Otsuki Takuya, Matsumoto Shigeyuki, Fujita Junso, Miyata Tomoko, Namba Keiichi, Kanada Ryo, Okuno Yasushi, Kamada Haruhiko, Ohno Hiroaki, Akiba Hiroki
Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto, Japan.
Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto, Japan.
J Biol Chem. 2025 Sep;301(9):110548. doi: 10.1016/j.jbc.2025.110548. Epub 2025 Jul 31.
Biparatopic antibodies (BpAbs) bind two different antigen epitopes to form characteristic immunocomplexes. Many BpAbs have been developed for enhanced crosslinking to induce signal transduction or cell internalization, whereas few have been reported with smaller immunocomplexes to suppress unwanted signaling. Here, we developed a strategy to induce 1:1 immunocomplex formation to maximize antagonistic function. Various peptide linkers were introduced into the hinge regions of IgG-like agonist BpAbs against tumor necrosis factor receptor 2. Loss of crosslinking activity was observed for one BpAb, allowing the conversion of its function from an agonist to an antagonist. However, crosslinking activity was retained for another agonist, BpAb, which binds to a different epitope pair. In a combined analysis of cryo-electron microscopy and coarse-grained molecular dynamics simulations, the effect of epitope combination on the stability of 1:1 complexes was observed. These results lead to an understanding of the mechanism and design of BpAbs to adopt a 1:1-binding mode.
双特异性抗体（BpAbs）结合两种不同的抗原表位以形成特征性免疫复合物。许多双特异性抗体已被开发用于增强交联以诱导信号转导或细胞内化，而报道中很少有形成较小免疫复合物以抑制不必要信号传导的双特异性抗体。在此，我们开发了一种策略来诱导1:1免疫复合物形成以最大化拮抗功能。将各种肽接头引入针对肿瘤坏死因子受体2的IgG样激动剂双特异性抗体的铰链区。观察到一种双特异性抗体的交联活性丧失，使其功能从激动剂转变为拮抗剂。然而，另一种与不同表位对结合的激动剂双特异性抗体保留了交联活性。在冷冻电子显微镜和粗粒度分子动力学模拟的联合分析中，观察到表位组合对1:1复合物稳定性的影响。这些结果有助于理解双特异性抗体采用1:1结合模式的机制和设计。
