Influence of salt solution on the physicochemical properties and in vitro/ in vivo expression of mRNA/LNP

Lipid nanoparticles (LNPs) have revolutionized nucleic acid delivery, enabling significant advances in mRNA-based therapeutics. While extensive research has focused on lipid composition, the impact of preparation solutions on LNP performance remains underexplored. This study systematically investigated the effects of pH, salt type, and concentration across key preparation solutions—mRNA aqueous, dilution, exchange, and storage solutions—on the physicochemical properties, stability, and expression efficiency of SM102-based mRNA/LNPs. Findings revealed that the pH of the mRNA aqueous solution was critical, with a pH of 4 optimizing encapsulation efficiency (EE) and cellular expression. The exchange solution's pH significantly influenced biodistribution, particularly liver-specific expression following intravenous and intramuscular administration. Sucrose was identified as essential for freeze-thaw stability, with a 300 mM concentration minimizing aggregation and mRNA leakage. Furthermore, preparation solutions were shown to influence the structural integrity of LNPs, impacting their in vivo and in vitro performance. These insights highlight the importance of preparation conditions in optimizing LNP formulations for clinical applications, offering a foundation for enhanced therapeutic design and delivery.