Effect of SARS-CoV-2 B.1.1.7 mutations on spike protein structure and function.
12 Aug 2021-Nature Structural & Molecular Biology (Springer Science and Business Media LLC)-Vol. 28, Iss: 9, pp 731-739
TL;DR: In this paper, the spike protein of SARS-CoV-2 was analyzed in the apo and ACE2-bound forms and three RBDs were found to be engaged in ACE2 binding.
Abstract: The B.1.1.7 variant of SARS-CoV-2 first detected in the UK harbors amino-acid substitutions and deletions in the spike protein that potentially enhance host angiotensin conversion enzyme 2 (ACE2) receptor binding and viral immune evasion. Here we report cryo-EM structures of the spike protein of B.1.1.7 in the apo and ACE2-bound forms. The apo form showed one or two receptor-binding domains (RBDs) in the open conformation, without populating the fully closed state. All three RBDs were engaged in ACE2 binding. The B.1.1.7-specific A570D mutation introduces a molecular switch that could modulate the opening and closing of the RBD. The N501Y mutation introduces a π-π interaction that enhances RBD binding to ACE2 and abolishes binding of a potent neutralizing antibody (nAb). Cryo-EM also revealed how a cocktail of two nAbs simultaneously bind to all three RBDs, and demonstrated the potency of the nAb cocktail to neutralize different SARS-CoV-2 pseudovirus strains, including B.1.1.7.
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TL;DR: The Delta variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a dominant strain worldwide as discussed by the authors, and the structure, func...
Abstract: The Delta variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has outcompeted previously prevalent variants and become a dominant strain worldwide. We report the structure, func...
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TL;DR: In this paper , the authors highlight some key points regarding mAb-based detection tests and treatments for the COVID-19 pandemic and highlight the use of mAbs as surveillance tools that can be used to prevent the spread of CoV-19.
Abstract: The coronavirus disease 2019 (COVID-19) pandemic is an exceptional public health crisis that demands the timely creation of new therapeutics and viral detection. Owing to their high specificity and reliability, monoclonal antibodies (mAbs) have emerged as powerful tools to treat and detect numerous diseases. Hence, many researchers have begun to urgently develop Ab-based kits for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and Ab drugs for use as COVID-19 therapeutic agents. The detailed structure of the SARS-CoV-2 spike protein is known, and since this protein is key for viral infection, its receptor-binding domain (RBD) has become a major target for therapeutic Ab development. Because SARS-CoV-2 is an RNA virus with a high mutation rate, especially under the selective pressure of aggressively deployed prophylactic vaccines and neutralizing Abs, the use of Ab cocktails is expected to be an important strategy for effective COVID-19 treatment. Moreover, SARS-CoV-2 infection may stimulate an overactive immune response, resulting in a cytokine storm that drives severe disease progression. Abs to combat cytokine storms have also been under intense development as treatments for COVID-19. In addition to their use as drugs, Abs are currently being utilized in SARS-CoV-2 detection tests, including antigen and immunoglobulin tests. Such Ab-based detection tests are crucial surveillance tools that can be used to prevent the spread of COVID-19. Herein, we highlight some key points regarding mAb-based detection tests and treatments for the COVID-19 pandemic.
115 citations
TL;DR: In this article , the authors highlight some key points regarding mAb-based detection tests and treatments for the COVID-19 pandemic and highlight the use of mAbs as surveillance tools that can be used to prevent the spread of CoV-19.
Abstract: The coronavirus disease 2019 (COVID-19) pandemic is an exceptional public health crisis that demands the timely creation of new therapeutics and viral detection. Owing to their high specificity and reliability, monoclonal antibodies (mAbs) have emerged as powerful tools to treat and detect numerous diseases. Hence, many researchers have begun to urgently develop Ab-based kits for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and Ab drugs for use as COVID-19 therapeutic agents. The detailed structure of the SARS-CoV-2 spike protein is known, and since this protein is key for viral infection, its receptor-binding domain (RBD) has become a major target for therapeutic Ab development. Because SARS-CoV-2 is an RNA virus with a high mutation rate, especially under the selective pressure of aggressively deployed prophylactic vaccines and neutralizing Abs, the use of Ab cocktails is expected to be an important strategy for effective COVID-19 treatment. Moreover, SARS-CoV-2 infection may stimulate an overactive immune response, resulting in a cytokine storm that drives severe disease progression. Abs to combat cytokine storms have also been under intense development as treatments for COVID-19. In addition to their use as drugs, Abs are currently being utilized in SARS-CoV-2 detection tests, including antigen and immunoglobulin tests. Such Ab-based detection tests are crucial surveillance tools that can be used to prevent the spread of COVID-19. Herein, we highlight some key points regarding mAb-based detection tests and treatments for the COVID-19 pandemic.
