ABSTRACT Heparin induced thrombocytopenia (HIT) is a severe thrombotic disorder initiated by ultralarge immune complexes (ULICs) containing IgG antibody to a multivalent antigen composed of platelet factor 4 (PF4) and heparin (H). Even today, patients with HIT remain at risk for death, amputation, recurrent thromboembolism and bleeding while receiving maximally tolerated doses of anti-Xa or anti-thrombin agents. Thus, there is an unmet need for a deeper insight into the pathobiology of thrombosis in HIT that will lead to targeted novel non- anticoagulant interventions to supplement contemporary therapy. Our published and pilot data demonstrate that activation of the complement pathway fulfills this gap. Specifically, we show that HIT ULICs: 1) interact with and bind activated complement components 2) generate soluble complement components via the classical pathway, 3) deposit C3 on platelets, neutrophils, monocytes and endothelial cells (ECs), 4) initiate cellular activation leading to neutrophil degranulation, monocyte tissue factor (TF) and procoagulant activity 5) activate complement in the presence of direct thrombin inhibitors, 6) trigger complement-mediated cellular activation upstream of C5 and 7) promote significant complement deposition in a murine thrombosis model. Based on these findings, we hypothesize that complement activation by HIT ULICs contributes to the prothrombotic state in HIT through EC injury mediated by soluble and surface expressed complement receptors (CRs) and dual amplification of FcγRs and CRs on cells expressing FcγRs. In the aims that follow, we will address the following questions related to complement in HIT: 1) Does complement stabilize HIT ULICs, prevent disassembly and amplify procoagulant responses by ECs that lack FcγRs? In this aim, we will test the hypothesis that complement stabilizes ULIC assembly and promotes EC injury and impairs complement regulatory function, leading to release of vWF multimers that amplify ULIC formation and complement activation. 2) Does complement amplify procoagulant responses by FcγR expressing cells and serve as a biomarker of incipient disease? In this aim, we will test the hypothesis that complement-coated ULICS amplify FcγRIIA signaling by promoting cooperativity of complement and FcγR receptors. In this aim, we will examine effects of ULIC composition on cellular activation, identify CRs involved in binding HIT ULICs, examine effects of HIT ULICs on soluble and cellular complement regulatory mechanisms and characterize complement activation in patients with and without symptomatic anti- PF4/heparin antibodies. 3) Can Complement inhibition serve as a therapeutic strategy for HIT? In this aim, we will use microfluidic assays and murine thrombosis models to test the hypothesis that the net effects of complement activation by HIT ULICs contributes to macrovascular thrombosis and its modulation can reduce need for antithrombotic therapy in HIT. Together, these studies will provi...