59 results on '"Czudnochowski N"'
Search Results
2. S2P6 Fab fragment bound to the SARS-CoV/SARS-CoV-2 spike stem helix peptide
- Author
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Snell, G., primary, Czudnochowski, N., additional, Croll, T.I., additional, Nix, J.C., additional, Corti, D., additional, Cameroni, E., additional, Pinto, D., additional, Beltramello, M., additional, Sauer, M.M., additional, and Veesler, D., additional
- Published
- 2021
- Full Text
- View/download PDF
3. SARS-CoV-2 spike receptor-binding domain (RBD) in complex with S2X35 Fab and S309 Fab
- Author
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Snell, G., primary, Czudnochowski, N., additional, Hernandez, P., additional, Nix, J.C., additional, Croll, T.I., additional, Corti, D., additional, Cameroni, E., additional, Pinto, D., additional, and Beltramello, M., additional
- Published
- 2021
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4. SARS-CoV-2 spike receptor-binding domain (RBD) in complex with S2E12 Fab, S309 Fab, and S304 Fab
- Author
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Snell, G., primary, Czudnochowski, N., additional, Croll, T.I., additional, Nix, J.C., additional, Corti, D., additional, Cameroni, E., additional, Pinto, D., additional, and Beltramello, M., additional
- Published
- 2021
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5. Broad betacoronavirus neutralization by a stem helix–specific human antibody
- Author
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Pinto, D, Sauer, MM, Czudnochowski, N, Low, JS, Alejandra Tortorici, M, Housley, MP, Noack, J, Walls, AC, Bowen, JE, Guarino, B, Rosen, LE, di Iulio, J, Jerak, J, Kaiser, H, Islam, S, Jaconi, S, Sprugasci, N, Culap, K, Abdelnabi, R, Foo, C, Coelmont, L, Bartha, I, Bianchi, S, Silacci-Fregni, C, Bassi, J, Marzi, R, Vetti, E, Cassotta, A, Ceschi, A, Ferrari, P ; https://orcid.org/0000-0002-6094-7592, Cippà, PE, Giannini, O, Ceruti, S, Garzoni, C, Riva, A, Benigni, F, Cameroni, E, Piccoli, L, Pizzuto, MS, Smithey, M, Hong, D, Telenti, A, Lempp, FA, Neyts, J, Havenar-Daughton, C, Lanzavecchia, A, Sallusto, F, Snell, G, Virgin, HW, Beltramello, M, Corti, D, Veesler, D, Pinto, D, Sauer, MM, Czudnochowski, N, Low, JS, Alejandra Tortorici, M, Housley, MP, Noack, J, Walls, AC, Bowen, JE, Guarino, B, Rosen, LE, di Iulio, J, Jerak, J, Kaiser, H, Islam, S, Jaconi, S, Sprugasci, N, Culap, K, Abdelnabi, R, Foo, C, Coelmont, L, Bartha, I, Bianchi, S, Silacci-Fregni, C, Bassi, J, Marzi, R, Vetti, E, Cassotta, A, Ceschi, A, Ferrari, P ; https://orcid.org/0000-0002-6094-7592, Cippà, PE, Giannini, O, Ceruti, S, Garzoni, C, Riva, A, Benigni, F, Cameroni, E, Piccoli, L, Pizzuto, MS, Smithey, M, Hong, D, Telenti, A, Lempp, FA, Neyts, J, Havenar-Daughton, C, Lanzavecchia, A, Sallusto, F, Snell, G, Virgin, HW, Beltramello, M, Corti, D, and Veesler, D
- Abstract
The spillovers of betacoronaviruses in humans and the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants highlight the need for broad coronavirus countermeasures. We describe five monoclonal antibodies (mAbs) cross-reacting with the stem helix of multiple betacoronavirus spike glycoproteins isolated from COVID-19 convalescent individuals. Using structural and functional studies, we show that the mAb with the greatest breadth (S2P6) neutralizes pseudotyped viruses from three different subgenera through the inhibition of membrane fusion, and we delineate the molecular basis for its cross-reactivity. S2P6 reduces viral burden in hamsters challenged with SARS-CoV-2 through viral neutralization and Fc-mediated effector functions. Stem helix antibodies are rare, oftentimes of narrow specificity, and can acquire neutralization breadth through somatic mutations. These data provide a framework for structure-guided design of pan-betacoronavirus vaccines eliciting broad protection.
- Published
- 2021
6. Circulating SARS-CoV-2 spike N439K variants maintain fitness while evading antibody-mediated immunity
- Author
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Thomson, EC, Rosen, LE, Shepherd, JG, Spreafico, R, da Silva Filipe, A, Wojcechowskyj, JA, Davis, C, Piccoli, L, Pascall, DJ, Dillen, J, Lytras, S, Czudnochowski, N, Shah, R, Meury, M, Jesudason, N, De Marco, A, Li, K, Bassi, J, O’Toole, A, Pinto, D, Colquhoun, RM, Culap, K, Jackson, B, Zatta, F, Rambaut, A, Jaconi, S, Sreenu, VB, Nix, J, Zhang, I, Jarrett, RF, Glass, WG, Beltramello, M, Nomikou, K, Pizzuto, M, Tong, L, Cameroni, E, Croll, TI, Johnson, N, Iulio, JD, Wickenhagen, A, Ceschi, A, Harbison, AM, Mair, D, Ferrari, P ; https://orcid.org/0000-0002-6094-7592, Smollett, K, Sallusto, F, Carmichael, S, Garzoni, C, Nichols, J, Galli, M, Hughes, J, Riva, A, Ho, A, Schiuma, M, Semple, MG, Openshaw, PJM, Fadda, E, Kenneth Baillie, J, Chodera, JD, Rihn, SJ, Lycett, SJ, Virgin, HW, Telenti, A, Corti, D, Robertson, DL, Snell, G, Connor, TR, Loman, NJ, Robson, SC, Golubchik, T, Estee Torok, M, Hamilton, WL, Bonsall, D, Awan, AR, Corden, S, Goodfellow, I, Smith, DL, Curran, MD, Parmar, S, Parker, MD, Moore, C, Fairley, DJ, Loose, MW, Watkins, J, Bull, M, Nicholls, S, Aanensen, DM, Glaysher, S, Bashton, M, Pacchiarini, N, Underwood, AP, de Silva, TI, Wang, D, Andersson, M, Chauhan, AJ, de Cesare, M, Ludden, C, Mahungu, TW, Dewar, R, Thomson, EC, Rosen, LE, Shepherd, JG, Spreafico, R, da Silva Filipe, A, Wojcechowskyj, JA, Davis, C, Piccoli, L, Pascall, DJ, Dillen, J, Lytras, S, Czudnochowski, N, Shah, R, Meury, M, Jesudason, N, De Marco, A, Li, K, Bassi, J, O’Toole, A, Pinto, D, Colquhoun, RM, Culap, K, Jackson, B, Zatta, F, Rambaut, A, Jaconi, S, Sreenu, VB, Nix, J, Zhang, I, Jarrett, RF, Glass, WG, Beltramello, M, Nomikou, K, Pizzuto, M, Tong, L, Cameroni, E, Croll, TI, Johnson, N, Iulio, JD, Wickenhagen, A, Ceschi, A, Harbison, AM, Mair, D, Ferrari, P ; https://orcid.org/0000-0002-6094-7592, Smollett, K, Sallusto, F, Carmichael, S, Garzoni, C, Nichols, J, Galli, M, Hughes, J, Riva, A, Ho, A, Schiuma, M, Semple, MG, Openshaw, PJM, Fadda, E, Kenneth Baillie, J, Chodera, JD, Rihn, SJ, Lycett, SJ, Virgin, HW, Telenti, A, Corti, D, Robertson, DL, Snell, G, Connor, TR, Loman, NJ, Robson, SC, Golubchik, T, Estee Torok, M, Hamilton, WL, Bonsall, D, Awan, AR, Corden, S, Goodfellow, I, Smith, DL, Curran, MD, Parmar, S, Parker, MD, Moore, C, Fairley, DJ, Loose, MW, Watkins, J, Bull, M, Nicholls, S, Aanensen, DM, Glaysher, S, Bashton, M, Pacchiarini, N, Underwood, AP, de Silva, TI, Wang, D, Andersson, M, Chauhan, AJ, de Cesare, M, Ludden, C, Mahungu, TW, and Dewar, R
- Abstract
SARS-CoV-2 can mutate and evade immunity, with consequences for efficacy of emerging vaccines and antibody therapeutics. Here, we demonstrate that the immunodominant SARS-CoV-2 spike (S) receptor binding motif (RBM) is a highly variable region of S and provide epidemiological, clinical, and molecular characterization of a prevalent, sentinel RBM mutation, N439K. We demonstrate N439K S protein has enhanced binding affinity to the hACE2 receptor, and N439K viruses have similar in vitro replication fitness and cause infections with similar clinical outcomes as compared to wild type. We show the N439K mutation confers resistance against several neutralizing monoclonal antibodies, including one authorized for emergency use by the US Food and Drug Administration (FDA), and reduces the activity of some polyclonal sera from persons recovered from infection. Immune evasion mutations that maintain virulence and fitness such as N439K can emerge within SARS-CoV-2 S, highlighting the need for ongoing molecular surveillance to guide development and usage of vaccines and therapeutics. Epidemiological, clinical, molecular, and structural characterization of the N439K mutation in the SARS-CoV-2 spike receptor binding motif demonstrates that it results in similar viral fitness compared to wild-type while conferring resistance against some neutralizing monoclonal antibodies and reducing the activity of some polyclonal antibody responses.