103 citations
TL;DR: In this paper , structural and biochemical aspects of SARS-CoV-2 were assessed using cryo-electron microscopy (cryo-EM), ACE2-binding and antibody neutralization analyses.
Abstract: The Delta and Kappa variants of SARS-CoV-2 co-emerged in India in late 2020, with the Delta variant underlying the resurgence of COVID-19, even in countries with high vaccination rates. In this study, we assess structural and biochemical aspects of viral fitness for these two variants using cryo-electron microscopy (cryo-EM), ACE2-binding and antibody neutralization analyses. Both variants demonstrate escape of antibodies targeting the N-terminal domain, an important immune hotspot for neutralizing epitopes. Compared to wild-type and Kappa lineages, Delta variant spike proteins show modest increase in ACE2 affinity, likely due to enhanced electrostatic complementarity at the RBD-ACE2 interface, which we characterize by cryo-EM. Unexpectedly, Kappa variant spike trimers form a structural head-to-head dimer-of-trimers assembly, which we demonstrate is a result of the E484Q mutation and with unknown biological implications. The combination of increased antibody escape and enhanced ACE2 binding provides an explanation, in part, for the rapid global dominance of the Delta variant.
50 citations
TL;DR: In this paper , Liu et al. describe the Omicron receptor-binding domain (RBD) mutational landscape using amino acid interaction (AAI) networks, which are well suited for interrogating constellations of mutations that function in an epistatic manner.
Abstract: •Network analyses offer distinct insights into RBD-neutralizing Ab interactions•Omicron mutations broadly and deeply perturb networks across RBD epitope classes•Networks capture indirect effects of Omicron mutations on Ab escape potential•Omicron mutations provide plausible structural rationale for enhanced transmission The Omicron variant features enhanced transmissibility and antibody escape. Here, we describe the Omicron receptor-binding domain (RBD) mutational landscape using amino acid interaction (AAI) networks, which are well suited for interrogating constellations of mutations that function in an epistatic manner. Using AAI, we map Omicron mutations directly and indirectly driving increased escape breadth and depth in class 1–4 antibody epitopes. Further, we present epitope networks for authorized therapeutic antibodies and assess perturbations to each antibody’s epitope. Since our initial modeling following the identification of Omicron, these predictions have been realized by experimental findings of Omicron neutralization escape from therapeutic antibodies ADG20, AZD8895, and AZD1061. Importantly, the AAI predicted escape resulting from indirect epitope perturbations was not captured by previous sequence or point mutation analyses. Finally, for several Omicron RBD mutations, we find evidence for a plausible role in enhanced transmissibility via disruption of RBD-down conformational stability at the RBDdown-RBDdown interface. The Omicron variant features enhanced transmissibility and antibody escape. Here, we describe the Omicron receptor-binding domain (RBD) mutational landscape using amino acid interaction (AAI) networks, which are well suited for interrogating constellations of mutations that function in an epistatic manner. Using AAI, we map Omicron mutations directly and indirectly driving increased escape breadth and depth in class 1–4 antibody epitopes. Further, we present epitope networks for authorized therapeutic antibodies and assess perturbations to each antibody’s epitope. Since our initial modeling following the identification of Omicron, these predictions have been realized by experimental findings of Omicron neutralization escape from therapeutic antibodies ADG20, AZD8895, and AZD1061. Importantly, the AAI predicted escape resulting from indirect epitope perturbations was not captured by previous sequence or point mutation analyses. Finally, for several Omicron RBD mutations, we find evidence for a plausible role in enhanced transmissibility via disruption of RBD-down conformational stability at the RBDdown-RBDdown interface. The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) Omicron (B.1.1.529) variant of concern (VOC) has spread globally due to an apparent fitness advantage over the Delta variant.1Pulliam J.R.C. Schalkwyk C. van Govender N. Gottberg A. von Cohen C. Groome M.J. Dushoff J. Mlisana K. Moultrie H. Increased risk of SARS-CoV-2 reinfection associated with emergence of the Omicron variant in South Africa.medRxiv. 