Visual Abstract - Cross-Linking GPVI-Fc by Anti-Fc Antibodies Potentiates Its Inhibition of Atherosclerotic Plaque- and Collagen-Induced Platelet Activation: 10.1016/j.jacbts.2016.03.008

• The GPVI collagen receptor mediates platelet activation to collagen exposed after rupture or injury of atherosclerotic plaques. Revacept®, a GPVI-Fc fusion protein binding to plaque collagen and concealing binding sites for platelet GPVI, acts as a lesion-focused antiplatelet drug. To optimize its specific inhibition of plaque-induced platelet activation, we cross-linked the Fc tails of GPVI-Fc with anti-human-Fc IgG or Fab2 antibodies.

• Cross-linking yielded oligomeric GPVI-Fc complexes, which inhibited atherosclerotic plaque- induced platelet aggregation in static and flow assays as efficiently as antibodies blocking GPVI receptors on platelets. Under arterial flow, GPVI-Fc cross-linking with anti-Fc-Fab2 was superior to cross-linking with anti-Fc IgG.

• Advanced optical imaging revealed a rapid and stable sheath-like coverage of collagen fibers by cross-linked GPVI-Fc complexes preventing platelet attachment to collagen.

• Cross-linked GPVI-Fc did not increase bleeding time in vitro.

• Transformation of GPVI into multivalent GPVI complexes may provide a new superior strategy to suppress atherothrombosis without increasing systemic bleeding risk.