- Published
- 2021
7. Antibodies to the SARS-CoV-2 receptor-binding domain that maximize breadth and resistance to viral escape
- Author
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Snell, G., primary, Czudnochowski, N., additional, Croll, T.I., additional, Nix, J.C., additional, Corti, D., additional, Cameroni, E., additional, Pinto, D., additional, and Beltramello, M., additional
- Published
- 2021
- Full Text
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8. Circulating SARS-CoV-2 spike N439K variants maintain fitness while evading antibody-mediated immunity
- Author
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Snell, G., primary, Czudnochowski, N., additional, Dillen, J., additional, Nix, J.C., additional, Croll, T.I., additional, and Corti, D., additional
- Published
- 2021
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9. Mapping neutralizing and immunodominant sites on the SARS-CoV-2 spike receptor-binding domain by structure-guided high-resolution serology
- Author
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Snell, G., primary, Czudnochowski, N., additional, Rosen, L.E., additional, Nix, J.C., additional, Corti, D., additional, Veesler, D., additional, Park, Y.J., additional, Walls, A.C., additional, Tortorici, M.A., additional, Cameroni, E., additional, Pinto, D., additional, and Beltramello, M., additional
- Published
- 2020
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10. An ultra-potent human neutralizing antibody locks the SARS-CoV-2 spike in the closed conformation
- Author
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Snell, G., primary, Czudnochowski, N., additional, and Ng, C., additional
- Published
- 2020
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11. Mapping Neutralizing and Immunodominant Sites on the SARS-CoV-2 Spike Receptor-Binding Domain by Structure-Guided High-Resolution Serology
- Author
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Piccoli, L, Park, YJ, Tortorici, MA, Czudnochowski, N, Walls, AC, Beltramello, M, Silacci-Fregni, C, Pinto, D, Rosen, LE, Bowen, JE, Acton, OJ, Jaconi, S, Guarino, B, Minola, A, Zatta, F, Sprugasci, N, Bassi, J, Peter, A, De Marco, A, Nix, JC, Mele, F, Jovic, S, Rodriguez, BF, Gupta, SV, Jin, F, Piumatti, G, Lo Presti, G, Pellanda, AF, Biggiogero, M, Tarkowski, M, Pizzuto, MS, Cameroni, E, Havenar-Daughton, C, Smithey, M, Hong, D, Lepori, V, Albanese, E, Ceschi, A, Bernasconi, E, Elzi, L, Ferrari, P ; https://orcid.org/0000-0002-6094-7592, Garzoni, C, Riva, A, Snell, G, Sallusto, F, Fink, K, Virgin, HW, Lanzavecchia, A, Corti, D, Veesler, D, Piccoli, L, Park, YJ, Tortorici, MA, Czudnochowski, N, Walls, AC, Beltramello, M, Silacci-Fregni, C, Pinto, D, Rosen, LE, Bowen, JE, Acton, OJ, Jaconi, S, Guarino, B, Minola, A, Zatta, F, Sprugasci, N, Bassi, J, Peter, A, De Marco, A, Nix, JC, Mele, F, Jovic, S, Rodriguez, BF, Gupta, SV, Jin, F, Piumatti, G, Lo Presti, G, Pellanda, AF, Biggiogero, M, Tarkowski, M, Pizzuto, MS, Cameroni, E, Havenar-Daughton, C, Smithey, M, Hong, D, Lepori, V, Albanese, E, Ceschi, A, Bernasconi, E, Elzi, L, Ferrari, P ; https://orcid.org/0000-0002-6094-7592, Garzoni, C, Riva, A, Snell, G, Sallusto, F, Fink, K, Virgin, HW, Lanzavecchia, A, Corti, D, and Veesler, D
- Abstract
Analysis of the specificity and kinetics of neutralizing antibodies (nAbs) elicited by SARS-CoV-2 infection is crucial for understanding immune protection and identifying targets for vaccine design. In a cohort of 647 SARS-CoV-2-infected subjects, we found that both the magnitude of Ab responses to SARS-CoV-2 spike (S) and nucleoprotein and nAb titers correlate with clinical scores. The receptor-binding domain (RBD) is immunodominant and the target of 90% of the neutralizing activity present in SARS-CoV-2 immune sera. Whereas overall RBD-specific serum IgG titers waned with a half-life of 49 days, nAb titers and avidity increased over time for some individuals, consistent with affinity maturation. We structurally defined an RBD antigenic map and serologically quantified serum Abs specific for distinct RBD epitopes leading to the identification of two major receptor-binding motif antigenic sites. Our results explain the immunodominance of the receptor-binding motif and will guide the design of COVID-19 vaccines and therapeutics.
- Published
- 2020
12. The structure of the C-terminus of virulence protein IncE from Chlamydia trachomatis bound to Mus musculus SNX5-PX domain
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Rosenberg, O., primary and Czudnochowski, N., additional
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- 2017
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13. Structure of the product complex of tRNA m1A58 methyltransferase with tRNA3Lys as substrate
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Finer-Moore, J., primary, Czudnochowski, N., additional, O'Connell III, J.D., additional, Wang, A.L., additional, and Stroud, R.M., additional
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- 2015
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14. Structure of an asymmetric tetramer of human tRNA m1A58 methyltransferase in a complex with SAH and tRNA3Lys
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Finer-Moore, J., primary, Czudnochowski, N., additional, O'Connell III, J.D., additional, Wang, A.L., additional, and Stroud, R.M., additional
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- 2015
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15. Crystal structure of human m1A58 methyltransferase in a complex with tRNA3Lys and SAH
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Finer-Moore, J., primary, Czudnochowski, N., additional, O'Connell III, J.D., additional, Wang, A.L., additional, and Stroud, R.M., additional
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- 2015
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16. Crystal structure of the catalytic domain of RluB in complex with a 21-nucleotide RNA substrate
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Czudnochowski, N., primary, Finer-Moore, J.S., additional, and Stroud, R.M., additional
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- 2013
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17. Crystal structure of the catalytic domain of RluB
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Czudnochowski, N., primary, Finer-Moore, J.S., additional, and Stroud, R.M., additional
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- 2013
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18. Crystal structure of the catalytic domain of human Pus1 with MES in the active site
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Czudnochowski, N., primary, Finer-Moore, J.S., additional, and Stroud, R.M., additional
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- 2013
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19. Crystal structure of the catalytic domain of human Pus1
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Czudnochowski, N., primary, Finer-Moore, J.S., additional, and Stroud, R.M., additional
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- 2013
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20. Tat competes with HEXIM1 to increase the active pool of P-TEFb for HIV-1 transcription
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Barboric, M., primary, Yik, J. H. N., additional, Czudnochowski, N., additional, Yang, Z., additional, Chen, R., additional, Contreras, X., additional, Geyer, M., additional, Matija Peterlin, B., additional, and Zhou, Q., additional
- Published
- 2007
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21. Circulating SARS-CoV-2 spike N439K variants maintain fitness while evading antibody-mediated immunity
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Emma Thomson, Rosen L, Shepherd J, Spreafico R, da Silva Filipe A, Wojcechowskyj J, Davis C, Piccoli L, Pascall D, Dillen J, Lytras S, Czudnochowski N, and Snell G
22. Insights into the activation of transcription elongation by lentiviruses: structure of the Cyclin T1-Tat-TAR RNA complex
- Author
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Schulte Antje, Vollmuth Friederike, Czudnochowski Nadine, Anand Kanchan, and Geyer Matthias
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Immunologic diseases. Allergy ,RC581-607 - Published
- 2009
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23. A potent pan-sarbecovirus neutralizing antibody resilient to epitope diversification.
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Rosen LE, Tortorici MA, De Marco A, Pinto D, Foreman WB, Taylor AL, Park YJ, Bohan D, Rietz T, Errico JM, Hauser K, Dang HV, Chartron JW, Giurdanella M, Cusumano G, Saliba C, Zatta F, Sprouse KR, Addetia A, Zepeda SK, Brown J, Lee J, Dellota E Jr, Rajesh A, Noack J, Tao Q, DaCosta Y, Tsu B, Acosta R, Subramanian S, de Melo GD, Kergoat L, Zhang I, Liu Z, Guarino B, Schmid MA, Schnell G, Miller JL, Lempp FA, Czudnochowski N, Cameroni E, Whelan SPJ, Bourhy H, Purcell LA, Benigni F, di Iulio J, Pizzuto MS, Lanzavecchia A, Telenti A, Snell G, Corti D, Veesler D, and Starr TN
- Subjects
- Humans, Animals, Spike Glycoprotein, Coronavirus immunology, Spike Glycoprotein, Coronavirus chemistry, Spike Glycoprotein, Coronavirus metabolism, Spike Glycoprotein, Coronavirus genetics, Cross Reactions immunology, Chiroptera virology, Chiroptera immunology, COVID-19 immunology, COVID-19 virology, Angiotensin-Converting Enzyme 2 metabolism, Angiotensin-Converting Enzyme 2 chemistry, SARS-CoV-2 immunology, SARS-CoV-2 genetics, Epitopes immunology, Epitopes chemistry, Antibodies, Neutralizing immunology, Antibodies, Neutralizing chemistry, Antibodies, Monoclonal immunology, Antibodies, Monoclonal chemistry, Antibodies, Viral immunology, Antibodies, Viral chemistry
- Abstract
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) evolution has resulted in viral escape from clinically authorized monoclonal antibodies (mAbs), creating a need for mAbs that are resilient to epitope diversification. Broadly neutralizing coronavirus mAbs that are sufficiently potent for clinical development and retain activity despite viral evolution remain elusive. We identified a human mAb, designated VIR-7229, which targets the viral receptor-binding motif (RBM) with unprecedented cross-reactivity to all sarbecovirus clades, including non-ACE2-utilizing bat sarbecoviruses, while potently neutralizing SARS-CoV-2 variants since 2019, including the recent EG.5, BA.2.86, and JN.1. VIR-7229 tolerates extraordinary epitope variability, partly attributed to its high binding affinity, receptor molecular mimicry, and interactions with RBM backbone atoms. Consequently, VIR-7229 features a high barrier for selection of escape mutants, which are rare and associated with reduced viral fitness, underscoring its potential to be resilient to future viral evolution. VIR-7229 is a strong candidate to become a next-generation medicine., Competing Interests: Declaration of interests L.E.R., A.D.M., D.P., D.B., T.R., J.M.E., K.H., H.V.D., M.G., G.C., C.S., F.Z., E.D., A.R., J.N., Q.T., Y.D., B.T., R.A., S.S., B.G., M.A.S., G. Schnell, J.L.M., F.A.L., N.C., E.C., L.A.P., F.B., J.d.I., M.S.P., A.L., A.T., G. Snell, and D.C. are current or previous employees of Vir Biotechnology and may hold shares in Vir Biotechnology. L.E.R., A.D.M., D.P., E.C., F.B., M.S.P., G. Snell, and D.C. are currently listed as inventors on multiple patent applications that disclose the subject matter described in this paper. J.W.C. is an employee and shareholder of ProtaBody. J.W.C. and ProtaBody have received funding from Vir Biotechnology related to the work described in this paper. I.Z., Z.L., S.P.J.W., G.D.d.M., L.K., H.B., and T.N.S. have received funding through sponsored research awards to their respective institutions from Vir Biotechnology related to the work described in this paper. I.Z. is a current employee of Bristol Myers Squibb. L.A.P. is a former employee and shareholder of Regeneron Pharmaceuticals and is a member of the Scientific Advisory Board AI-driven structure-enabled antiviral platform (ASAP). Regeneron provided no funding for this work. L.A.P. is a current employee of Third Rock Ventures. D.V. is named as inventor on patents for coronavirus vaccines filed by the University of Washington. The lab of T.N.S. has received sponsored research agreements unrelated to the present work from Aerium Therapeutics, Inc. and Invivyd, Inc., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)
- Published
- 2024
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24. Neutralization, effector function and immune imprinting of Omicron variants.
