Investigations Into the Influence of Ionizable Lipids on the Stability of Lipid Nanoparticle

Messenger RNA-loaded lipid nanoparticles (mRNA@LNPs) have achieved remarkable success in vaccine development, but their long-term preservation imposes stringent requirements for transportation and storage. To broaden application of this technology, it is essential to develop LNP formulations with enhanced stability. However, the fundamental rules behind LNP stability remains poorly understood. As a key lipid, the impact of the thermostability of ionizable lipids on LNP formulation's stability remains unexplored. In this study, we investigated the thermostability of two ionizable lipids—an in-house-developed lipid (A1-D1-5) and SM-102, the latter used in FDA-approved mRNA therapeutics—and assessed the stability of LNPs composed of these lipids under various storage conditions. Notably, we found that the size and polydispersity index (PDI) measured by dynamic light scattering (DLS) did not accurately reflect the stability of LNPs. While these indicators showed little change after 44 days of storage at 4°C, the mRNA activity sharply declined within just 14 days of preparation. Additionally, A1-D1-5 demonstrated greater thermostability compared to SM-102, leading to a slower decrease in mRNA activity. Importantly, our findings suggest that replacing ester bonds with amide bonds can significantly improve the thermostability of ionizable lipids. Overall, these results provide valuable insights into optimizing and evaluating the stability of mRNA@LNP formulations.