Hetero-bivalent nanobodies provide broad-spectrum protection against SARS-CoV-2 variants of concern including Omicron
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Abstract
SARS-CoV-2 variants with adaptive mutations have continued to emerge, causing fresh waves of infection even amongst vaccinated population. The development of broad-spectrum antivirals is thus urgently needed. We previously developed two hetero-bivalent nanobodies (Nbs), aRBD-2-5 and aRBD-2-7, with potent neutralization activity against the wild-type (WT) Wuhan isolated SARS-CoV-2, by fusing aRBD-2 with aRBD-5 and aRBD-7, respectively. Here, we resolved the crystal structures of these Nbs in complex with the receptor-binding domain (RBD) of the spike protein, and found that aRBD-2 contacts with highly-conserved RBD residues and retains binding to the RBD of the Alpha, Beta, Gamma, Delta, Delta plus, Kappa, Lambda, Omicron BA.1, and BA.2 variants. In contrast, aRBD-5 and aRBD-7 bind to less-conserved RBD epitopes non-overlapping with the epitope of aRBD-2, and do not show apparent binding to the RBD of some variants. However, when fused with aRBD-2, they effectively enhance the overall binding affinity. Consistently, aRBD-2-5-Fc and aRBD-2-7-Fc potently neutralized all of the tested authentic or pseudotyped viruses, including WT, Alpha, Beta, Gamma, Delta, and Omicron BA.1, BA.1.1 and BA.2. Furthermore, aRBD-2-5-Fc provided prophylactic protection against the WT and mouse-adapted SARS-CoV-2 in mice, and conferred protection against the Omicron BA.1 variant in hamsters prophylactically and therapeutically, indicating that aRBD-2-5-Fc could potentially benefit the prevention and treatment of COVID-19 caused by the emerging variants of concern. Our strategy provides new solutions in the development of broad-spectrum therapeutic antibodies for COVID-19.
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SciScore for 10.1101/2022.03.08.483381: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics not detected. Sex as a biological variable aRBD-2-5-Fc prophylactic protection in mice: Female K-18 hACE2 transgenic mice (The Jackson Laboratory) were administered with one dose of aRBD-2-5-Fc (10 mg/kg, i.p.) 24 hr before infected with 2 ×104 PFU of original SARS-CoV-2 (i.n.), and female A/J mice (The Jackson Laboratory) were administered with one dose of aRBD-2-5-Fc (1 mg/kg and 0.1 mg/kg, i.p.) 24 hr before infected with 1 ×105 PFU of mouse-adapted SARS-CoV-2 (intratracheally), body weight and death of the mice were monitored daily for 7 days post infection. Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
… SciScore for 10.1101/2022.03.08.483381: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics not detected. Sex as a biological variable aRBD-2-5-Fc prophylactic protection in mice: Female K-18 hACE2 transgenic mice (The Jackson Laboratory) were administered with one dose of aRBD-2-5-Fc (10 mg/kg, i.p.) 24 hr before infected with 2 ×104 PFU of original SARS-CoV-2 (i.n.), and female A/J mice (The Jackson Laboratory) were administered with one dose of aRBD-2-5-Fc (1 mg/kg and 0.1 mg/kg, i.p.) 24 hr before infected with 1 ×105 PFU of mouse-adapted SARS-CoV-2 (intratracheally), body weight and death of the mice were monitored daily for 7 days post infection. Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources After four washes with PBST (PBS containing 0.1% tween-20), bound Nb-Fc were detected with a HRP-conjugated anti-IgG1 Fc antibody (Sino Biological). anti-IgG1suggested: NoneAfter 2days of infection, the cells were fixed with 4% paraformaldehyde, permeabilized with 0.1% Triton-100, blocked with DMEM containing 10% fetal bovine serum, and stained with a rabbit monoclonal antibody against SARS-CoV-2 NP (GeneTex, GTX635679) and an Alexa Fluor 488-conjugated goat anti-mouse secondary antibody (Thermo Fisher Scientific) SARS-CoV-2 NPsuggested: NoneGTX635679suggested: (GeneTex Cat# GTX635679, RRID:AB_2888553)anti-mousesuggested: NoneExperimental Models: Cell Lines Sentences Resources The recombinant vectors were transiently transfected into HEK293F cells with polyethyleneimine (Polyscience). HEK293Fsuggested: RRID:CVCL_6642)Briefly, Vero E6 cells were seeded overnight in 24-well culture plates at 1.5 ×105 per well. Vero E6suggested: NoneExperimental Models: Organisms/Strains Sentences Resources aRBD-2-5-Fc prophylactic protection in mice: Female K-18 hACE2 transgenic mice (The Jackson Laboratory) were administered with one dose of aRBD-2-5-Fc (10 mg/kg, i.p.) 24 hr before infected with 2 ×104 PFU of original SARS-CoV-2 (i.n.), and female A/J mice (The Jackson Laboratory) were administered with one dose of aRBD-2-5-Fc (1 mg/kg and 0.1 mg/kg, i.p.) 24 hr before infected with 1 ×105 PFU of mouse-adapted SARS-CoV-2 (intratracheally), body weight and death of the mice were monitored daily for 7 days post infection. K-18 hACE2suggested: RRID:IMSR_GPT:T037657)A/Jsuggested: NoneRecombinant DNA Sentences Resources Briefly, the coding sequences for SARS-CoV-2 RBD (amino acids 321 to 591) of original WT, Alpha (N501Y), Beta (K417N, E484K, N501Y), Gamma (K417T, E484K, N501Y), Delta (L452R, T478K), Delta plus (K417N, L452R, T478K), Kappa (L452R, E484Q) and Lambda (L452R, F490S) variant were cloned into pTT5 vector, which the C terminal contain a TEV cleavage site and a human IgG1 Fc. pTT5suggested: RRID:Addgene_52326)Software and Algorithms Sentences Resources The data was analyzed using GraphPad Prism software. GraphPad Prismsuggested: (GraphPad Prism, RRID:SCR_002798)The total number of cells indicated by the nuclei staining and the infected cells indicated by the NP staining were quantified with the cellular analysis module of the Gen5 software (BioTek). Gen5suggested: (Gen5, RRID:SCR_017317)Subsequent models building and refinement were achieved using COOT and Phenix 62. COOTsuggested: (Coot, RRID:SCR_014222)All structural figures were prepared by PyMOL. PyMOLsuggested: (PyMOL, RRID:SCR_000305)Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
Results from scite Reference Check: We found no unreliable references.
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