No changes in hemostasis after COVID-19-heterologous vaccination schedule: A subanalysis of the phase 2 CombiVacS study

Abstract

Background: Several cases of unusual thrombotic events and thrombocytopenia were described after vaccination with recombinant adenoviral vectors encoding the spike protein antigen of SARS-CoV-2.

Objectives: The objective of this study was to elucidate the impact of a COVID-19 heterologous vaccination schedule, including priming with adenovirus vaccine, on hemostasis profiles.

Methods: The present study is a subanalysis of the CombiVacS clinical trial initiated in April 2021 that included adult participants previously vaccinated with a single dose of ChAdOx1-S. Between 8 and 12 weeks after vaccination, they were randomly assigned (2:1) to receive either BNT162b2 vaccine (intervention group, n = 99) or continue observation (control group, n = 50). Samples drawn before and 28 days after a vaccination with BNT162b2 were analyzed for platelet count and markers of hemostasis (D-dimer, anti-PF4 antibodies, cfDNA, PAI-1, thrombin generation, and serum capacity to activate platelets).

Results: Platelet count from all participants after receiving BNT162b2 was within the normal range. Anti-PF4 antibodies were present in 26% and 18% of the subjects from the control and intervention groups, respectively, at day 28. In most cases, the levels of anti-PF4 antibodies were high before receiving BNT162b2. Serum from these participants did not activate platelets from healthy controls. There were no differences between the groups in PAI-1 and cfDNA plasma levels. According to the D-dimer plasma concentration, the thrombin generation test showed that none of the participants had a procoagulant profile.

Conclusion: Our data suggest that the heterologous vaccination against COVID-19 with ChAdOx1-S and a second dose with BNT162b2 might be safe in terms of haemostasis.

Keywords: COVID-19 vaccine; CombiVacS study; anti-PF4 antibodies; coagulation; hemostasis; heterologous vaccination; thrombocytopenia.

Figure 1

(A) Platelet count (x10³/μL), (B) D-dimer (ng/mL), and (C) anti-PF4 antibodies (% of the positive control of the kit), measured in both intervention and control groups on days 0 (V1) and 28 (V4) or in10 healthy controls who did not have COVID-19 and were not vaccinated against SARS-CoV-2 (NonV). Gray zone represents normal range of platelet count (A). Solid gray line represents cutoff reference value of the kits used (B, C) and dashed gray line represents the strong positive cutoff value for anti-PF4 serum levels (C). Scatter dot plots with mean ± 95% CI represent the values obtained for NonV controls (A, B)

Figure 2

Effect of serum from participants with anti-PF4 antibodies titer beyond cutoff value on either day 0 (V1) or 28 (V4) or both (V1 and V4) on activation (% of P-selectin expression) of unstimulated (A) or 10 μmol/L TRAP-6 stimulated (B) platelets. Scatter dot plots with mean ± 95% CI represent the values obtained with serum from nonvaccinated (NonV) controls analyzed simultaneously with samples from trial participants (A, B). No significant differences in platelet activation were observed between this nonvaccinated group and participants at V1

Figure 3

Thrombin generation test determined by calibrated automated thrombinography. Endogenous thrombin potential (ETP) and Lag time measured in both intervention and control groups on days 0 (V1) and 28 (V4). Thrombin generation was triggered by 1 pmol/L TF and 4 μmol/L of phospholipids (PPP-Reagent LOW) (A and B); 1 pmol/L TF (PRP reagent) (C and D); and 4 μmol/L phospholipids (MP reagent) (E and F). Gray zone represents in-house reference range (5% and 95% percentiles) obtained from historical control data. Scatter dot plots with mean ± 95% CI represent the values obtained from nonvaccinated (NonV) controls analyzed simultaneously with samples from trial participants. No significant differences were observed in thrombin generation test variables between this nonvaccinated group and participants at V1

Figure 4

(A) Serum PAI-1 and (B) plasma cfDNA concentration measured in both intervention and control groups on days 0 (V1) and 28 (V4). Gray zone represents the range of normality established by kit manufacturers' instructions (A) or in-house reference range (5% and 95% percentiles) obtained from historical control data (B). Scatter dot plots with mean ± 95% CI represent the values obtained for nonvaccinated (NonV) controls analyzed simultaneously with samples from trial participants. Significant differences were obtained in plasma cfDNA levels (P < .05) but not in serum PAI-1 concentration between this nonvaccinated group and participants at V1