-15 表现出特异性，它们通过共享信号受体（IL-2Rß 和 γc）但通过私有 α 受体发挥作用； 2- 利用基于结构的蛋白质工程尝试创建具有不同信号特性和 T 细胞亚群偏好的 IL-2 变体，根据合作者在各种体内疾病模型中的评估，这可能是更有效的免疫治疗方法；目标 3-确定抗体对不同 T 细胞亚群增强 IL-2 活性的机制基础，并发现可以重塑野生型 IL-2 构象并改变其生物活性的新增强抗体。我们通过重建完整的全长细胞因子受体跨膜复合物，与细胞因子和细胞内 JAK 结合，通过晶体学和电子显微镜进行成像，继续寻求细胞因子受体细胞内片段如何与 Janus 激酶 (JAK) 分子结合的结构信息。，通过结合结构（X射线晶体学、电子显微镜和核磁共振）、蛋白质工程、信号传导和体内研究，我们建议从最初的参与中获得共享细胞因子受体信号传导的完整分子快照通过激活细胞内信号级联来调节配体。