2022 |
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828. | Pfaff-Kilgore, Jennifer M; Davidson, Edgar; Kadash-Edmondson, Kathryn; Hernandez, Mayda; Rosenberg, Erin; Chambers, Ross; Castelli, Matteo; Clementi, Nicola; Mancini, Nicasio; Bailey, Justin R; Crowe, James E; Law, Mansun; Doranz, Benjamin J: Sites of vulnerability in HCV E1E2 identified by comprehensive functional screening. In: Cell Rep, vol. 39, no. 8, pp. 110859, 2022, ISSN: 2211-1247. (Type: Journal Article | Abstract | Links)
@article{pmid35613596, The E1 and E2 envelope proteins of hepatitis C virus (HCV) form a heterodimer that drives virus-host membrane fusion. Here, we analyze the role of each amino acid in E1E2 function, expressing 545 individual alanine mutants of E1E2 in human cells, incorporating them into infectious viral pseudoparticles, and testing them against 37 different monoclonal antibodies (MAbs) to ascertain full-length translation, folding, heterodimer assembly, CD81 binding, viral pseudoparticle incorporation, and infectivity. We propose a model describing the role of each critical residue in E1E2 functionality and use it to examine how MAbs neutralize infection by exploiting functionally critical sites of vulnerability on E1E2. Our results suggest that E1E2 is a surprisingly fragile protein complex where even a single alanine mutation at 92% of positions disrupts its function. The amino-acid-level targets identified are highly conserved and functionally critical and can be exploited for improved therapies and vaccines.
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827. | Dacon, Cherrelle; Tucker, Courtney; Peng, Linghang; Lee, Chang-Chun D; Lin, Ting-Hui; Yuan, Meng; Cong, Yu; Wang, Lingshu; Purser, Lauren; Williams, Jazmean K; Pyo, Chul-Woo; Kosik, Ivan; Hu, Zhe; Zhao, Ming; Mohan, Divya; Cooper, Andrew; Peterson, Mary; Skinner, Jeff; Dixit, Saurabh; Kollins, Erin; Huzella, Louis; Perry, Donna; Byrum, Russell; Lembirik, Sanae; Zhang, Yi; Yang, Eun Sung; Chen, Man; Leung, Kwanyee; Weinberg, Rona S; Pegu, Amarendra; Geraghty, Daniel E; Davidson, Edgar; Douagi, Iyadh; Moir, Susan; Yewdell, Jonathan W; Schmaljohn, Connie; Crompton, Peter D; Holbrook, Michael R; Nemazee, David; Mascola, John R; Wilson, Ian A; Tan, Joshua: Broadly neutralizing antibodies target the coronavirus fusion peptide. 2022. (Type: Miscellaneous | Abstract | Links)
@misc{pmid35441178, The potential for future coronavirus outbreaks highlights the need to develop strategies and tools to broadly target this group of pathogens. Here, using an epitope-agnostic approach, we identified six monoclonal antibodies that bound to spike proteins from all seven human-infecting coronaviruses. Epitope mapping revealed that all six antibodies target the conserved fusion peptide region adjacent to the S2' cleavage site. Two antibodies, COV44-62 and COV44-79, broadly neutralize a range of alpha and beta coronaviruses, including SARS-CoV-2 Omicron subvariants BA.1 and BA.2, albeit with lower potency than RBD-specific antibodies. In crystal structures of Fabs COV44-62 and COV44-79 with the SARS-CoV-2 fusion peptide, the fusion peptide epitope adopts a helical structure and includes the arginine at the S2' cleavage site. Importantly, COV44-79 limited disease caused by SARS-CoV-2 in a Syrian hamster model. These findings identify the fusion peptide as the target of the broadest neutralizing antibodies in an epitope-agnostic screen, highlighting this site as a candidate for next-generation coronavirus vaccine development.nnONE-SENTENCE SUMMARY: Rare monoclonal antibodies from COVID-19 convalescent individuals broadly neutralize coronaviruses by targeting the fusion peptide.
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826. | Zhou, Panpan; Song, Ge; He, Wan-Ting; Beutler, Nathan; Tse, Longping V; Martinez, David R; Schäfer, Alexandra; Anzanello, Fabio; Yong, Peter; Peng, Linghang; Dueker, Katharina; Musharrafieh, Rami; Callaghan, Sean; Capozzola, Tazio; Yuan, Meng; Liu, Hejun; Limbo, Oliver; Parren, Mara; Garcia, Elijah; Rawlings, Stephen A; Smith, Davey M; Nemazee, David; Jardine, Joseph G; Wilson, Ian A; Safonova, Yana; Rogers, Thomas F; Baric, Ralph S; Gralinski, Lisa E; Burton, Dennis R; Andrabi, Raiees: Broadly neutralizing anti-S2 antibodies protect against all three human betacoronaviruses that cause severe disease. 2022. (Type: Miscellaneous | Abstract | Links)
@misc{pmid35291291, Pan-betacoronavirus neutralizing antibodies may hold the key to developing broadly protective vaccines against coronaviruses that cause severe disease, for anticipating novel pandemic-causing viruses, and to respond more effectively to SARS-CoV-2 variants. The emergence of the Omicron variant of SARS-CoV-2 has illustrated the limitations of solely targeting the receptor binding domain (RBD) of the envelope Spike (S)-protein. Here, we isolated a large panel of broadly neutralizing antibodies (bnAbs) from SARS-CoV-2 recovered-vaccinated donors that target a conserved S2 region in the fusion machinery on betacoronavirus spikes. Select bnAbs show broad protection against all three pathogenic betacoronaviruses, SARS-CoV-1, SARS-CoV-2 and MERS-CoV, that have spilled over into humans in the past 20 years to cause severe disease. The bnAbs provide new opportunities for antibody-based interventions and key insights for developing pan-betacoronavirus vaccines.
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825. | Spruit, Cindy M; Zhu, Xueyong; Tomris, Ilhan; Ríos-Carrasco, María; Han, Alvin X; Broszeit, Frederik; van der Woude, Roosmarijn; Bouwman, Kim M; Luu, Michel M T; Matsuno, Keita; Sakoda, Yoshihiro; Russell, Colin A; Wilson, Ian A; Boons, Geert-Jan; de Vries, Robert P: -Glycolylneuraminic Acid Binding of Avian and Equine H7 Influenza A Viruses. In: J Virol, vol. 96, no. 5, pp. e0212021, 2022, ISSN: 1098-5514. (Type: Journal Article | Abstract | Links)
@article{pmid35044215, Influenza A viruses (IAV) initiate infection by binding to glycans with terminal sialic acids on the cell surface. Hosts of IAV variably express two major forms of sialic acid, -acetylneuraminic acid (NeuAc) and -glycolylneuraminic acid (NeuGc). NeuGc is produced in most mammals, including horses and pigs, but is absent in humans, ferrets, and birds. The only known naturally occurring IAV that exclusively bind NeuGc are extinct highly pathogenic equine H7N7 viruses. We determined the crystal structure of a representative equine H7 hemagglutinin (HA) in complex with NeuGc and observed high similarity in the receptor-binding domain with an avian H7 HA. To determine the molecular basis for NeuAc and NeuGc specificity, we performed systematic mutational analyses, based on the structural insights, on two distant avian H7 HAs and an H15 HA. We found that the A135E mutation is key for binding α2,3-linked NeuGc but does not abolish NeuAc binding. The additional mutations S128T, I130V, T189A, and K193R converted the specificity from NeuAc to NeuGc. We investigated the residues at positions 128, 130, 135, 189, and 193 in a phylogenetic analysis of avian and equine H7 HAs. This analysis revealed a clear distinction between equine and avian residues. The highest variability was observed at key position 135, of which only the equine glutamic acid led to NeuGc binding. These results demonstrate that genetically distinct H7 and H15 HAs can be switched from NeuAc to NeuGc binding and vice versa after the introduction of several mutations, providing insights into the adaptation of H7 viruses to NeuGc receptors. Influenza A viruses cause millions of cases of severe illness and deaths annually. To initiate infection and replicate, the virus first needs to bind to a structure on the cell surface, like a key fitting in a lock. For influenza A viruses, these "keys" (receptors) on the cell surface are chains of sugar molecules (glycans). The terminal sugar on these glycans is often either -acetylneuraminic acid (NeuAc) or -glycolylneuraminic acid (NeuGc). Most influenza A viruses bind NeuAc, but a small minority bind NeuGc. NeuGc is present in species like horses, pigs, and mice but not in humans, ferrets, and birds. Here, we investigated the molecular determinants of NeuGc specificity and the origin of viruses that bind NeuGc.
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824. | Yuan, Meng; Zhu, Xueyong; He, Wan-Ting; Zhou, Panpan; Kaku, Chengzi I; Capozzola, Tazio; Zhu, Connie Y; Yu, Xinye; Liu, Hejun; Yu, Wenli; Hua, Yuanzi; Tien, Henry; Peng, Linghang; Song, Ge; Cottrell, Christopher A; Schief, William R; Nemazee, David; Walker, Laura M; Andrabi, Raiees; Burton, Dennis R; Wilson, Ian A: A broad and potent neutralization epitope in SARS-related coronaviruses. 2022. (Type: Miscellaneous | Abstract | Links)
@misc{pmid35313576, Many neutralizing antibodies (nAbs) elicited to ancestral SARS-CoV-2 through natural infection and vaccination generally have reduced effectiveness to SARS-CoV-2 variants. Here we show therapeutic antibody ADG20 is able to neutralize all SARS-CoV-2 variants of concern (VOCs) including Omicron (B.1.1.529) as well as other SARS-related coronaviruses. We delineate the structural basis of this relatively escape-resistant epitope that extends from one end of the receptor binding site (RBS) into the highly conserved CR3022 site. ADG20 can then benefit from high potency through direct competition with ACE2 in the more variable RBS and interaction with the more highly conserved CR3022 site. Importantly, antibodies that are able to target this site generally neutralize all VOCs, albeit with reduced potency against Omicron. Thus, this highly conserved and vulnerable site can be exploited for design of universal vaccines and therapeutic antibodies.
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823. | Yan, Nicholas L; Nair, Reji; Chu, Alan; Wilson, Ian A; Johnson, Kristen A; Morgan, Gareth J; Kelly, Jeffery W: Amyloidogenic immunoglobulin light chain kinetic stabilizers comprising a simple urea linker module reveal a novel binding sub-site. In: Bioorg Med Chem Lett, vol. 60, pp. 128571, 2022, ISSN: 1464-3405. (Type: Journal Article | Abstract | Links)
@article{pmid35065233, In immunoglobulin light chain (LC) amyloidosis, the misfolding, or misfolding and misassembly of LC a protein or fragments thereof resulting from aberrant endoproteolysis, causes organ damage to patients. A small molecule "kinetic stabilizer" drug could slow or stop these processes and improve prognosis. We previously identified coumarin-based kinetic stabilizers of LCs that can be divided into four components, including a "linker module" and "distal substructure". Our prior studies focused on characterizing carbamate, hydantoin, and spirocyclic urea linker modules, which bind in a solvent-exposed site at the V-V domain interface of the LC dimer. Here, we report structure-activity relationship data on 7-diethylamino coumarin-based kinetic stabilizers. This substructure occupies the previously characterized "anchor cavity" and the "aromatic slit". The potencies of amide and urea linker modules terminating in a variety of distal substructures attached at the 3-position of this coumarin ring were assessed. Surprisingly, crystallographic data on a 7-diethylamino coumarin-based kinetic stabilizer reveals that the urea linker module and distal substructure attached at the 3-position bind a solvent-exposed region of the full-length LC dimer distinct from previously characterized sites. Our results further elaborate the small-molecule binding surface of LCs that could be occupied by potent and selective LC kinetic stabilizers.
