Engineering mono-fucosylated IgGs to fine-tune antibody-mediated effector functions

工程化单岩藻糖基化 IgG 来微调抗体介导的效应功能

• 批准号: 10647938
• 负责人: ERIC JOHN SUNDBERG
• 金额: $ 19.56万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-03 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10647938
• 关键词: Affinity; Antibodies; Antigens; Autoimmunity; Binding; Biochemical Reaction; Biological; Biophysics; Cells; Chemical Structure; Clinical; Clinical effectiveness; Complement 1q; Complex; Complex Mixtures; Disease; Endoglycosidases; Endowment; Engineering; Equilibrium; Exhibits; FDA approved; Fab domain; Fc domain; Fucose; Generations; IgG Receptors; IgG1; Immune signaling; Immune system; Immunoglobulin Constant Region; Immunoglobulin G; Immunologic Receptors; Immunologic Stimulation; Immunotherapeutic agent; Infection; Inflammatory; Kinetics; Link; Malignant Neoplasms; Mannose; Measures; Mediating; Methods; Molecular; Monoclonal Antibodies; Nature; Pharmaceutical Preparations; Pharmacotherapy; Polysaccharides; Property; Reaction; Research; Resolution; Structure; Therapeutic Monoclonal Antibodies; Thermodynamics; Time; Toxic effect; Treatment-related toxicity; Variant; Work; antibody engineering; antibody-dependent cell cytotoxicity; antigen binding; cancer therapy; cell killing; clinical efficacy; clinically relevant; design; experimental study; glycosylation; human disease; in vivo; innovation; macromolecule; next generation; novel; recruit; sialylation; structural biology; sugar

Monoclonal IgG antibodies (mAbs) constitute a critically important class of drugs for the treatment of a wide range of diseases. Their abilities to recruit and stimulate immune system cells, which is required for their clinical effectiveness, especially in the immunotherapeutic treatment of cancer, are harbored in their Fc domains. For clinically relevant IgG antibodies, Fc domains engage Fc γ receptors (FcγRs) and complement C1q in order to induce antibody-mediated effector functions that direct the killing of cells in vivo. Methods to engineer IgG Fc domains to manipulate their in vivo killing capacities lag substantially behind those for customizing their antigen- binding Fab domains due to the presence of a conserved N-linked glycan in the Fc domain at residue Asn297 that is overwhelmingly the most important molecular determinant of FcγR and C1q binding. The next generation of immunotherapeutic mAbs depends on our ability to efficiently and rationally modify the chemical structure of this Asn297-linked glycan. The most important molecular feature of this glycan is a fucose sugar unit connected through an α-1,6 linkage to the Asn-proximal N-acetylglucosamine (GlcNac) saccharide. The absence of this core fucose moiety imparts Fc domains with increased binding affinity to FcγR3A, an activating FcγR, resulting in substantially increased antibody-mediated in vivo cellular killing. Naturally produced antibodies, though, are fucosylated and, thus, numerous methods have been developed to create antibodies without fucosylation, resulting in three afucosylated IgG1 antibodies that have recently been approved by the FDA. Antibodies with a fucose on the Asn297-linked glycan on one Fc chain but not on the other – mono-fucosylated antibodies – present a third fucosylation state that could provide fine-tuning of antibody-mediated effector functions, thereby expanding the ability to balance efficacy and toxicity for the treatment of a wide range of diseases. However, mono-fucosylated antibodies do not exist in nature and have never been produced or engineered. Here we propose methods to create mono-fucosylated IgG antibodies for the first time and to assess the biological consequences of antibody mono-fucosylation. We hypothesize that mono-fucosylated IgG antibodies will exhibit biophysical and functional properties distinct from those of fully afucosylated and di-fucosylated IgG antibodies, which can be leveraged to develop a new class of immunotherapeutic monoclonal antibodies that will induce levels of antibody-mediated effector functions optimal for certain clinical indications. This work is significant because it develops and explores an entirely new class of engineered antibodies that could become important for the immunotherapeutic treatment of cancer. The proposed studies are innovative in that they will investigate an entirely novel concept – mono-fucosylated antibodies – with the potential to manipulate antibody-mediated effector functions in ways that have never been explored before. The research plan will be accomplished by leveraging our expertise in IgG-specific glycan remodeling, molecular biophysics and structural biology.

单抗是一类非常重要的治疗多种疾病的药物。 一系列疾病。他们招募和刺激免疫系统细胞的能力，这是他们的临床所需 有效性，特别是在癌症的免疫治疗方面，隐藏在它们的Fc区域。为 临床上相关的免疫球蛋白抗体，Fc结构域与Fcγ受体(FcγRs)和补体C1q结合，以便 诱导抗体介导的效应器功能，指导体内细胞的杀伤。工程免疫球蛋白Fc的几种方法 用于操纵其体内杀伤能力的结构域大大落后于用于定制其抗原的结构域- 由于Fc结构域在Asn297残基上存在保守的N-连接糖链而导致Fab结构域结合 这是Fc、γ、R和C1q结合的最重要的分子决定因素。下一代 免疫治疗性单抗的成功与否取决于我们能否有效和合理地改变单抗的化学结构 这种Asn297连接的多糖。这种多糖最重要的分子特征是连接了一个岩藻糖单元 通过α-1，6连接到天冬氨酸近端的N-乙酰氨基葡萄糖(GlcNAc)糖。这一点的缺失 核心岩藻糖部分赋予Fc结构域与FcγR3A(激活FcγR)的结合亲和力增加，从而 显著增加抗体介导的体内细胞杀伤率。然而，自然产生的抗体是 岩藻糖化并因此开发了许多方法来产生没有岩藻糖化的抗体， 产生了三种果糖基化的IgG1抗体，最近已被FDA批准。带有A的抗体 一条Fc链上的Asn297连接的多糖上有岩藻糖，而另一条链上没有岩藻糖化的单链抗体- 呈现第三种岩藻糖化状态，可以提供抗体介导的效应器功能的微调，从而 扩大平衡治疗多种疾病的疗效和毒性的能力。然而， 单岩藻糖化抗体在自然界中并不存在，也从未被生产或改造过。在这里我们 首次提出了制备岩藻糖基化抗体的方法，并对其生物学活性进行了评价。 抗体单岩藻糖基化的后果。我们假设单岩藻糖基化的免疫球蛋白抗体将表现出 生物物理和功能特性不同于完全果糖基化和双岩藻糖基化的抗体， 它可以被用来开发一种新的免疫治疗性单抗，它将诱导 抗体介导的效应器功能水平对于某些临床适应症是最佳的。这项工作意义重大 因为它开发和探索了一种全新的工程抗体，这种抗体可能会成为重要的 用于癌症的免疫治疗。拟议的研究是创新的，因为他们将调查 一种全新的概念-单岩藻糖化抗体-有可能操纵抗体介导的 效应器以前所未有的方式发挥作用。研究计划将通过以下方式完成 利用我们在免疫球蛋白特异性糖链重塑、分子生物物理学和结构生物学方面的专业知识。