Patient-derived monoclonal antibodies directed towards beta2 glycoprotein-1 display lupus anticoagulant activity

Abstract

Background:Patients with antiphospholipid syndrome (APS) display a heterogeneous population of antibodies with beta2 glycoprotein-1 (β2GP1) as the major antigen. Objectives:We isolated and characterized human mAbs directed against β2GP1 from the immune repertoire of APS patients. Methods:Variable heavy chain repertoires from B cells from two APS patients with anti-β2GP1 antibodies were cloned into the pHEN1-VLrep vector. Constructed full-length IgG antibodies were tested for lupus anticoagulant (LAC) activity and binding to β2GP1 and its domains. Results:Two clones of each patient were selected on the basis of the reactivity of single chain Fv (scFv) fragments displayed on phages towards full-length β2GP1 and its isolated domain I. The affinity of selected antibodies for β2GP1 was lost when transforming from phages to monovalent scFvs, and was regained when antibodies were constructed as complete IgG, indicating a role for bivalency in binding to β2GP1. Both selected clones from patient 2 recognized domain I of β2GP1, and for both clones selected from patient 1, binding required the presence of both domain I and domain II. All mAbs displayed LAC activity in both activated partial thromboplastin time-based and dilute Russell's viper venom test-based clotting assays and in thrombin generation. Conclusions:In this study, we show successful cloning of patient-derived mAbs that require domain I of β2GP1 for binding, and that display LAC activity that is dependent on their affinity for β2GP1. These antibodies can help us to gain more insights into the pathogenesis of APS, and may facilitate standardization of APS diagnosis. © 2011 International Society on Thrombosis and Haemostasis.