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822. | Zhou, Panpan; Yuan, Meng; Song, Ge; Beutler, Nathan; Shaabani, Namir; Huang, Deli; He, Wan-Ting; Zhu, Xueyong; Callaghan, Sean; Yong, Peter; Anzanello, Fabio; Peng, Linghang; Ricketts, James; Parren, Mara; Garcia, Elijah; Rawlings, Stephen A; Smith, Davey M; Nemazee, David; Teijaro, John R; Rogers, Thomas F; Wilson, Ian A; Burton, Dennis R; Andrabi, Raiees: A human antibody reveals a conserved site on beta-coronavirus spike proteins and confers protection against SARS-CoV-2 infection. In: Sci Transl Med, vol. 14, no. 637, pp. eabi9215, 2022, ISSN: 1946-6242. (Type: Journal Article | Abstract | Links)
@article{pmid35133175, Broadly neutralizing antibodies (bnAbs) to coronaviruses (CoVs) are valuable in their own right as prophylactic and therapeutic reagents to treat diverse CoVs and as templates for rational pan-CoV vaccine design. We recently described a bnAb, CC40.8, from a CoV disease 2019 (COVID-19) convalescent donor that exhibits broad reactivity with human β-CoVs. Here, we showed that CC40.8 targets the conserved S2 stem helix region of the CoV spike fusion machinery. We determined a crystal structure of CC40.8 Fab with a SARS-CoV-2 S2 stem peptide at 1.6-Å resolution and found that the peptide adopted a mainly helical structure. Conserved residues in β-CoVs interacted with CC40.8 antibody, thereby providing a molecular basis for its broad reactivity. CC40.8 exhibited in vivo protective efficacy against SARS-CoV-2 challenge in two animal models. In both models, CC40.8-treated animals exhibited less weight loss and reduced lung viral titers compared to controls. Furthermore, we noted that CC40.8-like bnAbs are relatively rare in human COVID-19 infection, and therefore, their elicitation may require rational structure-based vaccine design strategies. Overall, our study describes a target on β-CoV spike proteins for protective antibodies that may facilitate the development of pan-β-CoV vaccines.
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821. | Beutler, Nathan; Pholcharee, Tossapol; Oyen, David; Flores-Garcia, Yevel; MacGill, Randall S; Garcia, Elijah; Calla, Jaeson; Parren, Mara; Yang, Linlin; Volkmuth, Wayne; Locke, Emily; Regules, Jason A; Dutta, Sheetij; Emerling, Daniel; Early, Angela M; Neafsey, Daniel E; Winzeler, Elizabeth A; King, C Richter; Zavala, Fidel; Burton, Dennis R; Wilson, Ian A; Rogers, Thomas F: A novel CSP C-terminal epitope targeted by an antibody with protective activity against Plasmodium falciparum. In: PLoS Pathog, vol. 18, no. 3, pp. e1010409, 2022, ISSN: 1553-7374. (Type: Journal Article | Abstract | Links)
@article{pmid35344575, Potent and durable vaccine responses will be required for control of malaria caused by Plasmodium falciparum (Pf). RTS,S/AS01 is the first, and to date, the only vaccine that has demonstrated significant reduction of clinical and severe malaria in endemic cohorts in Phase 3 trials. Although the vaccine is protective, efficacy declines over time with kinetics paralleling the decline in antibody responses to the Pf circumsporozoite protein (PfCSP). Although most attention has focused on antibodies to repeat motifs on PfCSP, antibodies to other regions may play a role in protection. Here, we expressed and characterized seven monoclonal antibodies to the C-terminal domain of CSP (ctCSP) from volunteers immunized with RTS,S/AS01. Competition and crystal structure studies indicated that the antibodies target two different sites on opposite faces of ctCSP. One site contains a polymorphic region (denoted α-ctCSP) and has been previously characterized, whereas the second is a previously undescribed site on the conserved β-sheet face of the ctCSP (denoted β-ctCSP). Antibodies to the β-ctCSP site exhibited broad reactivity with a diverse panel of ctCSP peptides whose sequences were derived from field isolates of P. falciparum whereas antibodies to the α-ctCSP site showed very limited cross reactivity. Importantly, an antibody to the β-site demonstrated inhibition activity against malaria infection in a murine model. This study identifies a previously unidentified conserved epitope on CSP that could be targeted by prophylactic antibodies and exploited in structure-based vaccine design.
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820. | He, Wan-Ting; Musharrafieh, Rami; Song, Ge; Dueker, Katharina; Tse, Longping V; Martinez, David R; Schäfer, Alexandra; Callaghan, Sean; Yong, Peter; Beutler, Nathan; Torres, Jonathan L; Volk, Reid M; Zhou, Panpan; Yuan, Meng; Liu, Hejun; Anzanello, Fabio; Capozzola, Tazio; Parren, Mara; Garcia, Elijah; Rawlings, Stephen A; Smith, Davey M; Wilson, Ian A; Safonova, Yana; Ward, Andrew B; Rogers, Thomas F; Baric, Ralph S; Gralinski, Lisa E; Burton, Dennis R; Andrabi, Raiees: Targeted isolation of panels of diverse human protective broadly neutralizing antibodies against SARS-like viruses. 2022. (Type: Miscellaneous | Abstract | Links)
@misc{pmid35169804, The emergence of current SARS-CoV-2 variants of concern (VOCs) and potential future spillovers of SARS-like coronaviruses into humans pose a major threat to human health and the global economy . Development of broadly effective coronavirus vaccines that can mitigate these threats is needed . Notably, several recent studies have revealed that vaccination of recovered COVID-19 donors results in enhanced nAb responses compared to SARS-CoV-2 infection or vaccination alone . Here, we utilized a targeted donor selection strategy to isolate a large panel of broadly neutralizing antibodies (bnAbs) to sarbecoviruses from two such donors. Many of the bnAbs are remarkably effective in neutralization against sarbecoviruses that use ACE2 for viral entry and a substantial fraction also show notable binding to non-ACE2-using sarbecoviruses. The bnAbs are equally effective against most SARS-CoV-2 VOCs and many neutralize the Omicron variant. Neutralization breadth is achieved by bnAb binding to epitopes on a relatively conserved face of the receptor binding domain (RBD) as opposed to strain-specific nAbs to the receptor binding site that are commonly elicited in SARS-CoV-2 infection and vaccination . Consistent with targeting of conserved sites, select RBD bnAbs exhibited protective efficacy against diverse SARS-like coronaviruses in a prophylaxis challenge model. The generation of a large panel of potent bnAbs provides new opportunities and choices for next-generation antibody prophylactic and therapeutic applications and, importantly, provides a molecular basis for effective design of pan-sarbecovirus vaccines.
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819. | Reincke, S Momsen; Yuan, Meng; Kornau, Hans-Christian; Corman, Victor M; van Hoof, Scott; Sánchez-Sendin, Elisa; Ramberger, Melanie; Yu, Wenli; Hua, Yuanzi; Tien, Henry; Schmidt, Marie Luisa; Schwarz, Tatjana; Jeworowski, Lara Maria; Brandl, Sarah E; Rasmussen, Helle Foverskov; Homeyer, Marie A; Stöffler, Laura; Barner, Martin; Kunkel, Désirée; Huo, Shufan; Horler, Johannes; von Wardenburg, Niels; Kroidl, Inge; Eser, Tabea M; Wieser, Andreas; Geldmacher, Christof; Hoelscher, Michael; Gänzer, Hannes; Weiss, Günter; Schmitz, Dietmar; Drosten, Christian; Prüss, Harald; Wilson, Ian A; Kreye, Jakob: SARS-CoV-2 Beta variant infection elicits potent lineage-specific and cross-reactive antibodies. In: Science, vol. 375, no. 6582, pp. 782–787, 2022, ISSN: 1095-9203. (Type: Journal Article | Abstract | Links)
@article{pmid35076281, Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Beta variant of concern (VOC) resists neutralization by major classes of antibodies from COVID-19 patients and vaccinated individuals. In this study, serum of Beta-infected patients revealed reduced cross-neutralization of wild-type virus. From these patients, we isolated Beta-specific and cross-reactive receptor-binding domain (RBD) antibodies. The Beta-specificity results from recruitment of VOC-specific clonotypes and accommodation of mutations present in Beta and Omicron into a major antibody class that is normally sensitive to these mutations. The Beta-elicited cross-reactive antibodies share genetic and structural features with wild type-elicited antibodies, including a public VH1-58 clonotype that targets the RBD ridge. These findings advance our understanding of the antibody response to SARS-CoV-2 shaped by antigenic drift, with implications for design of next-generation vaccines and therapeutics.
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818. | Qiang, Min; Ma, Peixiang; Li, Yu; Liu, Hejun; Harding, Adam; Min, Chenyu; Wang, Fulian; Liu, Lili; Yuan, Meng; Ji, Qun; Tao, Pingdong; Shi, Xiaojie; Li, Zhean; Li, Teng; Wang, Xian; Zhang, Yu; Wu, Nicholas C; Lee, Chang-Chun D; Zhu, Xueyong; Gilbert-Jaramillo, Javier; Zhang, Chuyue; Saxena, Abhishek; Huang, Xingxu; Wang, Hou; James, William; Dwek, Raymond A; Wilson, Ian A; Yang, Guang; Lerner, Richard A: Neutralizing Antibodies to SARS-CoV-2 Selected from a Human Antibody Library Constructed Decades Ago. In: Adv Sci (Weinh), vol. 9, no. 1, pp. e2102181, 2022, ISSN: 2198-3844. (Type: Journal Article | Abstract | Links)
@article{pmid34716683, Combinatorial antibody libraries not only effectively reduce antibody discovery to a numbers game, but enable documentation of the history of antibody responses in an individual. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has prompted a wider application of this technology to meet the public health challenge of pandemic threats in the modern era. Herein, a combinatorial human antibody library constructed 20 years before the coronavirus disease 2019 (COVID-19) pandemic is used to discover three highly potent antibodies that selectively bind SARS-CoV-2 spike protein and neutralize authentic SARS-CoV-2 virus. Compared to neutralizing antibodies from COVID-19 patients with generally low somatic hypermutation (SHM), these three antibodies contain over 13-22 SHMs, many of which are involved in specific interactions in their crystal structures with SARS-CoV-2 spike receptor binding domain. The identification of these somatically mutated antibodies in a pre-pandemic library raises intriguing questions about the origin and evolution of these antibodies with respect to their reactivity with SARS-CoV-2.