2021; https://doi.org/10.1101/2021.11.11.21266068Crossref Scopus (0) Google Scholar Several of Omicron’s spike mutations have been observed in other VOCs and are known to enhance transmissibility and confer varying degrees of escape from neutralizing antibodies.2Liu Y. Liu J. Johnson B.A. Xia H. Ku Z. Schindewolf C. Widen S.G. An Z. Weaver S.C. Menachery V.D. et al.Delta spike P681R mutation enhances SARS-CoV-2 fitness over Alpha variant.bioRxiv. 2021; https://doi.org/10.1101/2021.08.12.456173Crossref Scopus (0) Google Scholar, 3Liu Y. Liu J. Plante K.S. Plante J.A. Xie X. Zhang X. Ku Z. An Z. Scharton D. Schindewolf C. et al.The N501Y spike substitution enhances SARS-CoV-2 transmission.Nature. 2021; https://doi.org/10.1038/s41586-021-04245-0Crossref Scopus (36) Google Scholar, 4Zhou D. Dejnirattisai W. Supasa P. Liu C. Mentzer A.J. Ginn H.M. Zhao Y. Duyvesteyn H.M.E. Tuekprakhon A. Nutalai R. et al.Evidence of escape of SARS-CoV-2 variant B.1.351 from natural and vaccine-induced sera.Cell. 2021; 184: 2348-2361.e6Abstract Full Text Full Text PDF PubMed Scopus (408) Google Scholar, 5Greaney A.J. Starr T.N. Barnes C.O. Weisblum Y. Schmidt F. Caskey M. Gaebler C. Cho A. Agudelo M. Finkin S. et al.Mapping mutations to the SARS-CoV-2 RBD that escape binding by different classes of antibodies.Nat. Commun. 2021; 12 (2021 12:1 12, 1–14): 4196Crossref PubMed Scopus (61) Google Scholar, 6Ku Z. Xie X. Davidson E. Ye X. Su H. Menachery V.D. Li Y. Yuan Z. Zhang X. Muruato A.E. et al.Molecular determinants and mechanism for antibody cocktail preventing SARS-CoV-2 escape.Nat. Commun. 2021; 12: 469Crossref PubMed Scopus (61) Google Scholar However, numerous Omicron mutations have not been observed on previous VOCs nor characterized rigorously in terms of their functional effects. The position of many uncharacterized Omicron mutations within dominant antibody epitopes therefore prompted concerns that the efficacy of vaccines and therapeutic antibodies could be significantly reduced against Omicron, leading to policy decisions and research prioritizations with far-reaching consequences. In this study, we analyze the receptor-binding domain (RBD) mutational landscape of Omicron using amino acid interaction (AAI) networks.7Soundararajan V. Zheng S. Patel N. Warnock K. Raman R. Wilson I.A. Raguram S. Sasisekharan V. Sasisekharan R. Networks link antigenic and receptor-binding sites of influenza hemagglutinin: mechanistic insight into fitter strain propagation.Scientific Rep. 2011; 1: 200Crossref PubMed Scopus (36) Google Scholar,8Miller N.L. Clark T. Raman R. Sasisekharan R. An antigenic space framework for understanding antibody escape of SARS-CoV-2 variants.Viruses. 2021; 13: 2009Crossref PubMed Scopus (2) Google Scholar AAI network analysis is particularly well suited for understanding the impact of constellations of mutations residing within and adjacent to an antibody epitope as occurs on the Omicron variant. For example, AAI network analysis considers how mutation of a residue that does not directly interact with a given antibody paratope (e.g., via a hydrogen bond between side chain and antibody complementarity determining region [CDR]) may still perturb antibody binding if the residue plays a significant structural role in supporting other sites that interact directly with the antibody.8Miller N.L. Clark T. Raman R. Sasisekharan R. An antigenic space framework for understanding antibody escape of SARS-CoV-2 variants.Viruses. 2021; 13: 2009Crossref PubMed Scopus (2) Google Scholar,9Sethi A. Tian J. Derdeyn C.A. Korber B. Gnanakaran S. A mechanistic understanding of allosteric immune escape pathways in the HIV-1 envelope glycoprotein.PLoS Comput. Biol. 2013; 9: 1003046Crossref PubMed Scopus (45) Google Scholar Such indirect effects have been conceptualized as mutations outside of the direct antibody epitope that alter higher-order protein structure or perturb protein “breathing.”10Kalia V. Sarkar S. Gupta P. Montelaro R.C. Antibody neutralization escape mediated by point mutations in the intracytoplasmic tail of human immunodeficiency virus type 1 gp41.J. Virol. 2005; 79: 2097-2107Crossref PubMed Scopus (46) Google Scholar, 11Yewdell J.W. Antigenic drift: understanding COVID-19.Immunity. 2021; 54: 2681-2687Abstract Full Text Full Text PDF PubMed Scopus (5) Google Scholar, 12Kolawole A.O. Smith H.Q. Svoboda S.A. Lewis M.S. Sherman M.B. Lynch G.C. et al.Norovirus escape from broadly neutralizing antibodies is limited to allostery-like mechanisms.mSphere. 