A cross-reactive human IgA monoclonal antibody blocks SARS-CoV-2 spike-ACE2 interaction

Abstract

COVID-19 caused by SARS-CoV-2 has become a global pandemic requiring the development of interventions for the prevention or treatment to curtail mortality and morbidity. No vaccine to boost mucosal immunity, or as a therapeutic, has yet been developed to SARS-CoV-2. In this study, we discover and characterize a cross-reactive human IgA monoclonal antibody, MAb362. MAb362 binds to both SARS-CoV and SARS-CoV-2 spike proteins and competitively blocks ACE2 receptor binding, by overlapping the ACE2 structural binding epitope. Furthermore, MAb362 IgA neutralizes both pseudotyped SARS-CoV and SARS-CoV-2 in 293 cells expressing ACE2. When converted to secretory IgA, MAb326 also neutralizes authentic SARS-CoV-2 virus while the IgG isotype shows no neutralization. Our results suggest that SARS-CoV-2 specific IgA antibodies, such as MAb362, may provide effective immunity against SARS-CoV-2 by inducing mucosal immunity within the respiratory system, a potentially critical feature of an effective vaccine.

Fig. 1. Binding of MAb362 IgG and IgA to spikes of SARS-CoV and SARS-CoV-2.

MAb362 IgG and IgA bind to purified SARS-CoV S1 (S_1–590) and RBD (S_270–510) truncations a and SARS-CoV-2 S1 (S_1–604), RBD (S_319–541), and ectodomain trimer b. IgGs are red lines, IgAs are blue lines, and irrelevant IgGs are black. Affinity measurements of MAb362 IgG c, e, g and IgA d, f, h against the RBD truncations of S glycoprotein of SARS-CoV and SARS-CoV-2 c–f, as well as ectodomain trimer of SARS-CoV-2 g, h were conducted using bio-layer interferometry and demonstrate nano and sub-nanomolar affinities. Data are plotted as the mean ± s.d. from n = 3 independent experiments a, b. Source data are provided as a Source Data file.

Fig. 2. Mutationally guided molecular modeling of MAb362 binding to RBD.

a SARS-CoV-2 S1 was pre-incubated with MAb362 IgG (red circles) and IgA (blue squares) ranging from ~2 to 2000 nM. Both MAb362 isotypes demonstrated concentration-dependent inhibition of SARS-CoV-2 RBD binding to Vero E6 cells at concentrations >30 nM. Data are plotted as the mean ± s.d. from n = 3 independent experiments. b Mutational scanning was performed to better delineate the binding surface. Key residues were mutated and expressed as recombinant proteins. Identified critical residues (orange) were experimental confirmed by shifts in EC₅₀ values for MAb362 binding in ELISA relative to wild-type RBD (blue). EC₅₀ values calculated from n = 3 independent experiments. c Surface representation of the predicted molecular model of MAb362 SARS-CoV-2 RBD complex; the light chain of MAb362 (light yellow), the heavy chain (green), and the SARS-CoV-2 RBD (violet). d The predicted binding interface on SARS-CoV-2 RBD with MAb362. The residues identified by mutagenesis from b are labeled and colored according to influence degree; red represents strongest defects, orange for medium defects and yellow for subtle defects. Source data are provided as a Source Data file a, b.

Fig. 3. Predicted MAb362 structural epitope.

a Superposition of the space filling molecular model of MAb362 (green) complex on the crystal structure of the complex of ACE2 (orange) -SARS-CoV-2 RBD (violet) (6VW1) two views are rotated 180°. b The binding interface on SARS-CoV-2 RBD with ACE2 calculated from the co-crystal structure of the complex. The binding interface shown as darker shade is defined as having vdW contacts great than−0.5 kcal mol⁻¹. c Positioning of MAb362 on SARS-CoV-2 RBD (violet) relative to the binding of other currently published SARS-CoV-2 RBD-neutralizing antibodies: CR3022 (PDB: 6W41; orange); S309 (PDB: 6WPT; cyan); REGN10933 and REGN10987 (PDB: 6XDG; magenta and yellow); P2B-2F6 (PDB: 7BWJ; salmon); CB6 (PDB: 7C01; wheat) and B38 (PDB: 7BZ5; blue). MAb362 recognized a unique epitope overlapping with the binding interface of ACE2. d Predicted MAb362 molecular model on the spike trimer in open conformation with one RBD domain exposed 6VYB.

Fig. 4. IgA isotype switch enhances MAb362 neutralization of SARS-CoV-2.

MAb362 antibody-mediated neutralization of luciferase-encoding pseudovirions with spike proteins of SARS-CoV a and SARS-CoV-2 b. SARS-CoV and SARS-CoV-2 pseudovirions pre-incubated with serial dilutions of MAb362 were used to infect 293 cells expressing ACE2 receptor. Pseudoviral transduction was measured by luciferase activities in cell lysates 48 h post transduction to calculate neutralization (%) relative to non-antibody-treated controls. IC₅₀ values were calculated by nonlinear regression analysis using Prism version 8.1.1. Isotype switching improved SARS-CoV-2 IC50. Data are plotted as the mean ± s.d. from n = 3 independent experiments a, b. c Dose–response curve for PRNT with MAb362 at a starting concentration of 50 µg mL⁻¹ titrated 1:2. MAb362 sIgA had a 50% endpoint titer of 9.54 ± 5.88 µg mL⁻¹ calculated by Spearman–Kärber method, from n = 2 biologically independent experiments. Representative data are plotted with a Probit mid-point analysis curve ± 95% CI from one experiment with n = 2 technical replicates, using R programming language v3.5.3 and Library ggplot2 v3.3.0 for statistical computing and graphics^,. Source data are provided as a Source Data file a–c.