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Addetia A, Piccoli L, Case JB, Park YJ, Beltramello M, Guarino B, Dang H, de Melo GD, Pinto D, Sprouse K, Scheaffer SM, Bassi J, Silacci-Fregni C, Muoio F, Dini M, Vincenzetti L, Acosta R, Johnson D, Subramanian S, Saliba C, Giurdanella M, Lombardo G, Leoni G, Culap K, McAlister C, Rajesh A, Dellota E Jr, Zhou J, Farhat N, Bohan D, Noack J, Chen A, Lempp FA, Quispe J, Kergoat L, Larrous F, Cameroni E, Whitener B, Giannini O, Cippà P, Ceschi A, Ferrari P, Franzetti-Pellanda A, Biggiogero M, Garzoni C, Zappi S, Bernasconi L, Kim MJ, Rosen LE, Schnell G, Czudnochowski N, Benigni F, Franko N, Logue JK, Yoshiyama C, Stewart C, Chu H, Bourhy H, Schmid MA, Purcell LA, Snell G, Lanzavecchia A, Diamond MS, Corti D, and Veesler D
- Subjects
- Animals, Cricetinae, Humans, Mice, Angiotensin-Converting Enzyme 2 immunology, Angiotensin-Converting Enzyme 2 metabolism, Antibodies, Monoclonal chemistry, Antibodies, Monoclonal immunology, Cross Reactions, Immune Evasion, Membrane Fusion, Neutralization Tests, Mutation, Memory B Cells immunology, COVID-19 Vaccines immunology, Antibodies, Neutralizing chemistry, Antibodies, Neutralizing immunology, COVID-19 immunology, COVID-19 prevention & control, COVID-19 virology, SARS-CoV-2 classification, SARS-CoV-2 genetics, SARS-CoV-2 immunology
- Abstract
Currently circulating SARS-CoV-2 variants have acquired convergent mutations at hot spots in the receptor-binding domain
1 (RBD) of the spike protein. The effects of these mutations on viral infection and transmission and the efficacy of vaccines and therapies remains poorly understood. Here we demonstrate that recently emerged BQ.1.1 and XBB.1.5 variants bind host ACE2 with high affinity and promote membrane fusion more efficiently than earlier Omicron variants. Structures of the BQ.1.1, XBB.1 and BN.1 RBDs bound to the fragment antigen-binding region of the S309 antibody (the parent antibody for sotrovimab) and human ACE2 explain the preservation of antibody binding through conformational selection, altered ACE2 recognition and immune evasion. We show that sotrovimab binds avidly to all Omicron variants, promotes Fc-dependent effector functions and protects mice challenged with BQ.1.1 and hamsters challenged with XBB.1.5. Vaccine-elicited human plasma antibodies cross-react with and trigger effector functions against current Omicron variants, despite a reduced neutralizing activity, suggesting a mechanism of protection against disease, exemplified by S309. Cross-reactive RBD-directed human memory B cells remained dominant even after two exposures to Omicron spikes, underscoring the role of persistent immune imprinting., (© 2023. The Author(s).)- Published
- 2023
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25. Author Correction: A pan-influenza antibody inhibiting neuraminidase via receptor mimicry.
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Momont C, Dang HV, Zatta F, Hauser K, Wang C, di Iulio J, Minola A, Czudnochowski N, De Marco A, Branch K, Donermeyer D, Vyas S, Chen A, Ferri E, Guarino B, Powell AE, Spreafico R, Yim SS, Balce DR, Bartha I, Meury M, Croll TI, Belnap DM, Schmid MA, Schaiff WT, Miller JL, Cameroni E, Telenti A, Virgin HW, Rosen LE, Purcell LA, Lanzavecchia A, Snell G, Corti D, and Pizzuto MS
- Published
- 2023
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26. A pan-influenza antibody inhibiting neuraminidase via receptor mimicry.
- Author
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Momont C, Dang HV, Zatta F, Hauser K, Wang C, di Iulio J, Minola A, Czudnochowski N, De Marco A, Branch K, Donermeyer D, Vyas S, Chen A, Ferri E, Guarino B, Powell AE, Spreafico R, Yim SS, Balce DR, Bartha I, Meury M, Croll TI, Belnap DM, Schmid MA, Schaiff WT, Miller JL, Cameroni E, Telenti A, Virgin HW, Rosen LE, Purcell LA, Lanzavecchia A, Snell G, Corti D, and Pizzuto MS
- Subjects
- Animals, Humans, Mice, Antibodies, Monoclonal chemistry, Antibodies, Monoclonal immunology, Antibodies, Monoclonal therapeutic use, Arginine chemistry, Catalytic Domain, Hemagglutinins, Viral immunology, Influenza A Virus, H3N2 Subtype enzymology, Influenza A Virus, H3N2 Subtype immunology, Orthomyxoviridae Infections immunology, Orthomyxoviridae Infections prevention & control, Seasons, Sialic Acids chemistry, Antibodies, Viral chemistry, Antibodies, Viral immunology, Antibodies, Viral therapeutic use, Antibody Specificity immunology, Influenza A virus classification, Influenza A virus enzymology, Influenza A virus immunology, Influenza B virus classification, Influenza B virus enzymology, Influenza B virus immunology, Influenza Vaccines chemistry, Influenza Vaccines immunology, Influenza Vaccines therapeutic use, Influenza, Human immunology, Influenza, Human prevention & control, Neuraminidase antagonists & inhibitors, Neuraminidase chemistry, Neuraminidase immunology, Molecular Mimicry
- Abstract
Rapidly evolving influenza A viruses (IAVs) and influenza B viruses (IBVs) are major causes of recurrent lower respiratory tract infections. Current influenza vaccines elicit antibodies predominantly to the highly variable head region of haemagglutinin and their effectiveness is limited by viral drift
1 and suboptimal immune responses2 . Here we describe a neuraminidase-targeting monoclonal antibody, FNI9, that potently inhibits the enzymatic activity of all group 1 and group 2 IAVs, as well as Victoria/2/87-like, Yamagata/16/88-like and ancestral IBVs. FNI9 broadly neutralizes seasonal IAVs and IBVs, including the immune-evading H3N2 strains bearing an N-glycan at position 245, and shows synergistic activity when combined with anti-haemagglutinin stem-directed antibodies. Structural analysis reveals that D107 in the FNI9 heavy chain complementarity-determinant region 3 mimics the interaction of the sialic acid carboxyl group with the three highly conserved arginine residues (R118, R292 and R371) of the neuraminidase catalytic site. FNI9 demonstrates potent prophylactic activity against lethal IAV and IBV infections in mice. The unprecedented breadth and potency of the FNI9 monoclonal antibody supports its development for the prevention of influenza illness by seasonal and pandemic viruses., (© 2023. The Author(s).)- Published
- 2023
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27. Therapeutic and vaccine-induced cross-reactive antibodies with effector function against emerging Omicron variants.
- Author
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Addetia A, Piccoli L, Case JB, Park YJ, Beltramello M, Guarino B, Dang H, Pinto D, Scheaffer S, Sprouse K, Bassi J, Silacci-Fregni C, Muoio F, Dini M, Vincenzetti L, Acosta R, Johnson D, Subramanian S, Saliba C, Giurdanella M, Lombardo G, Leoni G, Culap K, McAlister C, Rajesh A, Dellota E, Zhou J, Farhat N, Bohan D, Noack J, Lempp FA, Cameroni E, Whitener B, Giannini O, Ceschi A, Ferrari P, Franzetti-Pellanda A, Biggiogero M, Garzoni C, Zappi S, Bernasconi L, Kim MJ, Schnell G, Czudnochowski N, Franko N, Logue JK, Yoshiyama C, Stewart C, Chu H, Schmid MA, Purcell LA, Snell G, Lanzavecchia A, Diamond M, Corti D, and Veesler D
- Abstract
Currently circulating SARS-CoV-2 variants acquired convergent mutations at receptor-binding domain (RBD) hot spots. Their impact on viral infection, transmission, and efficacy of vaccines and therapeutics remains poorly understood. Here, we demonstrate that recently emerged BQ.1.1. and XBB.1 variants bind ACE2 with high affinity and promote membrane fusion more efficiently than earlier Omicron variants. Structures of the BQ.1.1 and XBB.1 RBDs bound to human ACE2 and S309 Fab (sotrovimab parent) explain the altered ACE2 recognition and preserved antibody binding through conformational selection. We show that sotrovimab binds avidly to all Omicron variants, promotes Fc-dependent effector functions and protects mice challenged with BQ.1.1, the variant displaying the greatest loss of neutralization. Moreover, in several donors vaccine-elicited plasma antibodies cross-react with and trigger effector functions against Omicron variants despite reduced neutralizing activity. Cross-reactive RBD-directed human memory B cells remained dominant even after two exposures to Omicron spikes, underscoring persistent immune imprinting. Our findings suggest that this previously overlooked class of cross-reactive antibodies, exemplified by S309, may contribute to protection against disease caused by emerging variants through elicitation of effector functions.
- Published
- 2023
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28. Maturation of SARS-CoV-2 Spike-specific memory B cells drives resilience to viral escape.
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Marzi R, Bassi J, Silacci-Fregni C, Bartha I, Muoio F, Culap K, Sprugasci N, Lombardo G, Saliba C, Cameroni E, Cassotta A, Low JS, Walls AC, McCallum M, Tortorici MA, Bowen JE, Dellota EA Jr, Dillen JR, Czudnochowski N, Pertusini L, Terrot T, Lepori V, Tarkowski M, Riva A, Biggiogero M, Franzetti-Pellanda A, Garzoni C, Ferrari P, Ceschi A, Giannini O, Havenar-Daughton C, Telenti A, Arvin A, Virgin HW, Sallusto F, Veesler D, Lanzavecchia A, Corti D, and Piccoli L
- Abstract
Memory B cells (MBCs) generate rapid antibody responses upon secondary encounter with a pathogen. Here, we investigated the kinetics, avidity, and cross-reactivity of serum antibodies and MBCs in 155 SARS-CoV-2 infected and vaccinated individuals over a 16-month time frame. SARS-CoV-2-specific MBCs and serum antibodies reached steady-state titers with comparable kinetics in infected and vaccinated individuals. Whereas MBCs of infected individuals targeted both prefusion and postfusion Spike (S), most vaccine-elicited MBCs were specific for prefusion S, consistent with the use of prefusion-stabilized S in mRNA vaccines. Furthermore, a large fraction of MBCs recognizing postfusion S cross-reacted with human betacoronaviruses. The avidity of MBC-derived and serum antibodies increased over time resulting in enhanced resilience to viral escape by SARS-CoV-2 variants, including Omicron BA.1 and BA.2 sublineages, albeit only partially for BA.4 and BA.5 sublineages. Overall, the maturation of high-affinity and broadly reactive MBCs provides the basis for effective recall responses to future SARS-CoV-2 variants., Competing Interests: R.M., J.B., C.S.-F., I.B., F.M., K.C., N.S., G.L., C.S., E.C., E.A.D.J., J.R.D., N.C., C.H.-D., A.T., A.A., H.W.V., A.L., D.C., and L.Pi. are or were employees of Vir Biotechnology Inc. and may hold shares in Vir Biotechnology Inc. C.G. is an external scientific consultant to Humabs BioMed SA. The other authors declare no competing interests., (© 2022 The Authors.)
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- 2023
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29. Workshop-based learning and networking: a scalable model for research capacity strengthening in low- and middle-income countries.
