A Spike-destructing human antibody effectively neutralizes Omicron-included SARS-CoV-2 variants with therapeutic efficacy

Figures

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  Fig 1. Ab08 neutralizes SARS-CoV-2 Omicron-included variants and binds to RBD with pico-molar affinities. (A-B) Neutralization assay of Ab08 using pseudoviruses harboring the S proteins from SARS-CoV-2 Wuhan-Hu-1 and Omicron BA.1 (A) or D614G and Omicron subvariants as indicated (B). Data are plotted as mean ± SEM (SEM: standard error of mean; n = 3 biological replicates). Numbers in brackets indicate IC50 values in μg/mL. (C) Neutralization activity of Ab08 against authentic SARS-CoV-2. A Plaque Reduction Neutralization Test (PRNT) was applied. Data are plotted as mean ± SEM (n = 3, biological replicates). (D) Binding affinities of Ab08 to SARS-CoV-2 prototype and variant RBDs using bilayer interferometry (BLI). Biotinylated RBDs were immobilized on a Streptavidin sensor and Ab08 (scFv) was used as analytes at indicated concentrations (nM). Solid lines represent experimental data and dashed lines represent fitted curve.

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  Fig 2. Ab08 shows therapeutic efficacy in hACE2 mice infected with live SARS-CoV-2. (A) hACE2 mice intranasally infected with 105 PFU of SARS-CoV-2 were divided into two groups: the Ab08 group was intraperitoneally administered with 10 mg/kg Ab08 (n = 5) and the PBS group was injected with PBS (n = 5) at 4-hour, 28-hour and 52-hour post infection (red arrow). All mice were sacrificed on day 4 post infection. The mouse images are from Openclipart. (B) Weight change of mice was monitored daily (n = 5; Unpaired Student’s t test: *p<0.05; **p<0.01; symbols denote mean ± SEM.) (C) Percent survival was determined for the Ab08 and PBS groups. (n = 5; Kaplan-Meier survival analysis with Log-rank test: * p<0.05). (D) SARS-CoV-2 viral RNA loading in lungs of the Ab08 and PBS groups on day 4 post infection. Viral RNA copies were analyzed by real-time qPCR. (Unpaired Student’s t test: * p<0.05). (E) Representative H&E staining showing pathological changes in the mouse lungs from different groups (Top: Non-infected; Middle: PBS; Bottom: Ab08). The PBS group exhibited severe pneumonia with blocked terminal bronchioles, fibroplasia, and organization (black arrow), and peribronchial and perivascular…

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  Fig 3. Structural characterization informs mechanism for the relatively broad activity of Ab08. (A) The overall structure of the Ab08 (scFv) (cartoon) in complex with the high-chair-shaped RBD (grey surface). The Ab08 epitope is colored green. RBD-interacting CDRs are color-coded as indicated. (B) Interactions between RBD (grey) and Ab08. CDR residues are color-coded as in A. Ab08 residues are labeled black and RBD residues are labeled grey. A prime symbol indicates residues from the light chain. Dash lines indicate distances within 3.6 Å. (C) The β-sheet interaction between Ab08 and RBD. Backbone- and side-chain-mediated H-bonds are indicated by black and orange dash lines, respectively. (D, E) Ab08 is compatible with ACE2-binding. (D) Structural alignment of ACE2-RBD (PDB ID 6M0J) and Ab08-RBD (this work) reveals no clashes between ACE2 (yellow) and Ab08 (red and blue). (E) Ab08 and ACE2 simultaneously bind RBD. A sensor immobilized with RBD was soaked in 100 nM of ACE2 before being further soaked in ACE2-containing buffer with (black) or without (red) 100 nM of Ab08 for BLI signal recording. As a control, the Ab08-RBD interaction was monitored in the absence of ACE2 (blue). (F) The…

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  Fig 4. Ab08 neutralizes SARS-CoV-2 by destructing S trimer. (A-B) Structural alignment of Ab08 to the S trimer structure (PDB ID 7WLY) reveals severe clashes with the N-terminal domain (NTD) in cases of both the “down”-RBD (A) and the “up”-RBD conformation (B). The epitope of Ab08 is colored green. The three protomers of S and the RBD of the protomer A are color-coded as indicated in (i). In (ii), the RBD from protomer A is shown as surface representation, and Ab08 (magenta) and the NTD from protomer C (orange) are shown as ribbon representations. The two glycans are shown as cyan sticks. In (iii), the disruption of the interactions between NTD N165-linked glycans (Cyan) and RBD by Ab08 is highlighted. (C-E) 2D class averages of Negative-staining particles of S-2P showing typical S trimer features (C) were lost upon incubation with Ab08 (D), but not with an RBM-targeting control mAb (CB6, E) [22]. Raw images are in S7 Fig. In E, the 2D classes show binding of two CB6 molecules per S trimer and CB6 may induce more “up”-RBD population, a phenomenon that has been reported for other RBM-targeting antibodies [44].