Isolation and Characterization of Mouse Monoclonal Antibodies That Neutralize SARS-CoV-2 and Its Variants of Concern Alpha, Beta, Gamma and Delta by Binding Conformational Epitopes of Glycosylated RBD With High Potency
| Autor: | Maria Laura De Angelis, Antonella Marchi, Sabrina Mariotti, Valeria Esposito, Paola Di Bonito, Chiara Acchioni, Maria Franca Pirillo, Giulietta Venturi, Maria Vincenza Chiantore, Paul F. McKay, Fabio Tosini, Alessandra Gallinaro, Donatella R.M. Negri, Francesco Marino, Marco Sgarbanti, Andrea Canitano, Fabio Magurano, Roberto Nisini, Antonio Di Virgilio, Antonio Capocefalo, Paola Bucci, Andrea Cara, Angelo Iacobino, Melissa Baggieri, Silvia Sandini, Martina Borghi, Zuleika Michelini, Raffaela Teloni |
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| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
Glycosylation
Immunoprecipitation medicine.drug_class diagnosis Immunology Antibodies Viral Monoclonal antibody Epitope law.invention Epitopes Western blot Neutralization Tests law Cell Line Tumor Chlorocebus aethiops medicine Animals Humans Immunology and Allergy Vero Cells Original Research chemistry.chemical_classification SARS-COV-2 variants Mice Inbred BALB C therapy biology medicine.diagnostic_test SARS-CoV-2 pathogenesis epitopes expression Antibodies Monoclonal RC581-607 Antibodies Neutralizing Virology COVID-19 Drug Treatment HEK293 Cells chemistry Spike Glycoprotein Coronavirus Vero cell Recombinant DNA biology.protein Female Angiotensin-Converting Enzyme 2 Binding Sites Antibody Antibody neutralizing monoclonal antibodies Immunologic diseases. Allergy Glycoprotein |
| Zdroj: | Frontiers in Immunology, Vol 12 (2021) Frontiers in Immunology |
| ISSN: | 1664-3224 |
| DOI: | 10.3389/fimmu.2021.750386/full |
| Popis: | Antibodies targeting Receptor Binding Domain (RBD) of SARS-CoV-2 have been suggested to account for the majority of neutralizing activity in COVID-19 convalescent sera and several neutralizing antibodies (nAbs) have been isolated, characterized and proposed as emergency therapeutics in the form of monoclonal antibodies (mAbs). However, SARS-CoV-2 variants are rapidly spreading worldwide from the sites of initial identification. The variants of concern (VOC) B.1.1.7 (Alpha), B.1.351 (Beta), P.1 (Gamma) and B.1.167.2 (Delta) showed mutations in the SARS-CoV-2 spike protein potentially able to cause escape from nAb responses with a consequent reduction of efficacy of vaccines and mAbs-based therapy. We produced the recombinant RBD (rRBD) of SARS-CoV-2 spike glycoprotein from the Wuhan-Hu 1 reference sequence in a mammalian system, for mice immunization to isolate new mAbs with neutralizing activity. Here we describe four mAbs that were able to bind the rRBD in Enzyme-Linked Immunosorbent Assay and the transmembrane full-length spike protein expressed in HEK293T cells by flow cytometry assay. Moreover, the mAbs recognized the RBD in supernatants of SARS-CoV-2 infected VERO E6 cells by Western Blot under non-reducing condition or in supernatants of cells infected with lentivirus pseudotyped for spike protein, by immunoprecipitation assay. Three out of four mAbs lost their binding efficiency to completely N-deglycosylated rRBD and none was able to bind the same recombinant protein expressed in Escherichia coli, suggesting that the epitopes recognized by three mAbs are generated by the conformational structure of the glycosylated native protein. Of particular relevance, three mAbs were able to inhibit Wuhan SARS-CoV-2 infection of VERO E6 cells in a plaque-reduction neutralization test and the Wuhan SARS-CoV-2 as well as the Alpha, Beta, Gamma and Delta VOC in a pseudoviruses-based neutralization test. These mAbs represent important additional tools for diagnosis and therapy of COVID-19 and may contribute to the understanding of the functional structure of SARS-CoV-2 RBD. |
| Databáze: | OpenAIRE |
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