The Deglycosylated Form of 1E12, a Monoclonal Anti-PF4 IgG, Strongly Inhibits Antibody-Triggered Cellular Activation in Vaccine-Induced Thrombotic Thrombocytopenia, and Is a Potential New Treatment for Vιττ

• Published in: Blood 2021-11, Vol.138 (Supplement 1), p.582-582
• Main Authors: Vayne, Caroline, Palankar, Raghavendra, Billy, Sandra, Handtke, Stefan, Thiele, Thomas, Pouplard, Claire, Greinacher, Andreas, Gruel, Yves, Rollin, Jérôme
• Format: Article
• Language: English
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood-2021-147922

▪ Introduction Vaccine-induced thrombotic thrombocytopenia (VITT) is a severe complication of recombinant adenoviral vector vaccines used to prevent COVID-19, likely due to anti-platelet factor 4 (PF4) IgG antibodies. The specificity and platelet-activating activity of VITT antibodies strikingly resemble that of antibodies detected in “autoimmune” heparin-induced thrombocytopenia (HIT), but their features remain poorly characterized. In particular, a better knowledge of these antibodies should help to understand the mechanisms leading to hypercoagulability and the particular thrombotic events observed in VITT, but rarely in typical HIT. We have recently developed a chimeric IgG1 anti-PF4 antibody, 1E12, which strongly mimics “autoimmune” HIT antibodies in terms of specificity and cellular effects. Therefore, we assessed whether 1E12 could mimic VITT antibodies. We then evaluated the capability of DG-1E12, a deglycosylated form of 1E12 unable to bind FcγR, to inhibit cellular activation induced by VITT antibodies. Methods and Results Using a PF4-sensitized serotonin release assay (PF4-SRA) (Vayne C, New Engl J Med, 2021), we demonstrated that 1E12 (5 and 10 μg/mL) strongly activated platelets, with a pattern similar to that obtained with human VITT samples (n=7), i.e. in a PF4-dependent manner and without heparin. This platelet activation was inhibited by low heparin concentration (0.5 IU/mL), an effect also observed with VITT samples. Serotonin release induced by 1E12 was also fully inhibited by IV-3, a monoclonal antibody blocking FcγRIIa, or by IdeS, a bacterial protease that cleaves IgG and strongly inhibits the binding of IgG antibodies to FcγRIIa. This inhibitory effect of IV-3 and IdeS strongly supports that interactions between pathogenic anti-PF4 IgG and FcγRIIa play a central role in VITT. Incubation of 1E12 or VITT samples with isolated neutrophils (PMN) and platelets with PF4 (10 µg/mL) strongly induced DNA release and NETosis, supporting that PMN are involved in the processes leading to thrombosis in VITT. Furthermore, when whole blood from healthy donors incubated with 1E12 or VITT plasma was perfused in capillaries coated with von Willebrand Factor, numerous large platelet/leukocyte aggregates containing fibrin(ogen) were formed. To investigate whether 1E12 and VITT antibodies recognize overlapping epitopes on PF4, we then performed competitive assays with a deglycosylated form of 1E12 (DG-1E12), still able to bind PF4 but not to interact wit