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817. | Zhou, Panpan; Yuan, Meng; Song, Ge; Beutler, Nathan; Shaabani, Namir; Huang, Deli; He, Wan-Ting; Zhu, Xueyong; Callaghan, Sean; Yong, Peter; Anzanello, Fabio; Peng, Linghang; Ricketts, James; Parren, Mara; Garcia, Elijah; Rawlings, Stephen A; Smith, Davey M; Nemazee, David; Teijaro, John R; Rogers, Thomas F; Wilson, Ian A; Burton, Dennis R; Andrabi, Raiees: A human antibody reveals a conserved site on beta-coronavirus spike proteins and confers protection against SARS-CoV-2 infection. 2022. (Type: Miscellaneous | Abstract | Links)
@misc{pmid33821273, Broadly neutralizing antibodies (bnAbs) to coronaviruses (CoVs) are valuable in their own right as prophylactic and therapeutic reagents to treat diverse CoVs and, importantly, as templates for rational pan-CoV vaccine design. We recently described a bnAb, CC40.8, from a coronavirus disease 2019 (COVID-19)-convalescent donor that exhibits broad reactivity with human beta-coronaviruses (β-CoVs). Here, we showed that CC40.8 targets the conserved S2 stem-helix region of the coronavirus spike fusion machinery. We determined a crystal structure of CC40.8 Fab with a SARS-CoV-2 S2 stem-peptide at 1.6 Å resolution and found that the peptide adopted a mainly helical structure. Conserved residues in β-CoVs interacted with CC40.8 antibody, thereby providing a molecular basis for its broad reactivity. CC40.8 exhibited in vivo protective efficacy against SARS-CoV-2 challenge in two animal models. In both models, CC40.8-treated animals exhibited less weight loss and reduced lung viral titers compared to controls. Furthermore, we noted CC40.8-like bnAbs are relatively rare in human COVID-19 infection and therefore their elicitation may require rational structure-based vaccine design strategies. Overall, our study describes a target on β-CoV spike proteins for protective antibodies that may facilitate the development of pan-β-CoV vaccines.nnSUMMARY: A human mAb isolated from a COVID-19 donor defines a protective cross-neutralizing epitope for pan-β-CoV vaccine design strategies.
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2021 |
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816. | Omorodion, Oluwarotimi; Wilson, Ian A: Structural and Biochemical Characterization of Cysteinylation in Broadly Neutralizing Antibodies to HIV-1. In: J Mol Biol, vol. 433, no. 24, pp. 167303, 2021, ISSN: 1089-8638. (Type: Journal Article | Abstract | Links)
@article{pmid34666044, Antibodies with exceptional breadth and potency have been elicited in some individuals during natural HIV-1 infection. Elicitation and affinity maturation of broadly neutralizing antibodies (bnAbs) is therefore the central goal of HIV-1 vaccine development. The functional properties of bnAbs also make them attractive as immunotherapeutic agents, which has led to their production and optimization for passive immunotherapy. This process requires in vitro manufacturing and monitoring of any heterogeneous expression, especially when subpopulations of antibodies are produced with varying levels of biological activity. Post-translational modification (PTM) of antibodies can contribute to heterogeneity and is the focus of this study. Specifically, we have investigated cysteinylation in a bnAb lineage (PCDN family) targeting the N332-glycan supersite on the surface envelope glycoprotein (Env) of HIV-1. This PTM is defined by capping of unpaired cysteine residues with molecular cysteine. Through chromatography and mass spectrometry, we were able to characterize subpopulations of cysteinylated and non-cysteinylated antibodies when expressed in mammalian cells. The crystal structures of two PCDN antibodies represent the first structures of a cysteinylated antibody and reveal that the cysteinylation in this case is located in CDRH3. Biophysical studies indicate that cysteinylation of these HIV-1 antibodies does not interfere with antigen binding, which has been reported to occur in other cysteinylated antibodies. As such, these studies highlight the need for further investigation of cysteinylation in anti-HIV and other bnAbs.
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815. | Strohmeier, Shirin; Amanat, Fatima; Zhu, Xueyong; McMahon, Meagan; Deming, Meagan E; Pasetti, Marcela F; Neuzil, Kathleen M; Wilson, Ian A; Krammer, Florian: A Novel Recombinant Influenza Virus Neuraminidase Vaccine Candidate Stabilized by a Measles Virus Phosphoprotein Tetramerization Domain Provides Robust Protection from Virus Challenge in the Mouse Model. In: mBio, vol. 12, no. 6, pp. e0224121, 2021, ISSN: 2150-7511. (Type: Journal Article | Abstract | Links)
@article{pmid34809451, Current seasonal influenza virus vaccines do not induce robust immune responses to neuraminidase. Several factors, including immunodominance of hemagglutinin over neuraminidase, instability of neuraminidase in vaccine formulations, and variable, nonstandardized amounts of neuraminidase in the vaccines, may contribute to this effect. However, vaccines that induce strong antineuraminidase immune responses would be beneficial, as they are highly protective. Furthermore, antigenic drift is slower for neuraminidase than for hemagglutinin, potentially providing broader coverage. Here, we designed stabilized recombinant versions of neuraminidase by replacing the N-terminal cytoplasmic domain, transmembrane, and extracellular stalk with tetramerization domains from the measles or Sendai virus phosphoprotein or from an Arabidopsis thaliana transcription factor. The measles virus tetramerization domain-based construct, termed N1-MPP, was chosen for further evaluation, as it retained antigenicity, neuraminidase activity, and structural integrity and provided robust protection against lethal virus challenge in the mouse model. We tested N1-MPP as a standalone vaccine, admixed with seasonal influenza virus vaccines, or given with seasonal influenza virus vaccines but in the other leg of the mouse. Admixture with different formulations of seasonal vaccines led to a weak neuraminidase response, suggesting a dominant effect of hemagglutinin over neuraminidase when administered in the same formulation. However, administration of neuraminidase alone or with seasonal vaccine administered in the alternate leg of the mouse induced robust antibody responses. Thus, this recombinant neuraminidase construct is a promising vaccine antigen that may enhance and broaden protection against seasonal influenza viruses. Influenza virus infections remain a high risk to human health, causing up to 650,000 deaths worldwide every year, with an enormous burden on the health care system. Since currently available seasonal vaccines are only partially effective and often mismatched to the circulating strains, a broader protective influenza virus vaccine is needed. Here, we generated a recombinant influenza virus vaccine candidate based on the more conserved neuraminidase surface glycoprotein in order to induce a robust and broader protective immune response against a variety of circulating influenza virus strains.
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814. | Yin, Victor; Lai, Szu-Hsueh; Caniels, Tom G; Brouwer, Philip J M; Brinkkemper, Mitch; Aldon, Yoann; Liu, Hejun; Yuan, Meng; Wilson, Ian A; Sanders, Rogier W; van Gils, Marit J; Heck, Albert J R: Probing Affinity, Avidity, Anticooperativity, and Competition in Antibody and Receptor Binding to the SARS-CoV-2 Spike by Single Particle Mass Analyses. In: ACS Cent Sci, vol. 7, no. 11, pp. 1863–1873, 2021, ISSN: 2374-7943. (Type: Journal Article | Abstract | Links)
@article{pmid34845440, Determining how antibodies interact with the spike (S) protein of the SARS-CoV-2 virus is critical for combating COVID-19. Structural studies typically employ simplified, truncated constructs that may not fully recapitulate the behavior of the original complexes. Here, we combine two single particle mass analysis techniques (mass photometry and charge-detection mass spectrometry) to enable the measurement of full IgG binding to the trimeric SARS-CoV-2 S ectodomain. Our experiments reveal that antibodies targeting the S-trimer typically prefer stoichiometries lower than the symmetry-predicted 3:1 binding. We determine that this behavior arises from the interplay of steric clashes and avidity effects that are not reflected in common antibody constructs (i.e., Fabs). Surprisingly, these substoichiometric complexes are fully effective at blocking ACE2 binding despite containing free receptor binding sites. Our results highlight the importance of studying antibody/antigen interactions using complete, multimeric constructs and showcase the utility of single particle mass analyses in unraveling these complex interactions.
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813. | Gebert, Luca F R; Law, Mansun; MacRae, Ian J: A structured RNA motif locks Argonaute2:miR-122 onto the 5' end of the HCV genome. In: Nat Commun, vol. 12, no. 1, pp. 6836, 2021, ISSN: 2041-1723. (Type: Journal Article | Abstract | Links)
@article{pmid34824224, microRNAs (miRNAs) form regulatory networks in metazoans. Viruses engage miRNA networks in numerous ways, with Flaviviridae members exploiting direct interactions of their RNA genomes with host miRNAs. For hepatitis C virus (HCV), binding of liver-abundant miR-122 stabilizes the viral RNA and regulates viral translation. Here, we investigate the structural basis for these activities, taking into consideration that miRNAs function in complex with Argonaute (Ago) proteins. The crystal structure of the Ago2:miR-122:HCV complex reveals a structured RNA motif that traps Ago2 on the viral RNA, masking its 5' end from enzymatic attack. The trapped Ago2 can recruit host factor PCBP2, implicated in viral translation, while binding of a second Ago2:miR-122 competes with PCBP2, creating a potential molecular switch for translational control. Combined results reveal a viral RNA structure that modulates Ago2:miR-122 dynamics and repurposes host proteins to generate a functional analog of the mRNA cap-binding complex.
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812. | Wang, Yiquan; Yuan, Meng; Peng, Jian; Wilson, Ian A; Wu, Nicholas C: A large-scale systematic survey of SARS-CoV-2 antibodies reveals recurring molecular features. 2021. (Type: Miscellaneous | Abstract | Links)
@misc{pmid34873599, In the past two years, the global research in combating COVID-19 pandemic has led to isolation and characterization of numerous human antibodies to the SARS-CoV-2 spike. This enormous collection of antibodies provides an unprecedented opportunity to study the antibody response to a single antigen. From mining information derived from 88 research publications and 13 patents, we have assembled a dataset of ∼8,000 human antibodies to the SARS-CoV-2 spike from >200 donors. Analysis of antibody targeting of different domains of the spike protein reveals a number of common (public) responses to SARS-CoV-2, exemplified via recurring IGHV/IGK(L)V pairs, CDR H3 sequences, IGHD usage, and somatic hypermutation. We further present a proof-of-concept for prediction of antigen specificity using deep learning to differentiate sequences of antibodies to SARS-CoV-2 spike and to influenza hemagglutinin. Overall, this study not only provides an informative resource for antibody and vaccine research, but fundamentally advances our molecular understanding of public antibody responses to a viral pathogen.
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811. | Maisonnasse, Pauline; Aldon, Yoann; Marc, Aurélien; Marlin, Romain; Dereuddre-Bosquet, Nathalie; Kuzmina, Natalia A; Freyn, Alec W; Snitselaar, Jonne L; Gonçalves, Antonio; Caniels, Tom G; Burger, Judith A; Poniman, Meliawati; Bontjer, Ilja; Chesnais, Virginie; Diry, Ségolène; Iershov, Anton; Ronk, Adam J; Jangra, Sonia; Rathnasinghe, Raveen; Brouwer, Philip J M; Bijl, Tom P L; van Schooten, Jelle; Brinkkemper, Mitch; Liu, Hejun; Yuan, Meng; Mire, Chad E; van Breemen, Mariëlle J; Contreras, Vanessa; Naninck, Thibaut; Lemaître, Julien; Kahlaoui, Nidhal; Relouzat, Francis; Chapon, Catherine; Fang, Raphaël Ho Tsong; McDanal, Charlene; Osei-Twum, Mary; St-Amant, Natalie; Gagnon, Luc; Montefiori, David C; Wilson, Ian A; Ginoux, Eric; de Bree, Godelieve J; García-Sastre, Adolfo; Schotsaert, Michael; Coughlan, Lynda; Bukreyev, Alexander; van der Werf, Sylvie; Guedj, Jérémie; Sanders, Rogier W; van Gils, Marit J; Grand, Roger Le: COVA1-18 neutralizing antibody protects against SARS-CoV-2 in three preclinical models. In: Nat Commun, vol. 12, no. 1, pp. 6097, 2021, ISSN: 2041-1723. (Type: Journal Article | Abstract | Links)
@article{pmid34671037, Effective treatments against Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) are urgently needed. Monoclonal antibodies have shown promising results in patients. Here, we evaluate the in vivo prophylactic and therapeutic effect of COVA1-18, a neutralizing antibody highly potent against the B.1.1.7 isolate. In both prophylactic and therapeutic settings, SARS-CoV-2 remains undetectable in the lungs of treated hACE2 mice. Therapeutic treatment also causes a reduction in viral loads in the lungs of Syrian hamsters. When administered at 10 mg kg-1 one day prior to a high dose SARS-CoV-2 challenge in cynomolgus macaques, COVA1-18 shows very strong antiviral activity in the upper respiratory compartments. Using a mathematical model, we estimate that COVA1-18 reduces viral infectivity by more than 95% in these compartments, preventing lymphopenia and extensive lung lesions. Our findings demonstrate that COVA1-18 has a strong antiviral activity in three preclinical models and could be a valuable candidate for further clinical evaluation.
