Lyophilized mRNA-lipid nanoparticle vaccines with long-term stability and high antigenicity against SARS-CoV-2
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Abstract
Advanced mRNA vaccines play vital roles against SARS-CoV-2. However, most current mRNA delivery platforms need to be stored at −20 °C or −70 °C due to their poor stability, which severely restricts their availability. Herein, we develop a lyophilization technique to prepare SARS-CoV-2 mRNA-lipid nanoparticle vaccines with long-term thermostability. The physiochemical properties and bioactivities of lyophilized vaccines showed no change at 25 °C over 6 months, and the lyophilized SARS-CoV-2 mRNA vaccines could elicit potent humoral and cellular immunity whether in mice, rabbits, or rhesus macaques. Furthermore, in the human trial, administration of lyophilized Omicron mRNA vaccine as a booster shot also engendered strong immunity without severe adverse events, where the titers of neutralizing antibodies against Omicron BA.1/BA.2/BA.4 were increased by at least 253-fold after a booster shot following two doses of the commercial inactivated vaccine, CoronaVac. This lyophilization platform overcomes the instability of mRNA vaccines without affecting their bioactivity and significantly improves their accessibility, particularly in remote regions.
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SciScore for 10.1101/2022.02.10.479867: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics IACUC: Animal experiments were executed by certified staff in Center for Animal Experiments of Wuhan University, approved by the Institutional Animal Care and Use Committee (AUP #WP2021-0607). Sex as a biological variable 2.5 Cell culture: HEK 293T/17 and ACE2-expressing 293T cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 2 mM L-glutamine, 10% FBS (sigma-aldrich) and 1% penicillin/streptomycin (BI) at 37 °C and 5% CO2. 2.6 Animals: Female BALB/c mice age of 6-8 weeks and heterozygous B6/JGpt-H11em1Cin(K18-ACE2)/Gpt mice (K18-hACE2 KI mice) age of 5-7 weeks and body weight of 18-20 g were used. Randomization not detected. Blinding not detected. Power Analysis not … SciScore for 10.1101/2022.02.10.479867: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics IACUC: Animal experiments were executed by certified staff in Center for Animal Experiments of Wuhan University, approved by the Institutional Animal Care and Use Committee (AUP #WP2021-0607). Sex as a biological variable 2.5 Cell culture: HEK 293T/17 and ACE2-expressing 293T cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 2 mM L-glutamine, 10% FBS (sigma-aldrich) and 1% penicillin/streptomycin (BI) at 37 °C and 5% CO2. 2.6 Animals: Female BALB/c mice age of 6-8 weeks and heterozygous B6/JGpt-H11em1Cin(K18-ACE2)/Gpt mice (K18-hACE2 KI mice) age of 5-7 weeks and body weight of 18-20 g were used. Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources Following washes, the secondary antibody, goat Anti-Mouse IgG H&L conjugated HRP (Abcam), was incubated in plate at room temperature for 1 h. goat Anti-Mouse IgG H&L conjugated HRPsuggested: NoneAnti-Mouse IgGsuggested: None2.11 SARS-CoV-2 Delta virus neutralization assay: SARS-CoV-2 Delta-neutralizing antibody was determined by in vitro inhibition of cytopathic effect (CPE), and was performed in the ABSL-3 lab at Center of Laboratory Animal Sciences, Wuhan University (Wuhan, China). CPEsuggested: NoneExperimental Models: Cell Lines Sentences Resources 2.5 Cell culture: HEK 293T/17 and ACE2-expressing 293T cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 2 mM L-glutamine, 10% FBS (sigma-aldrich) and 1% penicillin/streptomycin (BI) at 37 °C and 5% CO2. 2.6 Animals: Female BALB/c mice age of 6-8 weeks and heterozygous B6/JGpt-H11em1Cin(K18-ACE2)/Gpt mice (K18-hACE2 KI mice) age of 5-7 weeks and body weight of 18-20 g were used. HEK 293T/17suggested: ATCC Cat# CRL-11268, RRID:CVCL_1926)293Tsuggested: NoneAfter that, the 293T-ACE2 cells are added and incubated for 24 h. 293T-ACE2suggested: NoneThe virus-serum mixtures were added to Vero-E6 cells seeded in 24-well plate and cultured in a 5% CO2 incubator at 37 °C for 3 days. Vero-E6suggested: NoneExperimental Models: Organisms/Strains Sentences Resources 2.5 Cell culture: HEK 293T/17 and ACE2-expressing 293T cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 2 mM L-glutamine, 10% FBS (sigma-aldrich) and 1% penicillin/streptomycin (BI) at 37 °C and 5% CO2. 2.6 Animals: Female BALB/c mice age of 6-8 weeks and heterozygous B6/JGpt-H11em1Cin(K18-ACE2)/Gpt mice (K18-hACE2 KI mice) age of 5-7 weeks and body weight of 18-20 g were used. BALB/csuggested: NoneB6/JGpt-H11em1Cin(K18-ACE2)/Gptsuggested: NoneK18-hACE2 KIsuggested: NoneSoftware and Algorithms Sentences Resources The mRNA integrity was measured with a gel retardation assay [15] and microfluidic capillary electrophoresis (Agilent Fragment Analyser) [16], as previously reported. Agilent Fragment Analysersuggested: (OMICtools, RRID:SCR_002250)Statistical analysis was mainly processed using Prism software version 8.3 Prismsuggested: (PRISM, RRID:SCR_005375)GraphPad Software Inc. GraphPadsuggested: (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 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 page 9. 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.
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