2017; 2 (e00334–17)Crossref PubMed Scopus (23) Google Scholar AAI networks quantitate these indirect relationships through which Omicron’s mutation constellation may substantially perturb the chemical and physical properties of RBD epitope surfaces. We apply the AAI network lens to map the potential impacts of Omicron RBD mutations on polyclonal antibody responses and therapeutic monoclonal antibodies. We further discuss the limitations in predicting efficacy of therapeutic antibodies against emerging variants based on existing in vitro data for isolated mutations. Finally, we present possible functional roles for Omicron RBD mutations that are not predicted to substantially enhance antibody evasion. Our analysis using a fixed-backbone Omicron homology model and a recently released Omicron spike structure suggests Omicron mutations may modulate RBD-up versus RBD-down conformational dynamics toward enhanced infectivity. Toward investigating the antigenic impact of the Omicron RBD mutations, we first mapped direct and indirect effects of Omicron mutations on RBD antibody epitopes using AAI networks. Our RBD epitope map included at least 10 antibodies from each of the four structural classes of anti-RBD antibodies13Barnes C.O. Jette C.A. Abernathy M.E. Dam K.M.A. Esswein S.R. Gristick H.B. Malyutin A.G. Sharaf N.G. Huey-Tubman K.E. Lee Y.E. et al.SARS-CoV-2 neutralizing antibody structures inform therapeutic strategies.Nature. 2020; 588: 682-687Crossref PubMed Scopus (459) Google Scholar and thus represents the dominant functional components of population-level polyclonal antibody responses.14Hastie K.M. Li H. Bedinger D. Schendel S.L. Moses Dennison S. Li K. Rayaprolu V. Yu X. Mann C. Zandonatti M. et al.Defining variant-resistant epitopes targeted by SARS-CoV-2 antibodies: a global consortium study.Science. 2021; 374: 472-478Crossref PubMed Scopus (48) Google Scholar First examining Omicron escape breadth, we found that Omicron mutations occur at RBD sites that interact with all antibodies examined and span the four antibody classes (Figure 1). In contrast, our analysis showed that the RBD mutations of the Beta and Delta variants are confined to sites within class 1 and 2 antibody epitopes, except for Beta N501Y, which interacts indirectly with certain class 3 antibodies. This is consistent with experimental evidence documenting Beta escape from class 1 and 2 antibodies15Wibmer C.K. Ayres F. Hermanus T. Madzivhandila M. Kgagudi P. Oosthuysen B. Lambson B.E. de Oliveira T. Vermeulen M. van der Berg K. et al.SARS-CoV-2 501Y.V2 escapes neutralization by South African COVID-19 donor plasma.Nat. Med. 2021; 27: 622-625Crossref PubMed Scopus (460) Google Scholar,16Yuan M. Liu H. Wu N.C. Lee C.C.D. Zhu X. Zhao F. Huang D. Yu W. Hua Y. Tien H. et al.Structural basis of a shared antibody response to SARS-CoV-2.Science. 2020; 369: 1119-1123Crossref PubMed Scopus (236) Google Scholar and Delta escape primarily from class 2 antibodies.17Cheng L. Song S. Fan Q. Shen S. Wang H. Zhou B. Ge X. Ju B. Zhang Z. Cross-neutralization of SARS-CoV-2 Kappa and Delta variants by inactivated vaccine-elicited serum and monoclonal antibodies.Cell Discov. 2021; 7: 112Crossref PubMed Scopus (3) Google Scholar Our analysis therefore suggests that Omicron’s increased antibody escape breadth as compared with previous variants is driven by mutations in class 3 and 4 antibody epitopes. Class 3 antibodies are potent neutralizers that are immunodominant for certain individuals,5Greaney A.J. Starr T.N. Barnes C.O. Weisblum Y. Schmidt F. Caskey M. Gaebler C. Cho A. Agudelo M. Finkin S. et al.Mapping mutations to the SARS-CoV-2 RBD that escape binding by different classes of antibodies.Nat. Commun. 2021; 12 (2021 12:1 12, 1–14): 4196Crossref PubMed Scopus (61) Google Scholar while class 4 antibodies tend to be weakly neutralizing.13Barnes C.O. Jette C.A. Abernathy M.E. Dam K.M.A. Esswein S.R. Gristick H.B. Malyutin A.G. Sharaf N.G. Huey-Tubman K.E. Lee Y.E. et al.SARS-CoV-2 neutralizing antibody structures inform therapeutic strategies.Nature. 