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Jessica Molhoek

Tuesday 5 December 2017

Looking into a paradox - Lupus anticoagulant and its relation to thrombosis in the antiphospholipid syndrome

Promotor: prof.dr. Ph.G. (Flip) de Groot
Supervisor: dr. R.T. (Rolf) Urbanus
Date: 5 December 2017
Time: 14.30 h

The antiphospholipid syndrome is an autoimmune disease characterized by thrombosis and pregnancy morbidity. One of the diagnostic hallmarks of the antiphospholipid syndrome is a prolonged clotting time. A prolonged clotting time is normally suggestive for bleeding tendency. The combination thrombosis in vivo and prolonged clotting assays in vitro indicates a paradox in the antiphospholipid syndrome. Patients with antiphospholipid syndrome have persistent so-called anti-phospholipid antibodies. A prolonged clotting time due to an inhibitor is known as lupus anticoagulant. The anti-phospholipid antibodies do not bind to phospholipids, but to the plasma protein β2-glycoprotein I (β2GPI) bound to phospholipids. β2GPI is an important player in the antiphospholipid syndrome, however, no physiological function has been described for β2GPI. The aim of this thesis is to understand lupus anticoagulant and its relation to thrombosis in the scope of the antiphospholipid syndrome.

We started to study the clinical consequence of lupus anticoagulant in stroke in a case-control study in Tanzania. In this case-control study we found a high correlation between lupus anticoagulant and the risk for stroke. Several studies identified the presence of anti-β2GPI antibodies upon an infection. Our group previously showed that injection of surface protein H of Streptococcus pyogenes, a subset of Streptococcus, induces anti-β2GPI antibodies in mice. However, this study did not investigate the effect of β2GPI on thrombus formation. We describe that anti-β2GPI antibodies against domain I that were generated upon protein H injection enhances thrombus formation.

Previous studies showed at least two different conformations of β2GPI. We developed a nanobody to distinguish between these conformations. The important sequences for a possible function for β2GPI are the cryptic epitopes in domain I and V. These epitopes are often involved in antibody and receptor/negative surfaces binding. Our nanobody recognizes an epitope only after the conformation of β2GPI has changed by binding to anionic surfaces.

We have found that β2GPI in the presence of the auto-antibodies strongly binds to factor V and factor Va. We describe the ability of β2GPI-anti-β2GPI antibody complexes to inhibit the activation of factor V by factor Xa. This inhibition partly explains lupus anticoagulant in antiphospholipid syndrome patients. β2GPI-anti-β2GPI antibody complexes interacting with factor V were able to inhibit the cofactor function of factor V in the inactivation of factor VIIIa by activated protein C. This could explain why these patients do have an increased thrombotic risk.

In this thesis we have studied auto-antibodies against β2GPI and its relation to thrombosis. There is found an association of lupus anticoagulant with thrombosis. We have found an alternative explanation why these antibodies prolonged in vitro clotting assays and we also showed that these antibodies induce protein C resistance.