This project seeks to help improve LNP-mRNA vaccine stability by developing and characterizing LNP-mRNA drug product reference materials that will mimic current COVID-19 vaccine characteristics, deploying LNP-mRNA reconstituted drug product physiochemical and functional critical quality attribute assays for characterization, formulation and stability assessment of reference materials.
Rensselaer and Carnegie Mellon will take the following steps during this project:
Additionally, the team will develop room temperature stable lyo LNP-mRNA drug product formulations and lyo processes which take into account the characteristics of the mRNA payload.
This project will demonstrate freeze-drying approaches for mRNA-based vaccines as a function of mRNA length and structure that will produce stable material at higher temperatures.
This may further impact domestic and global vaccine distribution by simplifying the cold-chain and reducing lost/wasted doses.
By implementing freeze-dried LNP mRNA formulation processes, organizations can significantly reduce cold-chain logistics costs, enabling global vaccine distribution without compromising quality or regulatory compliance. Improved stability minimizes reliance on ultra-cold storage, lowers dose loss from spoilage, and extends shelf life for better inventory management. Simplified storage requirements accelerate distribution to remote and resource-limited regions, supporting faster deployment at scale. Robust analytical validation ensures consistent potency and reliability across vaccine lots, safeguarding performance under varied conditions.
Developed and characterized three potential mRNA vaccine reference materials, (egfp)1, (egfp)2 and (egfp)4, which code for with mRNAs ranging in size from 1179, 1911 and 3375 nucleotides. These mRNAs code for one, two and four concatenated copies of an enhanced green fluorescent protein (EGFP). The largest mRNA is similar in size to the first-generation COVID-19 vaccines.
Developed assays based on micellar capillary electrophoresis, a technique that uses surfactants during electrophoresis, for the stability/ease of disruption of lipid nanoparticles (LNPs) containing mRNAs and for the direct determination of the length (physical integrity) of mRNA released by disrupted LNPs
Developed a mass spectrometry technique comprising partial mRNA digestion with selective enzymes followed by chromatographic separation of the resulting mRNA fragments and mass spectrometric fragment identifications to report on the structural integrity of mRNA in terms of revealing regions protected from digestion by the presence of structure and those regions that are unstructured/exposed
Freeze-drying demonstrated as a viable and potentially preferable alternative to solution and frozen state storage of LNP mRNA vaccine formulations
Generated (and continuing to generate) quantitative stability data in terms of the active half-lives of LNP mRNAs in solution, frozen and freeze-dried forms over the course of up to threemonths for storage at -20°C (standard freezer), 4°C (standard refrigerator), room temperature, and 37°C (elevated/body temperature) for mRNAs of three different lengths
Przybycien, T., Presenter, ARP-10 LNP-mRNA Vaccine Stability: Reference Materials, Rapid Stability Assessment and Lyophilized Formulation Development, NIIMBL National Meeting, Washington, D.C., July 28, 2022.
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Rensselaer Polytechnic Institute
Carnegie Mellon University
