Anti-Severe Acute Respiratory Syndrome Coronavirus 2 Hyperimmune Immunoglobulin Demonstrates Potent Neutralization and Antibody-Dependent Cellular Cytotoxicity and Phagocytosis Through N and S Proteins.

BACKGROUND: Although coronavirus disease 2019 (COVID-19) vaccinations have provided a significant reduction in infections, effective COVID-19 treatments remain an urgent need. METHODS: Functional characterization of anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) hyperimmune immunoglobulin (hIG) from human convalescent plasma was performed by different virus neutralization methodologies (plaque reduction, virus-induced cytotoxicity, median tissue culture infectious dose [TCID50] reduction, and immunofluorimetry) at different laboratories using geographically different SARS-CoV-2 isolates (USA [1], Italy [1], and Spain [2]; 2 containing the D614G mutation). Neutralization capacity against the original Wuhan SARS-CoV-2 strain and variants (D614G mutant, B.1.1.7, P.1, and B.1.351) was evaluated using a pseudovirus expressing the corresponding spike (S) protein. Antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP) was also evaluated. RESULTS: All SARS-CoV-2 isolates were potently neutralized by hIG as shown by all 4 methodologies. Wild-type SARS-CoV-2 and variants were effectively neutralized using the pseudovirus. The hIG (IgG type) induced ADCC and ADCP against SARS-CoV-2 N and S proteins but not E protein. Very low concentrations (25-100 µg IgG/mL) were required. A potent effect was triggered by antibodies in hIG solutions against the SARS-CoV-2 S and N proteins. CONCLUSIONS: Beyond neutralization, IgG Fc-dependent pathways may play a role in combatting SARS-CoV-2 infections using COVID-19 hIG. This could be especially relevant for the treatment of more neutralization-resistant SARS-CoV-2 variants.

Effective presence of antibodies against common human coronaviruses in immunoglobulin medicinal products.

BACKGROUND: Immunoglobulin products (for intravenous, intramuscular and subcutaneous administration) prepared from geographically diverse plasma pools were tested for activity against common human coronaviruses (HCoVs). Products from plasma obtained from Germany, Czech Republic, Slovak Republic, USA and Spain were tested for antibodies to common HCoVs: 229E, OC43, NL63 and HKU1. As these products are manufactured from pooled plasma from thousands of donors, the antibodies therein are representative of HCoV exposure in the population at large. METHODS: Immunoglobulin products were tested for antibodies to four common HCoVs by enzyme-linked immunosorbent assays (ELISAs). Neutralization assays were conducted using HCoV-229E cultured on to MRC5 cells. RESULTS: ELISAs showed that when expressed as specific activity (anti-HCoV activity/mg immunoglobulin), similar activity against the four common HCoVs was seen across the immunoglobulin products regardless of concentration or geographic origin. Highest anti-HCoV activity was seen against HCoV-229E, followed by HCoV-OC43, HCoV-NL63 and HCoV-HKU1. The neutralization assays showed similar potency for two immunoglobulin products prepared by different processes. CONCLUSIONS: To the authors' knowledge, this is the first demonstration of antibodies to common HCoVs in immunoglobulin products. These results may explain the cross-reactivity seen with pre-pandemic immunoglobulin products and severe acute respiratory syndrome coronavirus-2, and contribute to differences in severity of illness between patients.

Production of anti-SARS-CoV-2 hyperimmune globulin from convalescent plasma.

BACKGROUND: In late 2019, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus emerged in China and quickly spread into a worldwide pandemic. Prior to the development of specific drug therapies or a vaccine, more immediately available treatments were sought including convalescent plasma. A potential improvement from convalescent plasma could be the preparation of anti-SARS-CoV-2 hyperimmune globulin (hIVIG). STUDY DESIGN AND METHODS: Convalescent plasma was collected from an existing network of plasma donation centers. A caprylate/chromatography purification process was used to manufacture hIVIG. Initial batches of hIVIG were manufactured in a versatile, small-scale facility designed and built to rapidly address emerging infectious diseases. RESULTS: Processing convalescent plasma into hIVIG resulted in a highly purified immunoglobulin G (IgG) product with more concentrated neutralizing antibody activity. hIVIG will allow for the administration of greater antibody activity per unit of volume with decreased potential for several adverse events associated with plasma administration. IgG concentration and IgG specific to SARS-CoV-2 were increased over 10-fold from convalescent plasma to the final product. Normalized enzyme-linked immunosorbent assay activity (per mg/ml IgG) was maintained throughout the process. Protein content in these final product batches was 100% IgG, consisting of 98% monomer and dimer forms. Potentially hazardous proteins (IgM, IgA, and anti-A, anti-B, and anti-D) were reduced to minimal levels. CONCLUSIONS: Multiple batches of anti-SARS-CoV-2 hIVIG that met regulatory requirements were manufactured from human convalescent plasma. The first clinical study in which the hIVIG will be evaluated will be Inpatient Treatment with Anti-Coronavirus Immunoglobulin (ITAC) [NCT04546581].

Cross-neutralization activity against SARS-CoV-2 is present in currently available intravenous immunoglobulins.

Background: Cross-reactivity against human coronaviruses with Flebogamma® DIF and Gamunex®-C, two available intravenous immunoglobulins (IVIG), has been reported. In this study, these IVIG were tested for neutralization activity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), SARS-CoV and Middle East respiratory syndrome CoV (MERS-CoV). Materials & methods: Neutralization capacity of lots of IVIG manufactured prior to COVID-19 pandemic was assessed against these viruses in cell culture. Infectivity neutralization was quantified by percent reduction in plaque-forming units and/or cytopathic/cytotoxic methods. Results: All IVIG preparations showed neutralization of SARS-CoV-2 isolates. All IVIG lots produced neutralization of SARS-CoV. No IVIG preparation showed significant neutralizing activity against MERS-CoV. Conclusion: The tested IVIG contain antibodies with significant in vitro cross-neutralization capacity against SARS-CoV-2 and SARS-CoV, but not MERS-CoV. These preparations are currently under evaluation as potential therapies for COVID-19.

Currently available intravenous immunoglobulin contains antibodies reacting against severe acute respiratory syndrome coronavirus 2 antigens.

Aim: There is a critical need for effective therapies that are immediately available to control the spread of COVID-19 disease. Material & methods: Gamunex®-C and Flebogamma® DIF (Grifols) intravenous immunoglobulin (IVIG) products were tested using ELISA techniques for antibodies against several antigens of human common betacoronaviruses that may crossreact with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus. Results: Both IVIGs showed consistent reactivity to components of the tested viruses. Positive crossreactivity was seen in SARS-CoV, middle east respiratory syndrome-CoV and SARS-CoV-2. For SARS-CoV-2, positive reactivity was observed at IVIG concentrations ranging from 100 µg/ml with Gamunex-C to 1 mg/ml with Flebogamma 5% DIF. Conclusion: Gamunex-C and Flebogamma DIF contain antibodies reacting against SARS-CoV-2 antigens. Studies to confirm the utility of IVIG preparations for COVID-19 management may be warranted.