A Highly Immunogenic Measles Virus-based Th1-biased COVID-19 Vaccine

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The COVID-19 pandemic is caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and has spread world-wide with millions of cases and hundreds of thousands of deaths to date. The gravity of the situation mandates accelerated efforts to identify safe and effective vaccines. Here, we generated measles virus (MeV)-based vaccine candidates expressing the SARS-CoV-2 spike glycoprotein (S). Insertion of the full-length S protein gene in two different MeV genomic positions resulted in modulated S protein expression. The variant with lower S protein expression levels was genetically stable and induced high levels of effective Th1-biased antibody and T cell responses in mice after two immunizations. In addition to neutralizing IgG antibody responses in a protective range, multifunctional CD8 + and CD4 + T cell responses with S protein-specific killing activity were detected. These results are highly encouraging and support further development of MeV-based COVID-19 vaccines.

Author Contributions

CH performed research, analyzed data, and wrote the paper; CS performed research and analyzed data; AA performed research and analyzed data; AE performed research and analyzed data; SM performed research, analyzed data, and wrote the paper; MH developed the bioinformatics pipeline and analyzed data; BS contributed new reagents and concepts; MDM designed and supervised research, analyzed data and wrote the paper; all authors read, corrected and approved the final manuscript.

Significance Statement

The COVID-19 pandemic has caused hundreds of thousands of deaths, yet. Therefore, effective vaccine concepts are urgently needed. In search for such a concept, we have analysed a measles virus-based vaccine candidate targeting SARS-CoV-2. Using this well known, safe vaccine backbone, we demonstrate here induction of functional immune responses in both arms of adaptive immunity with the desired immune bias. Therefore, occurrence of immunopathologies such as antibody-dependent enhancement or enhanced respiratory disease is rather unlikely. Moreover, the candidate still induces immunity against the measles, recognized as a looming second menace, when countries are entrapped to stop routine vaccination campaigns in the face of COVID-19. Thus, a bivalent measles-based COVID-19 vaccine could be the solution for two significant public health threats.

