The SARS-CoV-2 receptor-binding domain elicits a potent neutralizing response without antibody-dependent enhancement
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Abstract
The SARS-coronavirus 2 (SARS-CoV-2) spike (S) protein mediates entry of SARS-CoV-2 into cells expressing the angiotensin-converting enzyme 2 (ACE2). The S protein engages ACE2 through its receptor-binding domain (RBD), an independently folded 197-amino acid fragment of the 1273-amino acid S-protein protomer. Antibodies to the RBD domain of SARS-CoV (SARS-CoV-1), a closely related coronavirus which emerged in 2002-2003, have been shown to potently neutralize SARS-CoV-1 S-protein-mediated entry, and the presence of anti-RBD antibodies correlates with neutralization in SARS-CoV-2 convalescent sera. Here we show that immunization with the SARS-CoV-2 RBD elicits a robust neutralizing antibody response in rodents, comparable to 100 µg/ml of ACE2-Ig, a potent SARS-CoV-2 entry inhibitor. Importantly, anti-sera from immunized animals did not mediate antibody-dependent enhancement (ADE) of S-protein-mediated entry under conditions in which Zika virus ADE was readily observed. These data suggest that an RBD-based vaccine for SARS-CoV-2 could be safe and effective.
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SciScore for 10.1101/2020.04.10.036418: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement IACUC: Immunizations and sera collection: All rats used in these studies were handled and maintained in accordance with NIH guidelines and approved by Institutional Animal Care and Use Committee (IACUC) of Scripps Research (Protocol 18-025). Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable Female Sprague Dawley rats were immunized with incremental increasing doses of antigen over seven days starting at day 0, and boosted with a similar regimen at day 30. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources hACE2 expression was confirmed by SARS1-PV and SARS2-PV entry assays and by … SciScore for 10.1101/2020.04.10.036418: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement IACUC: Immunizations and sera collection: All rats used in these studies were handled and maintained in accordance with NIH guidelines and approved by Institutional Animal Care and Use Committee (IACUC) of Scripps Research (Protocol 18-025). Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable Female Sprague Dawley rats were immunized with incremental increasing doses of antigen over seven days starting at day 0, and boosted with a similar regimen at day 30. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources hACE2 expression was confirmed by SARS1-PV and SARS2-PV entry assays and by immunofluorescence staining using mouse monoclonal antibody recognizing c-Myc. c-Mycsuggested: NoneAfter washing, cells were stained with anti-rabbit IgG-Alexa647 antibody for 45 minutes at 4°C, and mean fluorescence intensities were measured for each well by flow cytometry. anti-rabbit IgG-Alexa647suggested: NoneMeasurement of antibody-dependent enhancement: The ability of anti-SARS-CoV-2 RBD immune plasma to mediate antibody-dependent enhancement (ADE) was measured using HEK293T cells or HEK293T cells stably expressing human ACE2 (293T-ACE2 cells), transfected using the calcium phosphate transfection method to express the rat ortholog of FcγRI (CD64). anti-SARS-CoV-2suggested: NoneACE2suggested: NoneCD64suggested: NoneExperimental Models: Cell Lines Sentences Resources 293T-hACE2 cells were selected and maintained with medium containing puromycin (Sigma). 293T-hACE2suggested: NoneProtein Production: Expi293 cells (Thermo-Fisher) were transiently transfected using FectoPRO (Polyplus) with plasmids encoding SARS-CoV2 RBD with a human or rabbit-Fc fusion or a C-terminal C-tag (-EPEA). Expi293suggested: RRID:CVCL_D615)One hour later, 104 ACE2-239T cells were added along with DEAE-Dextran (final concentration 5 µg/ml), and media was exchanged 6 hours later with fresh media without rat sera. ACE2-239Tsuggested: NoneMeasurement of antibody-dependent enhancement: The ability of anti-SARS-CoV-2 RBD immune plasma to mediate antibody-dependent enhancement (ADE) was measured using HEK293T cells or HEK293T cells stably expressing human ACE2 (293T-ACE2 cells), transfected using the calcium phosphate transfection method to express the rat ortholog of FcγRI (CD64). HEK293Tsuggested: NoneThe human monocytic cell line K562 (ATCC CCL-243), which endogenously expresses FcγRII, was also used for ADE assays. K562suggested: ATCC Cat# CCL-243, RRID:CVCL_0004)Experimental Models: Organisms/Strains Sentences Resources Female Sprague Dawley rats were immunized with incremental increasing doses of antigen over seven days starting at day 0, and boosted with a similar regimen at day 30. Sprague Dawleysuggested: NoneSoftware and Algorithms Sentences Resources Statistical analyses: The statistical significance of differences between preimmune and immune sera, or ACE2-Ig, in their abilities to neutralize, bind the S protein, and mediate ADE was analyzed by two-way ANOVA calculated using GraphPad Prism 8.0. GraphPad Prismsuggested: (GraphPad Prism, RRID:SCR_002798)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 found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).
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.
- No funding statement was detected.
- No protocol registration statement was detected.
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