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810. | Cho, Hyeseon; Gonzales-Wartz, Kristina Kay; Huang, Deli; Yuan, Meng; Peterson, Mary; Liang, Janie; Beutler, Nathan; Torres, Jonathan L; Cong, Yu; Postnikova, Elena; Bangaru, Sandhya; Talana, Chloe Adrienna; Shi, Wei; Yang, Eun Sung; Zhang, Yi; Leung, Kwanyee; Wang, Lingshu; Peng, Linghang; Skinner, Jeff; Li, Shanping; Wu, Nicholas C; Liu, Hejun; Dacon, Cherrelle; Moyer, Thomas; Cohen, Melanie; Zhao, Ming; Lee, Frances Eun-Hyung; Weinberg, Rona S; Douagi, Iyadh; Gross, Robin; Schmaljohn, Connie; Pegu, Amarendra; Mascola, John R; Holbrook, Michael; Nemazee, David; Rogers, Thomas F; Ward, Andrew B; Wilson, Ian A; Crompton, Peter D; Tan, Joshua: Bispecific antibodies targeting distinct regions of the spike protein potently neutralize SARS-CoV-2 variants of concern. In: Sci Transl Med, vol. 13, no. 616, pp. eabj5413, 2021, ISSN: 1946-6242. (Type: Journal Article | Abstract | Links)
@article{pmid34519517, The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern threatens the efficacy of existing vaccines and therapeutic antibodies and underscores the need for additional antibody-based tools that potently neutralize variants by targeting multiple sites of the spike protein. We isolated 216 monoclonal antibodies targeting SARS-CoV-2 from plasmablasts and memory B cells collected from patients with coronavirus disease 2019. The three most potent antibodies targeted distinct regions of the receptor binding domain (RBD), and all three neutralized the SARS-CoV-2 Alpha and Beta variants. The crystal structure of the most potent antibody, CV503, revealed that it binds to the ridge region of SARS-CoV-2 RBD, competes with the angiotensin-converting enzyme 2 receptor, and has limited contact with key variant residues K417, E484, and N501. We designed bispecific antibodies by combining nonoverlapping specificities and identified five bispecific antibodies that inhibit SARS-CoV-2 infection at concentrations of less than 1 ng/ml. Through a distinct mode of action, three bispecific antibodies cross-linked adjacent spike proteins using dual N-terminal domain–RBD specificities. One bispecific antibody was greater than 100-fold more potent than a cocktail of its parent monoclonals in vitro and prevented clinical disease in a hamster model at a dose of 2.5 mg/kg. Two bispecific antibodies in our panel comparably neutralized the Alpha, Beta, Gamma, and Delta variants and wild-type virus. Furthermore, a bispecific antibody that neutralized the Beta variant protected hamsters against SARS-CoV-2 expressing the E484K mutation. Thus, bispecific antibodies represent a promising next-generation countermeasure against SARS-CoV-2 variants of concern.
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809. | Lv, Huibin; Tsang, Owen Tak-Yin; So, Ray T Y; Wang, Yiquan; Yuan, Meng; Liu, Hejun; Yip, Garrick K; Teo, Qi Wen; Lin, Yihan; Liang, Weiwen; Wang, Jinlin; Ng, Wilson W; Wilson, Ian A; Peiris, J S Malik; Wu, Nicholas C; Mok, Chris K P: Homologous and heterologous serological response to the N-terminal domain of SARS-CoV-2 in humans and mice. In: Eur J Immunol, vol. 51, no. 9, pp. 2296–2305, 2021, ISSN: 1521-4141. (Type: Journal Article | Abstract | Links)
@article{pmid34089541, The increasing numbers of infected cases of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) poses serious threats to public health and the global economy. Most SARS-CoV-2 neutralizing antibodies target the receptor binding domain (RBD) and some the N-terminal domain (NTD) of the spike protein, which is the major antigen of SARS-CoV-2. While the antibody response to RBD has been extensively characterized, the antigenicity and immunogenicity of the NTD protein are less well studied. Using 227 plasma samples from COVID-19 patients, we showed that SARS-CoV-2 NTD-specific antibodies could be induced during infection. As compared to the results of SARS-CoV-2 RBD, the serological response of SARS-CoV-2 NTD is less cross-reactive with SARS-CoV, a pandemic strain that was identified in 2003. Furthermore, neutralizing antibodies are rarely elicited in a mice model when NTD is used as an immunogen. We subsequently demonstrate that NTD has an altered antigenicity when expressed alone. Overall, our results suggest that while NTD offers a supplementary strategy for serology testing, it may not be suitable as an immunogen for vaccine development.
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808. | Yuan, Meng; Huang, Deli; Lee, Chang-Chun D; Wu, Nicholas C; Jackson, Abigail M; Zhu, Xueyong; Liu, Hejun; Peng, Linghang; van Gils, Marit J; Sanders, Rogier W; Burton, Dennis R; Reincke, S Momsen; Prüss, Harald; Kreye, Jakob; Nemazee, David; Ward, Andrew B; Wilson, Ian A: Structural and functional ramifications of antigenic drift in recent SARS-CoV-2 variants. In: Science, vol. 373, no. 6556, pp. 818–823, 2021, ISSN: 1095-9203. (Type: Journal Article | Abstract | Links)
@article{pmid34016740, Neutralizing antibodies (nAbs) elicited against the receptor binding site (RBS) of the spike protein of wild-type severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are generally less effective against recent variants of concern. RBS residues Glu, Lys, and Asn are mutated in variants first described in South Africa (B.1.351) and Brazil (P.1). We analyzed their effects on angiotensin-converting enzyme 2 binding, as well as the effects of two of these mutations (K417N and E484K) on nAbs isolated from COVID-19 patients. Binding and neutralization of the two most frequently elicited antibody families (IGHV3-53/3-66 and IGHV1-2), which can both bind the RBS in alternative binding modes, are abrogated by K417N, E484K, or both. These effects can be structurally explained by their extensive interactions with RBS nAbs. However, nAbs to the more conserved, cross-neutralizing CR3022 and S309 sites were largely unaffected. The results have implications for next-generation vaccines and antibody therapies.
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807. | Schoenle, Marta V; Li, Yang; Yuan, Meng; Clarkson, Michael W; Wilson, Ian A; Peti, Wolfgang; Page, Rebecca: NMR Based SARS-CoV-2 Antibody Screening. In: J Am Chem Soc, vol. 143, no. 21, pp. 7930–7934, 2021, ISSN: 1520-5126. (Type: Journal Article | Abstract | Links)
@article{pmid34018723, The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry into cells is a complex process that involves (1) recognition of the host entry receptor, angiotensin-converting enzyme 2 (ACE2), by the SARS-CoV-2 spike protein receptor binding domain (RBD), and (2) the subsequent fusion of the viral and cell membranes. Our long-term immune-defense is the production of antibodies (Abs) that recognize the SARS-CoV-2 RBD and successfully block viral infection. Thus, to understand immunity against SARS-CoV-2, a comprehensive molecular understanding of how human SARS-CoV-2 Abs recognize the RBD is needed. Here, we report the sequence-specific backbone assignment of the SARS-CoV-2 RBD and, furthermore, demonstrate that biomolecular NMR spectroscopy chemical shift perturbation (CSP) mapping successfully and rapidly identifies the molecular epitopes of RBD-specific mAbs. By incorporating NMR-based CSP mapping with other molecular techniques to define RBD-mAb interactions and then correlating these data with neutralization efficacy, structure-based approaches for developing improved vaccines and COVID-19 mAb-based therapies will be greatly accelerated.
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806. | McKitrick, Tanya R; Bernard, Steffen M; Noll, Alexander J; Collins, Bernard C; Goth, Christoffer K; McQuillan, Alyssa M; Heimburg-Molinaro, Jamie; Herrin, Brantley R; Wilson, Ian A; Cooper, Max D; Cummings, Richard D: Novel lamprey antibody recognizes terminal sulfated galactose epitopes on mammalian glycoproteins. In: Commun Biol, vol. 4, no. 1, pp. 674, 2021, ISSN: 2399-3642. (Type: Journal Article | Abstract | Links)
@article{pmid34083726, The terminal galactose residues of N- and O-glycans in animal glycoproteins are often sialylated and/or fucosylated, but sulfation, such as 3-O-sulfated galactose (3-O-SGal), represents an additional, but poorly understood modification. To this end, we have developed a novel sea lamprey variable lymphocyte receptor (VLR) termed O6 to explore 3-O-SGal expression. O6 was engineered as a recombinant murine IgG chimera and its specificity and affinity to the 3-O-SGal epitope was defined using a variety of approaches, including glycan and glycoprotein microarray analyses, isothermal calorimetry, ligand-bound crystal structure, FACS, and immunohistochemistry of human tissue macroarrays. 3-O-SGal is expressed on N-glycans of many plasma and tissue glycoproteins, but recognition by O6 is often masked by sialic acid and thus exposed by treatment with neuraminidase. O6 recognizes many human tissues, consistent with expression of the cognate sulfotransferases (GAL3ST-2 and GAL3ST-3). The availability of O6 for exploring 3-O-SGal expression could lead to new biomarkers for disease and aid in understanding the functional roles of terminal modifications of glycans and relationships between terminal sulfation, sialylation and fucosylation.
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805. | Tan, Timothy J C; Yuan, Meng; Kuzelka, Kaylee; Padron, Gilberto C; Beal, Jacob R; Chen, Xin; Wang, Yiquan; Rivera-Cardona, Joel; Zhu, Xueyong; Stadtmueller, Beth M; Brooke, Christopher B; Wilson, Ian A; Wu, Nicholas C: Sequence signatures of two public antibody clonotypes that bind SARS-CoV-2 receptor binding domain. In: Nat Commun, vol. 12, no. 1, pp. 3815, 2021, ISSN: 2041-1723. (Type: Journal Article | Abstract | Links)
@article{pmid34155209, Since the COVID-19 pandemic onset, the antibody response to SARS-CoV-2 has been extensively characterized. Antibodies to the receptor binding domain (RBD) on the spike protein are frequently encoded by IGHV3-53/3-66 with a short complementarity-determining region (CDR) H3. Germline-encoded sequence motifs in heavy chain CDRs H1 and H2 have a major function, but whether any common motifs are present in CDR H3, which is often critical for binding specificity, is not clear. Here, we identify two public clonotypes of IGHV3-53/3-66 RBD antibodies with a 9-residue CDR H3 that pair with different light chains. Distinct sequence motifs on CDR H3 are present in the two public clonotypes that seem to be related to differential light chain pairing. Additionally, we show that Y58F is a common somatic hypermutation that results in increased binding affinity of IGHV3-53/3-66 RBD antibodies with a short CDR H3. These results advance understanding of the antibody response to SARS-CoV-2.
