1. Introduction
Scientists have expedited vaccine development to combat COVID-19, and several types of vaccines have been developed: mRNA vaccines, non-replicative vector vaccines, inactivated and subunit vaccines.[1] Globally, as of June 1, 2021, more than 1.93 billion doses of COVID-19 vaccine have been administered.[1] From December 2020 through March 2021, based on randomized, blinded and controlled trials, the European Medicines Agency (EMA) approved four vaccines: two messenger RNA–based vaccines that encode the spike protein antigen of SARS-CoV-2, encapsulated in lipid nanoparticles: BNT162b2 (Pfizer–BioNTech) and mRNA-1273 (Moderna); the two other vaccines encode the SARS-CoV-2 spike glycoprotein: ChAdOx1 nCov-19 (AstraZeneca) which is a recombinant chimpanzee adenoviral vector and Ad26.COV2.S vaccine (Johnson & Johnson/Janssen) which is a recombinant adenovirus type 26 vector. Similarly, the United States Food and Drug Administration (FDA) authorized the use of the following vaccines: mRNA vaccines, BNT162b2 and mRNA-1273, and the adenovirus-vectored vaccine, Ad26. COV2.S.[2]
After that, many other countries have implemented their own vaccination programs to combat the COVID-19 pandemic. Analyses of clinical trials have demonstrated the efficacy and safety of COVID-19 vaccines and preliminary studies have shown that COVID-19 vaccines are effective in the prevention of SARS-CoV-2 infection in real-world settings.
However, several reports raised serious concerns regarding the risk of vaccine-related thrombosis named as vaccine-induced thrombosis and thrombocytopenia (VITT).[3] VITT or thrombosis with thrombocytopenia syndrome is a new, rare and potentially life-threatening disorder, following immunization with adenovirus vector COVID-19 vaccines ChAdOx1 nCOV-19 (AZD1222; AstraZeneca) and Ad26.COV2·S (Johnson & Johnson/Janssen).[4] As of June 1, 2021, the estimated incidence rate of VITT, based on the total number of first-dose vaccinations, was approximately 0.00086%, 0.000127%, 0.00028%, and 0.000087% in the UK, Canada, Australia, and central Europe, respectively.[4] More and more VITT cases have been reported across the UK, Europe, Canada, and Australia, but VITT remains a rare syndrome.[5] The United States Centers for Disease Control and Prevention has confirmed 28 cases among more than 8 million recipients of the Ad26.COV2.S vaccine.[4] VITT syndrome, that generally occurs 5 to 30 days after vaccine administration, is characterized by moderate to severe thrombocytopenia, thrombosis, including cerebral venous sinus thrombosis (CSVT)/splanchnic thrombosis, positive platelet factor 4 (PF4)-heparin enzyme-linked immunosorbent assay (ELISA) and platelet activation assays. Thrombosis occurs at unusual sites, particularly at the cerebral venous sinuses and splanchnic veins.[4, 6]
Thus, we aimed to review the up-to-date published literature to further understand and provide updated information on COVID-19 vaccine-related thrombosis and thrombocytopenia. Understanding VITT mechanisms and pathogenesis should provide a platform to orient future research and to optimize management of VITT.
