Antibody Cocktail Exhibits Broad Neutralization against SARS-CoV-2 and SARS-CoV-2 variants

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has precipitated multiple variants resistant to therapeutic antibodies. In this study, 12 high-affinity antibodies were generated from convalescent donors in early outbreaks using immune antibody phage display libraries. Of them, two RBD-binding antibodies (F61 and H121) showed high affinity neutralization against SARS-CoV-2, whereas three S2-target antibodies failed to neutralize SARS-CoV-2. Following structure analysis, F61 identified a linear epitope located in residues G446 - S494, which overlapped with angiotensin-converting enzyme 2 (ACE2) binding sites, while H121 recognized a conformational epitope located on the side face of RBD, outside from ACE2 binding domain. Hence the cocktail of the two antibodies achieved better performance of neutralization to SARS-CoV-2. Importantly, F61 and H121 exhibited efficient neutralizing activity against variants B.1.1.7 and B.1.351, those showed immune escape. Efficient neutralization of F61 and H121 against multiple mutations within RBD revealed a broad neutralizing activity against SARS-CoV-2 variants, which mitigated the risk of viral escape. Our findings defined the basis of therapeutic cocktails of F61 and H121 with broad neutralization and delivered a guideline for the current and future vaccine design, therapeutic antibody development, and antigen diagnosis of SARS-CoV-2 and its novel variants.

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Epitope-directed antibody selection by site-specific photocrosslinking

Science Advances ◽

10.1126/sciadv.aaz7825 ◽

2020 ◽

Vol 6 (14) ◽

pp. eaaz7825

Author(s):

Longxin Chen ◽

Chaoyang Zhu ◽

Hui Guo ◽

Runting Li ◽

Limeng Zhang ◽

...

Keyword(s):

Phage Display ◽

Conformational Epitope ◽

Phage Display Library ◽

Target Antigen ◽

Phage Display Libraries ◽

Antibody Phage ◽

Antibody Phage Display ◽

Antigen Epitope ◽

A Site ◽

Noncanonical Amino Acid

Currently, there are no methods available offering solutions to select and identify antibodies binding to a specific conformational epitope of an antigen. Here, we developed a method to allow epitope-directed antibody selection from a phage display library by photocrosslinking bound antibodies to a site that specifically incorporates a noncanonical amino acid, p-benzoyl-l-phenylalanine (pBpa), on the target antigen epitope. By one or two rounds of panning against antibody phage display libraries, those hits that covalently bind to the proximity site of pBpa on specific epitopes of target antigens after ultraviolet irradiation are enriched and selected. This method was applied to specific epitopes on human interleukin-1β and complement 5a. In both cases, more than one-third of hits identified bind to the target epitopes, demonstrating the feasibility and versatility of this method.

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Extremely potent monoclonal antibodies neutralize Omicron and other SARS-CoV-2 variants

10.1101/2022.01.12.22269023 ◽

2022 ◽

Author(s):

Zhaochun Chen ◽

Peng Zhang ◽

Yumiko Matsuoka ◽

Yaroslav Tsybovsky ◽

Kamille West ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Neutralizing Antibodies ◽

Spike Protein ◽

Structural Basis ◽

Hamster Model ◽

Golden Syrian Hamster ◽

Phage Display Libraries ◽

Antibody Phage ◽

Antibody Phage Display ◽

Early Isolate

The ongoing coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has triggered a devastating global health, social and economic crisis. The RNA nature and broad circulation of this virus facilitate the accumulation of mutations, leading to the continuous emergence of variants of concern with increased transmissibility or pathogenicity1. This poses a major challenge to the effectiveness of current vaccines and therapeutic antibodies1,2. Thus, there is an urgent need for effective therapeutic and preventive measures with a broad spectrum of action, especially against variants with an unparalleled number of mutations such as the recently emerged Omicron variant, which is rapidly spreading across the globe3. Here, we used combinatorial antibody phage-display libraries from convalescent COVID-19 patients to generate monoclonal antibodies against the receptor-binding domain of the SARS-CoV-2 spike protein with ultrapotent neutralizing activity. One such antibody, NE12, neutralizes an early isolate, the WA-1 strain, as well as the Alpha and Delta variants with half-maximal inhibitory concentrations at picomolar level. A second antibody, NA8, has an unusual breadth of neutralization, with picomolar activity against both the Beta and Omicron variants. The prophylactic and therapeutic efficacy of NE12 and NA8 was confirmed in preclinical studies in the golden Syrian hamster model. Analysis by cryo-EM illustrated the structural basis for the neutralization properties of NE12 and NA8. Potent and broadly neutralizing antibodies against conserved regions of the SARS-CoV-2 spike protein may play a key role against future variants of concern that evade immune control.

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Hevein-specific recombinant IgE antibodies from human single-chain antibody phage display libraries

Journal of Immunological Methods ◽

10.1016/s0022-1759(03)00070-x ◽

2003 ◽

Vol 278 (1-2) ◽

pp. 271-281 ◽

Cited By ~ 19

Author(s):

Marja-Leena Laukkanen ◽

Soili Mäkinen-Kiljunen ◽

Kirsi Isoherranen ◽

Tari Haahtela ◽

Hans Söderlund ◽

...

