Comprehensive Analytical Characterization of mRNA Influenza Vaccine Under Stress Conditions: Implications for Stability and Quality Control.

PURPOSE: This study aimed to characterize the physicochemical stability of mRNA-lipid nanoparticle (LNP) influenza vaccine candidates under various environmental stressors using forced degradation studies (FDSs) and to correlate these changes with in vitro protein expression. METHODS: FDSs were conducted to evaluate the effects of temperature, oxidation, hydrolysis, and agitation on mRNA-LNP formulations. Comprehensive analytical techniques were employed to monitor the purity and integrity of mRNA, mRNA encapsulation efficiency, in vitro expression, LNP components, and particle characteristics. RESULTS: The study revealed hydrolysis-induced degradation of distearoylphosphatidylcholine (DSPC), a key LNP component, and rapid oxidation-induced mRNA degradation within the LNPs. Agitation was found to increase LNP size, decrease mRNA encapsulation efficiency, and subsequently reduce in vitro protein expression. CONCLUSIONS: We established robust analytical methods to monitor the quality and stability of mRNA-LNP products. These findings provide a critical foundation for integrated analytical strategies to enhance the stability of mRNA-LNP therapeutics and vaccines during storage, transport, and clinical application.