ABSTRACT
Introduction: Severe COVID-19 patients present with a hypercoagulable state, complement activation and endothelial perturbation, which result from an excessive inflammatory response. Thromboinflammation is one important mechanism underlying the COVID-19-associated coagulopathy and the increased risk of thrombosis. Bergamo city is one of the first and most affected area by SARS-CoV-2 infection in the world. For this reason, since the beginning we were actively involved in recruiting convalescent COVID-19 patients, in a program of selection of candidates for convalescent plasma donation. In a large cohort of convalescent COVID-19 patients, we aimed to characterize markers of coagulation activation and endothelial perturbation, in order to explore whether the COVID-19-related hemostasis activation might persist afterwards and evaluate its possible association with the degree of severity of the previous infection, and/or with demographic characteristics, or anti-SARS-CoV-2 antibody levels. Methods: In 392 convalescent COVID-19 patients (216M/176F, median age: 46 years) plasma levels of fibrinogen, protein C, protein S, factor V, factor VIII, factor XIII, D-dimer, von Willebrand factor (vWF), prothrombin fragment F1+2 were measured at the recruitment, i.e. 1-5 months from recovery. Samples were tested for the anti-SARS-CoV-2 antibodies, including anti-S IgG (Anti-S Ab) and anti-N IgG (Anti-N Ab) antibodies at enrollment and at each scheduled subsequent visits. Results: Levels of fibrinogen, D-dimer, von Willebrand factor, protein S and protein C were significantly higher (p<0.05) in patients who were hospitalized for severe COVID-19 as compared to patients who were treated at home. There was no correlation between levels of coagulation biomarkers and days from end of symptoms. Male gender, age > 40 years, and severe form of COVID-19 were identified as independent predictors of high levels of both anti-S and anti-N Ab (p<0.001). Among hemostatic biomarkers, fibrinogen (p<0.01) and vWF (p<0.05) independently predict high levels of anti-S Ab. In particular, vWF levels positively correlated with anti-S Ab levels (vWFantigen r=0.188;vWF-activity r=0.241 and vWF-RiC of r=0.223, p<0.01). Evaluation of anti-SARS-CoV2 antibody levels at different time points during follow up revealed that 30% of patients displayed high levels of anti-S Ab until more than 8 months from the end of symptoms. Conclusions: Convalescent patients, with a history of severe COVID-19 had a persistent endothelium activation, despite of disease clinical remission even after 9 months from end of symptoms. Furthermore, fibrinogen and vWF levels predicted high levels of Anti-S Ab. Among demographic characteristics, gender, age and severe disease can be predictors of increased antibody response. These findings suggest that inflammation, coagulation and endothelial dysfunction may persist after recovery and may explain the findings of persistent clinical symptoms reported in these patients after healing from COVID-19.
ABSTRACT
[Formula presented] Introduction: Patients (pts) with immune thrombotic thrombocytopenic purpura (iTTP) are at high risk of severe COVID-19, therefore protection from SARS-CoV-2 by vaccination is particularly relevant in this setting, although concerns may exist on possible adverse reactions or disease relapse after vaccination. In this study, in a group of iTTP pts who received in-hospital COVID-19 vaccination in a special program for ‘fragile patients’, we prospectively evaluated over time the antibody response, the clinical and laboratory disease parameters and hemostatic biomarker levels. Methods: Twelve iTTP pts in clinical remission and regularly followed-up in our Center were enrolled in April 2021, all of them received 2 doses of BNT162b2 vaccine (Pfizer-BioNTech) over 21 days, and were followed-up for clinical and laboratory testing for 60 days. Blood samples were collected at enrollment (day 0, D0) before the 1 st vaccine dose;on day 21 (D21) before the 2 nd dose;and on day 60 (D60) after the 1 st dose. Blood cell counts, anti-Spike receptor-binding-domain protein (anti-S/RBD) IgG, ADAMTS-13 activity, and anti-ADAMTS-13 IgG (chromogenic assay and ELISA), were measured at each time point. Additionally, an extensive study of hemostatic markers (i.e. FVIII, von Willebrand Factor (vWF) antigen and activity, fibrinogen, D-dimer, tPA, PAI, and F1+2) was performed. Follow up is currently continuing. Results: Median age of our cohort was 65 years with M/F ratio of 4/8. Median time since last acute iTTP episode was 40 months, median follow up of the cohort was 71 months (95% CI 30-126). All pts were in clinical remission, except one patient (P1) who had an iTTP relapse after contracting SARS-CoV-2 infection, in Dec 2020, and was on low-dose steroids on D0. One patient (P2) had an ADAMTS-13 relapse in Jan 2021, and received pre-emptive rituximab. No other pts were on immunosuppressive therapy. Concerning the status of ADAMTS-13 activity on D0, 6 pts showed normal levels (>50%), while 5 had a moderate (50-20%) and 1 a complete (<10%) ADAMTS-13 deficiency. This latter patient (P3) had normal ADAMTS-13 activity before the pandemic. All patients were negative for anti-ADAMTS-13 inhibitor. Further, on D0, the anti-S/RBD IgG testing was positive in 3/12 pts (median 704,1 AU/mL), due to symptomatic infection in 1 case (P1), and asymptomatic in 2 (P3 and 1 pt with ADAMTS-13 activity of 54%, P4). The study of hemostatic markers on D0 showed an increase in median levels of FVIII and vWF antigen and activity. These parameters were altered in 7/12, 11/12 and 8/12 pts, respectively. Fibrinogen and D-dimer were increased in 3/12 and 2/12, respectively. Notably, P1, P3 and P4 presented the highest levels of FVIII and vWF antigen, associated with high levels of vWF activity in P1 and P3 (mean 233%);moreover, P3 showed higher levels of D-dimer (708 ng/mL) and tPA (13 ng/ml). After the 2 doses of BNT162b2, no significant clinical side effects were reported, and no changes in platelet counts. ADAMTS-13 activity and inhibitors did not significantly change on D21 and D60. A complete ADAMTS-13 activity deficiency persisted in P3 on D21 and D60, associated with anti-ADAMTS-13 IgG titer >15 U/ml, despite clinical remission. Overall, a significant increase in anti-S/RBD IgG level was observed on D21 (p = 0.0005) and D60 (p = 0.0005). Remarkably, only P2 did not show an increase in anti-S/RBD IgG titer after both doses of BNT162b2. Median levels of FVIII and vWF antigen did not significantly change during follow up, while increased vWF activity was seen on D60 (p = 0.05). Fibrinogen levels were stable, and an increase in D-dimer (>1000 ng/mL both on D21 and D60) was seen in P3. There were no changes in the other hemostatic parameters, and no thromboses were observed. Conclusions: In our cohort of iTTP pts, COVID-19 was associated with 1 clinical and 1 ADAMTS-13 relapse. Our data show that SARS-CoV-2 vaccination was effective in inducing an antibody response in all but one patient who received rituximab within 3 months before vaccinat on, confirming recent findings. Overall, vaccination had no relevant impact on the hemostatic profile of our pts, and did not appear to be a driver of iTTP relapses. However, anti-SARS-CoV-2 antibodies monitoring in iTTP pts may be useful after vaccination, as currently it is unknown how long the antibody titer may persist. Although small, this study is in favor of efficacy and safety of mRNA vaccines in pts with iTTP. Disclosures: Falanga: Bayer: Honoraria;Sanofi: Honoraria;Leo Pharma: Honoraria;Pfizer: Honoraria.