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804. | Tan, Joshua; Cho, Hyeseon; Pholcharee, Tossapol; Pereira, Lais S; Doumbo, Safiatou; Doumtabe, Didier; Flynn, Barbara J; Schön, Arne; Kanatani, Sachie; Aylor, Samantha O; Oyen, David; Vistein, Rachel; Wang, Lawrence; Dillon, Marlon; Skinner, Jeff; Peterson, Mary; Li, Shanping; Idris, Azza H; Molina-Cruz, Alvaro; Zhao, Ming; Olano, Lisa Renee; Lee, Patricia J; Roth, Alison; Sinnis, Photini; Barillas-Mury, Carolina; Kayentao, Kassoum; Ongoiba, Aissata; Francica, Joseph R; Traore, Boubacar; Wilson, Ian A; Seder, Robert A; Crompton, Peter D: Functional human IgA targets a conserved site on malaria sporozoites. In: Sci Transl Med, vol. 13, no. 599, 2021, ISSN: 1946-6242. (Type: Journal Article | Abstract | Links)
@article{pmid34162751, Immunoglobulin (Ig)A antibodies play a critical role in protection against mucosal pathogens. However, the role of serum IgA in immunity to nonmucosal pathogens, such as , is poorly characterized, despite being the second most abundant isotype in blood after IgG. Here, we investigated the circulating IgA response in humans to sporozoites that are injected into the skin by mosquitoes and migrate to the liver via the bloodstream to initiate malaria infection. We found that circulating IgA was induced in three independent sporozoite-exposed cohorts: individuals living in an endemic region in Mali, malaria-naïve individuals immunized intravenously with three large doses of irradiated sporozoites, and malaria-naïve individuals exposed to a single controlled mosquito bite infection. Mechanistically, we found evidence in an animal model that IgA responses were induced by sporozoites at dermal inoculation sites. From malaria-resistant individuals, we isolated several IgA monoclonal antibodies that reduced liver parasite burden in mice. One antibody, MAD2-6, bound to a conserved epitope in the amino terminus of the circumsporozoite protein, the dominant protein on the sporozoite surface. Crystal structures of this antibody revealed a unique mode of binding whereby two Fabs simultaneously bound either side of the target peptide. This study reveals a role for circulating IgA in malaria and identifies the amino terminus of the circumsporozoite protein as a target of functional antibodies.
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803. | Kumar, Sonu; Lin, Xiaohe; Ngo, Timothy; Shapero, Benjamin; Sou, Cindy; Allen, Joel D; Copps, Jeffrey; Zhang, Lei; Ozorowski, Gabriel; He, Linling; Crispin, Max; Ward, Andrew B; Wilson, Ian A; Zhu, Jiang: Neutralizing Antibodies Induced by First-Generation gp41-Stabilized HIV-1 Envelope Trimers and Nanoparticles. In: mBio, vol. 12, no. 3, pp. e0042921, 2021, ISSN: 2150-7511. (Type: Journal Article | Abstract | Links)
@article{pmid34156262, The immunogenicity of gp41-stabilized HIV-1 BG505 envelope (Env) trimers and nanoparticles (NPs) was recently assessed in mice and rabbits. Here, we combined Env-specific B-cell sorting and repertoire sequencing to identify neutralizing antibodies (NAbs) from immunized animals. A panel of mouse NAbs was isolated from mice immunized with a 60-meric I3-01 NP presenting 20 stabilized trimers. Three mouse NAbs potently neutralized BG505.T332N by recognizing a glycan epitope centered in the C3/V4 region on BG505 Env, as revealed by electron microscopy (EM), X-ray crystallography, and epitope mapping. A set of rabbit NAbs was isolated from rabbits immunized with a soluble trimer and a 24-meric ferritin NP presenting 8 trimers. Neutralization assays against BG505.T332N variants confirmed that potent rabbit NAbs targeted previously described glycan holes on BG505 Env and accounted for a significant portion of the autologous NAb response in both the trimer and ferritin NP groups. Last, we examined NAb responses that were induced by non-BG505 Env immunogens. We determined a 3.4-Å-resolution crystal structure for the clade C transmitted/founder (T/F) Du172.17 Env with a redesigned heptad repeat 1 (HR1) bend in gp41. This clade C Env, in a soluble trimer form and in a multivalent form with 8 trimers attached to ferritin NP, and the gp41-stabilized clade A Q482-d12 Env trimer elicited distinct NAb responses in rabbits, with notable differences in neutralization breadth. Although eliciting a broad NAb response remains a major challenge, our study provides valuable information on an HIV-1 vaccine design strategy that combines gp41 stabilization and NP display. Self-assembling protein nanoparticles (NPs) presenting BG505 envelope (Env) trimers can elicit tier 2 HIV-1-neutralizing antibody (NAb) responses more effectively than soluble trimers. In the present study, monoclonal NAbs were isolated from previously immunized mice and rabbits for structural and functional analyses, which revealed that potent mouse NAbs recognize the C3/V4 region and small NP-elicited rabbit NAbs primarily target known glycan holes on BG505 Env. This study validates the gp41 stabilization strategy for HIV-1 Env vaccine design and highlights the challenge in eliciting a broad NAb response.
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802. | Yechezkel, Itai; Law, Mansun; Tzarum, Netanel: From Structural Studies to HCV Vaccine Design. In: Viruses, vol. 13, no. 5, 2021, ISSN: 1999-4915. (Type: Journal Article | Abstract | Links)
@article{pmid34064532, Hepatitis C virus (HCV) is a serious and growing public health problem despite recent developments of antiviral therapeutics. To achieve global elimination of HCV, an effective cross-genotype vaccine is needed. The failure of previous vaccination trials to elicit an effective cross-reactive immune response demands better vaccine antigens to induce a potent cross-neutralizing response to improve vaccine efficacy. HCV E1 and E2 envelope (Env) glycoproteins are the main targets for neutralizing antibodies (nAbs), which aid in HCV clearance and protection. Therefore, a molecular-level understanding of the nAb responses against HCV is imperative for the rational design of cross-genotype vaccine antigens. Here we summarize the recent advances in structural studies of HCV Env and Env-nAb complexes and how they improve our understanding of immune recognition of HCV. We review the structural data defining HCV neutralization epitopes and conformational plasticity of the Env proteins, and the knowledge applicable to rational vaccine design.
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801. | Shi, Xiaojie; Wan, Yue; Wang, Nan; Xiang, Jiangchao; Wang, Tao; Yang, Xiaofeng; Wang, Ju; Dong, Xuxue; Dong, Liang; Yan, Lei; Li, Yu; Liu, Lili; Hou, Shinchen; Zhong, Zhenwei; Wilson, Ian A; Yang, Bei; Yang, Guang; Lerner, Richard A: Selection of a picomolar antibody that targets CXCR2-mediated neutrophil activation and alleviates EAE symptoms. In: Nat Commun, vol. 12, no. 1, pp. 2547, 2021, ISSN: 2041-1723. (Type: Journal Article | Abstract | Links)
@article{pmid33953162, Receptors and their ligands are important therapeutic targets for about one third of marketed drugs. Here, we describe an epitope-guided approach for selection of antibodies that modulate cellular signaling of targeted receptors. We chose CXC chemokine receptor 2 (CXCR2) in the G-protein coupled receptor superfamily as receptor and a CXCR2 N-terminal peptide for antibody selection. We obtain a highly selective, tight-binding antibody from a 10-member antibody library using combinatorial enrichment. Structural and Hydrogen-Deuterium-Exchange mass spectrometry analyses demonstrate antibody interaction with an N-terminal region of CXCR2 that is part of the IL-8 epitope. The antibody strongly inhibits IL-8-induced and CXCR2-mediated neutrophil chemotaxis in vitro and alleviates hCXCR2-dependent experimental autoimmune encephalomyelitis symptoms in mice. As inappropriate neutrophil migration accompanies many diseases including inflammatory bowel disease, glomerulonephritis, allergic asthma, chronic obstructive pulmonary disease, and cancer, this antibody has potential for development as a therapeutic agent, akin to anti-TNF antibodies. However, an important difference here is that the antibody targets the chemokine receptor and competes with natural ligand, rather than targeting the ligand itself.
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800. | He, Linling; Chaudhary, Anshul; Lin, Xiaohe; Sou, Cindy; Alkutkar, Tanwee; Kumar, Sonu; Ngo, Timothy; Kosviner, Ezra; Ozorowski, Gabriel; Stanfield, Robyn L; Ward, Andrew B; Wilson, Ian A; Zhu, Jiang: Single-component multilayered self-assembling nanoparticles presenting rationally designed glycoprotein trimers as Ebola virus vaccines. In: Nat Commun, vol. 12, no. 1, pp. 2633, 2021, ISSN: 2041-1723. (Type: Journal Article | Abstract | Links)
@article{pmid33976149, Ebola virus (EBOV) glycoprotein (GP) can be recognized by neutralizing antibodies (NAbs) and is the main target for vaccine design. Here, we first investigate the contribution of the stalk and heptad repeat 1-C (HR1) regions to GP metastability. Specific stalk and HR1 modifications in a mucin-deleted form (GPΔmuc) increase trimer yield, whereas alterations of HR1 exert a more complex effect on thermostability. Crystal structures are determined to validate two rationally designed GPΔmuc trimers in their unliganded state. We then display a modified GPΔmuc trimer on reengineered protein nanoparticles that encapsulate a layer of locking domains (LD) and a cluster of helper T-cell epitopes. In mice and rabbits, GP trimers and nanoparticles elicit cross-ebolavirus NAbs, as well as non-NAbs that enhance pseudovirus infection. Repertoire sequencing reveals quantitative profiles of vaccine-induced B-cell responses. This study demonstrates a promising vaccine strategy for filoviruses, such as EBOV, based on GP stabilization and nanoparticle display.
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799. | Zhou, Xiaojuan; Ma, Fengge; Xie, Jun; Yuan, Meng; Li, Yunqiao; Shaabani, Namir; Zhao, Fangzhu; Huang, Deli; Wu, Nicholas C; Lee, Chang-Chun D; Liu, Hejun; Li, Jiali; Chen, Zhonghui; Hong, Yazhen; Liu, Wen-Hsien; Xiao, Nengming; Burton, Dennis R; Tu, Haijian; Li, Hang; Chen, Xin; Teijaro, John R; Wilson, Ian A; Xiao, Changchun; Huang, Zhe: Diverse immunoglobulin gene usage and convergent epitope targeting in neutralizing antibody responses to SARS-CoV-2. In: Cell Rep, vol. 35, no. 6, pp. 109109, 2021, ISSN: 2211-1247. (Type: Journal Article | Abstract | Links)
@article{pmid33932326, It is unclear whether individuals with enormous diversity in B cell receptor repertoires are consistently able to mount effective antibody responses against SARS-CoV-2. We analyzed antibody responses in a cohort of 55 convalescent patients and isolated 54 potent neutralizing monoclonal antibodies (mAbs). While most of the mAbs target the angiotensin-converting enzyme 2 (ACE2) binding surface on the receptor binding domain (RBD) of SARS-CoV-2 spike protein, mAb 47D1 binds only to one side of the receptor binding surface on the RBD. Neutralization by 47D1 is achieved independent of interfering RBD-ACE2 binding. A crystal structure of the mAb-RBD complex shows that the IF motif at the tip of 47D1 CDR H2 interacts with a hydrophobic pocket in the RBD. Diverse immunoglobulin gene usage and convergent epitope targeting characterize neutralizing antibody responses to SARS-CoV-2, suggesting that vaccines that effectively present the receptor binding site on the RBD will likely elicit neutralizing antibody responses in a large fraction of the population.
