Multi-Angle Effector Function Analysis of Human Monoclonal IgG Glycovariants.
Multi-Angle Effector Function Analysis of Human Monoclonal IgG Glycovariants.
复制标题
DOI：
10.1371/journal.pone.0143520
复制
发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Schlothauer T
中科院分区：
文献类型：
--
作者：
Dashivets T;Thomann M;Rueger P;Knaupp A;Buchner J;Schlothauer T
重组抗体的治疗性能依赖于两种独立的机制：抗原识别和Fc介导的抗体效应功能。 Fc 片段与不同 FcR 的相互作用触发抗体依赖性细胞毒性和补体依赖性细胞毒性，并决定抗体在血清中的寿命。在治疗性抗体中，FcγR 发挥着最重要的作用。已证明，Fc 连接的糖部分对于 IgG 效应子功能至关重要，决定其与单个 FcγR 的亲和力，并决定与不同受体类别的结合：激活或抑制。在这项研究中，我们系统地分析了单克隆 IgG1 及其八种酶促糖基化变体的效应子功能。对单一糖变体以及免疫复合物形式的抗原结合抗体和 IgG 进行了糖变体与 FcR 相互作用的分析。除了功能特性之外，我们还讨论了糖基化对测试糖变体结构特性的影响。我们证明了糖基化模式对抗体稳定性以及与不同 FcγR 相互作用的明显影响。与之前的报道一致，去糖基化抗体未能结合所有 Fcγ 受体，高亲和力 FcγRI 除外。观察到 IgG1 的 FcγRII 和 FcγRIIIa 结合活性取决于半乳糖基化水平，并且高半乳糖基化抗体表现出增强的受体相互作用。唾液酸化不会降低测试 IgG 的 FcγR 结合；相反，抗体的唾液酸化改善了与 FcγRIIa 和 IIb 的结合。我们证明糖基化在一定程度上影响 IgG1 与 FcRn 的相互作用。然而，与糖基化模式无关，IgG1 与可溶性单体靶标的相互作用令人惊讶地导致受体结合受损。在这里，我们证明，由多聚体配体诱导的免疫复合物（IC）补偿了靶标结合抗体对FcR亲和力的降低，显示了IC形成对于FcR介导的效应器功能的重要性。
Therapeutic performance of recombinant antibodies relies on two independent mechanisms: antigen recognition and Fc-mediated antibody effector functions. Interaction of Fc-fragment with different FcR triggers antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity and determines longevity of the antibody in serum. In context of therapeutic antibodies FcγRs play the most important role. It has been demonstrated that the Fc-attached sugar moiety is essential for IgG effector functionality, dictates its affinity to individual FcγRs and determines binding to different receptor classes: activating or inhibitory. In this study, we systematically analyze effector functions of monoclonal IgG1 and its eight enzymatically engineered glycosylation variants. The analysis of interaction of glycovariants with FcRs was performed for single, as well as for antigen-bound antibodies and IgGs in a form of immune complex. In addition to functional properties we addressed impact of glycosylation on the structural properties of the tested glycovariants. We demonstrate a clear impact of glycosylation pattern on antibody stability and interaction with different FcγRs. Consistent with previous reports, deglycosylated antibodies failed to bind all Fcγ-receptors, with the exception of high affinity FcγRI. The FcγRII and FcγRIIIa binding activity of IgG1 was observed to depend on the galactosylation level, and hypergalactosylated antibodies demonstrated increased receptor interaction. Sialylation did not decrease the FcγR binding of the tested IgGs; in contrast, sialylation of antibodies improved binding to FcγRIIa and IIb. We demonstrate that glycosylation influences to some extent IgG1 interaction with FcRn. However, independent of glycosylation pattern the interaction of IgG1 with a soluble monomeric target surprisingly resulted in an impaired receptor binding. Here, we demonstrate, that immune complexes (IC), induced by multimeric ligand, compensated for the decreased affinity of target bound antibody towards FcRs, showing the importance of the IC-formation for the FcR- mediated effector functions.