2. Epidemiology of VITT
By late December 2020, mass vaccination campaigns have started around the world, but thromboembolic events have been reported after SARS-CoV-2 vaccines, in particular after ChAdOx1 n COV-19 vaccine and the Ad26.COV2.S.[7] In these cases, VITT has been described as severe characterized by thrombosis, particularly at unusual sites including cerebral/splanchnic thrombosis, mild to severe thrombocytopenia and positive PF4-heparin ELISA and platelet activation assays.[4] Available literature indicated that most of the investigated cases are associated with a clinical picture of moderate-to-severe thrombocytopenia and thrombotic complications beginning 10–15 days after vaccination, resembling severe heparin-induced thrombocytopenia (HIT).[6, 7] VITT symptoms include severe headache, blurred vision, seizure, severe and persistent abdominal pain, painful swelling of the lower leg, and chest pain or dyspnoea.[6, 7] Although a marked elevation of D-dimer levels is highly suggestive of thrombocytopenia and thrombotic syndrome (TTS), the PF4 antibody assay remains a confirmatory test. VITT is diagnosed in the first 30 days of vaccination and described as acute thrombosis and thrombocytopenia with elevated D-dimer, with the use of a D-dimer threshold of <2000 μg/L, which makes VITT unlikely, and >4000 μg/L for VITT, which would underpin a suspected VITT.[8] Scully and colleagues reported that 22 (96%) of 23 patients with VITT had antibodies against PF4. Such definition was approved by the scientific community since similar observations were made in other reports worldwide.[8]
VITT was described after vaccination with ChAdOx1 nCoV-19 or Ad26.COV2.S vaccines in patients not receiving heparin, and few of these individuals had risk factors for thrombosis, and cases were generally younger than 55 years and mostly females, and up to one-half of the reported patients died (Table 1).[3] The clinical picture of VITT was moderate to severe thrombocytopaenia associated with arterial and/or venous thrombosis, often in unusual locations.[4] A common denominator in all five patients was a high level of antibodies to PF4–polyanion complexes.[4] Another report showed that eight patients developed VITT after administration of ChAdOx1 nCoV-19. Autopsies showed arterial and venous thromboses in various organs and the occlusion of glomerular capillaries by hyaline thrombi. Sera from VITT patients contain high-titres of antibodies against PF4.[4] PF4 antibodies in patients with VITT induced significant increase in procoagulant markers (P-selectin and phosphatidylserine externalization) compared to healthy volunteers and healthy vaccinated volunteers.[9] Clinical and laboratory features have included thrombosis at unusual sites[10]; in intracranial venous sinus[11], pulmonary, portal, hepatic, renal and mesenteric veins, and cerebral thrombosis was reported in choroid plexuses, suggesting a very aggressive form of VITT.[12] Patients with VITT-associated CSVT had more intracranial veins thrombosed and more frequently had extracranial thrombosis compared with non-VITT.[13] Pottegård et al. assessed the rate of haemostatic events in the first 28 days after vaccination with the Oxford-AstraZeneca vaccine, in which the vaccinated cohort included 281,264 people.[14] They found higher than expected rates of venous thromboembolic events among the vaccinated cohort than general population; an excess of 11 events per 100,000 vaccinations.[15]
In the UK, by early June, 40 million first doses and 29 million second doses of ChAdOx1 nCOV-19 vaccine have been administered; nonetheless, 390 thrombotic events, including 71 fatal events have been reported. Interestingly, the cases reported low platelet counts with the presence of anti-PF4 antibodies, indicating an abnormal clotting reaction.[15] Nation-wide analysis of the Paul-Ehrlich Institute in Germany represents one of the largest cohorts of laboratory confirmed patients with VITT. The adverse drug reaction database was queried for VITT/thrombosis with thrombocytopenia syndrome (TTS) cases following ChAdOx1 nCoV-19 vaccination. [15] VITT has a high mortality and can present with isolated thrombocytopenia,
severe headache, and bleeding. Demonstration of platelet activating anti-PF4 immunoglobulin G (IgG) has high sensitivity for TTS and captures a wider spectrum of clinically relevant VITT, especially at low platelet counts <30,000/mL.[16] Thereafter, based on established criteria, case reports reported VITT from all over the world, cerebral venous thrombosis (CVT), CVST and TTS especially following ChAdOx1 nCoV-19 vaccine. A recent meta-analysis presented evidence suggesting the occurrence of VITT following the AstraZeneca vaccine. Clinical practice must, therefore, account for the possibility of VITT and subsequent embolic events in certain individuals’ post vaccination with adenovirus-based COVID-19 vaccines.[17]
