Anti-PF4 antibodies trigger a faster and stronger platelet procoagulant response than anti-heparin/PF4 antibodies

Vaccine-induced immune thrombocytopenia and thrombosis (VITT) and VITT-like syndromes are emerging causes of severe thrombocytopenia and thrombosis associated with immunoglobulin G (IgG) antibodies targeting platelet factor 4 (PF4). These syndromes are characterized by more frequent and severe thrombosis than heparin-induced thrombocytopenia (HIT), which is typically caused by antibodies targeting heparin-modified PF4 (anti-PF4/H). In this study, we compared the intrinsic prothrombotic properties of IgG anti-PF4 and anti-PF4/H to identify mechanisms that may contribute to the severity of VITT and VITT-like syndromes. Using the monoclonal IgG1 antibodies 1E12 (anti-PF4) and 5B9 (anti-PF4/H), we showed that IgG anti-PF4 exhibited an increased propensity to induce the formation of fibrin-rich platelet-leukocytes aggregates in a microfluidic model of whole-blood (WB) thrombosis. This phenomenon was related to a strong potentiation of thrombin generation in plasma by 1E12, but not by 5B9, and occurred only in the presence of platelets and coagulation factors V and XI. We also demonstrated a substantially faster and stronger ability of 1E12, compared with 5B9, to activate platelets using the serotonin release assay and confirmed these findings using human VITT and HIT samples. Furthermore, 1E12 was more likely than 5B9 to induce procoagulant platelets (CD62P+/phosphatidylserine+) in platelet-rich plasma (PRP) and to produce procoagulant microvesicles in WB and PRP. In conclusion, platelets are activated faster and exhibit more procoagulant activity in response to anti-PF4 than anti-PF4/H, which may contribute to the severity of VITT and VITT-like syndromes.