Anti-SARS-CoV-2 Spike Protein and Anti-Platelet Factor 4 Antibody Responses Induced by COVID-19 Disease and ChAdOx1 nCov-19 vaccination

Abstract Background: Some recipients of ChAdOx1 nCoV-19 COVID-19 Vaccine AstraZeneca develop antibody-mediated vaccine-induced thrombotic thrombocytopenia (VITT), associated with cerebral venous and other unusual thrombosis resembling autoimmune heparin-induced thrombocytopenia. A prothrombotic predisposition is also observed in Covid-19. We explored whether antibodies against the SARS-CoV-2 spike protein induced by Covid-19 cross-react with platelet factor 4 (PF4/CXLC4), the protein targeted in both VITT and autoimmune heparin-induced thrombocytopenia.Methods: Immunogenic epitopes of PF4 and SARS-CoV-2 spike protein were compared via prediction tools and 3D modelling software (IMED, SIM, MacMYPOL). Sera from 222 PCR-confirmed Covid-19 patients from five European centers were tested by PF4/heparin ELISA, heparin-dependent and PF4-dependent platelet activation assays. Immunogenic reactivity of purified anti-PF4 and anti-PF4/heparin antibodies from patients with VITT were tested against recombinant SARS-CoV-2 spike protein. Results: Three motifs within the spike protein sequence share a potential immunogenic epitope with PF4. Nineteen of 222 (8.6%) Covid-19 patient sera tested positive in the IgG-specific PF4/heparin ELISA, none of which showed platelet activation in the heparin-dependent activation assay, including 10 (4.5%) of the 222 Covid-19 patients who developed thromboembolic complications. Purified anti-PF4 and anti-PF4/heparin antibodies from two VITT patients did not show cross-reactivity to recombinant SARS-CoV-2 spike protein. Conclusions: The antibody responses to PF4 in SARS-CoV-2 infection and after vaccination with COVID-19 Vaccine AstraZeneca differ. Antibodies against SARS-CoV-2 spike protein do not cross-react with PF4 or PF4/heparin complexes through molecular mimicry. These findings make it very unlikely that the intended vaccine-induced immune response against SARS-CoV-2 spike protein would itself induce VITT.