Along with ongoing research on VITT, new clinical features are appearing and new tests and treatments are proposed. Cacciola and collaborators conducted and confirmed VITT by investigating the platelet and coagulation activation using new specialized tests.[17] In addition, new clinical features are reported such as ischaemic stroke as a manifestation of VITT secondary to ChAdOx1 nCoV-19 vaccine.[18]
In comparison to US and Europe, few studies investigated VITT in Eastern, African, Latino-American and Asian populations. Only two studies described VITT cases in the Middle East.[19] Also, few reports from Thailand described VITT after administration of the ChAdOx1 nCoV-19. Five VITT cases occurred after 15 million (10 million first doses and 5 million second doses) doses of ChAdOx1 n-CoV-19 have been administered in Thailand.[20] The incidence of VITT is estimated at 1 in 3 million. In other Asian countries, only a few cases of VITT have been reported. Current Australian data estimates that the risk of developing TTS/VITT is approximately 2–3 in 100,000 persons following the administration of an adenoviral vector vaccine.[20]
Several scientists assumed, although severe with aggressive outcomes, VITT remains a rare complication. A retrospective descriptive study, reported 54,571 adverse reactions, of which 28 were associated with thrombotic adverse reactions and many delayed reactions were classified as type III hypersensitivity reactions. The authors suggested that with 17 million people having had the ChAdOx1 nCoV-19 vaccine, these are extremely rare events. Accordingly, the European Medicines Agency's Pharmacovigilance Risk Assessment Committee concluded that the vaccine was safe and effective and that the benefits outweigh the risks.[20] A study using nationwide population-based data from Denmark, reported that the number of VITT cases caused by the ChAdOx1 nCoV-19 vaccine did not appear to increase beyond the expected incidence rate.[21] Low prevalence of both thrombocytopenia and antibodies to PF4/polyanion-complexes were reported among four hundred and ninety-two Norwegian health care workers after vaccination with ChAdOx1 nCoV-19.[22] Major limitation of such studies was the small number of potential thrombotic adverse events, thus questioning the clinical relevance of wide-spread antibody testing after vaccine administration.
There are less reports of VITT following other approved COVID-19 vaccines. Out of 13.6 million women aged ≤ 50 years exposed to at least one dose of COVID-19 mRNA vaccines in the US, a total of 68 thromboembolic events was reported. When compared to hormonal contraceptive use, the mRNA vaccines do not show disproportional reporting of thromboembolic events in younger women.[23]
Curcio et al. reported vaccine-induced massive pulmonary embolism and thrombocytopenia following a single dose of Janssen vaccination.[23] Reported rates for TTS were 3.83 per million vaccine doses (Ad26.COV2.S). The used TTS case definition excluded cases without thrombocytopenia and included cases with negative anti-PF4 antibody.[24] VITT syndrome occurred following vaccination with Ad26.COV2.S vaccine.[24]
In late February 2021, the first report for a total of 3,263,188 injections of ChAdOx1 vaccine showed 11,206 adverse reactions in France of which 2811 were serious, unexpected, thrombotic events. There was multi-site thrombosis, associated with or without thrombocytopenia or coagulation disorders, and the study reported eight fatalities, indicating a mortality rate of 30%.[25] Five cases of prothrombotic immune thrombocytopenia have been reported after exposure to the ChAdOx1 vaccine. The spectrum of clinical manifestations included CVST, splanchnic vein thrombosis, arterial cerebral thromboembolism, and thrombotic microangiopathy.[26]
Health Security Agency (formerly Public Health England) in UK reported that none of the 40 VITT secondary to ChAdOx1 vaccine in the UK had any relapse of symptoms or severe adverse reactions after receiving the second dose of the vaccine, regardless of the vaccine received.[27] None of the 29 patients with VITT who received a second vaccination dose with an mRNA COVID-19 vaccine developed new thromboses or relevant increase in anti-PF4/heparin IgG enzyme-immunoassay optical density , regardless whether PF4-dependent platelet-activating antibodies were still present. PF4-dependent platelet-activating antibodies are transient in most patients with VITT. Those can safely receive a second COVID-19 mRNA vaccine, as part of a heterologous vaccination strategy.[27]
3. Comparison between COVID-19 vaccines in relation to VITT
