NIEAs elicited by wild-type SARS-CoV-2 primary infection fail to enhance the infectivity of Omicron variants

Abstract

SARS-CoV-2 infection widely induces antibody response targeting diverse viral proteins, including typical representative N-terminal domain (NTD), receptor-binding domain (RBD), and S2 subunit of spike. A lot of NTD-, RBD-, and S2-specific monoclonal antibodies (mAbs) have been isolated from COVID-19 convalescents, some of which displaying potent activities to inhibit viral infection. However, a small portion of NTD-specific mAbs elicited by wild-type (WT) SARS-CoV-2 primary infection could facilitate the virus entry into target cells in vitro, so called NTD-targeting infection-enhancing antibodies (NIEAs). To date, SARS-CoV-2 has evolved to massive variants carrying various NTD mutations, especially recent Omicron BA.2.86 and JN.1. In this study, we investigated whether these WT-NIEAs could still enhance the infectivity of emerging Omicron variants. Nine novel WT-NIEAs with diverse germline gene usage were identified from 3 individuals, effectively enlarging available antibody panel of NIEAs. Bivalent binding of NIEAs to inter-spike contributed to their infection-enhancing activities. WT-NIEAs could enhance the infectivity of SARS-CoV-2 variants emerged before Omicron, but ineffective to Omicron variants including BA.2.86 and JN.1, which was because of their changed antigenicity of NTDs. Overall, these data clearly demonstrated the cross-reactivity of these pre-existed WT-NIEAs to a series of SARS-CoV-2 variants, helping to evaluate the risk of enhanced infection of emerging variants in future.

Keywords: Cross-reactivity; Infection-enhancing activity; NIEAs; NTD; SARS-CoV-2 variants.

Fig. 1

Enhancement activity and binding capacity of 9 NTD-targeting infection-enhancing antibodies (NIEAs). The infection-enhancing activity (a) and binding capacity (b) of P2S-2A5, P5-1B4, P5S-1B7, P5-1H4, P5S-2A8, P10S-1A10, P10S-1D9, P10S-2E8, and P5S-1D12 were measured by the pseudovirus infection assay and flow cytometry analysis. COV2-2490, COV2-2369, 8D2, and DH1052 were published NIEAs as positive controls. SA55 (anti-SARS-CoV-2 RBD), 76E1 (anti-SARS-CoV-2 S2), and VRC01 (anti-HIV-1) were non-NIEAs as negative controls. The infection-enhancing activity of IgG-, F(ab’)₂, and Fab-form P10S-1D9 were measured using HEK293T-hACE2 cells (c) and Huh7 cells (d). The experiments were performed twice and one representative result was shown

Fig. 2

Binding affinities of NIEAs to soluble WT SARS-CoV-2 NTD proteins by SPR. The association rate constant (K_on), dissociation rate constant (K_off), and dissociation constant (K_D) were calculated from three independent experiments and represented in mean values. One representative curve was shown

Fig. 3

The susceptibility and sequence alignment of SARS-CoV-2 variants to NIEAs. (a) Cross-reactivity of NIEAs to a series of SARS-CoV-2 variants including Alpha, Beta, Lota, Kappa, Delta, Mu, and C.1.2, as well as Omicron BA.1, BA.2, BA.2.75, XBB.1, BQ.1.1, EG.5.1, BA.2.86, and JN.1. Experiments were performed twice and data were shown in mean values. (b) Amino acid sequence alignment of NTDs from all tested SARS-CoV-2 pseudoviruses

Fig. 4

Identification of binding epitopes of NIEAs. (a) The infection-enhancing activity of NIEAs to a series of SARS-CoV-2 single-point mutated pseudoviruses. Experiments were performed twice and data were shown in mean values. (b) These infection-enhancing results were shown by different mutations and compared with that against WT. The positive rate, geometric mean EC₅₀, fold change, and significance of difference were labeled on the top. ‘‘-’’ represented decreased the infection-enhancing activity. The statistical significance was performed using two-tailed paired Wilcoxon test. **p < 0.01; *p < 0.05

Fig. 5

Structures of NIEAs bound to the NTD protein of SARS-CoV-2 variants. (a) The epitopes of NIEAs, namely DH1052 (PDB: 7LAB), 8D2 (PDB: 7DZX), and COV2-2490 (PDB: 2DZY) face the viral membrane. NIEAs are shown as cartoon, and colored orange, light goldenrod yellow, and light cyan, respectively. The other parts are shown as surface. The related NTD of NIEAs is colored yellow. (b) Footprint of DH1052 on spike NTD. The light chain and heavy chain of DH1052 are colored orchid and blue. The footprint is colored cadet blue and circled out by violet red line. Sites that affect and those that do not affect antibody-mediated enhancement of infection are colored red and green, respectively. (c) Mutation sites of 12 different SARS-CoV-2 variants are mapped onto the NTDs. The accession numbers for the NTD atomic model in PDB are 7R14 (Alpha), 7R16 (Beta), 7VXC (Kappa), 8HRI (Delta), 7YBJ (Mu), 7YBM (C.1.2), 8HHZ (BA.1), 7XIW (BA.2), 7YQT (BA.2.75), 8IOS (XBB.1), 8XMT (EG.5.1) and 8XLV (BA.2.86). The key amino acids are shown as sticks. Mutated amino acids are colored red, inserted amino acids are colored olive drab, and deleted amino acids are colored teal. The amino acids at both ends of the key amino acid sites that are missing in the atomic model of DH1052 are marked. The structure figures were prepared using ChimeraX (http://www.cgl.ucsf.edu/chimerax)