Isolation of a human monoclonal antibody specific for the receptor binding domain of SARS-CoV-2 using a competitive phage biopanning strategy

Abstract

The infection of the novel coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused more than 200 000 deaths, but no vaccine or therapeutic monoclonal antibody is currently available. SARS-CoV-2 relies on its spike protein, in particular the receptor-binding domain (RBD), to bind human cell receptor angiotensin-converting enzyme 2 (ACE2) for viral entry, and thus targeting RBD holds the promise for preventing SARS-CoV-2 infection. In this work, a competitive biopanning strategy of a phage display antibody library was applied to screen blocking antibodies against RBD. High-affinity antibodies were enriched after the first round using a standard panning process in which RBD-His was immobilized as a bait. At the next two rounds, immobilized ACE2-Fc and free RBD-His were mixed with the enriched phage antibodies. Antibodies binding to RBD at epitopes different from ACE2-binding site were captured by the immobilized ACE2-Fc, forming a "sandwich" complex. Only antibodies competed with ACE2 can bind to the free RBD-His in the supernatant and be subsequently separated by the nickel-nitrilotriacetic acid magnetic beads. rRBD-15 from the competitive biopanning of our synthetic antibody library, Lib AB1, was produced as the full-length IgG1 format. It was proved to competitively block the binding of RBD to ACE2 and potently inhibit SARS-CoV-2 pseudovirus infection with IC 50 values of 12 nM. Nevertheless, rRBD-16 from the standard biopanning can only bind to RBD in vitro, but not have the blocking or neutralization activity. Our strategy can efficiently isolate the blocking antibodies of RBD, and it would speed up the discovery of neutralizing antibodies against SARS-CoV-2. Statement of Significance:The authors developed a competitive phage screening strategy to isolate a high-affinity human antibody that binds the SARS-CoV-2 receptor binding domain and inhibits its interaction with human angiotensin-converting enzyme 2.

Development of humanized tri-specific nanobodies with potent neutralization for SARS-CoV-2

“…The COVID-19 pandemic has caused widespread health and social issues around the globe, and requires therapeutics that can effectively stop and prevent the infection of SARS-CoV-2. Several monoclonal antibodies against SARS-CoV-2 have been suggested and being tested as anti-viral therapies, either as individual agents or combination therapies; however, this is the first study that introduces and demonstrates the efficacy of multi-specific antibodies against SARS-CoV-2 to our knowledge [10][11][12][13][14][15]19,30 .…”

Receptor-binding domain-specific human neutralizing monoclonal antibodies against SARS-CoV and SARS-CoV-2

“…In a pseudovirus neutralization assay, rRBD-15 showed potent neutralization activity against SARS-CoV-2 pseudovirus infection with an IC 50 of 12.2 nM. 37 …”

“… 36 rRBD-15 IgG Human A synthetic human Fab antibody library RBD PsV neutralization: IC 50 = 12.2 nM. 37 VHH-72-Fc HCAb llama Animal immunization and sequencing RBD PsV neutralization: IC 50 ~ 0.2 µg/ml. 25 311mab–31B5 311mab–32D4 IgG Human B cells of convalescent patients RBD PsV neutralization: IC 50 = 33.8 ng/ml.…”

Potent mouse monoclonal antibodies that block SARS-CoV-2 infection

“…Isolation of multiple human neutralizing monoclonal antibodies against SARS-CoV-2 has been reported (Cao et al, 2020; Chen et al, 2020; Chi et al, 2020; Hassan et al, 2020; Ju et al, 2020; Liu et al, 2020; Pinto et al, 2020; Robbiani et al, 2020; Rogers et al, 2020; Shi et al, 2020; Wan et al, 2020; Wang et al, 2020; Wu et al, 2020; Zeng et al, 2020; Zost et al, 2020). These antibodies can avoid the potential risks of human-anti-mouse antibody responses and other side effects (Hansel, Kropshofer, Singer, Mitchell, & George, 2010).…”