RT Journal Article SR Electronic T1 Peptide-antibody Fusions Engineered by Phage Display Exhibit Ultrapotent and Broad Neutralization of SARS-CoV-2 Variants JF bioRxiv FD Cold Spring Harbor Laboratory SP 2021.11.29.470362 DO 10.1101/2021.11.29.470362 A1 Jonathan M. Labriola A1 Shane Miersch A1 Gang Chen A1 Chao Chen A1 Alevtina Pavlenco A1 Francesca Pisanu A1 Francesca Caccuri A1 Alberto Zani A1 Nitin Sharma A1 Annie Feng A1 Daisy W. Leung A1 Arnaldo Caruso A1 Gaya K. Amarasinghe A1 Sachdev S. Sidhu YR 2021 UL http://biorxiv.org/content/early/2021/11/30/2021.11.29.470362.abstract AB The COVID-19 pandemic has been exacerbated by the emergence of variants of concern (VoCs). Many VoC mutations are found in the viral spike protein (S-protein), and are thus implicated in host infection and response to therapeutics. Bivalent neutralizing antibodies (nAbs) targeting the S-protein receptor-binding domain (RBD) are promising therapeutics for COVID-19, but are limited due to low potency and vulnerability to RBD mutations found in VoCs. To address these issues, we used naïve phage-displayed peptide libraries to isolate and optimize 16-residue peptides that bind to the RBD or the N-terminal domain (NTD) of the S-protein. We fused these peptides to the N-terminus of a moderate affinity nAb to generate tetravalent peptide-IgG fusions, and showed that both classes of peptides were able to improve affinities for the S-protein trimer by >100-fold (apparent KD < 1 pM). Critically, cell-based infection assays with a panel of six SARS-CoV-2 variants demonstrate that an RBD-binding peptide was able to enhance the neutralization potency of a high-affinity nAb >100-fold. Moreover, this peptide-IgG was able to neutralize variants that were resistant to the same nAb in the bivalent IgG format. To show that this approach is general, we fused the same peptide to a clinically approved nAb drug, and showed that it rescued neutralization against a resistant variant. Taken together, these results establish minimal peptide fusions as a modular means to greatly enhance affinities, potencies, and breadth of coverage of nAbs as therapeutics for SARS-CoV-2.Competing Interest StatementThe authors have declared no competing interest.SARS-CoV-2severe acute respiratory syndrome coronavirus-2S-proteinSpike proteinRBDreceptor binding domainNTDN-terminal domainECDextracellular domainVoCvariants of concernACE2angiotensinogen converting enzyme 2R1-R4RBD binding peptide 1-4N1-N4NTD binding peptide 1-4nAbsneutralizing antibodiesHCIgG heavy chainLCIgG light chain33neutralizing antibody 1503333-7neutralizing antibody 15033-7NAVneutravidin