Potent and broadly neutralizing antibodies against sarbecoviruses induced by sequential COVID-19 vaccination

Figures

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  Fig. 1. Cross-reactive immune responses elicited by sequential vaccination. a Vaccination schedule overview. A healthy volunteer received a total of five doses of vaccine, starting with a two-dose mRNA vaccine (BNT162b2) followed by a three-dose inactivated whole-virion vaccine (BBIBP-CorV). Blood samples were collected after the 4th and 5th vaccination dose as illustrated in the diagram. b Neutralizing antibody titers of the sequential vaccination doner (4-dose: blue and 5-dose: red) compared to the 3-dose BBIBP vaccinees (black, n = 11). Neutralizing antibody titers represent serum dilution required to achieve 50% virus neutralization (ID50). The numbers indicate the fold of enhancement of ID50 values relative to mean titer measured among 3-dose BBIBP vaccinees.

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  Fig. 2. Identification of broadly neutralizing antibodies against severe human coronaviruses. a Schematic representation of the isolation process for potential broadly neutralizing antibodies using a combined method of single B-cell sorting and virtual screening. b Heatmap showing the binding EC50 values of the selected mAbs with S trimers of MERS-CoV, SARS-CoV, and SARS-CoV-2, respectively. c Heatmap showing the neutralization IC50 values of each mAb with indicated pseudotyped viruses. d Neutralization curves (left) and IC50 values (right) of indicated mAb with MERS-CoV, SARS-CoV, and SARS-CoV-2 pseudotyped viruses, respectively. The data are representative of one of at least three independent experiments and are presented as the mean ± SEM.

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  Fig. 3. Neutralization of pseudotyped and authentic SARS-CoV-2 variants, SARS-CoV and other sarbecoviruses. a Neutralization of SARS-CoV-2 variants of concern (Alpha, Beta, Gamma, Delta, and Omicron BA.1) by indicated mAb. b Neutralization of major SARS-CoV-2 Omicron subvariants, including BA.2, BA.5, BQ.1.1, and XBB, as well as its latest subvariants, such as XBB.1, XBB.1.16, XBB.2.3.3, EG.5.1, EU.1.1, and FY.4 by indicated mAb. c Neutralization of SARS-CoV-2-related sarbecoviruses (Pangolin CoV-GD and Bat CoV RaTG13) by indicated mAb. d Neutralization of SARS-CoV-related sarbecoviruses (Bat WIV-1 and SHC014) by indicated mAb. e IC50 neutralization values of each mAb against SARS-CoV-2 and its variants, and other related sarbecoviruses pseudoviruses as well as the indicated live viruses. The data are representative of one of at least three independent experiments and are presented as the mean ± SEM.

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  Fig. 4. Cryo-EM structure of PW5-570 IgG with Omicron BA.1 S trimer. a Cryo-EM structures of the BA.1 S trimer in complex with the antibody PW5-570 IgG. b Structure of BA.1-RBD–PW5-570. The RBD is displayed in tomato surface mode. The heavy chain and light chain of PW5-570 are shown as ribbons colored in dark green and light green, respectively. c Close-up view of the PW5-570 epitope on the RBD. ACE2-binding epitopes are represented by yellow dashed lines. d Detailed interactions between the PW5-570 and BA.1 S RBD. Hydrogen bonds are represented by dashed lines. Hydrophobic interactions are marked with red circles. e PW5-570 clash with the ACE2. ACE2 is marked yellow.

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  Fig. 5. Cryo-EM structure of PW5-5 IgG with Omicron XBB and SARS-CoV S trimer. a Cryo-EM structures of the XBB S in complex with the antibody PW5-5 IgG. b Structure of XBB RBD–PW5-5. The RBD is displayed in tomato. The heavy chain and light chain of PW5-5 are shown as ribbons colored in dark blue and light blue, respectively. c Close-up view of the PW5-535 epitope on XBB and SARS-CoV RBD. d Detailed interactions between the PW5-5 and XBB S RBD. e Detailed interactions between the PW5-5 and SARS-CoV S RBD. Hydrogen bonds are represented by dashed lines. Hydrophobic interactions are marked with red circles.

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  Fig. 6. Cryo-EM structure of PW5-535 IgG with Omicron XBB and SARS-CoV S trimer. a Cryo-EM structure of the XBB S in complex with the antibody PW5-535 IgG. b Cryo-EM structures of the SARS-CoV S in complex with the antibody PW5-535 IgG. c Structure of XBB RBD–PW5-535. The RBD is displayed in tomato. The heavy chain and light chain of PW5-535 are shown as ribbons colored in dark purple and light purple, respectively. d Close-up view of the PW5-535 epitope on the XBB RBD and SARS-CoV RBD. e Detailed interactions between the PW5-535 and XBB S RBD. f PW5-535 causes steric hindrance with ACE2. g Detailed interactions between the PW5-535 and SARS-CoV S RBD and SD1. Hydrogen bonds are represented by dashed lines. Hydrophobic interactions are marked with red circles.

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  Fig. 7. Prophylactic efficacy of PW5-5, PW5-535, and PW5-570 against SARS-CoV, SARS-CoV-2 Omicron BA.1, and XBB.1 in golden Syrian hamsters. a Schematic of the in vivo mAb efficacy test experiment in hamsters. b, c, e, f, h, i Viral load quantification of Omicron BA.1 (b, c), XBB.1 (e, f), and SARS-CoV (h, i) in golden hamster lung and nasal turbinate (NT) with or without treatments. Golden hamster lung and nasal turbinate samples were harvested on day 2 post SARS-CoV-2 Omicron BA.1, XBB.1, or SARS-CoV challenge and homogenized for qRT-PCR analysis and plaque assay titration (n = 6). d, g, j Immunofluorescence images of the whole section of infected hamster lungs with or without treatments. SARS-CoV-2 N protein was identified with a rabbit anti-SARS-CoV-2-N immune serum (green). Nuclei was identified with DAPI stain (blue). Scale bars in (d, g, j) represented 1000 μm for fourfold magnifications of the objective with tenfold magnification at the eyepiece. The experiments in (b, c and e, f and h, i) were repeated two times independently with similar results, respectively. Data represented means and SEMs from the indicated number of biological repeats. Statistical significance between…

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  Fig. 8. Therapeutic efficacy of PW5-5 and PW5-535 against XBB.1 in golden Syrian hamsters. a Schematic of the in vivo mAb efficacy test experiment in hamsters. b Viral load quantification of XBB.1 in golden hamster lung and nasal turbinate with or without treatments. Lung and nasal turbinate samples were harvested on day 4 post SARS-CoV-2 Omicron XBB.1 challenge and homogenized for plaque assay titration (n = 6). c Immunofluorescence images of the whole section of infected hamster lungs with or without treatments. SARS-CoV-2 N protein was identified with a rabbit anti-SARS-CoV-2-N immune serum (green). Nuclei was identified with DAPI stain (blue). Scale bars in c represented 1000 μm for fourfold magnifications of the objective with tenfold magnification at the eyepiece. d Representative H&E images of infected hamster lungs with or without treatments. A representative region of (1) bronchiole epithelium and (2) alveolar space was shown. Scale bars in d represented 500 or 100 μm for fourfold or 20-fold magnifications of the objective, respectively, with ×10 magnification at the eyepiece. The experiments in b, d were repeated two times independently with similar results, respectively. Data…