Selective mRNA Delivery to Activated Macrophages via Hyaluronic Acid-Functionalized Lipid Nanoparticles with Optimized PEGylation

Activated proinflammatory macrophages are associated with various inflammatory diseases, and due to their overexpression of the CD44 receptor, they may be targeted for therapy by hyaluronic acid (HA), its natural ligand. This study aimed to develop lipid nanoparticles (LNPs) functionalized with HA and stabilized with an optimized amount of poly(ethylene glycol) (PEG) for targeted mRNA delivery to activated macrophages. Using microfluidic mixing, LNPs were produced with either 1.5% PEG (LNP_1.5%PEG) or 0.5% PEG (LNP_0.5%PEG). HA-coated LNPs (HA-LNPs) were prepared by postinsertion of an HA-DPPE conjugate, and changes in size and zeta potential demonstrated a successful and efficient HA coating, which was quantified by spectrofluorimetry and nanoscale flow cytometry. In vitro studies showed that HA-LNP_0.5%PEG exhibited better uptake in activated macrophages while maintaining mRNA transfection efficiency, whereas HA-LNP_1.5%PEG did not improve its uptake, suggesting that excessive PEG can hinder targeting. Overall, HA-LNP_0.5%PEG effectively delivered mRNA to activated macrophages with enhanced selectivity.