Keyword(s):

Phage Display ◽

Single Chain ◽

Single Chain Antibody ◽

Ige Antibodies ◽

Phage Display Libraries ◽

Antibody Phage ◽

Antibody Phage Display

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Modular Construction of Large Non-Immune Human Antibody Phage-Display Libraries from Variable Heavy and Light Chain Gene Cassettes

Methods in Molecular Biology - Phage Display ◽

10.1007/978-1-4939-7447-4_4 ◽

2017 ◽

pp. 61-82 ◽

Cited By ~ 1

Author(s):

Nam-Kyung Lee ◽

Scott Bidlingmaier ◽

Yang Su ◽

Bin Liu

Keyword(s):

Phage Display ◽

Light Chain ◽

Chain Gene ◽

Human Antibody ◽

Modular Construction ◽

Light Chain Gene ◽

Phage Display Libraries ◽

Antibody Phage ◽

Variable Heavy ◽

Antibody Phage Display

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Engineering human ACE2 to optimize binding to the spike protein of SARS coronavirus 2

Science ◽

10.1126/science.abc0870 ◽

2020 ◽

Vol 369 (6508) ◽

pp. 1261-1265 ◽

Cited By ~ 45

Author(s):

Kui K. Chan ◽

Danielle Dorosky ◽

Preeti Sharma ◽

Shawn A. Abbasi ◽

John M. Dye ◽

...

Keyword(s):

Host Cells ◽

Viral Escape ◽

Spike Protein ◽

Converting Enzyme ◽

Angiotensin Converting Enzyme 2 ◽

High Affinity ◽

Decoy Receptors ◽

Catalytically Active ◽

Interaction Surface

The spike (S) protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) binds angiotensin-converting enzyme 2 (ACE2) on host cells to initiate entry, and soluble ACE2 is a therapeutic candidate that neutralizes infection by acting as a decoy. By using deep mutagenesis, mutations in ACE2 that increase S binding are found across the interaction surface, in the asparagine 90–glycosylation motif and at buried sites. The mutational landscape provides a blueprint for understanding the specificity of the interaction between ACE2 and S and for engineering high-affinity decoy receptors. Combining mutations gives ACE2 variants with affinities that rival those of monoclonal antibodies. A stable dimeric variant shows potent SARS-CoV-2 and -1 neutralization in vitro. The engineered receptor is catalytically active, and its close similarity with the native receptor may limit the potential for viral escape.

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Selection of high affinity human neutralizing antibodies to VEGFR2 from a large antibody phage display library for antiangiogenesis therapy

International Journal of Cancer ◽

10.1002/ijc.1634 ◽

2001 ◽

Vol 97 (3) ◽

pp. 393-399 ◽

Cited By ~ 90

Author(s):

Dan Lu ◽

Xenia Jimenez ◽

Haifan Zhang ◽

Peter Bohlen ◽

Larry Witte ◽

...

Keyword(s):

Phage Display ◽

Neutralizing Antibodies ◽

Phage Display Library ◽

High Affinity ◽

Antibody Phage ◽

Antibody Phage Display ◽

Antiangiogenesis Therapy ◽

Selection Of

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Neutralizing Human Fab Fragments against Measles Virus Recovered by Phage Display

Journal of Virology ◽

10.1128/jvi.76.1.251-258.2002 ◽

2002 ◽

Vol 76 (1) ◽

pp. 251-258 ◽

Cited By ~ 25

Author(s):

Cristina de Carvalho Nicacio ◽

R. Anthony Williamson ◽

Paul W. H. I. Parren ◽

Åke Lundkvist ◽

Dennis R. Burton ◽

...

Keyword(s):

Phage Display ◽

Measles Virus ◽

Protein Band ◽

Enzyme Linked Immunosorbent Assay ◽

Neutralization Capacity ◽

N Protein ◽

Fab Fragments ◽

Phage Display Libraries ◽

Antibody Phage ◽

Antibody Phage Display

ABSTRACT Five human recombinant Fab fragments (Fabs) specific for measles virus (MV) proteins were isolated from three antibody phage display libraries generated from RNAs derived from bone marrow or splenic lymphocytes from three MV-immune individuals. All Fabs reacted in an enzyme-linked immunosorbent assay with MV antigens. In radioimmunoprecipitation assays two of the Fabs, MV12 and MT14, precipitated an ⊘80-kDa protein band corresponding to the hemagglutinin (H) protein from MV-infected Vero cell cultures, while two other Fabs, MT64 and GL29, precipitated an ⊘60-kDa protein corresponding the nucleocapsid (N) protein. In competition studies with MV fusion, H- and N protein-specific monoclonal antibodies (MAbs), the H-specific Fabs predominantly blocked the binding of H-specific MAbs, while the N-specific Fabs blocked MAbs to N. In addition, N-specific Fabs bound to denatured MV N protein in Western blotting. The specificity of the fifth Fab, MV4, could not be determined. By plaque reduction assays, three of the five Fabs, MV4, MV12, and MT14, exhibited neutralizing activity (80% cutoff) against MV (LEC-KI strain) at concentrations ranging between ≈2 and 7 μg ml−1. Neutralization capacity against MV strains Edmonston and Schwarz was also detected, albeit at somewhat higher Fab concentrations. In conclusion, three neutralizing Fabs were isolated, two of them reactive against the H glycoprotein of MV and another reactive against an undefined epitope. This is the first study in which MV-neutralizing human recombinant Fab antibodies have been isolated from phage display libraries.

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