Engineering Hetero-Bivalent Ligands to Inhibit IgE Clustering on Mast Cells

描述（由申请人提供）：工程化异源二价配体以抑制肥大细胞上的IgE聚集项目概述/摘要过敏是一种免疫系统疾病，其由过敏原介导的免疫球蛋白E（IgE）分子在肥大细胞和嗜碱性粒细胞表面上的聚集引起。据估计，多达1200万美国人患有食物过敏，1000万患有过敏性哮喘，1000万患有特应性湿疹，3600万患有过敏性湿疹， 有过敏性鼻炎，而且患病率还在上升。目前的大多数治疗方法都是 治疗过敏相关的不良免疫反应涉及免疫系统的非特异性抑制。这导致感染性疾病的风险增加，突出表明需要开发有选择性的靶向治疗替代方案。 多价结合相互作用提供更高的亲和力（称为亲合力），并导致更强的相互作用。早期的研究已经确定，二价结合，如在抗体的情况下，可以增强相互作用的亲合力，比单价结合高六个数量级。我们在这个项目中的总体目标是设计异源二价配体，其将竞争性和选择性地抑制过敏原与肥大细胞上的IgE结合，从而抑制IgE聚集和过敏反应。这将通过设计同时靶向位于IgE的Fab结构域上的两个附近结合位点的异源二价分子来实现：抗原结合位点;和不那么熟知的“核苷酸结合位点”。同时与这两个位点的二价结合将为异源二价配体提供对靶IgE的增强的亲合力和选择性，并且将能够竞争性抑制过敏原与IgE的结合。这些异二价配体将被设计为具有两个部分-抗原分子本身（或模拟表位）和核苷酸结合位点靶向分子-通过柔性接头分子共价连接。具体地，我们将：i）鉴定将靶向 IgE上的非常规核苷酸结合位点; ii）设计和合成“半抗原/核苷酸-类似物”缀合物的异二价配体; iii）测定异二价配体- IgE相互作用的Kd并评价它们对过敏原与IgE结合的抑制作用和iv）在体外细胞测定中测定异二价配体抑制过敏反应的功效。

DESCRIPTION (provided by applicant): Engineering Hetero-Bivalent Ligands to Inhibit IgE Clustering on Mast Cells Project Summary/Abstract Allergy is a disorder of the immune system, which is elicited by the allergen-mediated-clustering of the immunoglobulin E (IgE) molecules on the surface of mast cells and basophils. It is estimated that up to 12 million Americans have food allergies, 10 million suffer from allergic asthma, 10 million suffer atopic eczema, 36 million have allergic rhinitis, and the prevalence is rising. The majority of the current therapies used to treat allergy related adverse immunologic responses involve non-specific suppression of the immune system. This results in the increased risk of infectious diseases, highlighting the need to develop selectively targeting treatment alternatives. Multivalent binding interactions provide higher affinity (termed avidity), and result in stronger interactions. Earlier studies have established that bivalent binding, as in the case of antibodies, can strengthen the avidity of an interaction up by six orders of magnitude over monovalent binding. Our overall goal in this proposed project is to engineer hetero-bivalent ligands that will competitively and selectively inhibit allergen binding to IgEs on mast cells, thereby inhibit IgE clustering and allergic responses. This will be accomplished via designing hetero-bivalent molecules that simultaneously target two nearby binding sites located on the Fab domain of an IgE: the antigen binding site; and the not-so-well-known "nucleotide binding site". Simultaneous bivalent binding to both these sites will provide the hetero-bivalent ligand with enhanced avidity and selectivity for the target IgE, and will enable competitive inhibition of allergen binding to the IgEs. These hetero-bivalent ligands will be designed to have two moieties - the antigenic molecule itself (or the mimotope), and the nucleotide binding site-targeting molecule - covalently linked by a flexible linker molecule. Specifically we will: i) identify nucleotide analogs that will target the unconventional nucleotide binding site on the IgE; ii) design and synthesize hetero-bivalent ligands of 'hapten/nucleotide-analog' conjugates; iii) determination of the Kd for hetero-bivalent ligand - IgE interactions and evaluation of their inhibitory effect on the allergen binding to IgEs and iv) determine the efficacy of the hetero-bivalent ligands to inhibit allergic responses in vitr cellular assays.