An observational study assessed the CVT rate attributed to four COVID-19 vaccines approved in Europe, namely BNT162b2, mRNA-1273, ChAdOx1 nCoV-19, and AD26.COV2.S. Data on thrombotic adverse events reported on Eudravigilance data base between January 1, 2021 and July 30, 2021, showed CVT occurring alongside thrombocytopenia for the four vaccines ranging from 3–5% to 31–44% for mRNA and adenoviral vaccines, respectively.[28] Another study from Eudravigilance European database up to April 16, 2021, compared severe adverse events related to thrombocytopenia in recipients of 20,869,192 ChAdOx1 nCoV-19 vaccine doses to 71,210,981 recipients of BNT162b2 vaccines. ChAdOx1 nCoV-19 administration was associated with a much higher frequency of severe adverse events as compared with that elicited by BNT162b2.[28] The highest risk following ChAdOx1 nCoV-19 vaccination was in young people and, likely, women of reproductive age.[28] The immune reaction promoted by the ChAdOx1 nCoV-19 vaccine may lead to not only thrombocytopenia and cerebral/splanchnic venous thrombosis but also other thrombotic and thromboembolic severe adverse events. These events are not induced by the BNT162b2 vaccine. The study may help in the evaluation of the benefit/risk profile of the ChAdOx1 nCoV-19 vaccine considering the epidemic curve present in a country.[28] Another comparison using data from VigiBase, which is the world’s largest pharmacovigilance database developed and maintained by the Uppsala Monitoring Centre in Sweden, assessed clinical features of arterial and venous thrombosis after injection of BNT162b2 (1197), mRNA-1273(325) and AZD1222 (639) until 16 March, 2021.[29] They suggested that thrombotic events, including CVT, might occur in association with all three vaccines. In all, 213 CVST cases were identified: 187 after ChAdOx1 nCov-19 vaccination and 26 after a mRNA vaccination (25 with BNT162b2 and one with mRNA-1273).[29] Thrombocytopenia was reported in 107/187 CVST cases in the ChAdOx1 nCov-19 group, in none in the mRNA vaccine group and in 7/100 in the pre-COVID-19 group.[30] Incidence rate of VITT in recipients of the ChAdOx1 vaccine is in excess to the general population, and that similar effects are not seen in recipients of BNT162b2,[30] although thrombocytopenia (without thrombosis) has been shown after BNT162b2.[30] These reports indicated that VITT is likely related to adenoviral vector-based DNA virus vaccines, but not mRNA/lipid-based vaccines. mRNA vaccines, mRNA-1273 and BNT162b2 are associated with thrombocytopenia, although typically with purpura and mucosal bleeding rather than thrombosis.[31]
A national cohort study in England assessed the risk after ChAdOx1 and BNT162b2 vaccines.[31] The incidence of events by dose in pre-defined post-vaccination risk periods relative to the unvaccinated cohort was estimated after adjustment for age, gender, co-morbidities, care home residency and health/social care worker status. The authors reported increased risk of thrombotic episodes and thrombocytopenia in adults under 65 years of age within a month of a first dose of ChAdOx1 vaccine but not after the BNT162b2.[31] Generally, estimates from such comparisons remain approximate since they are not matched for age, socioeconomic status and ethnic group.
4. Differences between VITT and HIT
A hallmark of patients with VITT is the presence of antibodies specific to PF4, resembling the immunological drug reaction HIT, which presents clinically as thrombocytopenia and thrombosis in patients who have previously been exposed to heparin.[32] The autoimmune element in VITT mimics autoimmune heparin-induced thrombocytopenia (aHIT), a well-known prothrombotic disorder caused by platelet-activating antibodies that recognise multimolecular complexes between cationic PF4 and anionic heparin.[33] In both VITT and HIT, IgG antibodies bind to PF4on the surface of platelets, resulting in widespread platelet activation. In HIT, monomeric IgG in high concentrations can trigger FcgR2A, thereby inhibiting platelet activation and aggregation by PF4/anti-PF4 immune complexes.[33] However, as most patients suffering from VITT/TTS were not previously exposed to heparin, the genesis of theanti-PF4 auto-antibodies is still controversial.[34] To determine the binding site on PF4 of antibodies from patients with VITT, a study used alanine-scanning mutagenesis, and found differences in anti-PF4 antibodies binding sites.[34] The authors reported that the binding of anti-PF4 antibodies from patients with VITT (n = 5) was restricted to eight surface amino acids on PF4, all of which were located within the heparin-binding site, and that the binding was inhibited by heparin.[34] By contrast, antibodies from patients with HIT (n = 10) bound to amino acids corresponding to two different sites on PF4. VITT antibodies can mimic the heparin effect by binding to a similar sites on PF4; this allows PF4 tetramers to cluster and form immune complexes, which in turn causes FcγRIIa-dependent platelet activation.[34] This finding explains the VITT-antibody-induced platelet activation and the resemblance of HIT.