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798. | Liu, Hejun; Yuan, Meng; Huang, Deli; Bangaru, Sandhya; Zhao, Fangzhu; Lee, Chang-Chun D; Peng, Linghang; Barman, Shawn; Zhu, Xueyong; Nemazee, David; Burton, Dennis R; van Gils, Marit J; Sanders, Rogier W; Kornau, Hans-Christian; Reincke, S Momsen; Prüss, Harald; Kreye, Jakob; Wu, Nicholas C; Ward, Andrew B; Wilson, Ian A: A combination of cross-neutralizing antibodies synergizes to prevent SARS-CoV-2 and SARS-CoV pseudovirus infection. In: Cell Host Microbe, vol. 29, no. 5, pp. 806–818.e6, 2021, ISSN: 1934-6069. (Type: Journal Article | Abstract | Links)
@article{pmid33894127, Coronaviruses have caused several human epidemics and pandemics including the ongoing coronavirus disease 2019 (COVID-19). Prophylactic vaccines and therapeutic antibodies have already shown striking effectiveness against COVID-19. Nevertheless, concerns remain about antigenic drift in SARS-CoV-2 as well as threats from other sarbecoviruses. Cross-neutralizing antibodies to SARS-related viruses provide opportunities to address such concerns. Here, we report on crystal structures of a cross-neutralizing antibody, CV38-142, in complex with the receptor-binding domains from SARS-CoV-2 and SARS-CoV. Recognition of the N343 glycosylation site and water-mediated interactions facilitate cross-reactivity of CV38-142 to SARS-related viruses, allowing the antibody to accommodate antigenic variation in these viruses. CV38-142 synergizes with other cross-neutralizing antibodies, notably COVA1-16, to enhance neutralization of SARS-CoV and SARS-CoV-2, including circulating variants of concern B.1.1.7 and B.1.351. Overall, this study provides valuable information for vaccine and therapeutic design to address current and future antigenic drift in SARS-CoV-2 and to protect against zoonotic SARS-related coronaviruses.
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797. | Yan, Nicholas L; Santos-Martins, Diogo; Nair, Reji; Chu, Alan; Wilson, Ian A; Johnson, Kristen A; Forli, Stefano; Morgan, Gareth J; Petrassi, H Michael; Kelly, Jeffery W: Discovery of Potent Coumarin-Based Kinetic Stabilizers of Amyloidogenic Immunoglobulin Light Chains Using Structure-Based Design. In: J Med Chem, vol. 64, no. 9, pp. 6273–6299, 2021, ISSN: 1520-4804. (Type: Journal Article | Abstract | Links)
@article{pmid33939422, In immunoglobulin light-chain (LC) amyloidosis, transient unfolding or unfolding and proteolysis enable aggregation of LC proteins, causing potentially fatal organ damage. A drug that kinetically stabilizes LCs could suppress aggregation; however, LC sequences are variable and have no natural ligands, hindering drug development efforts. We previously identified high-throughput screening hits that bind to a site at the interface between the two variable domains of the LC homodimer. We hypothesized that extending the stabilizers beyond this initially characterized binding site would improve affinity. Here, using protease sensitivity assays, we identified stabilizers that can be divided into four substructures. Some stabilizers exhibit nanomolar EC values, a 3000-fold enhancement over the screening hits. Crystal structures reveal a key π-π stacking interaction with a conserved tyrosine residue that was not utilized by the screening hits. These data provide a foundation for developing LC stabilizers with improved binding selectivity and enhanced physicochemical properties.
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796. | Montague, Zachary; Lv, Huibin; Otwinowski, Jakub; DeWitt, William S; Isacchini, Giulio; Yip, Garrick K; Ng, Wilson W; Tsang, Owen Tak-Yin; Yuan, Meng; Liu, Hejun; Wilson, Ian A; Peiris, J S Malik; Wu, Nicholas C; Nourmohammad, Armita; Mok, Chris Ka Pun: Dynamics of B cell repertoires and emergence of cross-reactive responses in patients with different severities of COVID-19. In: Cell Rep, vol. 35, no. 8, pp. 109173, 2021, ISSN: 2211-1247. (Type: Journal Article | Abstract | Links)
@article{pmid33991510, Individuals with the 2019 coronavirus disease (COVID-19) show varying severity of the disease, ranging from asymptomatic to requiring intensive care. Although monoclonal antibodies specific to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been identified, we still lack an understanding of the overall landscape of B cell receptor (BCR) repertoires in individuals with COVID-19. We use high-throughput sequencing of bulk and plasma B cells collected at multiple time points during infection to characterize signatures of the B cell response to SARS-CoV-2 in 19 individuals. Using principled statistical approaches, we associate differential features of BCRs with different disease severity. We identify 38 significantly expanded clonal lineages shared among individuals as candidates for responses specific to SARS-CoV-2. Using single-cell sequencing, we verify the reactivity of BCRs shared among individuals to SARS-CoV-2 epitopes. Moreover, we identify the natural emergence of a BCR with cross-reactivity to SARS-CoV-1 and SARS-CoV-2 in some individuals. Our results provide insights important for development of rational therapies and vaccines against COVID-19.
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795. | Cho, Hyeseon; Gonzales-Wartz, Kristina Kay; Huang, Deli; Yuan, Meng; Peterson, Mary; Liang, Janie; Beutler, Nathan; Torres, Jonathan L; Cong, Yu; Postnikova, Elena; Bangaru, Sandhya; Talana, Chloe Adrienna; Shi, Wei; Yang, Eun Sung; Zhang, Yi; Leung, Kwanyee; Wang, Lingshu; Peng, Linghang; Skinner, Jeff; Li, Shanping; Wu, Nicholas C; Liu, Hejun; Dacon, Cherrelle; Moyer, Thomas; Cohen, Melanie; Zhao, Ming; Lee, F Eun-Hyung; Weinberg, Rona S; Douagi, Iyadh; Gross, Robin; Schmaljohn, Connie; Pegu, Amarendra; Mascola, John R; Holbrook, Michael; Nemazee, David; Rogers, Thomas F; Ward, Andrew B; Wilson, Ian A; Crompton, Peter D; Tan, Joshua: Ultrapotent bispecific antibodies neutralize emerging SARS-CoV-2 variants. 2021. (Type: Miscellaneous | Abstract | Links)
@misc{pmid33821267, The emergence of SARS-CoV-2 variants that threaten the efficacy of existing vaccines and therapeutic antibodies underscores the urgent need for new antibody-based tools that potently neutralize variants by targeting multiple sites of the spike protein. We isolated 216 monoclonal antibodies targeting SARS-CoV-2 from plasmablasts and memory B cells of COVID-19 patients. The three most potent antibodies targeted distinct regions of the RBD, and all three neutralized the SARS-CoV-2 variants B.1.1.7 and B.1.351. The crystal structure of the most potent antibody, CV503, revealed that it binds to the ridge region of SARS-CoV-2 RBD, competes with the ACE2 receptor, and has limited contact with key variant residues K417, E484 and N501. We designed bispecific antibodies by combining non-overlapping specificities and identified five ultrapotent bispecific antibodies that inhibit authentic SARS-CoV-2 infection at concentrations of <1 ng/mL. Through a novel mode of action three bispecific antibodies cross-linked adjacent spike proteins using dual NTD/RBD specificities. One bispecific antibody was >100-fold more potent than a cocktail of its parent monoclonals and prevented clinical disease in a hamster model at a 2.5 mg/kg dose. Notably, six of nine bispecific antibodies neutralized B.1.1.7, B.1.351 and the wild-type virus with comparable potency, despite partial or complete loss of activity of at least one parent monoclonal antibody against B.1.351. Furthermore, a bispecific antibody that neutralized B.1.351 protected against SARS-CoV-2 expressing the crucial E484K mutation in the hamster model. Thus, bispecific antibodies represent a promising next-generation countermeasure against SARS-CoV-2 variants of concern.
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794. | He, Linling; Lin, Xiaohe; Wang, Ying; Abraham, Ciril; Sou, Cindy; Ngo, Timothy; Zhang, Yi; Wilson, Ian A; Zhu, Jiang: Single-component, self-assembling, protein nanoparticles presenting the receptor binding domain and stabilized spike as SARS-CoV-2 vaccine candidates. 2021. (Type: Miscellaneous | Abstract | Links)
@misc{pmid32995773, Vaccination against SARS-CoV-2 provides an effective tool to combat the COIVD-19 pandemic. Here, we combined antigen optimization and nanoparticle display to develop vaccine candidates for SARS-CoV-2. We first displayed the receptor-binding domain (RBD) on three self-assembling protein nanoparticle (SApNP) platforms using the SpyTag/SpyCatcher system. We then identified heptad repeat 2 (HR2) in S2 as the cause of spike metastability, designed an HR2-deleted glycine-capped spike (S2GΔHR2), and displayed S2GΔHR2 on SApNPs. An antibody column specific for the RBD enabled tag-free vaccine purification. In mice, the 24-meric RBD-ferritin SApNP elicited a more potent neutralizing antibody (NAb) response than the RBD alone and the spike with two stabilizing proline mutations in S2 (S2P). S2GΔHR2 elicited two-fold-higher NAb titers than S2P, while S2GΔHR2 SApNPs derived from multilayered E2p and I3-01v9 60-mers elicited up to 10-fold higher NAb titers. The S2GΔHR2-presenting I3-01v9 SApNP also induced critically needed T-cell immunity, thereby providing a promising vaccine candidate.
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793. | Montague, Zachary; Lv, Huibin; Otwinowski, Jakub; DeWitt, William S; Isacchini, Giulio; Yip, Garrick K; Ng, Wilson W; Tsang, Owen Tak-Yin; Yuan, Meng; Liu, Hejun; Wilson, Ian A; Peiris, J S Malik; Wu, Nicholas C; Nourmohammad, Armita; Mok, Chris Ka Pun: Dynamics of B-cell repertoires and emergence of cross-reactive responses in COVID-19 patients with different disease severity. 2021. (Type: Miscellaneous | Abstract | Links)
@misc{pmid32699862, COVID-19 patients show varying severity of the disease ranging from asymptomatic to requiring intensive care. Although a number of SARS-CoV-2 specific monoclonal antibodies have been identified, we still lack an understanding of the overall landscape of B-cell receptor (BCR) repertoires in COVID-19 patients. Here, we used high-throughput sequencing of bulk and plasma B-cells collected over multiple time points during infection to characterize signatures of B-cell response to SARS-CoV-2 in 19 patients. Using principled statistical approaches, we determined differential features of BCRs associated with different disease severity. We identified 38 significantly expanded clonal lineages shared among patients as candidates for specific responses to SARS-CoV-2. Using single-cell sequencing, we verified reactivity of BCRs shared among individuals to SARS-CoV-2 epitopes. Moreover, we identified natural emergence of a BCR with cross-reactivity to SARS-CoV-1 and SARS-CoV-2 in a number of patients. Our results provide important insights for development of rational therapies and vaccines against COVID-19.
