Glycoengineering of antibodies to modulate immune functions

项目摘要 抗体是一类重要的治疗药物，广泛用于治疗癌症，自身免疫 疾病和传染病。令人信服的实验数据表明，Fc糖基化是一种 调节抗体效应器功能的关键结构决定因素，包括抗体依赖 细胞毒性(ADCC)、补体依赖的细胞毒性(CDC)、激活细胞凋亡和抗 炎症活动。然而，了解抗体糖基化的功能作用的进展是 受制于Fc糖基化的巨大结构异质性。为了解决这个问题，我们有 开发了一种化学酶方法，允许位点特异性的Fc和Fab多糖重塑 均一抗体糖型。在这一应用中，我们建议开发位点特异性的化学酶 将糖链重塑方法作为关键平台技术来解决相关的三个重要问题 抗体的功能，如以下三个具体目标所强调的。目标1是了解FC如何 糖基化通过选择性修饰核心岩藻糖调节Fc受体结合和adcc活性，通过 进行结构研究，并通过合成新的糖形式，结合Fc受体和 ADCC检测。目的2是评价Fc糖链的位点特异性唾液酸化对抗体的影响。 通过构建不对称唾液酸化或多唾液酸化糖形式的炎症活性，结合 动物研究。目标3是通过构建增强抗体的补体依赖细胞毒(CDC) 结构明确的半乳糖/鼠李糖抗原抗体结合物募集天然Ig G和Ig M抗体 在流通中。这些研究将在糖免疫学方面产生重要的新知识，这将促进 开发更有效的基于抗体的疗法。

Project Summary Antibodies are an important class of therapeutics that are widely used for treatment of cancer, autoimmune diseases, and infectious diseases. Compelling experimental data have demonstrated that Fc glycosylation is a critical structural determinant for modulating antibody’s effector functions, including antibody-dependent cellular cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC), activation of apoptosis, and anti- inflammatory activities. However, progress in understanding the functional roles of antibody glycosylation is hampered by the tremendous structural heterogeneity of Fc glycosylation. To address this problem, we have developed a chemoenzymatic method that permits site-specific Fc and Fab glycan remodeling to generate homogeneous antibody glycoforms. In ths application, we propose to exploit the site-specific chemoenzymatic glycan remodeling method as a key platform technology to address three important questions related to antibody functions, as highlighted in the following three specific aims. Aim 1 is to understand how Fc glycosylation modulate Fc receptor binding and ADCC activity by selective modification of the core fucose, by performing structural studies, and by synthesizing novel glycoforms, coupled with Fc receptor binding and ADCC assays. Aim 2 is to evaluate how site-specific sialylation of Fc glycans affects antibody’s anti- inflammatory activity by constructing asymmetrically sialylated or multiply sialylated glycoforms, coupled with animal studies. Aim 3 is to augment antibody’s complement-dependent cytotoxicity (CDC) by constructing structurally well-defined Gal/rhamnose antigen-antibody conjugates to recruit natural IgG and IgM antibodies in circulation. These studies will yield important new knowledge in glyco-immunology, which will facilitate the development of more effective antibody-based therapeutics.