BA.2.12.1, BA.4 and BA.5 escape antibodies elicited by Omicron infection

  1. Yunlong Cao
  2. Ayijiang Yisimayi
  3. Fanchong Jian
  4. Weiliang Song
  5. Tianhe Xiao
  6. Lei Wang
  7. Shuo Du
  8. Jing Wang
  9. Qianqian Li
  10. Xiaosu Chen
  11. Yuanling Yu
  12. Peng Wang
  13. Zhiying Zhang
  14. Pulan Liu
  15. Ran An
  16. Xiaohua Hao
  17. Yao Wang
  18. Jing Wang
  19. Rui Feng
  20. Haiyan Sun
  21. Lijuan Zhao
  22. Wen Zhang
  23. Dong Zhao
  24. Jiang Zheng
  25. Lingling Yu
  26. Can Li
  27. Na Zhang
  28. Rui Wang
  29. Xiao Niu
  30. Sijie Yang
  31. Xuetao Song
  32. Yangyang Chai
  33. Ye Hu
  34. Yansong Shi
  35. Linlin Zheng
  36. Zhiqiang Li
  37. Qingqing Gu
  38. Fei Shao
  39. Weijin Huang
  40. Ronghua Jin
  41. Zhongyang Shen
  42. Youchun Wang
  43. Xiangxi Wang
  44. Junyu Xiao
  45. Xiaoliang Sunney Xie

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Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron sublineages BA.2.12.1, BA.4 and BA.5 exhibit higher transmissibility than the BA.2 lineage 1 . The receptor binding and immune-evasion capability of these recently emerged variants require immediate investigation. Here, coupled with structural comparisons of the spike proteins, we show that BA.2.12.1, BA.4 and BA.5 (BA.4 and BA.5 are hereafter referred collectively to as BA.4/BA.5) exhibit similar binding affinities to BA.2 for the angiotensin-converting enzyme 2 (ACE2) receptor. Of note, BA.2.12.1 and BA.4/BA.5 display increased evasion of neutralizing antibodies compared with BA.2 against plasma from triple-vaccinated individuals or from individuals who developed a BA.1 infection after vaccination. To delineate the underlying antibody-evasion mechanism, we determined the escape mutation profiles 2 , epitope distribution 3 and Omicron-neutralization efficiency of 1,640 neutralizing antibodies directed against the receptor-binding domain of the viral spike protein, including 614 antibodies isolated from people who had recovered from BA.1 infection. BA.1 infection after vaccination predominantly recalls humoral immune memory directed against ancestral (hereafter referred to as wild-type (WT)) SARS-CoV-2 spike protein. The resulting elicited antibodies could neutralize both WT SARS-CoV-2 and BA.1 and are enriched on epitopes on spike that do not bind ACE2. However, most of these cross-reactive neutralizing antibodies are evaded by spike mutants L452Q, L452R and F486V. BA.1 infection can also induce new clones of BA.1-specific antibodies that potently neutralize BA.1. Nevertheless, these neutralizing antibodies are largely evaded by BA.2 and BA.4/BA.5 owing to D405N and F486V mutations, and react weakly to pre-Omicron variants, exhibiting narrow neutralization breadths. The therapeutic neutralizing antibodies bebtelovimab 4 and cilgavimab 5 can effectively neutralize BA.2.12.1 and BA.4/BA.5, whereas the S371F, D405N and R408S mutations undermine most broadly sarbecovirus-neutralizing antibodies. Together, our results indicate that Omicron may evolve mutations to evade the humoral immunity elicited by BA.1 infection, suggesting that BA.1-derived vaccine boosters may not achieve broad-spectrum protection against new Omicron variants.

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  1. Version published to 10.1038/s41586-022-04980-y
  2. Version published to 10.1101/2022.04.30.489997v2 on bioRxiv
  3. ScreenIT

    SciScore for 10.1101/2022.04.30.489997: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    NIH rigor criteria are not applicable to paper type.

