TY - JOUR
T1 - Patient-derived monoclonal antibodies directed towards beta(2) glycoprotein-1 display lupus anticoagulant activity
AU - Dienava-Verdoold, I.
AU - Boon-Spijker, M. G.
AU - de Groot, Philip G.
AU - Brinkman, H. J. M.
AU - Voorberg, Jan
AU - Mertens, K.
AU - Derksen, Ronald H. W. M.
AU - de Laat, B.
PY - 2011/4
Y1 - 2011/4
N2 - Patients with antiphospholipid syndrome (APS) display a heterogeneous population of antibodies with beta(2) glycoprotein-1 (?(2)GP1) as the major antigen.We isolated and characterized human mAbs directed against ?(2)GP1 from the immune repertoire of APS patients.Variable heavy chain repertoires from B cells from two APS patients with anti-?(2)GP1 antibodies were cloned into the pHEN1-VLrep vector. Constructed full-length IgG antibodies were tested for lupus anticoagulant (LAC) activity and binding to ?(2)GP1 and its domains.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 ?(2)GP1 and its isolated domain I. The affinity of selected antibodies for ?(2)GP1 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 ?(2)GP1. Both selected clones from patient 2 recognized domain I of ?(2)GP1, 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.In this study, we show successful cloning of patient-derived mAbs that require domain I of ?(2)GP1 for binding, and that display LAC activity that is dependent on their affinity for ?(2)GP1. 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.
AB - Patients with antiphospholipid syndrome (APS) display a heterogeneous population of antibodies with beta(2) glycoprotein-1 (?(2)GP1) as the major antigen.We isolated and characterized human mAbs directed against ?(2)GP1 from the immune repertoire of APS patients.Variable heavy chain repertoires from B cells from two APS patients with anti-?(2)GP1 antibodies were cloned into the pHEN1-VLrep vector. Constructed full-length IgG antibodies were tested for lupus anticoagulant (LAC) activity and binding to ?(2)GP1 and its domains.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 ?(2)GP1 and its isolated domain I. The affinity of selected antibodies for ?(2)GP1 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 ?(2)GP1. Both selected clones from patient 2 recognized domain I of ?(2)GP1, 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.In this study, we show successful cloning of patient-derived mAbs that require domain I of ?(2)GP1 for binding, and that display LAC activity that is dependent on their affinity for ?(2)GP1. 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.
KW - antiphospholipid syndrome
KW - beta(2) glycoprotein-1
KW - lupus anticoagulants
KW - mAb
KW - phage display
U2 - 10.1111/j.1538-7836.2011.04212.x
DO - 10.1111/j.1538-7836.2011.04212.x
M3 - Article
C2 - 21255251
SN - 1538-7933
VL - 9
SP - 738
EP - 747
JO - Journal of Thrombosis and Haemostasis
JF - Journal of Thrombosis and Haemostasis
IS - 4
ER -