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  1. SciScore for 10.1101/2020.07.11.198291: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    RandomizationFor the experiments, animals were randomized for age- and sex-matched groups and housed in IVC cages in groups of 3 to 5 animals with nist packs as environmental enrichment at room temperature with regular 12 h day and night intervalls.
    Blindingnot detected.
    Power AnalysisGroup sizes were calculated based on statistical considerations to yield sufficient statistical power as authorized by the respective competent authority.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    The cells were then probed for 1 h with a polyclonal rabbit anti-SARS-CoV-2-S protein antibody (1:2,250; ab252690; Abcam, Cambridge, UK) or a rabbit anti-MeV N protein antibody (1:1,000, ab23974, Abcam) in PBS with 2% BSA.
    anti-SARS-CoV-2-S protein
    suggested: None
    anti-MeV N protein
    suggested: None
    The cells were washed 3 times with 1 ml PBS and subsequently incubated with the secondary HRP-coupled donkey anti-rabbit IgG(H+L) polyclonal antibody (1:1,000; 611-7202; Rockland, Gilbertsville, USA) for 1 h at 37°C.
    anti-rabbit IgG(H+L
    suggested: (Rockland Cat# 611-7202, RRID:AB_219746)
    Rabbit anti-SARS-S protein antibody (1:3,000; ab252690; Abcam), rabbit anti-MeV-N protein polyclonal antibody (1:5,000; ab23974; Abcam), and a mouse anti-ß-actin antibody (1:5,000; ab6276; Abcam) were used.
    anti-SARS-S protein
    suggested: None
    suggested: None
    suggested: (Abcam Cat# ab6276, RRID:AB_2223210)
    Donkey anti-rabbit IgG-HRP (H&L) polyclonal antibody (1:10,000; 611-7202; Rockland) and goat anti-mouse IgG-HRP (1:10,000; A2554-1ML; Merck, Darmstadt, Germany) served as secondary antibodies.
    anti-rabbit IgG-HRP
    suggested: None
    anti-mouse IgG-HRP
    suggested: None
    IFN-γ ELISpot Analysis: Murine interferon gamma (IFN-γ) enzyme-linked immunosorbent spot (ELISpot) assays were performed using the Mouse IFN-γ ELISPOT Pair kit including capture and detection antibody (BD Bioscience, Franklin Lakes, NJ, USA) and HRP Streptavidin (BD Bioscience) for ELISpot detection in combination with multiscreen immunoprecipitation (IP) ELISpot polyvinylidene difluoride (PVDF) 96-well plates (Merck Millipore, Darmstadt, Germany) according to the manufacturer’s instructions.
    suggested: None
    Stimulated cells were subsequently stained with CD3-PacBlue (1:50; clone 500A2; Invitrogen Life Technologies), CD8-APC (1:100; clone 53-6.7; ebioscience) and CD4-PE (1:2000; Cat. 553049; BD) antibodies and fixed with 1% PFA in PBS.
    suggested: None
    suggested: None
    suggested: (BD Biosciences Cat# 553049, RRID:AB_394585)
    Experimental Models: Cell Lines
    Cells: Vero (African green monkey kidney) (ATCC# CCL-81), Vero clone E6 (ATCC# CRL-1586), 293T (ATCC CRL-3216) and EL-4 (ATCC TIB-39) cell lines were purchased from ATCC (Manassas, VA, USA) and cultured in Dulbecco’s modified Eagle’s medium (DMEM, Biowest, Nuaillé, France) supplemented with 10% fetal bovine serum (FBS; Biochrom, Berlin, Germany) and 2 mM L-glutamine (L-Gln; Biochrom).
    Vero clone E6
    suggested: None
    suggested: None
    Subsequent purification by filtration and ultracentrifugation of supernatants yielded virus stocks were used to transduce murine DC cell lines, DC2.4 and JAWSII, as well as the murine T cell line EL-4, resulting in DC2.4-SARS2-S, JAWSII-SARS2-S, and EL-4green-SARS2-S, respectively, that express the SARS-CoV-2 S protein and GFP and present the respective peptides via MHC-I.
    suggested: None
    Single syncytia were picked and overlaid onto 50% confluent Vero cells cultured in 6-well plates and harvested as “passage 0” (P0) by scraping and freeze-thaw cycle of cells at the time of maximal infection.
    suggested: None
    It was propagated on Vero E6 cells and was titrated via TCID50 as described above for recombinant MeV.
    Vero E6
    suggested: None
    200 μL Medium containing 10 µg/ml Concanavalin A (Con A, Sigma-Aldrich), 10 μg/ml MeV bulk antigen (Virion Serion), or 5×103 DC2.4-SARS2-S cells were added to each well, and cultured for 6 days.
    suggested: None
    CTL killing assay: For re-stimulation of T cells isolated 3 weeks after the second immunization, 5×106 splenocytes were co-cultured with 5×104 DC2.4-SARS2-S cells for 6 days in 12-wells in RPMI 1640 supplemented with 10% FBS, 2 nM L-Glutamin, 1 mM HEPES, 1% penicillin/streptomycin, 100 μM 2-mercaptoethanol, and 100 U/ml murine rIL-2 (Peprotech, Hamburg, Germany). 5×103 EL-4red cells were labeled with 0.5 µM CFSE and mixed with 5×103 EL-4green-SARS2-S cells per well.
    suggested: None
    suggested: None
    Software and Algorithms
    NGS library preparation and sequencing: Total RNA was isolated from Vero cells after 4 days post infection using the Direct-zol RNA isolation kit (Zymo Research).
    suggested: (PM4NGS, RRID:SCR_019164)
    Mapping was performed with BWA mem v 0.7.12-r1039 (43), using default parameters unless stated otherwise.
    suggested: (BWA, RRID:SCR_010910)
    Host-derived reads were removed by mapping quality controlled reads against the African green monkey genome (Chlorocebus sabeus, RefSeq assembly GCA_000409795.2), specifying the minimum seed length (-k 31).
    suggested: (RefSeq, RRID:SCR_003496)
    Unmapped reads were extracted using samtools v1.7 (44) and bamToFastq v2.17.0 (45), and subsequently mapped to the plasmid reference genomes of either MeVvac2-SARS2-S(H) or MeVvac2-SARS2-S(P), as appropriate.
    suggested: (SAMTOOLS, RRID:SCR_002105)
    Host-free alignments were deduplicated using picard-tools MarkDuplicates (http://broadinstitute.github.io/picard) and left-aligned using GATK LeftAlignIndels v4.0 (46).
    suggested: (GATK, RRID:SCR_001876)
    Variant calling was performed with LoFreq v2.1.3 (47) using default parameters.
    suggested: (LoFreq, RRID:SCR_013054)

    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: Please consider improving the rainbow (“jet”) colormap(s) used on page 39. 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.
    • 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.

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