2020; 588: 682-687Crossref PubMed Scopus (459) Google Scholar Our network analysis associated Omicron mutations N440K, G446S, G496S, and Q498R most strongly with enhanced class 3 antibody escape based on these sites having the strongest network interactions with the antibodies surveyed. The RBD of PMS20, a research variant that escapes neutralization from most convalescent and polyclonal sera, features similar class 3 mutations to Omicron at sites 440 and 445, yet also features an R346K mutation that Omicron lacks.18Schmidt F. Weisblum Y. Rutkowska M. Poston D. da Silva J. Zhang F. Bednarski E. Cho A. Schaefer-Babajew D.J. Gaebler C. et al.High genetic barrier to SARS-CoV-2 polyclonal neutralizing antibody escape.Nature. 2021; 600: 512-516Crossref PubMed Scopus (43) Google Scholar Our analysis found that R346 is the most strongly networked residue for certain class 3 antibodies, including C135. This suggests that PMS20 may escape more effectively from the class 3 antibody component of sera than Omicron, highlighting a key caveat in comparisons between Omicron and PMS20. The AAI analysis suggested an additional set of Omicron mutations (G339D, S371L, S373P, S375F) may contribute to class 3 and 4 antibody epitopes via predominantly indirect mechanisms, with mutations at these sites appearing to affect nearly all class 4 antibodies in our panel, as well as select class 3 antibodies, including S309, S2M11, C110, CV38-142, Ab 812, and PDI-96. While mutational scanning of point mutations at these sites suggested these mutations are unlikely to confer significant class 3 and 4 antibody escape in isolation,5Greaney A.J. Starr T.N. Barnes C.O. Weisblum Y. Schmidt F. Caskey M. Gaebler C. Cho A. Agudelo M. Finkin S. et al.Mapping mutations to the SARS-CoV-2 RBD that escape binding by different classes of antibodies.Nat. Commun. 2021; 12 (2021 12:1 12, 1–14): 4196Crossref PubMed Scopus (61) Google Scholar it is plausible that the combined indirect effects of these four non-conservative mutations could meaningfully alter the local structure in this RBD region and thus perturb class 3 and 4 antibodies. In contrast to escape from class 3 antibodies, however, Omicron lacked mutation at a site that is strongly networked to multiple class 4 antibodies such as 368-9, 377-8, 384, or 434. Further, we observed that Omicron mutations may enhance escape depth from class 1 antibodies beyond that observed for the Beta variant due to accumulation of additional mutations in class 1 antibody epitopes. The polyclonal antibody response to infection and vaccination has been shown to broaden over the months following exposure due to persistent somatic mutation of antibody CDRs.19Wang Z. Muecksch F. Schaefer-Babajew D. Finkin S. Viant C. Gaebler C. Hoffmann H.H. Barnes C.O. Cipolla M. Ramos V. et al.Naturally enhanced neutralizing breadth against SARS-CoV-2 one year after infection.Nature. 2021; 595: 426-431Crossref PubMed Scopus (181) Google Scholar This broadening process can increase an antibody’s tolerance of escape mutations within its epitope,20Muecksch F. Weisblum Y. Barnes C.O. Schmidt F. Schaefer-Babajew D. Wang Z. Julio J.C. Flyak A.I. DeLaitsch A.T. Huey-Tubman K.E. et al.Affinity maturation of SARS-CoV-2 neutralizing antibodies confers potency, breadth, and resilience to viral escape mutations.Immunity. 2021; 54: 1853-1868.e7Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar resulting in polyclonal responses with enhanced neutralizing capacity against the Beta variant over time. We found that Omicron has accumulated multiple tightly clustered mutations and therefore may have enhanced escape from matured polyclonal responses that are tolerant of certain class 1 antibody escape mutations such as K417N and N501Y (see Figure 1). That is, Omicron’s accumulated class 1 mutations may reduce Omicron’s susceptibility to class 1 antibodies tolerant of canonical class 1 escape mutations such as K417N. Specifically, Omicron mutations at residues Q493, G496, Q498, and Y505 clustered closely with the Beta and Omicron escape mutations at residues K417 and N501 indicating class 1 escape depth. Such escape depth could have contributed to PMS20’s escape from convalescent and vaccinee sera,18Schmidt F. Weisblum Y. Rutkowska M. Poston D. da Silva J. Zhang F. Bednarski E. Cho A. Schaefer-Babajew D.J. Gaebler C. et al.High genetic barrier to SARS-CoV-2 polyclonal neutralizing antibody escape.Nature. 2021; 600: 512-516Crossref PubMed Scopus (43) Google Scholar as it can be seen in Figure 1 that PMS20 also features multiple class 1 mutations. Notably, however, PMS20 was shown not to fully escape from polyclonal responses generated from infection followed by vaccination,18Schmidt F. Weisblum Y. Rutkowska M. Poston D. da Silva J. Zhang F. Bednarski E. Cho A. Schaefer-Babajew D.J. Gaebler C. et al.High genetic barrier to SARS-CoV-2 polyclonal neutralizing antibody escape.Nature. 