In February 2021, shortly after the start of COVID-19 vaccination in Europe, hematologist Sabina Eichinger from the Medical University of Vienna encountered an unusual clinical case. A 49-year-old nurse developed thrombosis accompanied by uncontrolled bleeding after receiving the AstraZeneca vaccine. The patient died. An autopsy revealed no alternative cause, leading the physician to suspect a link to vaccination.
This clinical case resembled a rare complication of heparin therapy. That complication is caused by antibodies to PF4 – a protein involved in blood coagulation. These antibodies were also detected in the blood of the 49-year-old patient.
Soon, similar cases were reported in Europe among recipients of the AstraZeneca vaccine and in the United States among recipients of the Johnson & Johnson vaccine. The new syndrome was termed vaccine-induced immune thrombotic thrombocytopenia (VITT). Patients with VITT were likewise found to have antibodies to PF4.
VITT is rare – about one case per 200,000 vaccinated individuals. Nevertheless, the risk of VITT led to revisions in vaccination policies: many European countries discontinued or restricted AstraZeneca use in older adults, and the United States stopped using the Johnson & Johnson vaccine.
Mechanism of Vaccine-Induced Immune Thrombotic Thrombocytopenia
Several years later, scientists elucidated the mechanism underlying VITT. Both vaccines used an adenovirus as a vector to deliver the gene encoding the SARS-CoV-2 spike protein into cells. One of the adenoviral proteins can trigger the formation of abnormal antibodies, occurring in individuals with a particular combination of genetic factors and a specific mutation in antibody-producing B cells. Instead of recognizing the viral spike protein, these antibodies bind to the PF4 protein. This interaction initiates a cascade of reactions that leads to VITT.
Although both adenoviral COVID-19 vaccines are no longer in use, the technology remains important. An adenoviral Ebola vaccine is already in use, and similar platforms are being developed for other infections. In theory, such vaccines could also cause VITT, but the risk can be reduced by modifying the adenoviral vector itself.
Molecular Mechanism of VITT: How Anti-PF4 Antibodies Trigger Thrombosis
To understand the nature of anti-PF4 antibodies in VITT, researchers analyzed amino acid sequences from 21 patients. In all cases, a shared alteration was identified: at position 31 of the antibody light chain, either glutamic acid or aspartic acid – negatively charged amino acids – was present.
Analysis of genetic sequences in other cells from these patients showed that, under normal conditions, lysine – a positively charged amino acid – should occupy this position, indicating that the amino acid substitution occurred only in specific B cells that began producing abnormal antibodies.
Inherited variants of the light-chain gene also facilitated the development of VITT. In all patients, position 50 already contained a negatively charged amino acid. Together with the substitution at position 31, this increased the overall negative charge of the antibodies.
PF4, by contrast, carries a strong positive charge. The negative charge of the abnormal antibodies facilitates their binding to PF4. The resulting immune complexes activate platelets, which mediate blood clotting. Activated platelets release additional PF4, further enhancing complex formation and triggering a chain reaction. As a result, clots form while platelet reserves are depleted, leading to uncontrolled bleeding.
To test whether the amino acid substitution at position 31 truly contributes to VITT, researchers injected mice with anti-PF4 antibodies derived from patients. These antibodies induced symptoms in mice resembling VITT. The researchers then created a corrected version of the antibodies by restoring lysine at position 31, rather than glutamic acid. These antibodies still bound PF4, but much more weakly and caused far fewer thrombi in mice.
Trigger of the Abnormal Immune Response to Vaccination
The researchers identified the trigger of the abnormal immune response. Recombinant antibodies in VITT bind to adenoviral protein VII – pVII. A key region of this protein, consisting of 15 amino acids, forms an alpha helix – a structure similar to a region of the PF4 protein.
The researchers proposed the following hypothesis: patients with VITT had previously experienced an adenoviral infection that generated a pool of B cells recognizing pVII. Vaccination with an adenovirus-based vaccine activated these cells and initiated a mutation process that gives rise to new antibody variants. In some individuals, this process produced abnormal antibodies with a substitution introducing a negatively charged amino acid, which enhanced binding to positively charged PF4 and led to the development of VITT.
Conclusion
According to the European Medicines Agency, approximately 900 cases of VITT have been reported in Europe following vaccination with AstraZeneca and Johnson & Johnson, including about 200 fatal outcomes. At the same time, more than 3 billion doses of the AstraZeneca vaccine were administered worldwide, and it is estimated to have prevented millions of deaths.
Genetic variants that increase susceptibility to VITT are found in 40–60% of the population in most regions of the world, whereas their prevalence in East Asia is about 20%.
These findings are important for the development of future vaccines. Adenoviral vectors are used to create vaccines against influenza, malaria, meningitis, tuberculosis, and the Nipah virus. Adenoviral vaccines are relatively inexpensive to produce and easier to store, as they do not require extremely low temperatures. The new data will help improve their safety. Removing pVII from the virus is unlikely to be feasible, but its structure can potentially be modified to reduce its similarity to PF4.
Reference
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