Rapid discovery of diverse neutralizing SARS-CoV-2 antibodies from large-scale synthetic phage libraries

Abstract

Coronavirus disease 2019 (COVID-19) is an evolving global public health crisis in need of therapeutic options. Passive immunization of monoclonal antibodies (mAbs) represents a promising therapeutic strategy capable of conferring immediate protection from SARS-CoV-2 infection. Herein, we describe the discovery and characterization of neutralizing SARS-CoV-2 IgG and VHH antibodies from four large-scale phage libraries. Each library was constructed synthetically with shuffled complementarity-determining region loops from natural llama and human antibody repertoires. While most candidates targeted the receptor-binding domain of the S1 subunit of SARS-CoV-2 spike protein, we also identified a neutralizing IgG candidate that binds a unique epitope on the N-terminal domain. A select number of antibodies retained binding to SARS-CoV-2 variants Alpha, Beta, Gamma, Kappa and Delta. Overall, our data show that synthetic phage libraries can rapidly yield SARS-CoV-2 S1 antibodies with therapeutically desirable features, including high affinity, unique binding sites, and potent neutralizing activity in vitro , and a capacity to limit disease in vivo .

A synthetic bispecific antibody capable of neutralizing SARS-CoV-2 Delta and Omicron

“…SPR experiments performed using variant S1 monomers showed the same patterns, although apparent binding affinities were in the nanomolar range (Figure 2a). These data agree with previous biophysical and functional characterizations of TB202-03 28 (CoVIC-094 in Hastie et al 29 ) that showed reduced activity of TB202-03 against S proteins bearing the L452R mutation (namely, Delta and Epsilon).…”

“…In stark contrast to antibodies that target a highly conserved Sarbecovirus epitope (e.g., sotrovimab), TB202-03 binds toward the outer edge of the receptorbinding motif of SARS-CoV-2 (the RBD-4 community in Hastie et al 29 ), does not bind SARS-CoV S1, and competes with ACE2 for SARS-CoV-2 S binding. 28 The antiviral activity of RBT-0813 may be explained by the fact that none of the key residues in the TB202-03 epitopenamely, N450, I472, and F490 28 -are mutated in Omicron.…”

“…This bispecific antibody links together two humanized VHH antibodies -TB202-03 and TB339-031 -with constant heavy chain 2 (CH2) and 3 (CH3) Fc domains (Figure 1). TB202-03, which was discovered by panning the TB202 VHH library 28 against the S1 monomer of the SARS-CoV-2 WA1 strain, has been shown to effectively neutralize pseudoviruses encoding the Alpha, Beta, and Gamma S proteins, but not those encoding L452R-bearing S protein variants such as Delta and Epsilon (B.1.429). TB339-031 was discovered by panning the TB201 VHH library 28 against the Beta S1 (subdomain of S) and was found to bind and neutralize L452R-bearing S protein variants.…”

“…TB202-03, which was discovered by panning the TB202 VHH library 28 against the S1 monomer of the SARS-CoV-2 WA1 strain, has been shown to effectively neutralize pseudoviruses encoding the Alpha, Beta, and Gamma S proteins, but not those encoding L452R-bearing S protein variants such as Delta and Epsilon (B.1.429). TB339-031 was discovered by panning the TB201 VHH library 28 against the Beta S1 (subdomain of S) and was found to bind and neutralize L452R-bearing S protein variants. Here, we describe the biophysical and functional characterization of the bispecific RBT-0813 antibody, focusing on its binding and neutralization of Delta and Omicron variants.…”

Passive Immunotherapy Against SARS-CoV-2: From Plasma-Based Therapy to Single Potent Antibodies in the Race to Stay Ahead of the Variants

“…These camelid heavy chain variable domains, can be expressed as a single domain, typically referred to as a “nanobody,” with a molecular weight of about 12 kDa, roughly 1/12th that of the full-length IgG, and 1/80th the size of the IgM isotype described above. Other typical sources of single domain antibodies are camelid antibody libraries [ 339 ] and libraries of engineered human single domain antibodies [ 340 ].…”