Rationally designed negatively charged lipid nanoparticles reduce systemic reactogenicity of mRNA vaccines while...

Messenger RNA (mRNA) vaccines have emerged as versatile platforms for the prevention of infectious diseases and therapeutic cancer vaccination. However, strong reactogenicity remains a key challenge, with severe adverse events such as myocarditis. These concerns may limit the applications of mRNA vaccines. As lipid nanoparticles (LNP) are the primary driver of inflammation, optimizing LNP composition is a promising approach to reduce mRNA vaccine reactogenicity. However, the effects of tuning the physicochemical properties of LNP, particularly their surface charge, remain insufficiently characterized. In this study, we aimed to engineer negatively charged LNPs by incorporating anionic phospholipids and evaluate their biodistribution, immunogenicity, and adverse reactions in mice. Negatively charged LNPs significantly reduced systemic inflammatory responses and fever compared to a conventional LNP. Moreover, cardiac troponin I-a marker of myocarditis-was reduced in negatively charged LNPs. Despite reduced systemic reactogenicity, negatively charged LNPs maintained antigen-specific immunogenicity comparable to a conventional LNP. The negatively charged LNPs did not attenuate local inflammation at the injection site. This local inflammation depended on the fatty acyl chain of phospholipids in the LNP and was independent of the head group structure or negative charge. Surface charge modulation also reshaped biodistribution to distal organs, such as the spleen and liver. Correlation analysis linked reduced reactogenicity to decreased hepatic delivery of LNPs, and prior datasets supported this relationship. Taken together, our results establish phospholipid-driven surface-charge control as a promising approach to reduce systemic reactogenicity while maintaining the immunogenicity of mRNA vaccines.