Antibodies from convalescent plasma promote SARS-CoV-2 clearance in individuals with and without endogenous antibody response

Abstract

BACKGROUNDNeutralizing antibodies are considered a key correlate of protection by current SARS-CoV-2 vaccines. The manner in which human infections respond to therapeutic SARS-CoV-2 antibodies, including convalescent plasma therapy, remains to be fully elucidated.METHODSWe conducted a proof-of-principle study of convalescent plasma therapy based on a phase I trial in 30 hospitalized COVID-19 patients with a median interval between onset of symptoms and first transfusion of 9 days (IQR, 7-11.8 days). Comprehensive longitudinal monitoring of the virological, serological, and disease status of recipients allowed deciphering of parameters on which plasma therapy efficacy depends.RESULTSIn this trial, convalescent plasma therapy was safe as evidenced by the absence of transfusion-related adverse events and low mortality (3.3%). Treatment with highly neutralizing plasma was significantly associated with faster virus clearance, as demonstrated by Kaplan-Meier analysis (P = 0.034) and confirmed in a parametric survival model including viral load and comorbidity (adjusted hazard ratio, 3.0; 95% CI, 1.1-8.1; P = 0.026). The onset of endogenous neutralization affected viral clearance, but even after adjustment for their pretransfusion endogenous neutralization status, recipients benefitted from plasma therapy with high neutralizing antibodies (hazard ratio, 3.5; 95% CI, 1.1-11; P = 0.034).CONCLUSIONOur data demonstrate a clear impact of exogenous antibody therapy on the rapid clearance of viremia before and after onset of the endogenous neutralizing response, and point beyond antibody-based interventions to critical laboratory parameters for improved evaluation of current and future SARS-CoV-2 therapies.TRIAL REGISTRATIONClinicalTrials.gov NCT04869072.FUNDINGThis study was funded via an Innovation Pool project by the University Hospital Zurich; the Swiss Red Cross Glückskette Corona Funding; Pandemiefonds of the UZH Foundation; and the Clinical Research Priority Program "Comprehensive Genomic Pathogen Detection" of the University of Zurich.

Keywords: Adaptive immunity; COVID-19; Immunotherapy; Therapeutics.

Figure 1. Study design and clinical and virological assessment.

(A) Schematic depiction of the study design, including timeline of consecutive treatment with convalescent plasma units and clinical and laboratory assessments. PCR NPS, PCR from nasopharyngeal swab. Figure created with BioRender (biorender.com). (B) Study flow chart. CPT, convalescent plasma therapy. (C) Longitudinal clinical assessment of trial participants (n = 30), with a 7-category ordinal scale for pulmonary function. 1: Usual activities with minimal/no symptoms. 2: No supplemental oxygen; symptomatic and unable to undertake usual activities. 3: Supplemental oxygen <4 L/min. 4: Supplemental oxygen ≥4 L/min. 5: Noninvasive ventilation or high-flow oxygen. 6: Invasive ventilation, extracorporeal membrane oxygenation, mechanical circulatory support. 7: Death. (D and E) Assessment of viral load. Longitudinal viral RNA concentrations (copies/mL) in plasma (D) and NPS (E) in trial participants (n = 30).

Figure 2. Antibody characteristics of convalescent donor plasma.

(A) Profiling of donor plasma for neutralization activity and seroreactivity to SARS-CoV-2 antigens. Data of all banked donors (transfused, n = 30; not transfused, n = 45) are shown. Binding antibody reactivity was measured in the ABCORA test (readout median fluorescence intensity signal over cutoff [SOC]). Fifty percent neutralization titers (NT₅₀) were measured against Wuhan-Hu-1 pseudotyped virus. Donor plasmas are stratified into plasmas with high (NT₅₀ > 250) and low (NT₅₀ ≤ 250) neutralization potency. (B) Antibody binding titers in banked donor plasma measured by the Elecsys S assay (U/mL). (C) NT₅₀ titers against Wuhan-Hu-1 pseudotyped virus in banked plasma donors. (B and C) Two-sided, unpaired t test comparing the transfused and non-transfused plasma.

Figure 3. Treatment with highly neutralizing plasma leads to faster viral clearance.

(A and B) Assessment of the time (days) to viral clearance in NPSs in plasma recipients (n = 30) according to the level of neutralization potency of the received convalescent donor plasma. High neutralization activity is set as NT₅₀ > 250, low neutralization activity as NT₅₀ ≤ 250. (A) Kaplan-Meier curves compared by log-rank test. (B) Survival function estimate with a parametric model for interval-censored data. The parametric estimate is adjusted for the baseline NPS viral load and the presence of any comorbidity. Depicted survival curves of recipients of high- and low-neutralizing donor plasma correspond to the predicted viral clearance in individuals without comorbidity and with a baseline viral load (NPS) equal to the median viral load observed among the 30 patients. (C) Forest plot corresponding to B, showing the hazard ratios of the univariable (black) and the multivariable (red) model of time to viral clearance in NPSs for convalescent donor plasma neutralization level (low/high), baseline viral load, and the presence of comorbidity.

