Integration of Charged Peptide Amphiphiles within Lipid Nanoparticles (LNPs) for Enhanced Targeted mRNA Delivery

Abstract Lipid nanoparticles (LNPs) play a pivotal role in mRNA-based therapy; however, achieving targeted delivery remains challenging. We developed an LNP platform integrating peptide amphiphiles (PAs) to exploit their dual bioactivity and electrostatic tunability. Two PAs, cationic C16TAT (derived from a cell-penetrating peptide) and anionic C16HA2 (based on an endosomolytic peptide), were co-assembled into LNPs. In vitro, C16TAT/LNPs significantly enhanced mRNA delivery across multiple cell lines via improved cellular uptake. Due to the potent charge-modifying capacity of C16HA2, C16HA2/LNPs exhibit spleen-selective tropism in vivo, achieving >100-fold higher mRNA expression in the spleen than in the liver. While the commonly used surface-conjugated TAT-PEG-LNPs enhanced transfection in vitro, their in vivo performance was inferior relative to the C16TAT/LNP system due to the lack of stability, highlighting the advantage of PA integration. Critically, effective tissue targeting required substantial LNP charge alteration (pKa and zeta potential), which was achievable with C16HA2 but not C16TAT, likely due to the former’s higher aggregation tendency. Dual PA-PEG modifications may improve spleen targeting but required formulation optimization. This work establishes a simple PA/lipid co-assembly approach as a robust method for modulating LNP tropism, advancing the rational design of targeted mRNA delivery systems.