Other factors than heparin can trigger prothrombotic disorder that strongly resembles HIT, including certain polyanionic drugs. Also, both viral and bacterial infections can trigger prothrombotic syndrome in the absence of preceding exposure to any polyanionic medication.[35]
5. Potential causal factors favouring VITT
VITT pathogenesis is not fully understood and likely multifactorial. An important insight reported in investigations underpinned the multifactorial pathophysiology of VITT.[36] The presence of anti-PF4 antibodies was not sufficient to rouse clinically evident thrombosis. Generally, thrombocytopenia is attributed to infection, bone marrow suppression, lack of nutrition, genetic causes, or autoimmune processes. A large network using artificial intelligence by Geronikolou and collaborators identified three nodes that corresponded to genes (AURKA, CD46 and CD19) expressed only in VITT, whilst ADAM10, CDC20, SHC1 and STXBP2 are silenced in VITT, but are commonly expressed in both COVID 19 and thrombocytopenia.[37] The calculated average node degree was immense, illustrating the complexity of VITT and confirming the importance of cytokines and PYCARD, NLP3 and P2RX7 as key potential therapeutic targets for VITT. The authors concluded that common nodes appear to be key players in illness prevention, progression, and treatment.[37]
Based on previous studies describing vaccine-induced prothrombotic immune thrombocytopenia (VIPIT) or VITT that affects one individual in 1 million people around the world, Pai and colleagues suggested a possible presence of vaccinal antigens against the specific ADAMTS13 gene, responsible for creating a robust immune response in VITT.[37] Antibodies against PF4 have been found in VIPIT like the heparin complexes formed in heparin-induced thrombocytopenic purpura, suggesting analogous immune mechanisms.[37] Cases of thrombotic thrombocytopenic purpura were found in the elderly population following post-pneumococcal vaccination, with antigens present in the vaccine against the ADAMTS 13 gene.[37] Interestingly, hormonal factors could be among VITT causal factors. In younger females, thrombotic events are typically associated with ovarian hyperstimulation syndrome following reproductive treatment.[37]
Although the mechanism of VITT is still unclear, the absence of relapse suggests that this phenomenon is not related to spike protein immune responses. This conclusion is consistent with that reported by Greinacher et al. who showed that the anti-PF4 antibodies detected in patients with VITT do not interact with epitopes on the spike protein and appear to be independent of the spike antibody response. Scientists identified several other hereditary or naturally occurring thrombosis factors including pulmonary embolism, deep vein thrombosis, cancer, advanced age, trauma, smoking, inherited or acquired thrombophilic states, previous thromboembolism, and hospitalisation for congestive heart failure or acute exacerbation of chronic obstructive pulmonary disease.[38] Finally, considering the resemblance of VITT with thromboembolic events, it is also plausible that VITT may be multifactorial caused by similar triggers as thrombosis. However, given the small number of subjects with similar adverse events and the diversity of clinical outcomes, it remains difficult to draw final conclusions on associations.
6. Conclusion
We aimed to review the up-to-date published literature to further understand and provide updated information on COVID-19 vaccine-related thrombosis and thrombocytopenia. Understanding VITT mechanisms and pathogenesis should provide a platform to orient future research and to optimize management of VITT.
