Replacing Helper Lipids With Cationic Lipids Enhances mRNA Lipid Nanoparticles Stability in Solution

Abstract Messenger RNA lipid nanoparticles (mRNA‐LNPs) have achieved remarkable success in clinical vaccination efforts to curb the COVID‐19 pandemic, and have attracted tremendous interest from both industry and academia to broaden their biomedical applications. However, their storage and transportation rely heavily on the cold chain to prevent rapid degradation in non‐frozen solutions, posing significant challenges for logistics and accessibility. Therefore, enhancing the in‐solution stability of mRNA LNPs is crucial. Herein, the role of helper phospholipids is investigated in storage stability of mRNA LNPs. By ratiometrically replacing the helper phospholipids (DSPC) with other alternative lipids (DOPS, DOTAP, and DOPE) in commercially available mRNA LNP formulation, enhanced in‐solution stability was observed with cationic lipid (DOTAP) substitution at 42 °C, room temperature (22 °C), and 4 °C. Further correlation of stability with microstructure analysis using cryogenic electron microscopy revealed that partial replacement with DOTAP enhances structural stability by promoting the formation of intact LNPs and reducing the occurrence of “bleb”‐like structures. Overall, the study provides insights into the structure‐stability relationship in mRNA LNPs, and offers new strategies to address the storage limitations of mRNA‐LNP products in the future.