2021; 600: 512-516Crossref PubMed Scopus (43) Google Scholar suggesting that Omicron’s class 1 escape depth and class 3 escape breadth alone may not confer this ability. We subsequently applied AAI networks to examine the Omicron mutations in the context of their ability to evade neutralization by therapeutic antibodies of current clinical relevance (Figure 2). An important perspective offered by our network approach is the impact of mutations that are not directly located at the interface of antibody-antigen complexes, yet may still disrupt the antibody interaction. Specifically, it is important to consider the allosteric effects of the many Omicron mutations and how they may cooperatively affect antibody binding to the Omicron RBD by modulating the structural and chemical features of the epitope surface. Our results present an accessible and concise visualization of the network interactions between RBD sites mutated on Omicron and authorized therapeutic antibodies. We found that Omicron mutations occur at sites that are strongly directly networked to REGN10987 + REGN10933 (casirivimab + imdevimab; sites 417, 440, 446, 484, 493, 496, 498) and LY-CoV016 + LY-CoV555 (bamlanivimab + etesevimab; sites 417, 484, 493, 501, 505), suggesting the binding of these antibodies will be directly perturbed by Omicron mutations. In contrast, AAI networking revealed Omicron mutations do not appear to strongly directly interact with S309 (sotrovimab) or AZD8895 + AZD1061 (tixagevimab + cilgavimab). Our findings on the basis of direct networking between epitope residues in Omicron variant and paratope residues in the neutralizing antibodies align with existing analyses and commentary for these therapeutic antibodies derived from mutagenesis screens based on the binding perturbation induced by the Omicron mutations individually and in isolation.5Greaney A.J. Starr T.N. Barnes C.O. Weisblum Y. Schmidt F. Caskey M. Gaebler C. Cho A. Agudelo M. Finkin S. et al.Mapping mutations to the SARS-CoV-2 RBD that escape binding by different classes of antibodies.Nat. Commun. 2021; 12 (2021 12:1 12, 1–14): 4196Crossref PubMed Scopus (61) Google Scholar,21Cathcart A.L. Havenar-Daughton C. Lempp F.A. Ma D. Schmid M.A. Agostini M.L. Guarino B. iulio J. di Rosen L.E. Tucker H. et al.The dual function monoclonal antibodies VIR-7831 and VIR-7832 demonstrate potent in vitro and in vivo activity against SARS-CoV-2.bioRxiv. 2021; https://doi.org/10.1101/2021.03.09.434607Crossref Scopus (0) Google Scholar, 22REGENERON Regeneron evaluating REGEN-COV® and next generation antibodies against new Omicron COVID-19 variant. 1–8.https://investor.regeneron.com/static-files/969bdb0b-53f5-46c7-94fb-7473ee7f5be3Date: 2021Google Scholar, 23Liu L. Iketani S. Guo Y. Chan J.F.-W. Wang M. Liu L. Luo Y. Chu H. Huang Y. Nair M.S. et al.Striking antibody evasion manifested by the Omicron variant of SARS-CoV-2.Nature. 2021; https://doi.org/10.1038/s41586-021-04388-0Crossref Scopus (113) Google Scholar However, point mutation analyses and AAI direct networking do not explain interactions observed between these antibodies and the Omicron variant in the full context of the set of Omicron RBD mutations. Specifically, analysis of the AZD1061 network found that Omicron mutated sites are highly networked to sites directly networked to AZD1061, resulting in significant indirect networking (Figures 2 and S1A). Meanwhile, we observed that the Omicron mutated sites result in a large number of moderate direct and indirect interactions for AZD8895 (Figure S1B). Since the submission of this analysis, multiple studies reported significant reductions in pseudovirus neutralization for AZD8895 and AZD1061,23Liu L. Iketani S. Guo Y. Chan J.F.-W. Wang M. Liu L. Luo Y. Chu H. Huang Y. Nair M.S. et al.Striking antibody evasion manifested by the Omicron variant of SARS-CoV-2.Nature. 2021; https://doi.org/10.1038/s41586-021-04388-0Crossref Scopus (113) Google Scholar,24Cao Y. Wang J. Jian F. Xiao T. Song W. Yisimayi A. Huang W. Li Q. Wang P. An R. et al.Omicron escapes the majority of existing SARS-CoV-2 neutralizing antibodies.Nature. 