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Perier C, Nasinghe E, Charles I, Ssetaba LJ, Ahyong V, Bangs D, Beatty PR, Czudnochowski N, Diallo A, Dugan E, Fabius JM, Fong Baker H, Gardner J, Isaacs S, Joanah B, Kalantar K, Kateete D, Knight M, Krasilnikov M, Krogan NJ, Langelier C, Lee E, Li LM, Licht D, Lien K, Lyons Z, Mboowa G, Mwebaza I, Mwesigwa S, Nalwadda G, Nichols R, Penaranda ME, Petnic S, Phelps M, Popper SJ, Rape M, Reingold A, Robbins R, Rosenberg OS, Savage DF, Schildhauer S, Settles ML, Sserwadda I, Stanley S, Tato CM, Tsitsiklis A, Van Dis E, Vanaerschot M, Vinden J, Cox JS, Joloba ML, and Schaletzky J
- Subjects
- Capacity Building, Humans, Poverty, Students, Universities, Developing Countries, Global Health
- Abstract
Science education and research have the potential to drive profound change in low- and middle-income countries (LMICs) through encouraging innovation, attracting industry, and creating job opportunities. However, in LMICs, research capacity is often limited, and acquisition of funding and access to state-of-the-art technologies is challenging. The Alliance for Global Health and Science (the Alliance) was founded as a partnership between the University of California, Berkeley (USA) and Makerere University (Uganda), with the goal of strengthening Makerere University's capacity for bioscience research. The flagship program of the Alliance partnership is the MU/UCB Biosciences Training Program, an in-country, hands-on workshop model that trains a large number of students from Makerere University in infectious disease and molecular biology research. This approach nucleates training of larger and more diverse groups of students, development of mentoring and bi-directional research partnerships, and support of the local economy. Here, we describe the project, its conception, implementation, challenges, and outcomes of bioscience research workshops. We aim to provide a blueprint for workshop implementation, and create a valuable resource for bioscience research capacity strengthening in LMICs.
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- 2022
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30. Maturation of SARS-CoV-2 Spike-specific memory B cells drives resilience to viral escape.
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Marzi R, Bassi J, Silacci-Fregni C, Bartha I, Muoio F, Culap K, Sprugasci N, Lombardo G, Saliba C, Cameroni E, Cassotta A, Low JS, Walls AC, McCallum M, Tortorici MA, Bowen JE, Dellota EA Jr, Dillen JR, Czudnochowski N, Pertusini L, Terrot T, Lepori V, Tarkowski M, Riva A, Biggiogero M, Pellanda AF, Garzoni C, Ferrari P, Ceschi A, Giannini O, Havenar-Daughton C, Telenti A, Arvin A, Virgin HW, Sallusto F, Veesler D, Lanzavecchia A, Corti D, and Piccoli L
- Abstract
Memory B cells (MBCs) generate rapid antibody responses upon secondary encounter with a pathogen. Here, we investigated the kinetics, avidity and cross-reactivity of serum antibodies and MBCs in 155 SARS-CoV-2 infected and vaccinated individuals over a 16-month timeframe. SARS-CoV-2-specific MBCs and serum antibodies reached steady-state titers with comparable kinetics in infected and vaccinated individuals. Whereas MBCs of infected individuals targeted both pre- and postfusion Spike (S), most vaccine-elicited MBCs were specific for prefusion S, consistent with the use of prefusion-stabilized S in mRNA vaccines. Furthermore, a large fraction of MBCs recognizing postfusion S cross-reacted with human betacoronaviruses. The avidity of MBC-derived and serum antibodies increased over time resulting in enhanced resilience to viral escape by SARS-CoV-2 variants, including Omicron BA.1 and BA.2 sub-lineages, albeit only partially for BA.4 and BA.5 sublineages. Overall, the maturation of high-affinity and broadly-reactive MBCs provides the basis for effective recall responses to future SARS-CoV-2 variants.
- Published
- 2022
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31. Omicron spike function and neutralizing activity elicited by a comprehensive panel of vaccines.
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Bowen JE, Addetia A, Dang HV, Stewart C, Brown JT, Sharkey WK, Sprouse KR, Walls AC, Mazzitelli IG, Logue JK, Franko NM, Czudnochowski N, Powell AE, Dellota E Jr, Ahmed K, Ansari AS, Cameroni E, Gori A, Bandera A, Posavad CM, Dan JM, Zhang Z, Weiskopf D, Sette A, Crotty S, Iqbal NT, Corti D, Geffner J, Snell G, Grifantini R, Chu HY, and Veesler D
- Subjects
- Humans, Immunization, Secondary, Antibodies, Neutralizing blood, Antibodies, Neutralizing immunology, Antibodies, Viral blood, Antibodies, Viral immunology, COVID-19 blood, COVID-19 prevention & control, COVID-19 Vaccines immunology, SARS-CoV-2 immunology, Spike Glycoprotein, Coronavirus genetics, Spike Glycoprotein, Coronavirus immunology
- Abstract
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant of concern comprises several sublineages, with BA.2 and BA.2.12.1 having replaced the previously dominant BA.1 and with BA.4 and BA.5 increasing in prevalence worldwide. We show that the large number of Omicron sublineage spike mutations leads to enhanced angiotensin-converting enzyme 2 (ACE2) binding, reduced fusogenicity, and severe dampening of plasma neutralizing activity elicited by infection or seven clinical vaccines relative to the ancestral virus. Administration of a homologous or heterologous booster based on the Wuhan-Hu-1 spike sequence markedly increased neutralizing antibody titers and breadth against BA.1, BA.2, BA.2.12.1, BA.4, and BA.5 across all vaccines evaluated. Our data suggest that although Omicron sublineages evade polyclonal neutralizing antibody responses elicited by primary vaccine series, vaccine boosters may provide sufficient protection against Omicron-induced severe disease.
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- 2022
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32. Structural basis of SARS-CoV-2 Omicron immune evasion and receptor engagement.
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McCallum M, Czudnochowski N, Rosen LE, Zepeda SK, Bowen JE, Walls AC, Hauser K, Joshi A, Stewart C, Dillen JR, Powell AE, Croll TI, Nix J, Virgin HW, Corti D, Snell G, and Veesler D
- Subjects
- Amino Acid Substitution, Angiotensin-Converting Enzyme 2 metabolism, Antibodies, Monoclonal chemistry, Antibodies, Monoclonal immunology, Antibodies, Monoclonal metabolism, Antibodies, Viral immunology, Antibodies, Viral metabolism, Antigenic Drift and Shift, Broadly Neutralizing Antibodies chemistry, Broadly Neutralizing Antibodies immunology, Broadly Neutralizing Antibodies metabolism, Cryoelectron Microscopy, Crystallography, X-Ray, Humans, Models, Molecular, Mutation, Protein Binding, Protein Conformation, Protein Domains genetics, Protein Interaction Domains and Motifs genetics, Receptors, Coronavirus metabolism, SARS-CoV-2 genetics, SARS-CoV-2 physiology, Spike Glycoprotein, Coronavirus genetics, Spike Glycoprotein, Coronavirus immunology, Spike Glycoprotein, Coronavirus metabolism, Angiotensin-Converting Enzyme 2 chemistry, Antibodies, Viral chemistry, Immune Evasion, Receptors, Coronavirus chemistry, SARS-CoV-2 chemistry, SARS-CoV-2 immunology, Spike Glycoprotein, Coronavirus chemistry
- Abstract
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant of concern evades antibody-mediated immunity that comes from vaccination or infection with earlier variants due to accumulation of numerous spike mutations. To understand the Omicron antigenic shift, we determined cryo-electron microscopy and x-ray crystal structures of the spike protein and the receptor-binding domain bound to the broadly neutralizing sarbecovirus monoclonal antibody (mAb) S309 (the parent mAb of sotrovimab) and to the human ACE2 receptor. We provide a blueprint for understanding the marked reduction of binding of other therapeutic mAbs that leads to dampened neutralizing activity. Remodeling of interactions between the Omicron receptor-binding domain and human ACE2 likely explains the enhanced affinity for the host receptor relative to the ancestral virus.
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- 2022
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33. Poor neutralization and rapid decay of antibodies to SARS-CoV-2 variants in vaccinated dialysis patients.
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Bassi J, Giannini O, Silacci-Fregni C, Pertusini L, Hitz P, Terrot T, Franzosi Y, Muoio F, Saliba C, Meury M, Dellota EA Jr, Dillen JR, Hernandez P, Czudnochowski N, Cameroni E, Beria N, Ventresca M, Badellino A, Lavorato-Hadjeres S, Lecchi E, Bonora T, Mattiolo M, Trinci G, Garzoni D, Bonforte G, Forni-Ogna V, Giunzioni D, Berwert L, Gupta RK, Ferrari P, Ceschi A, Cippà P, Corti D, Lanzavecchia A, and Piccoli L
- Subjects
- Animals, Antibodies, Neutralizing blood, Antibody Affinity, CHO Cells, COVID-19 Vaccines immunology, Case-Control Studies, Cricetulus, Dose-Response Relationship, Immunologic, Follow-Up Studies, HEK293 Cells, Humans, Immunoglobulin G blood, Risk Factors, mRNA Vaccines immunology, Antibodies, Neutralizing immunology, Neutralization Tests, Renal Dialysis, SARS-CoV-2 immunology, Vaccination
- Abstract
Patients on dialysis are at risk of severe course of SARS-CoV-2 infection. Understanding the neutralizing activity and coverage of SARS-CoV-2 variants of vaccine-elicited antibodies is required to guide prophylactic and therapeutic COVID-19 interventions in this frail population. By analyzing plasma samples from 130 hemodialysis and 13 peritoneal dialysis patients after two doses of BNT162b2 or mRNA-1273 vaccines, we found that 35% of the patients had low-level or undetectable IgG antibodies to SARS-CoV-2 Spike (S). Neutralizing antibodies against the vaccine-matched SARS-CoV-2 and Delta variant were low or undetectable in 49% and 77% of patients, respectively, and were further reduced against other emerging variants. The fraction of non-responding patients was higher in SARS-CoV-2-naïve hemodialysis patients immunized with BNT162b2 (66%) than those immunized with mRNA-1273 (23%). The reduced neutralizing activity correlated with low antibody avidity. Patients followed up to 7 months after vaccination showed a rapid decay of the antibody response with an average 21- and 10-fold reduction of neutralizing antibodies to vaccine-matched SARS-CoV-2 and Delta variant, which increased the fraction of non-responders to 84% and 90%, respectively. These data indicate that dialysis patients should be prioritized for additional vaccination boosts. Nevertheless, their antibody response to SARS-CoV-2 must be continuously monitored to adopt the best prophylactic and therapeutic strategy., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: J.B., C.S.-F., F.M., C.S., M.Me., E.A.D.J., N.C., E.C., D.C. A.L., and L.Pi. are employees of Vir Biotechnology Inc. and may hold shares in Vir Biotechnology Inc. R.K.G. has received consulting fees from Johnson and Johnson and GlaxoSmithKline for educational activities. The other authors declare no competing interests. This does not alter our adherence to PLOS ONE policies on sharing data and materials.
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- 2022
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34. Broadly neutralizing antibodies overcome SARS-CoV-2 Omicron antigenic shift.
