Mutations of SARS-CoV-2 variants of concern escaping Spike-specific T cells

  1. Nina Le Bert
  2. Anthony Tan
  3. Kamini Kunasegaran
  4. Adeline Chia
  5. Nicole Tan
  6. Qi Chen
  7. Shou Kit Hang
  8. Martin DC Qui
  9. Bianca SW Chan
  10. Jenny GH Low
  11. Barnaby Young
  12. Kee Chong Ng
  13. Derrick Wei Shih Chan
  14. David Chien Lye
  15. Antonio Bertoletti

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Abstract

The amino acid (AA) mutations that characterise the different variants of concern (VOCs), which replaced the ancestral SARS-CoV-2 Wuhan-Hu-1 isolate worldwide, provide biological advantages such as increased infectivity and partial escape from humoral immunity. Here we analysed the impact of these mutations on vaccination- and infection-induced Spike-specific T cells. We confirmed that, in the majority of infected or vaccinated individuals, different mutations present in a single VOC (Delta) or a combined mosaic of more than 30 AA substitutions and deletions found in Alpha, Beta, Gamma, Delta and Omicron VOCs cause modest alteration in the global Spike-specific T cell response. However, distinct numerically dominant Spike-specific CD4 and CD8 T cells preferentially targeted regions affected by AA mutations and do not recognise the mutated peptides. Importantly, some of these mutations, such as N501Y (present in Alpha, Beta, Gamma, and Omicron) and L452R (present in Delta), known to provide biological advantage to SARS-CoV-2 in terms of infectivity also abolished CD8 T cell recognition.

Taken together, our data show that while global mRNA vaccine- and infection-induced Spike-specific T cells largely tolerate the diverse mutations present in VOCs, single Spike-specific T cells might contribute to the natural selection of SARS-CoV-2 variants.

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  1. ScreenIT

    SciScore for 10.1101/2022.01.20.477163: (What is this?)

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

    Table 1: Rigor

    EthicsConsent: Ethics statement: All donors provided written consent.
    IRB: The study was conducted in accordance with the Declaration of Helsinki and approved by the NUS institutional review board (H-20-006) and the SingHealth Centralised Institutional Review Board (reference CIRB/F/2018/2387).
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    ELISpot assay: ELISpot plates (Millipore) were coated with human IFN-γ antibody (1-D1K, Mabtech; 5 μg/ml) overnight at 4°C. 400,000 PBMC were seeded per well and stimulated for 18h with pools of SARS-CoV-1/2 peptides (2 μg/ml).
    IFN-γ
    suggested: None
    anti-CD4 (clone SK3; 3:50), and anti-CD8 (clone SK1; 3:50) antibodies.
    anti-CD4
    suggested: None
    Cells were subsequently fixed and permeabilized using the Cytofix/Cytoperm kit (BD Biosciences-Pharmingen) and stained with anti-IFN-γ (clone 25723, R&D Systems; 1:25) and anti-TNF-α (clone MAb11; 1:25) antibodies and analyzed on a BD-LSR II FACS Scan.
    anti-TNF-α
    suggested: None
    , anti-human CD3 and CD8 antibodies (BD Bioscience).
    CD8
    suggested: None
    The cells were stained with Live/Dead in 1 × PBS for 10 min at room temperature and then stained with anti-human CD3 and CD8 antibodies for 30 min at 4°C.
    anti-human CD3
    suggested: None
    Intracellular cytokine staining was performed with anti-human IFNγ and TNFα (BD Biosciences) antibodies for 30 min at room temperature, followed by washing and analysis by flow cytometry.
    anti-human IFNγ
    suggested: None
    TNFα
    suggested: None
    Software and Algorithms
    SentencesResources
    Data were analyzed by FlowJo (Tree Star Inc.)
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    Spike 491-510-specific TCR α and β chain genes were subcloned into T7 expression vector (p-VAX1), the SARS-CoV-2-TCR mRNA was transcribed in vitro using the mMESSAGE mMACHINE™ T7 ULTRA Transcription Kit (ThermoFisher Scientitic) following the manufacturer’s protocols.
    ThermoFisher Scientitic
    suggested: None

    Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

    Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:
    There are some limitations to this study. We tested the impact of AA substitutions on Spike-specific T cells utilizing peptide-pulsed target cells and not infected cells. This method might overestimate the quantity of T cells specific for SARS-CoV-2 because low affinity T cells might have been quantified and be more sensitive to AA mutations. On the other hand, peptide-pulsed target cells cannot be used to evaluate the impact that AA substitutions might have on the processing of T cell epitopes37. AA outside the T cell epitopes can alter their generation39 and as such we might have underestimate the impact of the mutations on the Spike-specific cellular immunity.

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).

    Results from JetFighter: Please consider improving the rainbow (“jet”) colormap(s) used on pages 24, 25 and 28. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.

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

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

    About SciScore

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  2. Version published to 10.1101/2022.01.20.477163v1 on bioRxiv