2021; : 1-9Google Scholar which is consistent with the cumulative Omicron mutations resulting in perturbation of these epitopes. Examination of the S309 AAI network showed indirect interactions between S309 and the Omicron mutated sites 339, 373, 440, and 446 (Figure S2). Since the submission of this manuscript, multiple studies have reported an approximately 3-fold reduction in neutralization for S309 against Omicron.21Cathcart A.L. Havenar-Daughton C. Lempp F.A. Ma D. Schmid M.A. Agostini M.L. Guarino B. iulio J. di Rosen L.E. Tucker H. et al.The dual function monoclonal antibodies VIR-7831 and VIR-7832 demonstrate potent in vitro and in vivo activity against SARS-CoV-2.bioRxiv. 2021; https://doi.org/10.1101/2021.03.09.434607Crossref Scopus (0) Google Scholar,23Liu L. Iketani S. Guo Y. Chan J.F.-W. Wang M. Liu L. Luo Y. Chu H. Huang Y. Nair M.S. et al.Striking antibody evasion manifested by the Omicron variant of SARS-CoV-2.Nature. 2021; https://doi.org/10.1038/s41586-021-04388-0Crossref Scopus (113) Google Scholar, 24Cao Y. Wang J. Jian F. Xiao T. Song W. Yisimayi A. Huang W. Li Q. Wang P. An R. et al.Omicron escapes the majority of existing SARS-CoV-2 neutralizing antibodies.Nature. 2021; : 1-9Google Scholar, 25Cameroni E. Bowen J.E. Rosen L.E. Saliba C. Zepeda S.K. Culap K. Pinto D. VanBlargan L.A. De Marco A. di Iulio J. et al.Broadly neutralizing antibodies overcome SARS-CoV-2 Omicron antigenic shift.Nature. 2021; https://doi.org/10.1038/s41586-021-04386-2Crossref Scopus (114) Google Scholar This reduction is consistent with the AAI model prediction that the cumulative indirect effects of the Omicron mutations moderately perturb the S309 binding surface. Building on this observation, we noted that the AAI network indicated R346 is important to the epitope network of S309, and R346 K/S mutations had been detected in Omicron subclades.23Liu L. Iketani S. Guo Y. Chan J.F.-W. Wang M. Liu L. Luo Y. Chu H. Huang Y. Nair M.S. et al.Striking antibody evasion manifested by the Omicron variant of SARS-CoV-2.Nature. 2021; https://doi.org/10.1038/s41586-021-04388-0Crossref Scopus (113) Google Scholar Further AAI analysis showed that R346 is networked both directly and indirectly to the S309 paratope in the context of the other Omicron mutations. While isolated mutations at R346 only minorly affect S309 binding and neutralization,21Cathcart A.L. Havenar-Daughton C. Lempp F.A. Ma D. Schmid M.A. Agostini M.L. Guarino B. iulio J. di Rosen L.E. Tucker H. et al.The dual function monoclonal antibodies VIR-7831 and VIR-7832 demonstrate potent in vitro and in vivo activity against SARS-CoV-2.bioRxiv. 2021; https://doi.org/10.1101/2021.03.09.434607Crossref Scopus (0) Google Scholar,26Starr T.N. Czudnochowski N. Liu Z. Zatta F. Park Y.J. Addetia A. Pinto D. Beltramello M. Hernandez P. Greaney A.J. et al.SARS-CoV-2 RBD antibodies that maximize breadth and resistance to escape.Nature. 2021; 597: 97-102Crossref PubMed Scopus (68) Google Scholar the AAI network suggests R346 mutations are likely to have a greater impact in the context of the other Omicron mutations. Liu et al. and Cameroni et al. further tested Omicron + R346K and found no significant additional reduction in neutralization.23Liu L. Iketani S. Guo Y. Chan J.F.-W. Wang M. Liu L. Luo Y. Chu H. Huang Y. Nair M.S. et al.Striking antibody evasion manifested by the Omicron variant of SARS-CoV-2.Nature. 2021; https://doi.org/10.1038/s41586-021-04388-0Crossref Scopus (113) Google Scholar,25Cameroni E. Bowen J.E. Rosen L.E. Saliba C. Zepeda S.K. Culap K. Pinto D. VanBlargan L.A. De Marco A. di Iulio J. et al.Broadly neutralizing antibodies overcome SARS-CoV-2 Omicron antigenic shift.Nature. 2021; https://doi.org/10.1038/s41586-021-04386-2Crossref Scopus (114) Google Scholar Importantly, R→K is a conservative mutation, highlighting how the physiochemical features of a mutation are critical to understanding whether the mutation will result in a significant perturbation of the AAI network with effect on antibody function. Therefore, it is important to monitor whether there is a further reduction in the susceptibility (>5-fold change as defined in the sotrovimab emergency use authorization [EUA] fact sheet27GSKFDAFact sheet for healthcare providers emergency use authorization (EUA) OF sotrovimab authorized use.https://www.fda.gov/emergency-preparedness-and-response/mcm-legal-Date: 2021Google Scholar) of subclades with other R346 mutations to neutralization by S309, as R→S has also been observed on Omicron subclades and could result in more substantial perturbation of the epitope network. Similarly, our analysis suggested that such indirect effects may affect the recently identified anti-RBD antibodies such as ADG-2 that target the conserved epitope shared across clade 1a and 1b coronaviruses.28Garrett Rappazzo C. Tse L.v. Kaku C.I. Wrapp D. Sakharkar M. Huang D. Deveau L.M. Yockachonis T.J. Herbert A.S. Battles M.B. et al.Broad and potent activity against SARS-like viruses by an engineered human monoclonal antibody.Science. 