B-CELL RESPONSES TO CLOTTING FACTOR VIII

描述(由申请人提供)：作为一种致病的B细胞反应，第八因子抑制剂是一个主要的临床挑战，包括血友病A患者的同种抗体或获得性血友病的自身抗体。对不同结构域上有限数量的主要表位的反应是多克隆的，而IgG4反应通常占主导地位。然而，大多数关于因子抑制剂的研究都是在代表单个时间点的样本上进行的，集中在高滴度抗体的峰值反应，并定义了与整个领域的表位反应的组份。我们建议研究针对因子VII1 C区域上的表面表位簇的多克隆反应的一部分，以表征受试者之间的可变性和个体患者随时间的稳定性(或变化)。我们将研究三组患者：(1)重度血友病A中的异体抗体，(2)获得性血友病A中的自身抗体，以及(3)轻度血友病患者中自身和异体抗体的组合。C区表位仍然是最复杂和最不了解的。亲和纯化的抗体部分与C区表位簇结合将被表征。我们已经表达了C2和C1C2以及几个突变体。将评估与C2、其突变体或C1C2反应的抑制和/或结合患者抗体的相对量，以及在患者之间和在给定患者样本中纵向分布的免疫球蛋白同型分布。特定的血友病C1C2突变将表征使用抑制剂的中度重度患者的反应。作为最后的组成部分，我们将定义C1C2‘S表面的结构元素，以及C2上残基的构象灵活性，即当整个C结构域存在时，表面残基的不同程度。结果将为治疗出血发作和诱导耐受或预防同种异体免疫的策略提供洞察力。

DESCRIPTION (provided by applicant): As a pathogenic B cell response, factor VIII inhibitors represent a major clinical challenge including alloantibodies in hemophilia A patients or autoantibodies in acquired hemophilia. Responses are polyclonal to a limited number of major epitopes on different domains and IgG4 responses often predominate. However, most studies to characterize factor VIII inhibitors have been on samples representing single points in time, focusing upon high titer antibodies at their peak response and defining fractions reacting with epitopes throughout entire domains. We propose to study fractions of the polyclonal responses that are directed to clusters of surface epitopes on factor VIIl's C domains to characterize variability between subjects and stability (or changes) across time in individual patients. We will study three sets of patient groups: (1) allo antibodies in severe hemophilia A, (2) auto antibodies in acquired hemophilia A and (3) combined auto and allo antibodies in patients with mild hemophilia A. C domain epitopes remain the most complex and least well understood. Affinity-purified antibody fractions binding to C domain epitope clusters will be characterized. We have expressed C2 and C1C2 and several mutants. Relative amounts of inhibiting and/or binding patient antibodies reacting to C2, its mutants or C1C2 will be assessed as will distribution of IgG isotypes, both between patients and longitudinally in a given patient's samples. Specific hemophilic C1C2 mutants will characterize responses in mildly severe patients with inhibitors. As a final component, we will define structural elements of C1C2's surface, and the extent of flexibility in conformations of residues on C2, surface residues that differ when the entire C domain is present. Results will provide insights into strategies for therapy of bleeding episodes and the induction of tolerance or prevention of alloimmunization.