The Storage and In-Use Stability of mRNA Vaccines and Therapeutics: Not A Cold Case

Blenke, Erik Oude; Örnskov, Eivor; Schöneich, Christian; Nilsson, Gunilla Åkesson; Volkin, David B.; Mastrobattista, Enrico; Almarsson, Ӧrn; Crommelin, Daan J.A.

doi:10.1016/j.xphs.2022.11.001

Cited by 75 publications

(50 citation statements)

References 122 publications

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Section: Resultsmentioning

confidence: 99%

“…These NPs are "softer" than the standard bilayer liposomes or other LNPs with fuller membrane coverage, which may explain the diluted vaccine's limited stability on ambient temperature. [42][43][44] Despite its softness, and, hence, limited stability, the vaccine is clearly very effective in inducing an immune response, raising the possibility that the above newly recognized structural features are contributing to, or may be essential for its immunogenic function. Our data may therefore lay the grounds for a paradigm shift in understanding the structure-function relationship in mRNA vaccines, as well as contribute to developing better imaging of LNP core structure.…”

Section: Discussionmentioning

confidence: 99%

“…These NPs are “softer” than the standard bilayer liposomes or other LNPs with fuller membrane coverage, which may explain the diluted vaccine’s limited stability on ambient temperature. 42–44…”

Section: Discussionmentioning

confidence: 99%

“…Beyond the acidic milieu, the other likely contributing factor to LNP disintegration is the limited stability of the vaccine in water, as reflected in the brief (up to 6h) shelf-life of Comirnaty on RT after dilution. [42][43][44] (including proton) concentration. Changes and stability of intra-NP proton concentration can be determined for the pH gradient (NP pH << medium pH).…”

Section: Dls Analysis Of the Hydrodynamic Size And Size Distribution ...mentioning

confidence: 99%

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New insights into the structure of Comirnaty Covid-19 vaccine: A theory on soft nanoparticles with mRNA-lipid supercoils stabilized by hydrogen bonds

Szebeni

Kiss

Bozó

et al. 2022

Preprint

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Despite the worldwide success of mRNA-LNP Covid-19 vaccines, the nanoscale structure of these formulations is still poorly understood. To fill this gap, we used a combination of atomic force microscopy (AFM), dynamic light scattering (DLS), transmission electron microscopy (TEM), cryogenic transmission electron microscopy (cryo-TEM) and the determination of LNP pH gradient to analyze the nanoparticles (NPs) in BNT162b2 (Comirnaty), comparing it with the well characterized pegylated liposomal doxorubicin (Doxil). Comirnaty NPs had similar size to Doxil, however, unlike Doxil liposomes, wherein the stable ammonium and pH gradient enables accumulation of 14C-methylamine in the intraliposomal aqueous phase, Comirnaty LNPs lack such pH gradient in spite of the fact that the pH 4, at which LNPs are prepared, is raised to pH 7.2 after loading of the mRNA. Mechanical manipulation of Comirnaty NPs with AFM revealed soft, compliant structures. The sawtooth-like force transitions seen during cantilever retraction implies that molecular strands, corresponding to mRNA, can be pulled out of NPs, and the process is accompanied by stepwise rupture of mRNA-lipid bonds. Unlike Doxil, cryo-TEM of Comirnaty NPs revealed a granular, solid core enclosed by mono- and bilayers. Negative staining TEM shows 2-5 nm electron-dense spots in the liposom's interior that are aligned into strings, semicircles, or labyrinth-like networks, which may imply crosslink-stabilized supercoils. The neutral intra-LNP core questions the dominance of ionic interactions holding together this scaffold, raising the alternative possibility of hydrogen bonding between the mRNA and the lipids. Such interaction, described previously for another mRNA/lipid complex, is consistent with the steric structure of ionizable lipid in Comirnaty, ALC-0315, displaying free =O and -OH groups. It is hypothesized that the latter groups can get into steric positions that enable hydrogen bonding with the nitrogenous bases in the mRNA. These newly recognized structural features of mRNA-LNP may be important for the vaccine's efficacy.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Dls Analysis Of the Hydrodynamic Size And Size Distribution ...mentioning

confidence: 99%

See 2 more Smart Citations

New insights into the structure of Comirnaty Covid-19 vaccine: A theory on soft nanoparticles with mRNA-lipid supercoils stabilized by hydrogen bonds

Szebeni

Kiss

Bozó

et al. 2022

Preprint

View full text Add to dashboard Cite

show abstract

“…alternative poly(A) tail designs 13 ) or the potential introduction of hidden open reading frames within the gene of interest or during construct design and optimization 28 . In the formulated vaccine drug product, the lipid nanoparticle protects the mRNA from hydrolytic fragmentation; however, additional mRNA degradation mechanisms (e.g., through interactions with lipid excipients in the formulated drug product 29 , 30 ) can potentially affect antigen translation. While not in scope of the present study, these non-hydrolytic degradation mechanisms should be explored and appropriately controlled in the final drug product.…”

Section: Discussionmentioning

confidence: 99%

Characterization of BNT162b2 mRNA to Evaluate Risk of Off-Target Antigen Translation

Patel

Zhang

Utegg

et al. 2023

Journal of Pharmaceutical Sciences

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An Ultrahydrating Polymer that Protects Protein Therapeutics and RNA‐Lipid Nanoparticles Against Freezing, Heat and Lyophilization Stress

Herrmann,

Abbina,

Bathula

et al. 2024

Adv Funct Materials

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RNA and protein‐based therapeutics constitute almost half of recent drug approvals and receive considerable attention within biotechnology industries. Ensuring their stability and longevity in the context of heat, freezing, and lyophilization processes are paramount to a successful deployment. However, the advancement of formulations designed to achieve this goal is still in its nascent phase. To address these challenges, a new class of semi‐dendritic hydrophilic polymer with extended linear units is reported, which showcase very high hydration. These novel polymers demonstrated exceptional efficacy in preserving messenger RNA‐ (mRNA‐) and self‐amplifying RNA‐ (saRNA‐) lipid nanoparticles during freezing and lyophilization. Additionally, they have been found to protect therapeutic proteins against external stressors such as freezing, heat, and lyophilization. These polymers are non‐toxic, which enables their utilization at high concentrations and eliminates the requirement for removal prior to administration. It is found that their unique topology contributes to the high hydration. These excipients are anticipated to create new prospects in biotechnology, food science, and cryopreservation.

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The Storage and In-Use Stability of mRNA Vaccines and Therapeutics: Not A Cold Case

Cited by 75 publications

References 122 publications

New insights into the structure of Comirnaty Covid-19 vaccine: A theory on soft nanoparticles with mRNA-lipid supercoils stabilized by hydrogen bonds

New insights into the structure of Comirnaty Covid-19 vaccine: A theory on soft nanoparticles with mRNA-lipid supercoils stabilized by hydrogen bonds

Characterization of BNT162b2 mRNA to Evaluate Risk of Off-Target Antigen Translation

An Ultrahydrating Polymer that Protects Protein Therapeutics and RNA‐Lipid Nanoparticles Against Freezing, Heat and Lyophilization Stress

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