A German researcher thinks he knows what has caused some people to develop blood clots after receiving coronavirus vaccines. University of Greifswald hematologist Andreas Greinacher believes the reaction is linked to EDTA, a preservative found in the AstraZeneca vaccine.
The AstraZeneca vaccine contains more than 1,000 proteins that have been derived from human proteins — and the EDTA helps them stay in the bloodstream. There, they bind to platelet factor 3, which forms structures that kickstart antibody production. Greinacher’s hypothesis is that when the vaccine elicits an inflammatory response, it triggers an ancient immune defense mechanism that runs amok and causes clotting and bleeding. It’s like “awakening a sleeping dragon,” he told.
From Greinacher’s data, published in Research Square, the following sequence of events appears to mediate VITT.
In Step 1, a neo-antigen is generated: following intramuscular injection, vaccine components and platelets come into contact, resulting in platelet activation.
ChAdOx1 nCov-19 vaccine activates platelet by multiple mechanisms including platelet interaction with adenovirus, cell-culture derived proteins (currently, it is unknown which of the > 1,000 proteins identied in the vaccine are involved in platelet activation), and EDTA. Activated platelets release PF4. As shown by TEM, released PF4 binds to constituents of the vaccine forming multimolecular aggregates, which also include virus proteins, resulting in particles formation of ≥ 120 nm size.
Step 2 generates an inammatory co-signal that further stimulates the immune response: EDTA in the vaccine increases capillary leakage at the inoculation site, likely by endothelial (VE)-cadherin disassembly. Proteins found in the vaccine include virus proteins, but also proteins originating from the human kidney-derived production cell line T-REx HEK-293. Increased vascular permeability facilitates dissemination of these proteins into the blood. Blood dissemination of vaccine components is not unique to ChAdOx1 nCov-19. A ChAdOx1 vector variant (with a hepatitis B vector insert) was detectable by PCR in multiple organs, including liver, heart, and lymph nodes at days 2 and 29 after intramuscular injection in mice.
Within the circulation, vaccine constituents including its complexes with PF4 are recognized by preformed natural immunoglobulin G antibodies,23 presumably resulting in immune complexes. This contributes to clinical symptoms within 8 to 24 hours following inoculation that are reminiscent of systemic inflammation (fever, chills, large joint arthralgia, occasionally skin lesions, probably reecting a similar process as known in serum sickness or serum sicknesslike illnes).
Such symptoms have also been observed as acute vasculitis like reaction when a column used for immunoadsorption leaked protein A with bound antibodies. This inflammatory response likely provides an important co-signal that stimulates antibody production by preformed B-cells capable of producing anti-PF4 antibodies, as is known to occur in the pathogenesis of “classical” HIT. Multimolecular complexes containing PF4 also activate the complement system. Complement bound to the aggregates subsequently allows binding of the complexes to B-cells via their complement receptor.
Step 3 leads to prothrombotic reactions: high avidity anti-PF4 antibodies among the anti-PF4 antibodies in VITT patient blood bind and cluster PF4 on the platelet surface, likely involving polyanions such as cell surface chondroitin sulfate or exposed polyphosphate. Clustering of PF4 by high-avidity autoantibodies is also crucial for platelet activation in autoimmune heparin-induced thrombocytopenia. The resulting PF4/IgG immune complexes activate platelets, which release additional PF4 and polyphosphate. Crosstalk of PF4, activated platelets and antibodies with neutrophils subsequently leads to NETosis. Extracellular DNA in NETs binds PF4 and resulting DNA/PF4 complexes further recruit anti-PF4 antibodies with lower avidity, which require the polyanion cofactor (DNA).
This culminates in massive Fcγ receptor-dependent activation of neutrophils, platelets and, most likely (by analogy with HIT), monocytes and endothelial cells.
An important potential natural regulator of this process are extracellular DNases, which degrade NETs. DNase activity in VITT patients with thrombosis was markedly reduced, likely facilitating accumulation of NETs and DNA. Ultimately, ChAdOx1 nCov-19 vaccine-triggered VITT culminates in marked activation of the coagulation system.
Broadened reactivity of antibodies in a boosted immune response is a hallmark of certain disorders besides VITT. For example, autoimmune heparin-induced thrombocytopenia features heparin-dependent reactivity that extends to include heparin-independent reactivity.
Similarly, post-transfusion purpura (PTP) reects a strong alloimmune response that progresses to include autoreactive properties.
In summary, this study provides a mechanism by which an adenoviral vector vaccine can trigger an immune response leading to highly reactive anti-PF4 antibodies with downstream FcγIIa receptor-dependent amplification recruiting neutrophils and triggering NETosis with prothrombotic consequences.