ABSTRACT
Background. COVID-19 disease severity and outcomes have been linked to high antibody titers and a dysregulated neutrophil immune response. Here we query associations and connections between the endogenous SARS-CoV-2 antibody response and neutrophil activation in COVID-19. Methods. Baseline serum or plasma samples from 57 patients hospitalized on oxygen with COVID-19 were used to perform;1) quantitative measurements of SARS-CoV-2 specific antibodies using a luciferase-based immunoprecipitation system assay, 2) quantitative measurements of neutrophil specific biomarkers using Luminex technology, and 3) neutrophil extracellular traps (NETs) as measured by myeloperoxidase-DNA (MPO-DNA) complexes by ELISA. Absolute neutrophil count (ANC) and immature granulocyte count (IGC) were measured from complete blood counts (CBC). Antibody levels were compared by disease severity using Wilcoxon rank-sum test and correlations were generated between antibody levels and neutrophil activation markers using Spearman's correlation (SC). Results. In a cohort of hospitalized patients, severe/critical COVID-19 was associated with higher levels of nucleocapsid-IgA (p=0.011) as well as spike-IgG (p= 0.0007) compared tomoderate disease,while spike-IgA and nucleocapsid-IgG showed similar associations, trending towards significance (Figure 1A). Levels of IgG-spike and IgG-nucleocapsid both had significant correlations with the ANC (SC 0.33, p = 0.029;SC 0.38 p = 0.012). All four antibody titers showed strong correlations with IGC, lactoferrin and lipocalin-2, evidence of emergency granulopoiesis. Further, S100A9, a component calprotectin correlated with spike-IgG and nucleocapsid-IgA levels (SC 0.29, p = 0.030, SC 0.29 p = 0.029). Lastly, we found circulating NETs correlated with spike IgA levels (SC 0.38 p = 0.006), and its correlations with IgG-spike and IgA-nucleocapsid additionally approached significance with NETs levels as well (Figure 1B). Antibody Levels Correlate with Disease Severity and Neutrophil Activation Markers Figure 1: A) Levels of anti-Spike and anti-Nucleocapsid IgA and IgG levels measured in the serum of 57 unvaccinated hospitalized COVID-19 patients. Moderate illness represents ordinal scale 5 requiring low flow oxygen, while severe/critical patients represent ordinal scale 6 and 7, requiring high flow oxygen, non-invasive or mechanical ventilation, respectively. P values are compared by a Wilcoxon ranked sum test. B) Heatmap showing Spearman correlations between levels of anti-Spike and anti-Nucleocapsid IgA and IgG and markers of neutrophil activation. P values for individual correlations are represented in parentheses. MPO (myeloperoxidase), ANC (absolute neutrophil count), S100A9 (S100 calcium binding protein A9). Conclusion. Higher anti-spike and anti-nucleocapsid IgG and IgA levels associate with more severe COVID-19 illness. Further, endogenous SARS-CoV-2 specific antibody levels associate with markers of emergency granulopoiesis and neutrophil activation. Inhibiting antibody mediated neutrophil activation may improve outcomes in COVID-19.
ABSTRACT
The ability of patients with hematologic malignancies (HM) to develop an effective humoral immune response after COVID-19 is unknown. A prospective study was performed to monitor the immune response to SARS-CoV-2 of patients with follicular lymphoma (FL), diffuse large B-cell lymphoma (DLBCL), chronic lymphoproliferative disorders (CLD), multiple myeloma (MM), or myelodysplastic/myeloproliferative syndromes (MDS/MPN). Antibody (Ab) levels to the SARS-CoV-2 nucleocapsid (N) and spike (S) protein were measured at +1, +3, +6 months after nasal swabs became PCR-negative. Forty-five patients (9 FL, 8 DLBCL, 8 CLD, 10 MM, 10 MDS/MPS) and 18 controls were studied. Mean anti-N and anti-S-Ab levels were similar between HM patients and controls, and shared the same behavior, with anti-N Ab levels declining at +6 months and anti-S-Ab remaining stable. Seroconversion rates were lower in HM patients than in controls. In lymphoma patients mean Ab levels and seroconversion rates were lower than in other HM patients, primarily because all nine patients who had received rituximab within 6 months before COVID-19 failed to produce anti-N and anti-S-Ab. Only one patient requiring hematological treatment after COVID-19 lost seropositivity after 6 months. No reinfections were observed. These results may inform vaccination policies and clinical management of HM patients.
Subject(s)
COVID-19/immunology , Hematologic Neoplasms/immunology , Immunity, Humoral/drug effects , Rituximab/pharmacology , SARS-CoV-2/immunology , Adult , Aged , Aged, 80 and over , Antibodies, Viral/drug effects , Antibodies, Viral/metabolism , Antibody Formation/drug effects , Antibody Formation/physiology , Antibody Specificity/drug effects , COVID-19/complications , COVID-19/epidemiology , COVID-19/therapy , Case-Control Studies , Female , Follow-Up Studies , Hematologic Neoplasms/complications , Hematologic Neoplasms/drug therapy , Hematologic Neoplasms/epidemiology , Hospitalization , Humans , Italy/epidemiology , Longitudinal Studies , Male , Middle Aged , Rituximab/therapeutic useABSTRACT
BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the cause of coronavirus disease 2019 (COVID-19), is associated with respiratory-related disease and death. Assays to detect virus-specific antibodies are important to understand the prevalence of infection and the course of the immune response. METHODS: Quantitative measurements of plasma or serum antibodies to the nucleocapsid and spike proteins were analyzed using luciferase immunoprecipitation system assays in 100 cross-sectional or longitudinal samples from patients with SARS-CoV-2 infection. A subset of samples was tested both with and without heat inactivation. RESULTS: At 14 days after symptom onset, antibodies against SARS-CoV-2 nucleocapsid protein showed 100% sensitivity and 100% specificity, whereas antibodies to spike protein were detected with 91% sensitivity and 100% specificity. Neither antibody levels nor the rate of seropositivity were significantly reduced by heat inactivation of samples. Analysis of daily samples from 6 patients with COVID-19 showed anti-nucleocapsid and spike protein antibodies appearing between days 8 and 14 after initial symptoms. Immunocompromised patients generally had a delayed antibody response to SARS-CoV-2, compared with immunocompetent patients. CONCLUSIONS: Antibody to the nucleocapsid protein of SARS-CoV-2 is more sensitive than spike protein antibody for detecting early infection. Analyzing heat-inactivated samples with a luciferase immunoprecipitation system assay is a safe and sensitive method for detecting SARS-CoV-2 antibodies.