A conserved immunogenic and vulnerable site on the coronavirus spike protein delineated by cross-reactive monoclonal antibodies

The coronavirus spike glycoprotein, located on the virion surface, is the key mediator of cell entry and the focus for development of protective antibodies and vaccines. Structural studies show exposed sites on the spike trimer that might be targeted by antibodies with cross-species specificity. Here we isolated two human monoclonal antibodies from immunized humanized mice that display a remarkable cross-reactivity against distinct spike proteins of betacoronaviruses including SARS-CoV, SARS-CoV-2, MERS-CoV and the endemic human coronavirus HCoV-OC43. Both cross-reactive antibodies target the stem helix in the spike S2 fusion subunit which, in the prefusion conformation of trimeric spike, forms a surface exposed membrane-proximal helical bundle. Both antibodies block MERS-CoV infection in cells and provide protection to mice from lethal MERS-CoV challenge in prophylactic and/or therapeutic models. Our work highlights an immunogenic and vulnerable site on the betacoronavirus spike protein enabling elicitation of antibodies with unusual binding breadth.

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For passive immunization and protection tests in mice, each group includes five to eight mice. The sample size was sufficient for a good statistical analysis based on previous experience (PMID: 30938227 No data were excluded from the analyses.
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20-30-week old female K18 TghDpp4 mice expressing the receptor for the human MERS-CoV and 16-week old female Balb-C mice were used to test the in vivo prophylactic and/or therapeutic efficacy of antibodies against MERS-CoV and SARS-CoV infection, respectively. Transgenic H2L2 mice from Harbour company that encode the human immunoglobulin repertoire were used for immunization at 8 weeks of age to develop fully human antibodies. Both female and male H2L2 mice were equally used . Male mice nature research | reporting summary April 2020 Experiments of concern Does the work involve any of these experiments of concern: No Yes Confirm that both raw and final processed data have been deposited in a public database such as GEO.
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HEK-293T cells were seeded at a density of 2.5×10E5 cells per ml in a T75 flask. After reaching 70~80% confluency, cells were transfected with an expression plasmid C-terminally fused to the GFP -using Lipofectamine 2000 (Invitrogen). Two days post transfection, cells were dissociated by cell dissociation solution (Sigma-aldrich, Merck KGaA; cat. no. C5914). Cells were then fixed by incubation with 2% paraformaldehyde in PBS for 20 min at room temperature and subject to antibody staining.