Anti–platelet factor 4 antibodies causing VITT do not cross-react with SARS-CoV-2 spike protein

Abstract

Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a severe adverse effect of ChAdOx1 nCoV-19 COVID-19 vaccine (Vaxzevria) and COVID-19 vaccine Janssen (Ad26.COV2.S), and associated with unusual thrombosis. VITT is caused by anti-platelet factor 4 (PF4) antibodies activating platelets through their FcgRIIa receptors. Antibodies activating platelets through FcgRIIa receptors have also been identified in COVID-19 patients. These findings raise concern that vaccination-induced antibodies against anti-SARS-CoV-2 spike protein cause thrombosis by cross-reacting with PF4. Immunogenic epitopes of PF4 and SARS-CoV-2 spike protein were compared using in-silico prediction tools and 3D-modelling. The SARS-CoV-2 spike protein and PF4 share at least one similar epitope. Reactivity of purified anti-PF4 antibodies from patients with VITT was tested against recombinant SARS-CoV-2 spike protein. However, none of the affinity-purified anti-PF4 antibodies from 14 VITT patients cross-reacted with SARS-CoV-2 spike protein. Sera from 222 PCR-confirmed COVID-19 patients from five European centers were tested by PF4/heparin ELISA and PF4-dependent platelet activation assays. We found anti-PF4 antibodies in 19 of 222 (8.6%) COVID-19 patient sera. However, only four showed weak to moderate platelet activation in the presence of PF4, and none of these patients developed thrombotic complications. Among 10 of 222 (4.5%) COVID-19 patients with thrombosis, none showed PF4-dependent platelet-activating antibodies. In conclusion, antibodies against PF4 induced by vaccination do not cross-react with the SARS-CoV-2 spike protein, indicating that the intended vaccine-induced immune response against SARS-CoV-2 spike protein is not the trigger of VITT. PF4-reactive antibodies found in COVID-19 patients of the present study were not associated with thrombotic complications.

Interactions of Spike-RBD of SARS-CoV-2 and Platelet Factor 4: New Insights in the Etiopathogenesis of Thrombosis

“…In this study we evidenced the ability of polyclonal anti-PF4 antibodies to recognize the Spike-RBD, highlighting a possible direct association of Spike to platelets misregulation. These results are in contrast with those previously reported [20], showing a lack of cross-reactivity of affinitypurified anti-PF4 antibodies isolated from 14 VITT patients for SARS-CoV-2 spike protein, however we cannot exclude (as stated as well by the authors of that study) that eventual antibodies involved in VITT were bound in complexes and thus they were not accessible for purification or not able to bind to other molecules. We used instead a purified untreated polyclonal anti-PF4 antibody free to bind to all ligands and not deprived of eventual populations trapped in complexes with platelets or PF4.…”

Complicated Long Term Vaccine Induced Thrombotic Immune Thrombocytopenia—A Case Report

“…How far epitope similarities between PF4 and the spike-protein of SARS-CoV-2 contribute remains controversial. Members of our group did not find any cross-reactivity between VITT antibodies and the SARS-CoV-2 spike-protein [12].…”

Coagulopathies after Vaccination against SARS-CoV-2 May Be Derived from a Combined Effect of SARS-CoV-2 Spike Protein and Adenovirus Vector-Triggered Signaling Pathways

“…While a previous study by Greinacher’s team has shown no cross-reactivity of platelet-activating antibodies isolated from VITT patients with the coronavirus spike protein (and only a few out of 200 patients recovered from COVID-19 reacting weakly with PF4) [ 104 ], they now show in a recent preprint close proximity between PF4 molecule, platelets, and adenovirus hexon capsid protein-positive staining particles. This association in the complex could be displaced by high concentrations of (highly negatively charged) heparin.…”