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Cameroni E, Bowen JE, Rosen LE, Saliba C, Zepeda SK, Culap K, Pinto D, VanBlargan LA, De Marco A, di Iulio J, Zatta F, Kaiser H, Noack J, Farhat N, Czudnochowski N, Havenar-Daughton C, Sprouse KR, Dillen JR, Powell AE, Chen A, Maher C, Yin L, Sun D, Soriaga L, Bassi J, Silacci-Fregni C, Gustafsson C, Franko NM, Logue J, Iqbal NT, Mazzitelli I, Geffner J, Grifantini R, Chu H, Gori A, Riva A, Giannini O, Ceschi A, Ferrari P, Cippà PE, Franzetti-Pellanda A, Garzoni C, Halfmann PJ, Kawaoka Y, Hebner C, Purcell LA, Piccoli L, Pizzuto MS, Walls AC, Diamond MS, Telenti A, Virgin HW, Lanzavecchia A, Snell G, Veesler D, and Corti D
- Subjects
- Angiotensin-Converting Enzyme 2 metabolism, Animals, Antibodies, Monoclonal therapeutic use, Antibodies, Monoclonal, Humanized immunology, Antibodies, Neutralizing immunology, Antibodies, Viral blood, Antigenic Drift and Shift genetics, COVID-19 Vaccines immunology, Cell Line, Convalescence, Epitopes, B-Lymphocyte immunology, Humans, Immune Evasion, Mice, SARS-CoV-2 chemistry, SARS-CoV-2 classification, SARS-CoV-2 genetics, Spike Glycoprotein, Coronavirus chemistry, Spike Glycoprotein, Coronavirus genetics, Spike Glycoprotein, Coronavirus immunology, Spike Glycoprotein, Coronavirus metabolism, Vesiculovirus genetics, Antibodies, Monoclonal immunology, Antibodies, Viral immunology, Antigenic Drift and Shift immunology, Broadly Neutralizing Antibodies immunology, Neutralization Tests, SARS-CoV-2 immunology
- Abstract
The recently emerged SARS-CoV-2 Omicron variant encodes 37 amino acid substitutions in the spike protein, 15 of which are in the receptor-binding domain (RBD), thereby raising concerns about the effectiveness of available vaccines and antibody-based therapeutics. Here we show that the Omicron RBD binds to human ACE2 with enhanced affinity, relative to the Wuhan-Hu-1 RBD, and binds to mouse ACE2. Marked reductions in neutralizing activity were observed against Omicron compared to the ancestral pseudovirus in plasma from convalescent individuals and from individuals who had been vaccinated against SARS-CoV-2, but this loss was less pronounced after a third dose of vaccine. Most monoclonal antibodies that are directed against the receptor-binding motif lost in vitro neutralizing activity against Omicron, with only 3 out of 29 monoclonal antibodies retaining unaltered potency, including the ACE2-mimicking S2K146 antibody
1 . Furthermore, a fraction of broadly neutralizing sarbecovirus monoclonal antibodies neutralized Omicron through recognition of antigenic sites outside the receptor-binding motif, including sotrovimab2 , S2X2593 and S2H974 . The magnitude of Omicron-mediated immune evasion marks a major antigenic shift in SARS-CoV-2. Broadly neutralizing monoclonal antibodies that recognize RBD epitopes that are conserved among SARS-CoV-2 variants and other sarbecoviruses may prove key to controlling the ongoing pandemic and future zoonotic spillovers., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)- Published
- 2022
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35. Broadly neutralizing antibodies overcome SARS-CoV-2 Omicron antigenic shift.
- Author
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Cameroni E, Saliba C, Bowen JE, Rosen LE, Culap K, Pinto D, VanBlargan LA, De Marco A, Zepeda SK, Iulio JD, Zatta F, Kaiser H, Noack J, Farhat N, Czudnochowski N, Havenar-Daughton C, Sprouse KR, Dillen JR, Powell AE, Chen A, Maher C, Yin L, Sun D, Soriaga L, Bassi J, Silacci-Fregni C, Gustafsson C, Franko NM, Logue J, Iqbal NT, Mazzitelli I, Geffner J, Grifantini R, Chu H, Gori A, Riva A, Giannini O, Ceschi A, Ferrari P, Cippà P, Franzetti-Pellanda A, Garzoni C, Halfmann PJ, Kawaoka Y, Hebner C, Purcell LA, Piccoli L, Pizzuto MS, Walls AC, Diamond MS, Telenti A, Virgin HW, Lanzavecchia A, Veesler D, Snell G, and Corti D
- Abstract
The recently emerged SARS-CoV-2 Omicron variant harbors 37 amino acid substitutions in the spike (S) protein, 15 of which are in the receptor-binding domain (RBD), thereby raising concerns about the effectiveness of available vaccines and antibody therapeutics. Here, we show that the Omicron RBD binds to human ACE2 with enhanced affinity relative to the Wuhan-Hu-1 RBD and acquires binding to mouse ACE2. Severe reductions of plasma neutralizing activity were observed against Omicron compared to the ancestral pseudovirus for vaccinated and convalescent individuals. Most (26 out of 29) receptor-binding motif (RBM)-directed monoclonal antibodies (mAbs) lost in vitro neutralizing activity against Omicron, with only three mAbs, including the ACE2-mimicking S2K146 mAb
1 , retaining unaltered potency. Furthermore, a fraction of broadly neutralizing sarbecovirus mAbs recognizing antigenic sites outside the RBM, including sotrovimab2 , S2X2593 and S2H974 , neutralized Omicron. The magnitude of Omicron-mediated immune evasion and the acquisition of binding to mouse ACE2 mark a major SARS-CoV-2 mutational shift. Broadly neutralizing sarbecovirus mAbs recognizing epitopes conserved among SARS-CoV-2 variants and other sarbecoviruses may prove key to controlling the ongoing pandemic and future zoonotic spillovers.- Published
- 2021
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36. Broad betacoronavirus neutralization by a stem helix-specific human antibody.
- Author
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Pinto D, Sauer MM, Czudnochowski N, Low JS, Tortorici MA, Housley MP, Noack J, Walls AC, Bowen JE, Guarino B, Rosen LE, di Iulio J, Jerak J, Kaiser H, Islam S, Jaconi S, Sprugasci N, Culap K, Abdelnabi R, Foo C, Coelmont L, Bartha I, Bianchi S, Silacci-Fregni C, Bassi J, Marzi R, Vetti E, Cassotta A, Ceschi A, Ferrari P, Cippà PE, Giannini O, Ceruti S, Garzoni C, Riva A, Benigni F, Cameroni E, Piccoli L, Pizzuto MS, Smithey M, Hong D, Telenti A, Lempp FA, Neyts J, Havenar-Daughton C, Lanzavecchia A, Sallusto F, Snell G, Virgin HW, Beltramello M, Corti D, and Veesler D
- Subjects
- Animals, Antibodies, Monoclonal isolation & purification, Antibodies, Neutralizing isolation & purification, Convalescence, Cricetinae, Cross Reactions, Humans, Immunoglobulin Fab Fragments immunology, Immunoglobulin Fc Fragments immunology, Jurkat Cells, Lung immunology, Membrane Fusion immunology, Neutralization Tests, Peptide Mapping, Protein Conformation, alpha-Helical, SARS-CoV-2 immunology, Spike Glycoprotein, Coronavirus genetics, Viral Load immunology, Antibodies, Monoclonal immunology, Antibodies, Neutralizing immunology, Betacoronavirus immunology, Spike Glycoprotein, Coronavirus chemistry, Spike Glycoprotein, Coronavirus immunology, Viral Vaccines immunology, Virus Internalization
- Abstract
The spillovers of betacoronaviruses in humans and the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants highlight the need for broad coronavirus countermeasures. We describe five monoclonal antibodies (mAbs) cross-reacting with the stem helix of multiple betacoronavirus spike glycoproteins isolated from COVID-19 convalescent individuals. Using structural and functional studies, we show that the mAb with the greatest breadth (S2P6) neutralizes pseudotyped viruses from three different subgenera through the inhibition of membrane fusion, and we delineate the molecular basis for its cross-reactivity. S2P6 reduces viral burden in hamsters challenged with SARS-CoV-2 through viral neutralization and Fc-mediated effector functions. Stem helix antibodies are rare, oftentimes of narrow specificity, and can acquire neutralization breadth through somatic mutations. These data provide a framework for structure-guided design of pan-betacoronavirus vaccines eliciting broad protection.
- Published
- 2021
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37. Broad sarbecovirus neutralization by a human monoclonal antibody.
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Tortorici MA, Czudnochowski N, Starr TN, Marzi R, Walls AC, Zatta F, Bowen JE, Jaconi S, Di Iulio J, Wang Z, De Marco A, Zepeda SK, Pinto D, Liu Z, Beltramello M, Bartha I, Housley MP, Lempp FA, Rosen LE, Dellota E Jr, Kaiser H, Montiel-Ruiz M, Zhou J, Addetia A, Guarino B, Culap K, Sprugasci N, Saliba C, Vetti E, Giacchetto-Sasselli I, Fregni CS, Abdelnabi R, Foo SC, Havenar-Daughton C, Schmid MA, Benigni F, Cameroni E, Neyts J, Telenti A, Virgin HW, Whelan SPJ, Snell G, Bloom JD, Corti D, Veesler D, and Pizzuto MS
- Subjects
- Animals, Antibodies, Monoclonal chemistry, Antibodies, Viral chemistry, Antibodies, Viral therapeutic use, Broadly Neutralizing Antibodies chemistry, COVID-19 immunology, COVID-19 virology, Cross Reactions immunology, Disease Models, Animal, Female, Humans, Immune Evasion genetics, Immune Evasion immunology, Mesocricetus immunology, Mesocricetus virology, Mutation, Neutralization Tests, SARS-CoV-2 chemistry, SARS-CoV-2 genetics, Viral Zoonoses immunology, Viral Zoonoses prevention & control, Viral Zoonoses virology, Antibodies, Monoclonal immunology, Antibodies, Monoclonal therapeutic use, Antibodies, Viral immunology, Broadly Neutralizing Antibodies immunology, Broadly Neutralizing Antibodies therapeutic use, COVID-19 prevention & control, SARS-CoV-2 classification, SARS-CoV-2 immunology
- Abstract
The recent emergence of SARS-CoV-2 variants of concern
1-10 and the recurrent spillovers of coronaviruses11,12 into the human population highlight the need for broadly neutralizing antibodies that are not affected by the ongoing antigenic drift and that can prevent or treat future zoonotic infections. Here we describe a human monoclonal antibody designated S2X259, which recognizes a highly conserved cryptic epitope of the receptor-binding domain and cross-reacts with spikes from all clades of sarbecovirus. S2X259 broadly neutralizes spike-mediated cell entry of SARS-CoV-2, including variants of concern (B.1.1.7, B.1.351, P.1, and B.1.427/B.1.429), as well as a wide spectrum of human and potentially zoonotic sarbecoviruses through inhibition of angiotensin-converting enzyme 2 (ACE2) binding to the receptor-binding domain. Furthermore, deep-mutational scanning and in vitro escape selection experiments demonstrate that S2X259 possesses an escape profile that is limited to a single substitution, G504D. We show that prophylactic and therapeutic administration of S2X259 protects Syrian hamsters (Mesocricetus auratus) against challenge with the prototypic SARS-CoV-2 and the B.1.351 variant of concern, which suggests that this monoclonal antibody is a promising candidate for the prevention and treatment of emergent variants and zoonotic infections. Our data reveal a key antigenic site that is targeted by broadly neutralizing antibodies and will guide the design of vaccines that are effective against all sarbecoviruses., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)- Published
- 2021
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38. SARS-CoV-2 RBD antibodies that maximize breadth and resistance to escape.
