OC 79.1 - Targeting factor XII attenuates vasculitis and systemic inflammatory organ injury in extracorporeal membrane oxygenation

Background: Factor XII (FXII) is the zymogen of the plasma protease FXIIa that activates the intrinsic coagulation pathway and the kallikrein kinin-system. The role of FXII in inflammation has been obscure.

Aims: To examine the impact of FXII inhibition and deletion on vasculitis and inflammatory injury in extracorporeal membrane oxygenation (ECMO).

Methods: Single variable heavy chain domain (nanobody) was produced by immunizing an alpaca with human FXII. FXII inhibition and gene-deletion was examined in murine models of vasculitis and ECMO.

Results: A novel nanobody was obtained which recognized FXII (Kd=0.41 nM) in a conformation-dependent manner and interfered with FXIIa formation. Its bivalent construct (Nb-Fc), derived by fusion with an Fc segment of human antibody, dose-dependently inhibited arterial thrombosis in mice to a level comparable to FXII knockout, without affecting hemostasis. Nb-Fc treatment and FXII knockout both markedly ameliorated immune complex-induced local vasculitis, suppressing neutrophil infiltration, and they also attenuated anti-neutrophil cytoplasmic antibody-induced systemic vasculitis. In mice that underwent 2-hour ECMO, FXII inhibition or knockout decreased thrombus deposition on oxygenator membranes and circulating TNF-α, and, strikingly, they substantially decreased liver and kidney microvascular injury and reduced circulating levels of D-dimer, alkaline phosphatase, and creatinine, at 24-hour after ECMO. Furthermore, intravital confocal imaging of the microcirculation revealed that ECMO process triggered neutrophil adherence, platelet aggregation and neutrophil-platelet aggregation and that FXII knockout or Nb-Fc treatment attenuated these vascular inflammatory responses. Consistently, FXII blockade suppressed thioglycolate-induced peritoneal migration of neutrophils in mice. Nb-Fc selectively inhibited in vitro migration of neutrophils among total human blood leukocytes. In human blood microfluidic analysis, Nb-Fc treatment prevented collagen induced platelet/fibrin deposition and neutrophil activation.

Conclusion(s): FXII critically mediates the inflammatory responses in vasculitis and ECMO, culminating at neutrophil activation. Nb-Fc bears a promise for ameliorating vasculitis and systemic inflammatory organ injury in ECMO.