    Table 2: Resources

    Antibodies
    SentencesResources
    B cells were then stained with FITC anti-human CD19 antibody (BioLegend, 392508)
    anti-human CD19
    suggested: (BioLegend Cat# 392508, RRID:AB_2750099)
    FITC anti-human CD20 antibody (BioLegend, 302304)
    anti-human CD20
    suggested: (BioLegend Cat# 302304, RRID:AB_314252)
    Brilliant Violet 421™ anti-human CD27 antibody (BioLegend, 302824)
    anti-human CD27
    suggested: (BioLegend Cat# 302824, RRID:AB_11150782)
    anti-human IgM antibody (BioLegend, 314532)
    anti-human IgM
    suggested: (BioLegend Cat# 314532, RRID:AB_2566485)
    B cells were then stained with FITC anti-human CD20 antibody, Brilliant Violet 421™ anti-human CD27 antibody, PE/Cyanine7 anti-human IgM antibody, PE/Cyanine7 anti-human IgD antibody(BioLegend, 348210), biotinylated SARS-CoV-2 BA.1
    anti-human IgD
    suggested: (BioLegend Cat# 348210, RRID:AB_10680462)
    High-throughput antibody-escape mutation profiling: The previously described high-throughput MACS (magnetic-activated cell sorting)-based antibody-escape mutation profiling system3, 17 was used to characterize mutation escape profile for neutralizing antibodies.
    antibody-escape
    suggested: None
    antibody-escape mutation profiling system3
    suggested: None
    A, E, n are parameters, where E is the desired EC50 value for the specific antibody and antigen.
    antigen .
    suggested: None
    After blocking, the plates were washed five times, and the mixture of ACE2-biotin (Sino Biological, 10108-H27B-B) and serially diluted competitor antibodies was added followed by 30min incubation at RT.
    ACE2-biotin
    suggested: None
    10108-H27B-B
    suggested: None
    For protein production, these expression plasmids, as well as the plasmids encoding the antigen-binding fragments (Fabs) of the antibodies described in this paper, were transfected into the HEK293F cells using polyethylenimine (Polysciences).
    antigen-binding
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    G*ΔG-VSV virus (VSV G pseudotyped virus, Kerafast) is used to infect 293T cells (American Type Culture Collection [ATCC], CRL-3216), and spike protein expressing plasmid was used for transfection at the same time.
    293T
    suggested: None
    After incubation at 5% CO2 and 37℃ for 1 h, digested Huh-7 cell (Japanese Collection of Research Bioresources [JCRB]
    Huh-7
    suggested: CLS Cat# 300156/p7178_HuH7, RRID:CVCL_0336)
    , 0403) or 293T-hACE2 cells (AmericanTypeCultureCollection[ATCC],CRL-3216) were seeded.
    293T-hACE2
    suggested: None
    Healthy HEK293 cells were passaged into a new cell culture and grown in suspension at 37 °C, 120 RPM, 8% CO2 to logarithmic growth phase and transfected with the recombinant constructs by using liposomal vesicles as DNA carrier.
    HEK293
    suggested: None
    For protein production, these expression plasmids, as well as the plasmids encoding the antigen-binding fragments (Fabs) of the antibodies described in this paper, were transfected into the HEK293F cells using polyethylenimine (Polysciences).
    HEK293F
    suggested: RRID:CVCL_6642)
    Recombinant DNA
    SentencesResources
    Pseudovirus neutralization assay: SARS-CoV-2 spike (GenBank: MN908947) , Pangolin-GD spike (GISAID: EPI_ISL_410721), RaTG13 spike (GISAID: EPI_ISL_402131), SARS-CoV-1 spike (GenBank: AY278491), Omicron BA.1 spike (A67V, H69del, V70del, T95I, G142D, V143del, Y144del, Y145del, N211del, L212I, ins214EPE, G339D, S371L, S373P, S375F, K417N, N440K, G446S, S477N, T478K, E484A, Q493R, G496S, Q498R, N501Y, Y505H, T547K, D614G, H655Y, N679K, P681H, N764K, D796Y, N856K, Q954H, N969K, L981F), BA.2 spike (GISAID: EPI_ISL_7580387, T19I, L24S, del25-27, G142D, V213G, G339D, S371F, S373P, S375F, T376A, D405N, R408S, K417N, N440K, G446S, S477N, T478K, E484A, Q493R, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, N969K), BA.1.1 spike (BA.1+R346K), BA.3 spike (A67V, del69-70, T95I, G142D, V143del, Y144del, Y145del, N211del, L212I, G339D, S371F, S373P, S375F, D405N, K417N, N440K, G446S, S477N, T478K, E484A, Q493R, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, N969K), BA.2.12.1 spike (BA.2+L452Q+S704L), BA.2.13 spike (BA.2+L452M), BA.4 spike (T19I, L24S, del25-27, del69-70, G142D, V213G, G339D, S371F, S373P, S375F, T376A, D405N, R408S, K417N, N440K, G446S, L452R, S477N, T478K, E484A, F486V, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, N969K) plasmid is constructed into pcDNA3.1 vector.
    pcDNA3.1
    suggested: RRID:Addgene_79663)
    The purified PCR products were ligated to the secretory expression vector pCMV3 with CMV promoter, and then transformed into E. coli competent cells XL1-blue.
    pCMV3
    suggested: RRID:Addgene_161029)
    Software and Algorithms
    SentencesResources
    FACS data were analyzed using FlowJo™ v10.8 (BD Biosciences)
    FlowJo™
    suggested: (FlowJo, RRID:SCR_008520)
    V(D)J gene annotation was performed using NCBI IgBlast (v1.17.1) with the IMGT reference.
    IgBlast
    suggested: (IgBLAST, RRID:SCR_002873)
    The V-J pairs were visualized by R package circlize (v0.4.10).
    circlize
    suggested: (circlize, RRID:SCR_002141)
    VarianceThreshold of scikit-learn Python package (v0.24.2) with the variance threshold as 0.1.
    Python
    suggested: (IPython, RRID:SCR_001658)
    KMeans of scikit-learn in the resulting D-dimensional feature space.
    scikit-learn
    suggested: (scikit-learn, RRID:SCR_002577)
    All t-SNE plots were generated by R package ggplot2 (v3.3.3)
    ggplot2
    suggested: (ggplot2, RRID:SCR_014601)
    To improve the density surrounding the RBD-Fab region, UCSF Chimera49 and Relion50 were used to generate the masks, and local refinement was then performed using cryoSPARC.
    cryoSPARC
    suggested: (cryoSPARC, RRID:SCR_016501)
    Figures were prepared using USCF ChimeraX53 and Pymol (Schrödinger, LLC.).
    Pymol
    suggested: (PyMOL, RRID:SCR_000305)
    Raw Illumina and PacBio sequencing data are available on NCBI Sequence Read Archive BioProject PRJNA804413.
    NCBI Sequence Read Archive BioProject
    suggested: None

    Results from OddPub: Thank you for sharing your code and data.

    Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

    Results from JetFighter: We did not find any issues relating to colormaps.

    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
    • No protocol registration statement was detected.

    Results from scite Reference Check: We found no unreliable references.

    About SciScore

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  4. Version published to 10.1101/2022.04.30.489997v1 on bioRxiv
  5. Version published to 10.21203/rs.3.rs-1611421/v1 on Research Square