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Starr TN, Czudnochowski N, Liu Z, Zatta F, Park YJ, Addetia A, Pinto D, Beltramello M, Hernandez P, Greaney AJ, Marzi R, Glass WG, Zhang I, Dingens AS, Bowen JE, Tortorici MA, Walls AC, Wojcechowskyj JA, De Marco A, Rosen LE, Zhou J, Montiel-Ruiz M, Kaiser H, Dillen JR, Tucker H, Bassi J, Silacci-Fregni C, Housley MP, di Iulio J, Lombardo G, Agostini M, Sprugasci N, Culap K, Jaconi S, Meury M, Dellota E Jr, Abdelnabi R, Foo SC, Cameroni E, Stumpf S, Croll TI, Nix JC, Havenar-Daughton C, Piccoli L, Benigni F, Neyts J, Telenti A, Lempp FA, Pizzuto MS, Chodera JD, Hebner CM, Virgin HW, Whelan SPJ, Veesler D, Corti D, Bloom JD, and Snell G
- Subjects
- Adult, Aged, Animals, Antibodies, Monoclonal chemistry, Antibodies, Monoclonal immunology, Antibodies, Viral chemistry, Antibodies, Viral immunology, Antibody Affinity, Broadly Neutralizing Antibodies chemistry, COVID-19 immunology, COVID-19 Vaccines chemistry, COVID-19 Vaccines immunology, Cell Line, Cricetinae, Epitopes, B-Lymphocyte chemistry, Epitopes, B-Lymphocyte genetics, Epitopes, B-Lymphocyte immunology, Female, Humans, Male, Mesocricetus, Middle Aged, Models, Molecular, SARS-CoV-2 chemistry, SARS-CoV-2 genetics, Spike Glycoprotein, Coronavirus genetics, Vaccinology, COVID-19 Drug Treatment, Broadly Neutralizing Antibodies immunology, COVID-19 virology, Cross Reactions immunology, Immune Evasion genetics, Immune Evasion immunology, SARS-CoV-2 classification, SARS-CoV-2 immunology, Spike Glycoprotein, Coronavirus chemistry, Spike Glycoprotein, Coronavirus immunology
- Abstract
An ideal therapeutic anti-SARS-CoV-2 antibody would resist viral escape
1-3 , have activity against diverse sarbecoviruses4-7 , and be highly protective through viral neutralization8-11 and effector functions12,13 . Understanding how these properties relate to each other and vary across epitopes would aid the development of therapeutic antibodies and guide vaccine design. Here we comprehensively characterize escape, breadth and potency across a panel of SARS-CoV-2 antibodies targeting the receptor-binding domain (RBD). Despite a trade-off between in vitro neutralization potency and breadth of sarbecovirus binding, we identify neutralizing antibodies with exceptional sarbecovirus breadth and a corresponding resistance to SARS-CoV-2 escape. One of these antibodies, S2H97, binds with high affinity across all sarbecovirus clades to a cryptic epitope and prophylactically protects hamsters from viral challenge. Antibodies that target the angiotensin-converting enzyme 2 (ACE2) receptor-binding motif (RBM) typically have poor breadth and are readily escaped by mutations despite high neutralization potency. Nevertheless, we also characterize a potent RBM antibody (S2E128 ) with breadth across sarbecoviruses related to SARS-CoV-2 and a high barrier to viral escape. These data highlight principles underlying variation in escape, breadth and potency among antibodies that target the RBD, and identify epitopes and features to prioritize for therapeutic development against the current and potential future pandemics., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)- Published
- 2021
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39. Structural basis for broad sarbecovirus neutralization by a human monoclonal antibody.
- Author
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Tortorici MA, Czudnochowski N, Starr TN, Marzi R, Walls AC, Zatta F, Bowen JE, Jaconi S, di Iulio J, Wang Z, De Marco A, Zepeda SK, Pinto D, Liu Z, Beltramello M, Bartha I, Housley MP, Lempp FA, Rosen LE, Dellota E Jr, Kaiser H, Montiel-Ruiz M, Zhou J, Addetia A, Guarino B, Culap K, Sprugasci N, Saliba C, Vetti E, Giacchetto-Sasselli I, Silacci Fregni C, Abdelnabi R, Caroline Foo SY, Havenar-Daughton C, Schmid MA, Benigni F, Cameroni E, Neyts J, Telenti A, Snell G, Virgin HW, Whelan SPJ, Bloom JD, Corti D, Veesler D, and Pizzuto MS
- Abstract
The recent emergence of SARS-CoV-2 variants of concern (VOC) and the recurrent spillovers of coronaviruses in the human population highlight the need for broadly neutralizing antibodies that are not affected by the ongoing antigenic drift and that can prevent or treat future zoonotic infections. Here, we describe a human monoclonal antibody (mAb), designated S2X259, recognizing a highly conserved cryptic receptor-binding domain (RBD) epitope and cross-reacting with spikes from all sarbecovirus clades. S2X259 broadly neutralizes spike-mediated entry of SARS-CoV-2 including the B.1.1.7, B.1.351, P.1 and B.1.427/B.1.429 VOC, as well as a wide spectrum of human and zoonotic sarbecoviruses through inhibition of ACE2 binding to the RBD. Furthermore, deep-mutational scanning and in vitro escape selection experiments demonstrate that S2X259 possesses a remarkably high barrier to the emergence of resistance mutants. We show that prophylactic administration of S2X259 protects Syrian hamsters against challenges with the prototypic SARS-CoV-2 and the B.1.351 variant, suggesting this mAb is a promising candidate for the prevention and treatment of emergent VOC and zoonotic infections. Our data unveil a key antigenic site targeted by broadly-neutralizing antibodies and will guide the design of pan-sarbecovirus vaccines.
- Published
- 2021
- Full Text
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40. Antibodies to the SARS-CoV-2 receptor-binding domain that maximize breadth and resistance to viral escape.
- Author
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Starr TN, Czudnochowski N, Zatta F, Park YJ, Liu Z, Addetia A, Pinto D, Beltramello M, Hernandez P, Greaney AJ, Marzi R, Glass WG, Zhang I, Dingens AS, Bowen JE, Wojcechowskyj JA, De Marco A, Rosen LE, Zhou J, Montiel-Ruiz M, Kaiser H, Tucker H, Housley MP, di Iulio J, Lombardo G, Agostini M, Sprugasci N, Culap K, Jaconi S, Meury M, Dellota E, Cameroni E, Croll TI, Nix JC, Havenar-Daughton C, Telenti A, Lempp FA, Pizzuto MS, Chodera JD, Hebner CM, Whelan SPJ, Virgin HW, Veesler D, Corti D, Bloom JD, and Snell G
- Abstract
An ideal anti-SARS-CoV-2 antibody would resist viral escape
1-3 , have activity against diverse SARS-related coronaviruses4-7 , and be highly protective through viral neutralization8-11 and effector functions12,13 . Understanding how these properties relate to each other and vary across epitopes would aid development of antibody therapeutics and guide vaccine design. Here, we comprehensively characterize escape, breadth, and potency across a panel of SARS-CoV-2 antibodies targeting the receptor-binding domain (RBD), including S3094 , the parental antibody of the late-stage clinical antibody VIR-7831. We observe a tradeoff between SARS-CoV-2 in vitro neutralization potency and breadth of binding across SARS-related coronaviruses. Nevertheless, we identify several neutralizing antibodies with exceptional breadth and resistance to escape, including a new antibody (S2H97) that binds with high affinity to all SARS-related coronavirus clades via a unique RBD epitope centered on residue E516. S2H97 and other escape-resistant antibodies have high binding affinity and target functionally constrained RBD residues. We find that antibodies targeting the ACE2 receptor binding motif (RBM) typically have poor breadth and are readily escaped by mutations despite high neutralization potency, but we identify one potent RBM antibody (S2E12) with breadth across sarbecoviruses closely related to SARS-CoV-2 and with a high barrier to viral escape. These data highlight functional diversity among antibodies targeting the RBD and identify epitopes and features to prioritize for antibody and vaccine development against the current and potential future pandemics.- Published
- 2021
- Full Text
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41. Circulating SARS-CoV-2 spike N439K variants maintain fitness while evading antibody-mediated immunity.
- Author
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Thomson EC, Rosen LE, Shepherd JG, Spreafico R, da Silva Filipe A, Wojcechowskyj JA, Davis C, Piccoli L, Pascall DJ, Dillen J, Lytras S, Czudnochowski N, Shah R, Meury M, Jesudason N, De Marco A, Li K, Bassi J, O'Toole A, Pinto D, Colquhoun RM, Culap K, Jackson B, Zatta F, Rambaut A, Jaconi S, Sreenu VB, Nix J, Zhang I, Jarrett RF, Glass WG, Beltramello M, Nomikou K, Pizzuto M, Tong L, Cameroni E, Croll TI, Johnson N, Di Iulio J, Wickenhagen A, Ceschi A, Harbison AM, Mair D, Ferrari P, Smollett K, Sallusto F, Carmichael S, Garzoni C, Nichols J, Galli M, Hughes J, Riva A, Ho A, Schiuma M, Semple MG, Openshaw PJM, Fadda E, Baillie JK, Chodera JD, Rihn SJ, Lycett SJ, Virgin HW, Telenti A, Corti D, Robertson DL, and Snell G
- Subjects
- Angiotensin-Converting Enzyme 2 chemistry, Antibodies, Neutralizing genetics, Antibodies, Neutralizing immunology, Antibodies, Viral immunology, COVID-19 virology, Humans, Mutation, Phylogeny, SARS-CoV-2 chemistry, SARS-CoV-2 pathogenicity, Spike Glycoprotein, Coronavirus chemistry, Virulence, COVID-19 immunology, Genetic Fitness, Immune Evasion, SARS-CoV-2 genetics, Spike Glycoprotein, Coronavirus genetics
- Abstract
SARS-CoV-2 can mutate and evade immunity, with consequences for efficacy of emerging vaccines and antibody therapeutics. Here, we demonstrate that the immunodominant SARS-CoV-2 spike (S) receptor binding motif (RBM) is a highly variable region of S and provide epidemiological, clinical, and molecular characterization of a prevalent, sentinel RBM mutation, N439K. We demonstrate N439K S protein has enhanced binding affinity to the hACE2 receptor, and N439K viruses have similar in vitro replication fitness and cause infections with similar clinical outcomes as compared to wild type. We show the N439K mutation confers resistance against several neutralizing monoclonal antibodies, including one authorized for emergency use by the US Food and Drug Administration (FDA), and reduces the activity of some polyclonal sera from persons recovered from infection. Immune evasion mutations that maintain virulence and fitness such as N439K can emerge within SARS-CoV-2 S, highlighting the need for ongoing molecular surveillance to guide development and usage of vaccines and therapeutics., Competing Interests: Declaration of interests L.E.R., R. Spreafico, J.A.W., L.P., J.D., N.C., M.M., A.D.M., J.B., D.P., K.C., F.Z., S.J., M.B., M.P., E.C., J.D.I., H.W.V., A.T., D.C., and G.S. are or were employees of Vir Biotechnology and may hold shares in Vir Biotechnology. C.G. is an external scientific advisor for Humabs BioMed SA. J. Nix and T.I.C. are consultants with Vir Biotechnology. M.G.S. declares interest in Integrum Scientific, Greensboro, NC, outside the scope of this work. J.D.C. is a current member of the Scientific Advisory Board of OpenEye Scientific Software and is a scientific consultant to Foresite Labs. The Chodera laboratory (I.Z., W.G.G., and J.D.C.) receives or has received funding from multiple sources, including the NIH, the National Science Foundation, the Parker Institute for Cancer Immunotherapy, Relay Therapeutics, Entasis Therapeutics, Silicon Therapeutics, EMD Serono (Merck KGaA), AstraZeneca, Vir Biotechnology, XtalPi, the Molecular Sciences Software Institute, the Starr Cancer Consortium, the Open Force Field Consortium, Cycle for Survival, a Louis V. Gerstner Young Investigator Award, and the Sloan Kettering Institute. A complete funding history for the Chodera lab can be found at https://www.choderalab.org/funding. The other authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)
- Published
- 2021
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42. Ultrapotent human antibodies protect against SARS-CoV-2 challenge via multiple mechanisms.