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792. | Montague, Zachary; Lv, Huibin; Otwinowski, Jakub; DeWitt, William S; Isacchini, Giulio; Yip, Garrick K; Ng, Wilson W; Tsang, Owen Tak-Yin; Yuan, Meng; Liu, Hejun; Wilson, Ian A; Peiris, J S Malik; Wu, Nicholas C; Nourmohammad, Armita; Mok, Chris Ka Pun: Dynamics of B-cell repertoires and emergence of cross-reactive responses in COVID-19 patients with different disease severity. 2021, ISSN: 2331-8422. (Type: Miscellaneous | Abstract)
@misc{pmid32699813, COVID-19 patients show varying severity of the disease ranging from asymptomatic to requiring intensive care. Although a number of SARS-CoV-2 specific monoclonal antibodies have been identified, we still lack an understanding of the overall landscape of B-cell receptor (BCR) repertoires in COVID-19 patients. Here, we used high-throughput sequencing of bulk and plasma B-cells collected over multiple time points during infection to characterize signatures of B-cell response to SARS-CoV-2 in 19 patients. Using principled statistical approaches, we determined differential features of BCRs associated with different disease severity. We identified 38 significantly expanded clonal lineages shared among patients as candidates for specific responses to SARS-CoV-2. Using single-cell sequencing, we verified reactivity of BCRs shared among individuals to SARS-CoV-2 epitopes. Moreover, we identified natural emergence of a BCR with cross-reactivity to SARS-CoV-1 and SARS-CoV-2 in a number of patients. Our results provide important insights for development of rational therapies and vaccines against COVID-19.
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791. | Chen, Fang; Tzarum, Netanel; Lin, Xiaohe; Giang, Erick; Velázquez-Moctezuma, Rodrigo; Augestad, Elias H; Nagy, Kenna; He, Linling; Hernandez, Mayda; Fouch, Mallorie E; Grinyó, Ariadna; Chavez, Deborah; Doranz, Benjamin J; Prentoe, Jannick; Stanfield, Robyn L; Lanford, Robert; Bukh, Jens; Wilson, Ian A; Zhu, Jiang; Law, Mansun: Functional convergence of a germline-encoded neutralizing antibody response in rhesus macaques immunized with HCV envelope glycoproteins. In: Immunity, vol. 54, no. 4, pp. 781–796.e4, 2021, ISSN: 1097-4180. (Type: Journal Article | Abstract | Links)
@article{pmid33675683, Human IGHV1-69-encoded broadly neutralizing antibodies (bnAbs) that target the hepatitis C virus (HCV) envelope glycoprotein (Env) E2 are important for protection against HCV infection. An IGHV1-69 ortholog gene, VH1.36, is preferentially used for bnAbs isolated from HCV Env-immunized rhesus macaques (RMs). Here, we studied the genetic, structural, and functional properties of VH1.36-encoded bnAbs generated by vaccination, in comparison to IGHV1-69-encoded bnAbs from HCV patients. Global B cell repertoire analysis confirmed the expansion of VH1.36-derived B cells in immunized animals. Most E2-specific, VH1.36-encoded antibodies cross-neutralized HCV. Crystal structures of two RM bnAbs with E2 revealed that the RM bnAbs engaged conserved E2 epitopes using similar molecular features as human bnAbs but with a different binding mode. Longitudinal analyses of the RM antibody repertoire responses during immunization indicated rapid lineage development of VH1.36-encoded bnAbs with limited somatic hypermutation. Our findings suggest functional convergence of a germline-encoded bnAb response to HCV Env with implications for vaccination in humans.
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790. | Law, Mansun: Antibody Responses in Hepatitis C Infection. In: Cold Spring Harb Perspect Med, vol. 11, no. 3, 2021, ISSN: 2157-1422. (Type: Journal Article | Abstract | Links)
@article{pmid32341067, Antibody responses in hepatitis C virus (HCV) have been a rather mysterious research topic for many investigators working in the field. Chronic HCV infection is often associated with dysregulation of immune functions particularly in B cells, leading to abnormal lymphoproliferation or the production of autoantibodies that exacerbate inflammation and extrahepatic diseases. When considering the antiviral function of antibody, it was difficult to endorse its role in HCV protection, whereas T-cell response has been shown unequivocally critical for natural recovery. Recent breakthroughs in the study of HCV and antigen-specific antibody responses provide important insights into viral vulnerability to antibodies and the immunogenetic and structural properties of the neutralizing antibodies. The new knowledge reinvigorates HCV vaccine research by illuminating a new path for the rational design of vaccine antigens to elicit broadly neutralizing antibodies for protection.
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789. | He, Linling; Lin, Xiaohe; Wang, Ying; Abraham, Ciril; Sou, Cindy; Ngo, Timothy; Zhang, Yi; Wilson, Ian A; Zhu, Jiang: Single-component, self-assembling, protein nanoparticles presenting the receptor binding domain and stabilized spike as SARS-CoV-2 vaccine candidates. In: Sci Adv, vol. 7, no. 12, 2021, ISSN: 2375-2548. (Type: Journal Article | Abstract | Links)
@article{pmid33741598, Vaccination against SARS-CoV-2 provides an effective tool to combat the COVID-19 pandemic. Here, we combined antigen optimization and nanoparticle display to develop vaccine candidates for SARS-CoV-2. We first displayed the receptor-binding domain (RBD) on three self-assembling protein nanoparticle (SApNP) platforms using the SpyTag/SpyCatcher system. We then identified heptad repeat 2 (HR2) in S2 as the cause of spike metastability, designed an HR2-deleted glycine-capped spike (S2GΔHR2), and displayed S2GΔHR2 on SApNPs. An antibody column specific for the RBD enabled tag-free vaccine purification. In mice, the 24-meric RBD-ferritin SApNP elicited a more potent neutralizing antibody (NAb) response than the RBD alone and the spike with two stabilizing proline mutations in S2 (S2P). S2GΔHR2 elicited twofold higher NAb titers than S2P, while S2GΔHR2 SApNPs derived from multilayered E2p and I3-01v9 60-mers elicited up to 10-fold higher NAb titers. The S2GΔHR2-presenting I3-01v9 SApNP also induced critically needed T cell immunity, thereby providing a promising vaccine candidate.
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788. | Lee, Chang-Chun D; Watanabe, Yasunori; Wu, Nicholas C; Han, Julianna; Kumar, Sonu; Pholcharee, Tossapol; Seabright, Gemma E; Allen, Joel D; Lin, Chih-Wei; Yang, Ji-Rong; Liu, Ming-Tsan; Wu, Chung-Yi; Ward, Andrew B; Crispin, Max; Wilson, Ian A: A cross-neutralizing antibody between HIV-1 and influenza virus. In: PLoS Pathog, vol. 17, no. 3, pp. e1009407, 2021, ISSN: 1553-7374. (Type: Journal Article | Abstract | Links)
@article{pmid33750987, Incessant antigenic evolution enables the persistence and spread of influenza virus in the human population. As the principal target of the immune response, the hemagglutinin (HA) surface antigen on influenza viruses continuously acquires and replaces N-linked glycosylation sites to shield immunogenic protein epitopes using host-derived glycans. Anti-glycan antibodies, such as 2G12, target the HIV-1 envelope protein (Env), which is even more extensively glycosylated and contains under-processed oligomannose-type clusters on its dense glycan shield. Here, we illustrate that 2G12 can also neutralize human seasonal influenza A H3N2 viruses that have evolved to present similar oligomannose-type clusters on their HAs from around 20 years after the 1968 pandemic. Using structural biology and mass spectrometric approaches, we find that two N-glycosylation sites close to the receptor binding site (RBS) on influenza hemagglutinin represent the oligomannose cluster recognized by 2G12. One of these glycan sites is highly conserved in all human H3N2 strains and the other emerged during virus evolution. These two N-glycosylation sites have also become crucial for fitness of recent H3N2 strains. These findings shed light on the evolution of the glycan shield on influenza virus and suggest 2G12-like antibodies can potentially act as broad neutralizers to target human enveloped viruses.
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787. | Kalagiri, Rajasree; Stanfield, Robyn L; Meisenhelder, Jill; Clair, James J La; Fuhs, Stephen R; Wilson, Ian A; Hunter, Tony: Structural basis for differential recognition of phosphohistidine-containing peptides by 1-pHis and 3-pHis monoclonal antibodies. In: Proc Natl Acad Sci U S A, vol. 118, no. 6, 2021, ISSN: 1091-6490. (Type: Journal Article | Abstract | Links)
@article{pmid33547238, In 2015, monoclonal antibodies (mAbs) that selectively recognize the 1-pHis or 3-pHis isoforms of phosphohistidine were developed by immunizing rabbits with degenerate Ala/Gly peptides containing the nonhydrolyzable phosphohistidine (pHis) analog- phosphotriazolylalanine (pTza). Here, we report structures of five rabbit mAbs bound to cognate pTza peptides: SC1-1 and SC50-3 that recognize 1-pHis, and their 3-pHis-specific counterparts, SC39-4, SC44-8, and SC56-2. These cocrystal structures provide insights into the binding modes of the pTza phosphate group that are distinct for the 1- and 3-pHis mAbs with the selectivity arising from specific contacts with the phosphate group and triazolyl ring. The mode of phosphate recognition in the 3-pHis mAbs recapitulates the Walker A motif, as present in kinases. The complementarity-determining regions (CDRs) of four of the Fabs interact with the peptide backbone rather than peptide side chains, thus conferring sequence independence, whereas SC44-8 shows a proclivity for binding a GpHAGA motif mediated by a sterically complementary CDRL3 loop. Specific hydrogen bonding with the triazolyl ring precludes recognition of pTyr and other phosphoamino acids by these mAbs. Kinetic binding experiments reveal that the affinity of pHis mAbs for pHis and pTza peptides is submicromolar. Bound pHis mAbs also shield the pHis peptides from rapid dephosphorylation. The epitope-paratope interactions illustrate how these anti-pHis antibodies are useful for a wide range of research techniques and this structural information can be utilized to improve the specificity and affinity of these antibodies toward a variety of pHis substrates to understand the role of histidine phosphorylation in healthy and diseased states.
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786. | Liu, Hejun; Yuan, Meng; Huang, Deli; Bangaru, Sandhya; Lee, Chang-Chun D; Peng, Linghang; Zhu, Xueyong; Nemazee, David; van Gils, Marit J; Sanders, Rogier W; Kornau, Hans-Christian; Reincke, S Momsen; Prüss, Harald; Kreye, Jakob; Wu, Nicholas C; Ward, Andrew B; Wilson, Ian A: A combination of cross-neutralizing antibodies synergizes to prevent SARS-CoV-2 and SARS-CoV pseudovirus infection. 2021. (Type: Miscellaneous | Abstract | Links)
@misc{pmid33594361, Coronaviruses have caused several epidemics and pandemics including the ongoing coronavirus disease 2019 (COVID-19). Some prophylactic vaccines and therapeutic antibodies have already showed striking effectiveness against COVID-19. Nevertheless, concerns remain about antigenic drift in SARS-CoV-2 as well as threats from other sarbecoviruses. Cross-neutralizing antibodies to SARS-related viruses provide opportunities to address such concerns. Here, we report on crystal structures of a cross-neutralizing antibody CV38-142 in complex with the receptor binding domains from SARS-CoV-2 and SARS-CoV. Our structural findings provide mechanistic insights into how this antibody can accommodate antigenic variation in these viruses. CV38-142 synergizes with other cross-neutralizing antibodies, in particular COVA1-16, to enhance neutralization of SARS-CoV-2 and SARS-CoV. Overall, this study provides valuable information for vaccine and therapeutic design to address current and future antigenic drift in SARS-CoV-2 and to protect against zoonotic coronaviruses.