2021; 371: 823-829Crossref PubMed Scopus (90) Google Scholar Using the AAI network model, we established that Omicron mutations at sites 496, 498, 501, and 505 are networked to the critical ADG-2 epitope residues D405, G502, G504, and Y505 (Figure S3). Rappazzo et al. further reported that Y505 C/N/S knocked down ADG-2 binding potency.28Garrett Rappazzo C. Tse L.v. Kaku C.I. Wrapp D. Sakharkar M. Huang D. Deveau L.M. Yockachonis T.J. Herbert A.S. Battles M.B. et al.Broad and potent activity against SARS-like viruses by an engineered human monoclonal antibody.Science. 2021; 371: 823-829Crossref PubMed Scopus (90) Google Scholar Single point mutation analysis on an epitope does not consider the extent to which mutations at proximal residues may affect the epitope-paratope binding interaction. For example, ADG-2 does not bind the RaTG13 RBD, and RatG13 shares sequence identity with Omicron at the four critical ADG-2 binding sites 405, 502, 504, and 505, demonstrating the importance of the network context of these four critical sites. Since the submission of this manuscript, Adagio reported a 300-fold knockdown of ADG-2 neutralization potency against Omicron in both pseudovirus and live virus experiments,29Adagio Therapeutics Adagio therapeutics reports reduction in in vitro neutralizing activity of ADG20 against Omicron SARS-CoV-2 variant.https://investors.adagiotx.com/news-releases/news-release-details/adagio-therapeutics-reports-reduction-vitro-neutralizingDate: 2021Google Scholar consistent with AAI network observations and highlighting the importance of indirect effects in assessing monoclonal antibody function when multiple mutations occur within or adjacent to an epitope. Given the substantial transmission advantage of Omicron over Delta, it is plausible that Omicron RBD mutations contribute to enhanced transmission by mechanisms other than or in addition to antibody escape. Above (Figure 1), we identified several Omicron mutations that do not appear to play a major role as polyclonal epitope constituents or whose predicted contribution to antibody evasion is unlikely to confer a substantial fitness advantage due to the mutations occurring in class 4 antibody epitopes. Here, we highlight two mechanisms by which these RBD mutations may contribute to Omicron’s fitness via enhanced transmissibility. There is existing evidence that certain Omicron mutations enhance ACE-2 binding, and this is a leading hypothesis to explain Omicron’s enhanced transmission. In particular, Zahradník et al. identified enhanced ACE2 binding by RBD’s bearing Q498R and S477N when combined with the N501Y mutation.30Zahradník J. Marciano S. Shemesh M. Zoler E. Harari D. Chiaravalli J. Meyer B. Rudich Y. Li C. Marton I. et al.SARS-CoV-2 variant prediction and antiviral drug design are enabled by RBD in vitro evolution.Nat. Microbiol. 2021; 6: 1188-1198Crossref PubMed Scopus (73) Google Scholar Importantly, these mutation combinations resulted in a synergistic effect extending beyond the increased ACE-2 binding predicted when the effect of these mutations assayed individually was summed. It is therefore likely that a similar synergy effect occurs on the Omicron variant at these three sites and may also include additional Omicron mutations. Using AAI networks to examine the local residue dependencies in this vicinity, we observed a network extending from the known synergistic pair 498 + 501 to position 505, suggesting Omicron H505 may modulate the synergistic ACE-2 binding effect for Q498R + N501Y. While Omicron’s mutations at sites 371, 373, and 375 may contribute to escape from class 3 and 4 antibodies, evolution of such a triplet mutation toward this escape function is not parsimonious given that other single mutations have previously been identified as conferring greater class 3 and 4 antibody escape.5Greaney A.J. Starr T.N. Barnes C.O. Weisblum Y. Schmidt F. Caskey M. Gaebler C. Cho A. Agudelo M. Finkin S. et al.Mapping mutations to the SARS-CoV-2 RBD that escape binding by different classes of antibodies.Nat. Commun. 2021; 12 (2021 12:1 12, 1–14): 4196Crossref PubMed Scopus (61) Google Scholar We therefore hypothesized that mutations S371L, S373P, and S375F might provide a fitness advantage oth
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