Partial Characterization of an Autoantibody Recognizing the Secondary Binding Site(s) of Thrombin in a Patient with Recurrent Spontaneous Arterial Thrombosis

Abstract

SummaryA patient with an 18 year history of recurrent arterial thrombosis and no evidence of atherosclerosis or embolism of cardiac origin presented with a prolonged thrombin clotting time when performed with human thrombin. The bovine thrombin clotting time was only slightly prolonged. During 30 months of follow-up, the thrombin time fluctuated, but remained prolonged. The patient has been treated with an oral anticoagulant for the past 8 years, with no thrombotic recurrence.The inhibitor activity was due to the presence of polyclonal IgGs which bound to thrombin-Sepharose. The influence of IgGs purified from the patient’s serum was compared to the influence of normal IgGs in several systems exploring the catalytic activity of thrombin and the binding of the enzyme to macromolecular substrates through secondary binding site(s). We found that the IgGs did not impair the catalytic activity toward small synthetic substrates, but inhibited the binding of thrombin to fibrinogen, thrombomodulin and heparin cofactor II. Such proteins are known to require a secondary binding site of thrombin to interact with the enzyme.The anti-thrombin antibody might have resulted from an abnormal generation of thrombin. This would be the consequence of the process favouring thrombosis. Alternatively, the autoantibody might have favoured thrombosis primarily, by impairing natural antithrombotic mechanisms triggered by thrombin.

Identification of a Monoclonal Thrombin Inhibitor Associated with Multiple Myeloma and a Severe Bleeding Disorder

“…One could envisage how an antibody binding to one domain of the molecule could inhibit the procoagulant effect of thrombin and result in bleeding as reported here and elsewhere (Scully et al, 1982;Sié et al, 1991;La Spada et al, 1995). Alternatively, antibody binding to a different site could enhance the procoagulant activity of thrombin resulting in thrombosis that requires oral anticoagulant therapy (Costa et al, 1992;Arnaud et al, 1994).…”

“…Significantly, the IgG does not bind human prothrombin or other vitamin K-dependent coagulation factors (Fig 5), implying that the epitope is not present in these related molecules. In one report of an antibody which resulted in recurrent arterial thrombosis, the IgG recognized anion-binding exosite I of thrombin and interfered with the interaction of thrombin with fibrinogen, thrombomodulin and heparin cofactor II (Costa et al, 1992;Arnaud et al, 1994). In the patient reported here, the IgG does not bind prothrombin, in which exosite I is exposed, and the clinical outcome was haemorrhage rather than thrombosis, so that it is likely that the binding site is elsewhere in the thrombin molecule.…”

Topical thrombin and acquired coagulation factor inhibitors: Clinical spectrum and laboratory diagnosis

“…A subset of patients with factor V inhibitors develop a severe hemorrhagic diathesis, however, and may die in spite of aggressive therapeutic intervention [ 171. Acquired inhibitors to thrombin are also relatively rare, and, as with factor V inhibitors, carry a variable clinical presentation [18][19][20]. One type of autoantibody to prothrombin that does occur with slightly more frequency, however, is the clearing antiprothrombin antibody that is seen in association with the lupus anticoagulant [21,22].…”

Beta2‐glycoprotein I protects thrombin from inhibition by heparin cofactor II: Potentiation of this effect in the presence of anti–β2‐glycoprotein I autoantibodies

“…The effect of ␤ 2 GPI on HCII inactivation of thrombin was assessed in the presence of heparin or dermatan sulfate. The assay was first performed with heparin as previously described (37), with some modifications. Briefly, 0.1-1 M human native ␤ 2 GPI or 1 M bovine serum albumin (BSA) in 50 mM Tris buffered saline (TBS; Tris HCl, 150 mM NaCl [pH 7.5]) containing 0.5% polyethylene glycol (TBSP) was incubated with 75 nM thrombin for 1 hour at 25°C, and incubated further in 30 nM HCII/0.3 units/ml heparin for 17 minutes at 37°C.…”