Directing an mRNA-LNP vaccine toward lymph nodes improves humoral and cellular immunity against SARS-CoV-2
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Abstract
The exploration and identification of safe and effective vaccines for the SARS-CoV-2 pandemic has captured the world’s attention and remains an ongoing issue in order to protect against emerging variants of concern (VoCs) while generating long lasting immunity. Here, we report the synthesis of a novel messenger ribonucleic acid (mRNA) encoding the spike protein in a lipid nanoparticle formulation (LNP) (STI-7264) that generates robust humoral and cellular immunity following immunization of C57Bl6 mice. In efforts to continually improve immunity, a lymphatic drug delivery device (MuVaxx) was engineered and tested to modulate immune cells at the injection site (epidermis and dermis) and draining lymph node (LN) to elicit adaptive immunity. Using MuVaxx, immune responses were elicited and maintained at a 10-fold dose reduction compared to traditional intramuscular (IM) administration as measured by anti-spike antibodies, cytokine producing CD8 T cells, and neutralizing antibodies against the Washington (Wild Type, WT) and South African (beta) variants. Remarkably, a 4-fold elevated T cell response was observed in MuVaxx administered vaccination as compared to that of IM administered vaccination. Thus, these data support further investigation into STI-7264 and lymphatic mediated delivery using MuVaxx for SARS-CoV-2 and VoCs vaccines.
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SciScore for 10.1101/2021.08.25.457699: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics IACUC: All protocols were approved by the Institutional Animal Care and Use Committee (IACUC). Sex as a biological variable not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources 24 hours post-transfection, the cells were collected and stained with anti-Spike antibody STI-2020 in FACS buffer (DPBS+0.5%BSA) for 30 minutes on ice. anti-Spikesuggested: NoneThereafter, cells were washed twice in FACS buffer and incubated with rat anti-human Fc antibody conjugated to allophycocyanin (Cat: 410712, BioLegend) for 15 minutes on ice. anti-human Fcsuggested: (BioLegend Cat# 410712, RRID:AB_256…SciScore for 10.1101/2021.08.25.457699: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics IACUC: All protocols were approved by the Institutional Animal Care and Use Committee (IACUC). Sex as a biological variable not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources 24 hours post-transfection, the cells were collected and stained with anti-Spike antibody STI-2020 in FACS buffer (DPBS+0.5%BSA) for 30 minutes on ice. anti-Spikesuggested: NoneThereafter, cells were washed twice in FACS buffer and incubated with rat anti-human Fc antibody conjugated to allophycocyanin (Cat: 410712, BioLegend) for 15 minutes on ice. anti-human Fcsuggested: (BioLegend Cat# 410712, RRID:AB_2565790)allophycocyaninsuggested: NoneELISA assays: To asses spike specific antibodies, S1 (Cat: 40591-V08H Sino Biological) or RBD (Cat: 40592-V08B, Sino Biological) protein was coated on half-area high binding plates (Cat: N503 Thermo) at 1 μg/mL overnight at 4°C. S1suggested: NoneSecondary antibody of horse radish peroxidase (HRP)-conjugated rabbit anti-mouse IgM (μ chain) anti-mouse IgM ( μ chain)suggested: NoneExperimental Models: Cell Lines Sentences Resources Cell culture media was removed from cells and sera/virus premix was added to VeroE6 cells at 100 μl/well and incubated for 1 hour at 37°C. VeroE6suggested: JCRB Cat# JCRB1819, RRID:CVCL_YQ49)Experimental Models: Organisms/Strains Sentences Resources Animals and in vivo studies: 6- to 12-week old C57Bl6 mice were purchased from the Jackson Laboratory. C57Bl6suggested: NoneRecombinant DNA Sentences Resources In vitro transcription and purification of RNA: To generate the template for RNA synthesis, the sequence of the SARS-Cov-2 Spike protein (GenBank: QHD43416.1) was codon optimized and cloned into pVAX1-based backbone which features 5′-UTR, 3′-UTR and Poly-A tail. pVAX1-basedsuggested: NoneSoftware and Algorithms Sentences Resources Whole blood was stimulated for 6 hours with 1 μg/mL of SIINFEKL (Sigma) or overnight with 1 μg/peptide per well of spike associated (Cat: 130-127-951, Miltenyi Biotec Peptivator) peptides at 37°C, 5% CO2 in the presence of brefeldin A (Biolegend) and monensin (Biolegend). Miltenyi Biotec Peptivatorsuggested: NoneCells were then run on a Beckman Coulter CytoFLEX instrument and analyzed via FlowJo V10 software. FlowJosuggested: (FlowJo, RRID:SCR_008520)Statistics: Statistical significance of differences between experimental groups was determined with Prism software (Graphpad). Prismsuggested: (PRISM, RRID:SCR_005375)Graphpadsuggested: (GraphPad, RRID:SCR_000306)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.
Results from scite Reference Check: We found no unreliable references.
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