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785. | Koenig, Paul-Albert; Das, Hrishikesh; Liu, Hejun; Kümmerer, Beate M; Gohr, Florian N; Jenster, Lea-Marie; Schiffelers, Lisa D J; Tesfamariam, Yonas M; Uchima, Miki; Wuerth, Jennifer D; Gatterdam, Karl; Ruetalo, Natalia; Christensen, Maria H; Fandrey, Caroline I; Normann, Sabine; Tödtmann, Jan M P; Pritzl, Steffen; Hanke, Leo; Boos, Jannik; Yuan, Meng; Zhu, Xueyong; Schmid-Burgk, Jonathan L; Kato, Hiroki; Schindler, Michael; Wilson, Ian A; Geyer, Matthias; Ludwig, Kerstin U; Hällberg, B Martin; Wu, Nicholas C; Schmidt, Florian I: Structure-guided multivalent nanobodies block SARS-CoV-2 infection and suppress mutational escape. In: Science, vol. 371, no. 6530, 2021, ISSN: 1095-9203. (Type: Journal Article | Abstract | Links)
@article{pmid33436526, The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to spread, with devastating consequences. For passive immunization efforts, nanobodies have size and cost advantages over conventional antibodies. In this study, we generated four neutralizing nanobodies that target the receptor binding domain of the SARS-CoV-2 spike protein. We used x-ray crystallography and cryo-electron microscopy to define two distinct binding epitopes. On the basis of these structures, we engineered multivalent nanobodies with more than 100 times the neutralizing activity of monovalent nanobodies. Biparatopic nanobody fusions suppressed the emergence of escape mutants. Several nanobody constructs neutralized through receptor binding competition, whereas other monovalent and biparatopic nanobodies triggered aberrant activation of the spike fusion machinery. These premature conformational changes in the spike protein forestalled productive fusion and rendered the virions noninfectious.
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784. | Pholcharee, Tossapol; Oyen, David; Flores-Garcia, Yevel; Gonzalez-Paez, Gonzalo; Han, Zhen; Williams, Katherine L; Volkmuth, Wayne; Emerling, Daniel; Locke, Emily; King, C Richter; Zavala, Fidel; Wilson, Ian A: Structural and biophysical correlation of anti-NANP antibodies with in vivo protection against P. falciparum. In: Nat Commun, vol. 12, no. 1, pp. 1063, 2021, ISSN: 2041-1723. (Type: Journal Article | Abstract | Links)
@article{pmid33594061, The most advanced P. falciparum circumsporozoite protein-based malaria vaccine, RTS,S/AS01 (RTS,S), confers partial protection but with antibody titers that wane relatively rapidly, highlighting the need to elicit more potent and durable antibody responses. Here, we elucidate crystal structures, binding affinities and kinetics, and in vivo protection of eight anti-NANP antibodies derived from an RTS,S phase 2a trial and encoded by three different heavy-chain germline genes. The structures reinforce the importance of homotypic Fab-Fab interactions in protective antibodies and the overwhelmingly dominant preference for a germline-encoded aromatic residue for recognition of the NANP motif. In this study, antibody apparent affinity correlates best with protection in an in vivo mouse model, with the more potent antibodies also recognizing epitopes with repeating secondary structural motifs of type I β- and Asn pseudo 3 turns; such insights can be incorporated into design of more effective immunogens and antibodies for passive immunization.
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783. | Yuan, Meng; Huang, Deli; Lee, Chang-Chun D; Wu, Nicholas C; Jackson, Abigail M; Zhu, Xueyong; Liu, Hejun; Peng, Linghang; van Gils, Marit J; Sanders, Rogier W; Burton, Dennis R; Reincke, S Momsen; Prüss, Harald; Kreye, Jakob; Nemazee, David; Ward, Andrew B; Wilson, Ian A: Structural and functional ramifications of antigenic drift in recent SARS-CoV-2 variants. 2021. (Type: Miscellaneous | Abstract | Links)
@misc{pmid33619487, The protective efficacy of neutralizing antibodies (nAbs) elicited during natural infection with SARS-CoV-2 and by vaccination based on its spike protein has been compromised with emergence of the recent SARS-CoV-2 variants. Residues E484 and K417 in the receptor-binding site (RBS) are both mutated in lineages first described in South Africa (B.1.351) and Brazil (B.1.1.28.1). The nAbs isolated from SARS-CoV-2 patients are preferentially encoded by certain heavy-chain germline genes and the two most frequently elicited antibody families (IGHV3-53/3-66 and IGHV1-2) can each bind the RBS in two different binding modes. However, their binding and neutralization are abrogated by either the E484K or K417N mutation, whereas nAbs to the cross-reactive CR3022 and S309 sites are largely unaffected. This structural and functional analysis illustrates why mutations at E484 and K417 adversely affect major classes of nAbs to SARS-CoV-2 with consequences for next-generation COVID-19 vaccines.
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782. | Tan, Timothy J C; Yuan, Meng; Kuzelka, Kaylee; Padron, Gilberto C; Beal, Jacob R; Chen, Xin; Wang, Yiquan; Rivera-Cardona, Joel; Zhu, Xueyong; Stadtmueller, Beth M; Brooke, Christopher B; Wilson, Ian A; Wu, Nicholas C: Sequence signatures of two IGHV3-53/3-66 public clonotypes to SARS-CoV-2 receptor binding domain. 2021. (Type: Miscellaneous | Abstract | Links)
@misc{pmid33532781, Since the COVID-19 pandemic onset, the antibody response to SARS-CoV-2 has been extensively characterized. Antibodies to the receptor binding domain (RBD) on the spike protein are frequently encoded by IGHV3-53/3-66 with a short CDR H3. Germline-encoded sequence motifs in CDRs H1 and H2 play a major role, but whether any common motifs are present in CDR H3, which is often critical for binding specificity, have not been elucidated. Here, we identify two public clonotypes of IGHV3-53/3-66 RBD antibodies with a 9-residue CDR H3 that pair with different light chains. Distinct sequence motifs on CDR H3 are present in the two public clonotypes that appear to be related to differential light chain pairing. Additionally, we show that Y58F is a common somatic hypermutation that results in increased binding affinity of IGHV3-53/3-66 RBD antibodies with a short CDR H3. Overall, our results advance fundamental understanding of the antibody response to SARS-CoV-2.
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781. | Yuan, Meng; Liu, Hejun; Wu, Nicholas C; Wilson, Ian A: Recognition of the SARS-CoV-2 receptor binding domain by neutralizing antibodies. In: Biochem Biophys Res Commun, vol. 538, pp. 192–203, 2021, ISSN: 1090-2104. (Type: Journal Article | Abstract | Links)
@article{pmid33069360, Immediately from the outset of the COVID-19 pandemic, researchers from diverse biomedical and biological disciplines have united to study the novel pandemic virus, SARS-CoV-2. The antibody response to SARS-CoV-2 has been a major focus of COVID-19 research due to its clinical relevance and importance in vaccine and therapeutic development. Isolation and characterization of antibodies to SARS-CoV-2 have been accumulating at an unprecedented pace. Most of the SARS-CoV-2 neutralizing antibodies to date target the spike (S) protein receptor binding domain (RBD), which engages the host receptor ACE2 for viral entry. Here we review the binding sites and molecular features of monoclonal antibodies that target the SARS-CoV-2 RBD, including a few that also cross-neutralize SARS-CoV.
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780. | Wilson, Ian A; Stanfield, Robyn L: 50 Years of structural immunology. In: J Biol Chem, vol. 296, pp. 100745, 2021, ISSN: 1083-351X. (Type: Journal Article | Abstract | Links)
@article{pmid33957119, Fifty years ago, the first landmark structures of antibodies heralded the dawn of structural immunology. Momentum then started to build toward understanding how antibodies could recognize the vast universe of potential antigens and how antibody-combining sites could be tailored to engage antigens with high specificity and affinity through recombination of germline genes (V, D, J) and somatic mutation. Equivalent groundbreaking structures in the cellular immune system appeared some 15 to 20 years later and illustrated how processed protein antigens in the form of peptides are presented by MHC molecules to T cell receptors. Structures of antigen receptors in the innate immune system then explained their inherent specificity for particular microbial antigens including lipids, carbohydrates, nucleic acids, small molecules, and specific proteins. These two sides of the immune system act immediately (innate) to particular microbial antigens or evolve (adaptive) to attain high specificity and affinity to a much wider range of antigens. We also include examples of other key receptors in the immune system (cytokine receptors) that regulate immunity and inflammation. Furthermore, these antigen receptors use a limited set of protein folds to accomplish their various immunological roles. The other main players are the antigens themselves. We focus on surface glycoproteins in enveloped viruses including SARS-CoV-2 that enable entry and egress into host cells and are targets for the antibody response. This review covers what we have learned over the past half century about the structural basis of the immune response to microbial pathogens and how that information can be utilized to design vaccines and therapeutics.
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779. | Wu, Nicholas C; Yuan, Meng; Bangaru, Sandhya; Huang, Deli; Zhu, Xueyong; Lee, Chang-Chun D; Turner, Hannah L; Peng, Linghang; Yang, Linlin; Burton, Dennis R; Nemazee, David; Ward, Andrew B; Wilson, Ian A: A natural mutation between SARS-CoV-2 and SARS-CoV determines neutralization by a cross-reactive antibody. In: PLoS Pathog, vol. 16, no. 12, pp. e1009089, 2020, ISSN: 1553-7374. (Type: Journal Article | Abstract | Links)
@article{pmid33275640, Epitopes that are conserved among SARS-like coronaviruses are attractive targets for design of cross-reactive vaccines and therapeutics. CR3022 is a SARS-CoV neutralizing antibody to a highly conserved epitope on the receptor binding domain (RBD) on the spike protein that is able to cross-react with SARS-CoV-2, but with lower affinity. Using x-ray crystallography, mutagenesis, and binding experiments, we illustrate that of four amino acid differences in the CR3022 epitope between SARS-CoV-2 and SARS-CoV, a single mutation P384A fully determines the affinity difference. CR3022 does not neutralize SARS-CoV-2, but the increased affinity to SARS-CoV-2 P384A mutant now enables neutralization with a similar potency to SARS-CoV. We further investigated CR3022 interaction with the SARS-CoV spike protein by negative-stain EM and cryo-EM. Three CR3022 Fabs bind per trimer with the RBD observed in different up-conformations due to considerable flexibility of the RBD. In one of these conformations, quaternary interactions are made by CR3022 to the N-terminal domain (NTD) of an adjacent subunit. Overall, this study provides insights into antigenic variation and potential cross-neutralizing epitopes on SARS-like viruses.
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