Discovery and engineering of novel anti-IgE disruptive inhibitors

项目摘要 大多数过敏反应是由抗过敏原IgE抗体引起的，其使过敏效应物敏感 细胞如肥大细胞和嗜碱性粒细胞。与IgG抗体相反，IgE以亚纳摩尔亲和力结合至IgG抗体。 高亲和力IgE受体（FceRI），并在过敏效应细胞上以受体结合形式持续数月 甚至在高亲和力抗IgE抗体抑制剂如奥马珠单抗存在下。我们已经证明 抗体和设计的锚蛋白重复蛋白（DARPins）都能够动力学地加速 IgE通过两种机制从FceRI解离： 受体邻近抑制剂，以促进受体解离和变构机制，限制IgE的一个 与受体结合不相容的构象。我们开发了一种酵母展示方法， 能够破坏受体复合物的抗IgE抑制剂，并证明了我们选择更有效的 奥马珠单抗的抗IgE变体。此外，我们已经证明了工程更有效的途径， 由破坏性抗IgE结构域和非竞争性IgE锚定结构域组成的二价破坏性抑制剂 域在这里，我们建议将这些方法应用于询问抗IgE免疫文库，以分离更多 有效的破坏性抑制剂，探索破坏性抑制剂的机制多样性，并开发 与未来临床开发相容的超强效二价抗IgE抑制剂。

Project Summary The majority of allergic reactions are caused by anti-allergen IgE antibodies, which sensitize allergic effector cells such as mast cells and basophils. In contrast to IgG antibodies, IgE binds with subnanomolar affinity to the high affinity IgE receptor (FceRI) and persists in a receptor-bound form on allergic effector cells for months even in the presence of high affinity anti-IgE antibody inhibitors, such as omalizumab. We have shown that both antibodies and Designed Ankyrin Repeat Proteins (DARPins) are able to kinetically accelerate the dissociation of IgE from FceRI through two mechanisms: a facilitated dissociation mechanism that allows receptor-adjacent inhibitors to promote receptor dissociation and an allosteric mechanism that restricts IgE to a conformation incompatible with receptor binding. We have developed a yeast display approach to selecting anti-IgE inhibitors capable of disrupting receptor complexes and demonstrated our ability to select more potent anti-IgE variants of omalizumab. In addition, we have demonstrated pathways to engineering more potent bivalent disruptive inhibitors consisting of a disruptive anti-IgE domain and a non-competitive IgE anchoring domain. Here we propose to apply these approaches to interrogate an anti-IgE immune library, to isolate more potent disruptive inhibitors, to explore the mechanistic diversity of disruptive inhibitors and to develop ultrapotent bivalent anti-IgE inhibitors compatible with future clinical development.