Figure 4. Spike-specific binding and neutralizing antibodies in convalescent donor plasma are linked with rapid viral clearance.

(A) Impact of convalescent donor plasma antibody parameters on the time to viral clearance in recipients (n = 30) was assessed by multivariable parametric survival models. Hazard ratios for individual antibody reactivities adjusted for the presence of comorbidity and the baseline viral load (NPS) are shown. Significant results are marked in red. Low and high binding activity for each individual binding antibody parameter measured in the Elecsys S test (total RBD) and for the ABCORA test parameters is stratified by the respective median binding reactivity. Low and high neutralization activity of transfused convalescent donor plasma is stratified by an NT₅₀ of 250. (B) Forest plot depicting hazard ratios of univariable (black) and multivariable (red) models of time to viral clearance including total S1 (sum of ABCORA IgG, IgA, and IgM reactivity with S1) stratified by the median binding reactivity. Multivariable analyses are corrected for baseline viral load (NPS) and the presence of comorbidity.

Figure 5. High-neutralizing plasma leads to faster virus decay in NPSs.

(A and B) Censored regression model estimating decay rate of viral load (log₁₀ viral load) in NPSs in recipients (n = 30) from time of treatment initiation according to the received convalescent donor plasma with respect to neutralizing antibody content (low neutralization, NT₅₀ ≤ 250, light green; high neutralization, NT₅₀ > 250, purple) (A) or level of binding antibodies as defined by the ABCORA test total S1 values (sum of IgG, IgA, and IgM reactivity with S1) (B). Low and high total S1 binding is stratified by the median binding reactivity. Significance was assessed using a 2-sided t test.

Figure 6. Recipients’ endogenous neutralizing antibodies efficiently control plasma viremia.

(A) Recipients’ pretransfusion endogenous 50% plasma neutralization titers (NT₅₀) against Wuhan-Hu-1 pseudovirus. For each time point (days 0, 9, 72) the number of patients with available sample is indicated. Box plots depict the interquartile ranges, with vertical lines (whiskers) representing a distance of 1.5 times the interquartile range below the first quartile and above the third quartile. (B) Recipients’ longitudinal binding antibody activity at baseline (day 0), day 9, and day 72 assessed with the multiplex SARS-CoV-2 ABCORA 2 test. Sample numbers per time point are as shown in A. Signal over cutoff (SOC) values of IgG, IgA, and IgM against RBD and S1 are shown. Box plots indicate the interquartile ranges,with vertical lines (whiskers) representing a distance of 1.5 times the interquartile range below the first quartile and above the third quartile. (C) Spearman’s correlation matrix assessing correlation between age; comorbidities; viral load in NPS (copies/mL) and blood (binary yes/no); and neutralization titer (NT₅₀) and total S1 (SOC values) at day 0 and day 9. Levels of significance were assessed by asymptotic t approximation of Spearman’s rank correlation. Color shading denotes correlation coefficient. Levels of significance: *P < 0.05, **P < 0.01, ***P < 0.001. (D) Group comparison of plasma viral load from recipients stratified by presence of pretransfusion endogenous neutralization activity (baseline d = 0) (no neutralization, NT₅₀ ≤ 100; neutralization activity, NT₅₀ > 100). Levels of significance were calculated by Wilcoxon’s rank sum test.

Figure 7. Influence of recipients’ endogenous and convalescent plasma antibodies on viral clearance.

(A) Analysis of recipients’ plasma neutralization activity before and after transfusion of convalescent plasma. NT₅₀ titers against Wuhan-Hu-1 pseudotyped virus at baseline (day 0, n = 26), day 9 (n = 26), and day 72 (n = 26) are depicted. Recipients are stratified by neutralizing levels of transfused convalescent donor plasma (left: low NT₅₀ donor plasma ≤ 250; right: high NT₅₀ > 250). Levels of significance were calculated by 2-sided, paired t test. (B) Longitudinal comparison of NT₅₀ activity in low/high donor plasma NT₅₀ groups at days 0, 9, and 72. Levels of significance were calculated by 2-sided, unpaired t test. (C) Longitudinal comparison of the evolution of recipient neutralization activity (no neutralization, NT₅₀ ≤ 100; neutralization activity, NT₅₀ > 100). Levels of significance were calculated by unpaired t test. (D) Alteration in NT₅₀ between days 9 and 72 (ratio) according to the donor and recipient baseline neutralization. Levels of significance were calculated by 2-sided, unpaired t test. (E and F) Forest plot showing hazard ratios of univariable and multivariable survival models of time to viral clearance (n = 26). Both E and F test for baseline viral load and comorbidity; E tests additionally for neutralization level in donor plasma and recipients’ pretransfusion plasma, and F tests additionally for S1 antibody level in donor plasma and in recipients’ pretransfusion plasma.