Virus Induced Lymphocytes (VIL) as a novel viral antigen-specific T cell therapy for COVID-19 and potential future pandemics

  1. Rohan Sivapalan
  2. Jinyan Liu
  3. Krishnendu Chakraborty
  4. Elisa Arthofer
  5. Modassir Choudhry
  6. Philip S. Barie
  7. Dan H. Barouch
  8. Tom Henley

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Abstract

The a priori T cell repertoire and immune response against SARS-CoV-2 viral antigens may explain the varying clinical course and prognosis of patients having a mild COVID-19 infection as opposed to those developing more fulminant multisystem organ failure and associated mortality. Using a novel SARS-Cov-2-specific artificial antigen presenting cell (aAPC), coupled with a rapid expansion protocol (REP) as practiced in tumor infiltrating lymphocytes (TIL) therapy, we generate an immune catalytic quantity of Virus Induced Lymphocytes (VIL). Using T cell receptor (TCR)-specific aAPCs carrying co-stimulatory molecules and major histocompatibility complex (MHC) class-I immunodominant SARS-CoV-2 peptide-pentamer complexes, we expand virus-specific VIL derived from peripheral blood mononuclear cells (PBMC) of convalescent COVID-19 patients up to 1000-fold. This is achieved in a clinically relevant 7-day vein-to-vein time-course as a potential adoptive cell therapy (ACT) for COVID-19. We also evaluate this approach for other viral pathogens using Cytomegalovirus (CMV)-specific VIL from donors as a control. Rapidly expanded VIL are enriched in virus antigen-specificity and show an activated, polyfunctional cytokine profile and T effector memory phenotype which may contribute to a robust immune response. Virus-specific T cells can also be delivered allogeneically via MHC-typing and patient human leukocyte antigen (HLA)-matching to provide pragmatic treatment in a large-scale therapeutic setting. These data suggest that VIL may represent a novel therapeutic option that warrants further clinical investigation in the armamentarium against COVID-19 and other possible future pandemics.

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  1. Version published to 10.1038/s41598-021-94654-y
  2. ScreenIT

    SciScore for 10.1101/2020.11.26.400390: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Preparation of VIPR Particles: Micro-aAPC VIPR particles were constructed by direct conjugation of biotin labelled peptide-MHC-Pentamers and biotinylated MHC class II Tetramers to streptavidin Dynabeads in combination with biotinylated anti-CD28 antibodies.
    anti-CD28
    suggested: None
    These biotin-labelled MHC-peptide Pentamers and biotinylated MHC-peptide Monomer complexes, and mouse anti-human CD28 antibody (BD Biosciences) were conjugated to M270 Streptavidin Dynabeads (Thermo Scientific) at defined molar ratios of Pentamer:anti-CD28 and Monomer:anti-CD28 (calculated to account for tetramer formation of these monomers), and both in molar excess of the number of streptavidin molecules per Dynabead.
    anti-human CD28
    suggested: None
    Flow cytometry analysis of T cell phenotype: For flow cytometric analysis of the antigen-specific T cell population and cell surface marker expression, cells were harvested from culture plates and washed using PBS with 1% Bovine Serum Albumen (Thermo Scientific) and were then stained with monoclonal antibodies specific for CD8 (HIT8A, 1:100), CD4 (OKT4, 1:100), HLA-DR (L243 1:80), LAG-3 (11C3C65, 1:80), TIGIT (VSTM3, 1:40), CD45RO (UCHL1, 1:40), CD45RA (HI100, 1:80), TIM3 (F38-2E2, 1:40), CD62L (DREG-56, 1:40), CD57 (QA17A04, 1:80), PD-1 (EH12.1, 1:40), OX-40 (Ber-ACT35, 1:40), CD25 (MA2-51, 1:40), 41BB (4B4-1, 1:40), (Biolegend) or specific for CD8 (RPA-T8, 1:100) (BD Bioscience), or TNF-a (MAb11, 1:40) and CD3 (UCHT1, 1:100) (ThermoFisher).
    CD8
    suggested: (BD Biosciences Cat# 340574, RRID:AB_400477)
    HIT8A
    suggested: None
    CD4
    suggested: (Cell Sciences Cat# 873.019.050, RRID:AB_10048825)
    OKT4
    suggested: None
    HLA-DR
    suggested: (MabTag GmbH Cat# hHLADR-04, RRID:AB_11128965)
    LAG-3 (11C3C65
    suggested: None
    VSTM3
    suggested: (Thermo Fisher Scientific Cat# 51-9501-82, RRID:AB_10719115)
    CD45RO
    suggested: (GeneTex Cat# GTX19741, RRID:AB_423713)
    UCHL1
    suggested: None
    CD45RA
    suggested: None
    TIM3
    suggested: None
    CD62L
    suggested: None
    CD57
    suggested: None
    PD-1
    suggested: None
    CD25
    suggested: None
    CD3
    suggested: None
    UCHT1
    suggested: None
    Cells were then stained for surface markers followed by intracellular cytokines using antibodies specific for IFN-y (4S.B3, 1:40) (Biolegend) IL-2 (MQ1-17H12, 1:40) (BD Bioscience), or TNF-a (MAb11, 1:40) (ThermoFisher).
    IL-2
    suggested: None
    MQ1-17H12
    suggested: None
    TNF-a
    suggested: None
    Software and Algorithms
    SentencesResources
    After staining, cells were resuspended in PBS with 2% Human Heat Inactivated AB Serum (Sigma) and 0.1M EDTA pH 8.0 (Invitrogen) before analysis on a Fortessa flow cytometer (BD Bioscience) and data analyzed using FlowJo 10 software (BD Biosciences).
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    T cells were then harvested, and cells fixed and permeabilized using BD Cytofix/Cytoperm Fixation/Permeabilization Solution (ThermoFisher).
    BD Cytofix/Cytoperm Fixation/Permeabilization Solution
    suggested: None
    Statistical analyses: Statistical differences between two sample groups, where appropriate, was analyzed by a standard Student’s two-tailed, non-paired, t test using GraphPad Prism Software version 8.
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)

    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: 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.
    • No funding statement was detected.
    • No protocol registration statement was detected.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  3. Version published to 10.1101/2020.11.26.400390v1 on bioRxiv