Abstract 5298: Staphylococcal Superantigen-like 5 Induces Platelet Activation and Thrombosis via Binding to GPIbα and GPVI
Staphylococcus aureus (S. aureus) is a common and dangerous bacterial pathogen capable of causing life-threatening thromboembolic disease such as disseminated intravascular coagulation (DIC). S. aureus is known to secrete potent toxins that include Staphylococcal superantigen-like protein 5 (SSL5), which was recently shown to bind P-selectin glycoprotein ligand-1 (PSGL-1). As PSGL-1 is structurally and functionally related to the glycoprotein (GP) Ibα platelet receptor, this study assessed the ability of SSL5 to interact with GPIbα as well as other platelet glycoprotein receptors. Using flow cytometry, immunoprecipitation, surface plasmon resonance, platelet adhesion and aggregometry assays, recombinantly produced SSL5 was found to directly bind to platelet GPIbα via the sLacNac residues that terminate its glycan chains. SSL5 was also found to bind to the platelet collagen receptor GPVI. This binding of SSL5 to platelets causes platelet activation and aggregation as demonstrated in in vitro assays. SSL5 was subsequently tested for its prothrombotic effects in C57BL/6 mice. Pulmonary thrombi developed in 8 of 11 intraveneously SSL5-injected mice but were absent from 5 mice injected with a functionally-inactive SSL5 protein carrying a T175P point mutation. Further expriments using flow cytometry showed that SSL5-induced platelet activation was significantly inhibited by an anti-SSL5 monoclonal antibody designated 5E3 (10μg/ml, P < 0.001), as well as sLex (IC50 < 5μM, P < 0.001) and sLacNac glycans (IC50 < 50μM, P < 0.001). Moreover, the injection of 5E3 in SSL5-treated mice (n = 8) compared with mice only injected with SSL5 (n = 8), was found to inhibit SSL5-induced thrombus formation (P < 0.05). Taken together, our data show that SSL5 binds to GPIbα and GPVI and as a consequence of this activates platelets and induces thrombosis. Thus, we describe a novel pathogenic mechanism potentially active in S. aureus infections. Furthermore, blockade of SSL5-induced platelet activation and thrombosis by either 5E3 or inhibitory glycans defines a novel treatment strategy and identifies suitable drug candidates for prevention and treatment of thrombotic diseases associated with S. aureus infections.