Lupus‐like anticoagulant properties of murine monoclonal antibodies to β₂‐glycoprotein I

Abstract

The lipid-binding inhibitor of coagulation, beta 2-glycoprotein I (beta 2GPI), has been shown to form the antigen to which some autoantibodies against anionic phospholipids (aPL) are directed. Six murine monoclonal antibodies (MAbs) of the IgG1 isotype were raised against human beta 2GPI and could be subdivided into three groups on the basis of mutual competition experiments. MAbs 9G1 and 8C3 (group A) markedly inhibited the binding of immunoglobulins from aPL-positive sera to beta 2GPI-coated wells. Using a lipid-based solid-phase radioimmunoassay, the MAbs interacted with both anionic phospholipids and phosphatidylethanolamine, but not phosphatidylcholine, in a beta 2GPI-dependent manner. A cross-reaction between beta 2GPI from several (including bovine) species was seen with one of the MAbs (9G1). All six MAbs induced dose-dependent prolongation of the DAPTT, DRVVT, KCT and TTI clotting times of human plasma, whereas 9G1 was the sole antibody to be inhibitory with plasma from bovine origin. Synergistic inhibitory effects were observed with MAbs used in pairs provided that they did not compete with each other for beta 2GPI binding. The anticoagulant activity of the MAbs was fully neutralized by the addition of freeze-thawed platelets. The MAbs described here resemble lupus anticoagulants in several respects which makes them valuable to study the involvement of beta 2GPI in the autoimmune thrombotic pathophysiology.

Anticardiolipin IgG subclasses association of IgG2 with arterial and/or venous thrombosis

“…The importance of immunoglobulin receptors as potential initiators of thrombosis is supported by 1) emerging evidence that aCL binding to platelets and neutrophils leads to activation via cellular FcyR (33,34), 2) the precedent of an FcyR-dependent mechanism of thrombosis in heparin-induccd thrombocytopenia (35), and 3) the case reports of successful treatment of APS with high-dose intravenous gamma globulin (36,37). The presence of IgG2 aCL in association with FcyRIIA-H131 may therefore be a useful clinical predictor of increased thrombotic risk in patients with autoimmune IgG aCL.…”

TLR2 Is One of the Endothelial Receptors for β2-Glycoprotein I

“…Two anti-b2-gpI mAbs were supplied by Josiane Arvieux (Brest, France): the 9G1 mAb that recognizes both human and bovine b2-gpI, and the 8C3 mAb that is specific for human b2-gpI (35). The 6C4 mAb (36) directed to Lactoferin-binding protein A and specific for late endosomes was donated by Jean Gruenberg (Geneva, Switzerland).…”

“…An ELISA was developed to quantitate specifically human b2-gpI (35). It combined the capture human-specific anti-b2-gpI 8C3 mAb, a rabbit anti-b2-gpI Ab, and the developing HRP-goat anti-rabbit Ig Ab (Jackson ImmunoResearch Laboratories).…”

“…It combined the capture human-specific anti-b2-gpI 8C3 mAb, a rabbit anti-b2-gpI Ab, and the developing HRP-goat anti-rabbit Ig Ab (Jackson ImmunoResearch Laboratories). Using serial dilutions of human b2-gpI, a standard curve was drawn (35,38). Aliquots of human b2-gpI and Cohn Fraction II (CFII) were biotinylated using the EZ-link kit (Pierce, Rockford, IL).…”

A monoclonal IgG anticardiolipin antibody from a patient with the antiphospholipid syndrome is thrombogenic in mice.

“…Some studies showed that the binding of ACA to cardiolipin (CL) was markedly enhanced by a plasma protein, (32 glycoprotein 1 (f32GP1), suggesting that ACA reacted with phospholipidf32GP1 complexes- (11)(12)(13)(14)(15). In contrast, other laboratories reported that these antibodies bound f32GP1 alone (16)(17)(18)(19)(20)(21)(22). However, some laboratories failed to detect similar direct anti-(32GP1 binding (15,23).…”