- Author
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Tortorici MA, Beltramello M, Lempp FA, Pinto D, Dang HV, Rosen LE, McCallum M, Bowen J, Minola A, Jaconi S, Zatta F, De Marco A, Guarino B, Bianchi S, Lauron EJ, Tucker H, Zhou J, Peter A, Havenar-Daughton C, Wojcechowskyj JA, Case JB, Chen RE, Kaiser H, Montiel-Ruiz M, Meury M, Czudnochowski N, Spreafico R, Dillen J, Ng C, Sprugasci N, Culap K, Benigni F, Abdelnabi R, Foo SC, Schmid MA, Cameroni E, Riva A, Gabrieli A, Galli M, Pizzuto MS, Neyts J, Diamond MS, Virgin HW, Snell G, Corti D, Fink K, and Veesler D
- Subjects
- Amino Acid Motifs immunology, Angiotensin-Converting Enzyme 2, Animals, Antibodies, Neutralizing administration & dosage, Antibodies, Neutralizing isolation & purification, Antibodies, Viral administration & dosage, Antibodies, Viral isolation & purification, CHO Cells, COVID-19, Coronavirus Infections therapy, Cricetinae, Cricetulus, Cryoelectron Microscopy, HEK293 Cells, Humans, Immunodominant Epitopes chemistry, Immunodominant Epitopes immunology, Microscopy, Electron, Pneumonia, Viral therapy, Protein Domains immunology, SARS-CoV-2, Spike Glycoprotein, Coronavirus chemistry, Spike Glycoprotein, Coronavirus immunology, Antibodies, Neutralizing immunology, Antibodies, Viral immunology, Betacoronavirus immunology, Coronavirus Infections prevention & control, Pandemics prevention & control, Peptidyl-Dipeptidase A immunology, Pneumonia, Viral prevention & control, Spike Glycoprotein, Coronavirus antagonists & inhibitors
- Abstract
Efficient therapeutic options are needed to control the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that has caused more than 922,000 fatalities as of 13 September 2020. We report the isolation and characterization of two ultrapotent SARS-CoV-2 human neutralizing antibodies (S2E12 and S2M11) that protect hamsters against SARS-CoV-2 challenge. Cryo-electron microscopy structures show that S2E12 and S2M11 competitively block angiotensin-converting enzyme 2 (ACE2) attachment and that S2M11 also locks the spike in a closed conformation by recognition of a quaternary epitope spanning two adjacent receptor-binding domains. Antibody cocktails that include S2M11, S2E12, or the previously identified S309 antibody broadly neutralize a panel of circulating SARS-CoV-2 isolates and activate effector functions. Our results pave the way to implement antibody cocktails for prophylaxis or therapy, circumventing or limiting the emergence of viral escape mutants., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)
- Published
- 2020
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43. Mapping Neutralizing and Immunodominant Sites on the SARS-CoV-2 Spike Receptor-Binding Domain by Structure-Guided High-Resolution Serology.
- Author
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Piccoli L, Park YJ, Tortorici MA, Czudnochowski N, Walls AC, Beltramello M, Silacci-Fregni C, Pinto D, Rosen LE, Bowen JE, Acton OJ, Jaconi S, Guarino B, Minola A, Zatta F, Sprugasci N, Bassi J, Peter A, De Marco A, Nix JC, Mele F, Jovic S, Rodriguez BF, Gupta SV, Jin F, Piumatti G, Lo Presti G, Pellanda AF, Biggiogero M, Tarkowski M, Pizzuto MS, Cameroni E, Havenar-Daughton C, Smithey M, Hong D, Lepori V, Albanese E, Ceschi A, Bernasconi E, Elzi L, Ferrari P, Garzoni C, Riva A, Snell G, Sallusto F, Fink K, Virgin HW, Lanzavecchia A, Corti D, and Veesler D
- Subjects
- Angiotensin-Converting Enzyme 2, Antibodies, Monoclonal chemistry, Antibodies, Monoclonal genetics, Antibodies, Monoclonal immunology, Antibodies, Neutralizing blood, Antibodies, Neutralizing chemistry, Antibodies, Viral blood, Antibodies, Viral chemistry, Antibodies, Viral immunology, Antigen-Antibody Reactions, Betacoronavirus immunology, Betacoronavirus isolation & purification, Betacoronavirus metabolism, Binding Sites, COVID-19, Coronavirus Infections pathology, Coronavirus Infections virology, Epitopes chemistry, Epitopes immunology, Humans, Immunoglobulin A blood, Immunoglobulin A immunology, Immunoglobulin G blood, Immunoglobulin G immunology, Immunoglobulin M blood, Immunoglobulin M immunology, Kinetics, Molecular Dynamics Simulation, Pandemics, Peptidyl-Dipeptidase A chemistry, Peptidyl-Dipeptidase A metabolism, Pneumonia, Viral pathology, Pneumonia, Viral virology, Protein Binding, Protein Domains immunology, Protein Structure, Quaternary, SARS-CoV-2, Spike Glycoprotein, Coronavirus chemistry, Spike Glycoprotein, Coronavirus genetics, Spike Glycoprotein, Coronavirus metabolism, Antibodies, Neutralizing immunology, Epitope Mapping methods, Spike Glycoprotein, Coronavirus immunology
- Abstract
Analysis of the specificity and kinetics of neutralizing antibodies (nAbs) elicited by SARS-CoV-2 infection is crucial for understanding immune protection and identifying targets for vaccine design. In a cohort of 647 SARS-CoV-2-infected subjects, we found that both the magnitude of Ab responses to SARS-CoV-2 spike (S) and nucleoprotein and nAb titers correlate with clinical scores. The receptor-binding domain (RBD) is immunodominant and the target of 90% of the neutralizing activity present in SARS-CoV-2 immune sera. Whereas overall RBD-specific serum IgG titers waned with a half-life of 49 days, nAb titers and avidity increased over time for some individuals, consistent with affinity maturation. We structurally defined an RBD antigenic map and serologically quantified serum Abs specific for distinct RBD epitopes leading to the identification of two major receptor-binding motif antigenic sites. Our results explain the immunodominance of the receptor-binding motif and will guide the design of COVID-19 vaccines and therapeutics., Competing Interests: Declaration of Interests L.P., N.C., M. Beltramello, C.S.-F., D.P., L.E.R., F.Z., N.S., J.B., A.P., S. Jaconi, B.G., A.M., A.D.M., M.S.P., E.C., S.V.G., F.J., C.H.-D., M.S., D.H., G.S., K.F., H.W.V., A.L., and D.C. are employees of Vir Biotechnology Inc. and may hold shares in Vir Biotechnology Inc. D.C. is currently listed as an inventor on multiple patent applications, which disclose the subject matter described in this manuscript. The Veesler laboratory has received a sponsored research agreement from Vir Biotechnology Inc. The other authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)
- Published
- 2020
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44. The structure of the endogenous ESX-3 secretion system.
- Author
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Poweleit N, Czudnochowski N, Nakagawa R, Trinidad DD, Murphy KC, Sassetti CM, and Rosenberg OS
- Subjects
- Bacterial Proteins genetics, Bacterial Proteins ultrastructure, Chromosomes chemistry, Chromosomes genetics, Epitopes chemistry, Epitopes genetics, Mycobacterium smegmatis ultrastructure, Operon genetics, Type VII Secretion Systems genetics, Type VII Secretion Systems ultrastructure, Bacterial Proteins chemistry, Mycobacterium smegmatis chemistry, Protein Transport genetics, Type VII Secretion Systems chemistry
- Abstract
The ESX (or Type VII) secretion systems are protein export systems in mycobacteria and many Gram-positive bacteria that mediate a broad range of functions including virulence, conjugation, and metabolic regulation. These systems translocate folded dimers of WXG100-superfamily protein substrates across the cytoplasmic membrane. We report the cryo-electron microscopy structure of an ESX-3 system, purified using an epitope tag inserted with recombineering into the chromosome of the model organism Mycobacterium smegmatis . The structure reveals a stacked architecture that extends above and below the inner membrane of the bacterium. The ESX-3 protomer complex is assembled from a single copy of the EccB
3 , EccC3 , and EccE3 and two copies of the EccD3 protein. In the structure, the protomers form a stable dimer that is consistent with assembly into a larger oligomer. The ESX-3 structure provides a framework for further study of these important bacterial transporters., Competing Interests: NP, NC, RN, DT, KM, CS, OR No competing interests declared, (© 2019, Poweleit et al.)- Published
- 2019
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45. Two Accessory Proteins Govern MmpL3 Mycolic Acid Transport in Mycobacteria.
