Mechanism and engineering of IgG-based monoclonal antibody/receptor interactions

基于 IgG 的单克隆抗体/受体相互作用的机制和工程

• 批准号: 9300976
• 负责人: Adam Wesley Barb
• 金额: $ 28.74万
• 依托单位: IOWA STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2020-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9300976
• 关键词: Affect; Affinity; Amino Acids; Anti-Inflammatory Agents; Anti-inflammatory; Antibodies; Asparagine; Autoimmune Diseases; Binding; Binding Sites; Biochemistry; Carbohydrates; Cells; Development; Disease; Engineering; Fc Receptor; Glycoproteins; Goals; Graft Rejection; Hematologic Neoplasms; Human; Immune; Immune system; Immunoglobulin G; Immunoglobulins; Inflammatory; Knowledge; Laboratories; Lead; Libraries; Link; Malignant Neoplasms; Mass Spectrum Analysis; Measures; Modification; Molecular; Molecular Conformation; Monoclonal Antibodies; Motion; Mutation; N-Glycosylation Site; NMR Spectroscopy; Patients; Pharmacologic Substance; Polysaccharides; Positioning Attribute; Preparation; Process; Production; Property; Protein Engineering; Proteins; Public Health; Publishing; Reporting; Resolution; Rheumatoid Arthritis; Role; Sampling; Site; Solid Neoplasm; Stable Isotope Labeling; Structure; Surface; System; Systemic Lupus Erythematosus; Techniques; Testing; Therapeutic Monoclonal Antibodies; Therapeutic antibodies; Transplant-Related Disorder; Variant; Work; X-Ray Crystallography; antibody-dependent cell cytotoxicity; base; cancer therapy; design; experimental study; improved; leukemia/lymphoma; novel; novel drug class; novel therapeutics; pathogen; personalized medicine; polypeptide; public health relevance; receptor; receptor binding; response; therapeutic evaluation; tool

 DESCRIPTION (provided by applicant): The fragment crystallizable (Fc) region links the dual pathogen identification and destruction properties of immunoglobulin G (IgG). Pathogen opsonization positions Fcs to activate pro-inflammatory Fc¿ receptors (FcgRs) on immune cells. Asparagine-linked (N)-glycan attached to Fc is required for productive engagement of the low-affinity FcgRs, though it is not known how the Fc N-glycan contributes to FcγR binding because the N-glycan does not directly contact the FcgRs. It has been suggested that the N-glycan provides optimal spacing of two Fc domains, stabilizing the Fc quaternary structure to bind the FCγR. Evidence from our laboratory points to a different hypothesis. We determined that Fc N-glycan motion, increased by Fc amino acid mutations far from the FcγR binding site, negatively correlated with Fc¿RIIIa affinity. Only a single region of Fc, the CE polypeptide loop that contains the site of N-glycosylation, was perturbed as a result of these Fc mutations. This result led to the proposal that the N-glycan affects FcγR affinity by pre-organizing the CE loop, and not by optimizing Fc domain orientation. Here we will directly test our hypothesis by measuring the structure and motion of the CE loop, in multiple forms stabilized through glycan or protein engineering, using solution nuclear magnetic resonance spectroscopy. The knowledge of CE loop structure and motion will be applied to redesign the Fc polypeptide to generate aglycosylated Fc variants that maintain high affinity for FcgRs. The molecular details of immune system activation that will emerge from these studies will be important to understand the process of multiple diseases and will be critical to enhance therapeutic monoclonal antibody function through engineering to treat cancer, transplant and autoimmune disease patients.

 描述（由申请方提供）：可结晶片段（Fc）区将免疫球蛋白G（IgG）的双重病原体识别和破坏特性联系起来。病原体调理作用定位Fc以激活免疫细胞上的促炎性Fc受体（FcgR）。与Fc连接的天冬酰胺连接（N）-聚糖是低亲和力FcgR有效结合所必需的，但尚不清楚Fc N-聚糖如何促进FcγR结合，因为N-聚糖不直接接触FcgR。已表明N-聚糖提供了两个Fc结构域的最佳间距，稳定了Fc四级结构以结合FCγR。我们实验室的证据指向一个不同的假设。我们确定，Fc N-聚糖的运动，通过远离FcγR结合位点的Fc氨基酸突变而增加，与Fc <$RIIIa亲和力呈负相关。这些Fc突变仅干扰了Fc的单个区域，即含有N-糖基化位点的CE多肽环。这一结果导致提出N-聚糖通过预组织CE环影响FcγR亲和力， 通过优化Fc结构域方向。在这里，我们将通过测量CE环的结构和运动来直接测试我们的假设，通过聚糖或蛋白质工程以多种形式稳定，使用溶液核磁共振光谱。CE环结构和运动的知识将被应用于重新设计Fc多肽以产生维持对FcgR的高亲和力的无糖基化Fc变体。从这些研究中出现的免疫系统激活的分子细节对于理解多种疾病的过程将是重要的，并且对于通过工程改造来增强治疗性单克隆抗体功能以治疗癌症、移植和自身免疫性疾病患者将是至关重要的。