Epitope mapping of severe acute respiratory syndrome coronavirus 2 neutralizing receptor binding domain-specific monoclonal antibodies

Faezeh Maghsood; Mohammad Mehdi Amiri; Amir-Hassan Zarnani; Vahid Salimi; Gholam Ali Kardar; Jalal Khoshnoodi; Maryam Mobini; Hengameh Ahmadi Zare; Abbas Ghaderi; Mahmood Jeddi-Tehrani; Sylvie Schmidt; Géraldine Laumond; Christiane Moog; Fazel Shokri

doi:10.3389/fmed.2022.973036

Epitope mapping of severe acute respiratory syndrome coronavirus 2 neutralizing receptor binding domain-specific monoclonal antibodies

Front Med (Lausanne). 2022 Sep 6:9:973036. doi: 10.3389/fmed.2022.973036. eCollection 2022.

Affiliations

¹ Department of Immunology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran.
² Department of Virology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran.
³ Immunology, Asthma and Allergy Research Institute, Children's Medical Center, Tehran University of Medical Sciences, Tehran, Iran.
⁴ Shiraz Institute for Cancer Research, School of Medicine, Shiraz University of Medical Science, Shiraz, Iran.
⁵ Monoclonal Antibody Research Center, Avicenna Research Institute, Academic Center for Education, Culture and Research (ACECR), Tehran, Iran.
⁶ Laboratoire d'ImmunoRhumatologie Moléculaire, Institut National de la Santé et de la Recherche Médicale (INSERM) UMR_S 1109, Institut Thématique Interdisciplinaire (ITI) de Médecine de Précision de Strasbourg, Transplantex NG, Faculté de Médecine, Fédération Hospitalo-Universitaire OMICARE, Fédération de Médecine Translationnelle de Strasbourg (FMTS), Strasbourg, France.

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the outbreak led to the coronavirus disease 2019 (COVID-19) pandemic. Receptor binding domain (RBD) of spike (S) protein of SARS-CoV-2 is considered as a major target for immunotherapy and vaccine design. Here, we generated and characterized a panel of anti-RBD monoclonal antibodies (MAbs) isolated from eukaryotic recombinant RBD-immunized mice by hybridoma technology. Epitope mapping was performed using a panel of 20-mer overlapping peptides spanning the entire sequence of the RBD protein from wild-type (WT) Wuhan strain by enzyme-linked immunosorbent assay (ELISA). Several hybridomas showed reactivity toward restricted RBD peptide pools by Pepscan analysis, with more focus on peptides encompassing aa 76-110 and 136-155. However, our MAbs with potent neutralizing activity which block SARS-CoV-2 spike pseudovirus as well as the WT virus entry into angiotensin-converting enzyme-2 (ACE2) expressing HEK293T cells showed no reactivity against these peptides. These findings, largely supported by the Western blotting results suggest that the neutralizing MAbs recognize mainly conformational epitopes. Moreover, our neutralizing MAbs recognized the variants of concern (VOC) currently in circulation, including alpha, beta, gamma, and delta by ELISA, and neutralized alpha and omicron variants at different levels by conventional virus neutralization test (CVNT). While the neutralization of MAbs to the alpha variant showed no substantial difference as compared with the WT virus, their neutralizing activity was lower on omicron variant, suggesting the refractory effect of mutations in emerging variants against this group of neutralizing MAbs. Also, the binding reactivity of our MAbs to delta variant showed a modest decline by ELISA, implying that our MAbs are insensitive to the substitutions in the RBD of delta variant. Our data provide important information for understanding the immunogenicity of RBD, and the potential application of the novel neutralizing MAbs for passive immunotherapy of SARS-CoV-2 infection.

Keywords: COVID-19; SARS-CoV-2; monoclonal antibodies; neutralization; omicron.