- Author
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Fay A, Czudnochowski N, Rock JM, Johnson JR, Krogan NJ, Rosenberg O, and Glickman MS
- Subjects
- Cell Membrane metabolism, Cell Wall metabolism, Mycobacterium smegmatis genetics, Mycobacterium tuberculosis genetics, Bacterial Proteins metabolism, Membrane Transport Proteins metabolism, Mycobacterium smegmatis metabolism, Mycobacterium tuberculosis metabolism, Mycolic Acids metabolism
- Abstract
Mycolic acids are the signature lipid of mycobacteria and constitute an important physical component of the cell wall, a target of mycobacterium-specific antibiotics and a mediator of Mycobacterium tuberculosis pathogenesis. Mycolic acids are synthesized in the cytoplasm and are thought to be transported to the cell wall as a trehalose ester by the MmpL3 transporter, an antibiotic target for M. tuberculosis However, the mechanism by which mycolate synthesis is coupled to transport, and the full MmpL3 transport machinery, is unknown. Here, we identify two new components of the MmpL3 transport machinery in mycobacteria. The protein encoded by MSMEG_0736 / Rv0383c is essential for growth of Mycobacterium smegmatis and M. tuberculosis and is anchored to the cytoplasmic membrane, physically interacts with and colocalizes with MmpL3 in growing cells, and is required for trehalose monomycolate (TMM) transport to the cell wall. In light of these findings, we propose MSMEG_0736/Rv0383c be named "TMM transport factor A", TtfA. The protein encoded by MSMEG_5308 also interacts with the MmpL3 complex but is nonessential for growth or TMM transport. However, MSMEG_5308 accumulates with inhibition of MmpL3-mediated TMM transport and stabilizes the MmpL3/TtfA complex, indicating that it may stabilize the transport system during stress. These studies identify two new components of the mycobacterial mycolate transport machinery, an emerging antibiotic target in M. tuberculosis IMPORTANCE The cell envelope of Mycobacterium tuberculosis , the bacterium that causes the disease tuberculosis, is a complex structure composed of abundant lipids and glycolipids, including the signature lipid of these bacteria, mycolic acids. In this study, we identified two new components of the transport machinery that constructs this complex cell wall. These two accessory proteins are in a complex with the MmpL3 transporter. One of these proteins, TtfA, is required for mycolic acid transport and cell viability, whereas the other stabilizes the MmpL3 complex. These studies identify two new components of the essential cell envelope biosynthetic machinery in mycobacteria., (Copyright © 2019 Fay et al.)
- Published
- 2019
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46. Chlamydia interfere with an interaction between the mannose-6-phosphate receptor and sorting nexins to counteract host restriction.
- Author
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Elwell CA, Czudnochowski N, von Dollen J, Johnson JR, Nakagawa R, Mirrashidi K, Krogan NJ, Engel JN, and Rosenberg OS
- Subjects
- Animals, Bacterial Proteins chemistry, Bacterial Proteins genetics, Crystallography, X-Ray, DNA Mutational Analysis, Mice, Models, Molecular, Protein Conformation, Protein Interaction Mapping, Receptor, IGF Type 2 chemistry, Receptor, IGF Type 2 genetics, Sorting Nexins chemistry, Sorting Nexins genetics, Bacterial Proteins metabolism, Chlamydia trachomatis immunology, Chlamydia trachomatis physiology, Host-Pathogen Interactions, Immune Evasion, Receptor, IGF Type 2 metabolism, Sorting Nexins metabolism
- Abstract
Chlamydia trachomatis is an obligate intracellular pathogen that resides in a membrane-bound compartment, the inclusion. The bacteria secrete a unique class of proteins, Incs, which insert into the inclusion membrane and modulate the host-bacterium interface. We previously reported that IncE binds specifically to the Sorting Nexin 5 Phox domain (SNX5-PX) and disrupts retromer trafficking. Here, we present the crystal structure of the SNX5-PX:IncE complex, showing IncE bound to a unique and highly conserved hydrophobic groove on SNX5. Mutagenesis of the SNX5-PX:IncE binding surface disrupts a previously unsuspected interaction between SNX5 and the cation-independent mannose-6-phosphate receptor (CI-MPR). Addition of IncE peptide inhibits the interaction of CI-MPR with SNX5. Finally, C. trachomatis infection interferes with the SNX5:CI-MPR interaction, suggesting that IncE and CI-MPR are dependent on the same binding surface on SNX5. Our results provide new insights into retromer assembly and underscore the power of using pathogens to discover disease-related cell biology.
- Published
- 2017
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47. The mechanism of pseudouridine synthases from a covalent complex with RNA, and alternate specificity for U2605 versus U2604 between close homologs.
- Author
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Czudnochowski N, Ashley GW, Santi DV, Alian A, Finer-Moore J, and Stroud RM
- Subjects
- Apoenzymes chemistry, Arginine chemistry, Catalytic Domain, Escherichia coli Proteins metabolism, Intramolecular Transferases metabolism, Models, Molecular, Nucleic Acid Conformation, Protein Conformation, RNA, Ribosomal, 23S metabolism, Substrate Specificity, Tyrosine chemistry, Uridine analogs & derivatives, Uridine chemistry, Uridine metabolism, Water chemistry, Escherichia coli Proteins chemistry, Intramolecular Transferases chemistry, RNA, Ribosomal, 23S chemistry
- Abstract
RluB catalyses the modification of U2605 to pseudouridine (Ψ) in a stem-loop at the peptidyl transferase center of Escherichia coli 23S rRNA. The homolog RluF is specific to the adjacent nucleotide in the stem, U2604. The 1.3 Å resolution crystal structure of the complex between the catalytic domain of RluB and the isolated substrate stem-loop, in which the target uridine is substituted by 5-fluorouridine (5-FU), reveals a covalent bond between the isomerized target base and tyrosine 140. The structure is compared with the catalytic domain alone determined at 2.5 Å resolution. The RluB-bound stem-loop has essentially the same secondary structure as in the ribosome, with a bulge at A2602, but with 5-FU2605 flipped into the active site. We showed earlier that RluF induced a frame-shift of the RNA, moving A2602 into the stem and translating its target, U2604, into the active site. A hydrogen-bonding network stabilizes the bulge in the RluB-RNA but is not conserved in RluF and so RluF cannot stabilize the bulge. On the basis of the covalent bond between enzyme and isomerized 5-FU we propose a Michael addition mechanism for pseudouridine formation that is consistent with all experimental data.
- Published
- 2014
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48. In human pseudouridine synthase 1 (hPus1), a C-terminal helical insert blocks tRNA from binding in the same orientation as in the Pus1 bacterial homologue TruA, consistent with their different target selectivities.
- Author
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Czudnochowski N, Wang AL, Finer-Moore J, and Stroud RM
- Subjects
- Bacteria genetics, Bacteria metabolism, Catalytic Domain, Humans, Hydro-Lyases chemistry, Models, Molecular, Molecular Docking Simulation, Protein Binding, Protein Conformation, Protein Folding, Protein Interaction Domains and Motifs, Pseudouridine biosynthesis, RNA, Transfer chemistry, Hydro-Lyases metabolism, RNA, Transfer metabolism
- Abstract
Human pseudouridine (Ψ) synthase Pus1 (hPus1) modifies specific uridine residues in several non-coding RNAs: tRNA, U2 spliceosomal RNA, and steroid receptor activator RNA. We report three structures of the catalytic core domain of hPus1 from two crystal forms, at 1.8Å resolution. The structures are the first of a mammalian Ψ synthase from the set of five Ψ synthase families common to all kingdoms of life. hPus1 adopts a fold similar to bacterial Ψ synthases, with a central antiparallel β-sheet flanked by helices and loops. A flexible hinge at the base of the sheet allows the enzyme to open and close around an electropositive active-site cleft. In one crystal form, a molecule of Mes [2-(N-morpholino)ethane sulfonic acid] mimics the target uridine of an RNA substrate. A positively charged electrostatic surface extends from the active site towards the N-terminus of the catalytic domain, suggesting an extensive binding site specific for target RNAs. Two α-helices C-terminal to the core domain, but unique to hPus1, extend along the back and top of the central β-sheet and form the walls of the RNA binding surface. Docking of tRNA to hPus1 in a productive orientation requires only minor conformational changes to enzyme and tRNA. The docked tRNA is bound by the electropositive surface of the protein employing a completely different binding mode than that seen for the tRNA complex of the Escherichia coli homologue TruA., (Copyright © 2013 Elsevier Ltd. All rights reserved.)
- Published
- 2013
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49. Serine-7 but not serine-5 phosphorylation primes RNA polymerase II CTD for P-TEFb recognition.
- Author
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Czudnochowski N, Bösken CA, and Geyer M
- Subjects
- Amino Acid Motifs, Amino Acid Sequence, Humans, Molecular Sequence Data, Phosphorylation, Positive Transcriptional Elongation Factor B genetics, Protein Binding, Protein Structure, Tertiary, RNA Polymerase II genetics, RNA-Binding Proteins genetics, RNA-Binding Proteins metabolism, Serine genetics, Transcription Factors, Positive Transcriptional Elongation Factor B metabolism, RNA Polymerase II chemistry, RNA Polymerase II metabolism, Serine metabolism
- Abstract
Phosphorylation of RNA polymerase II carboxy-terminal domain (CTD) in hepta-repeats YSPTSPS regulates eukaryotic transcription. Whereas Ser5 is phosphorylated in the initiation phase, Ser2 phosphorylation marks the elongation state. Here we show that the positive transcription elongation factor P-TEFb is a Ser5 CTD kinase that is unable to create Ser2/Ser5 double phosphorylations, while it exhibits fourfold higher activity on a CTD substrate pre-phosphorylated at Ser7 compared with the consensus hepta-repeat or the YSPTSPK variant. Mass spectrometry reveals an equal number of phosphorylations to the number of hepta-repeats provided, yet the mechanism of phosphorylation is distributive despite the repetitive nature of the substrate. Inhibition of P-TEFb activity is mediated by two regions in Hexim1 that act synergistically on Cdk9 and Cyclin T1. HIV-1 Tat/TAR abrogates Hexim1 inhibition to stimulate transcription of viral genes but does not change the substrate specificity. Together, these results provide insight into the multifaceted pattern of CTD phosphorylation.
- Published
- 2012
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50. Mechanistic insights into the translocation of full length HIV-1 Tat across lipid membranes.
- Author
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Boll A, Jatho A, Czudnochowski N, Geyer M, and Steinem C
- Subjects
- Microscopy, Confocal, Microscopy, Fluorescence, Phosphatidylcholines chemistry, Protein Transport, Unilamellar Liposomes chemistry, tat Gene Products, Human Immunodeficiency Virus chemistry
- Abstract
The mechanism of how full length Tat (aa 1-86) crosses artificial lipid membranes was elucidated by means of fluorescence spectroscopy and fluorescence microscopy. It was shown that full length Tat (aa 1-86) neither forms pores in large unilamellar vesicles (LUVs) nor in giant unilamellar vesicles (GUVs) composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC). In contrast, an N-terminally truncated Tat protein (aa 35-86) that lacks the structurally defined proline- and cysteine-rich region as well as the highly conserved tryptophan residue at position 11 generates pores in artificial POPC-membranes, through which a water-soluble dye up to a size of 10kDa can pass. By means of fluorescence microscopy, the transfer of fluorescently labeled full length Tat across POPC-bilayers was unambiguously visualized with a concomitant accumulation of the protein in the membrane interface. However, if the dye was attached to the protein, also pore formation was induced. The size of the pores was, however smaller than the protein size, i.e. the labeled protein with a mass of 11.6kDa passed the membrane, while a fluorescent dye with a mass of 10kDa was excluded from the vesicles' interior. The results demonstrate that pore formation is not the prime mechanism by which full length Tat crosses a membrane., (Copyright © 2011 Elsevier B.V. All rights reserved.)
- Published
- 2011
- Full Text
- View/download PDF
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