Enhanced mRNA Delivery in ARPE-19 Cells by RVG29-Modified Ready-to-Use LNP

The efficient and safe delivery of nucleic acids to retinal pigment epithelial (RPE) cells remains a significant challenge in ocular therapeutics. Lipid nanoparticles (LNPs) represent a promising platform for nucleic acid delivery; however, conventional LNPs often exhibit limited transfection efficiency in RPE cells. Here, we report RVG-modified, ready-to-use (RTU) LNPs for enhanced mRNA delivery in ARPE-19 cells. RVG29, a 29-amino acid peptide derived from rabies virus glycoprotein (RVG), was conjugated to LNP to facilitate receptor-mediated uptake via nicotinic acetylcholine receptors (nAChRs). The RVG-LNPs were formulated using a nonconventional post-encapsulation method. The resulting RVG-RTU LNP demonstrated an eGFP mRNA encapsulation efficiency (EE) over 95% with a particle size of approximately 96 nm. Compared with unmodified LNP prepared by the conventional method, RTU processes alone increased eGFP expression by [Formula: see text]3.6-fold (RTU-LNP) while RVG29 modification alone (RVG-LNP) by [Formula: see text]7.4-fold. Combining both strategies resulted in a 12.8-fold increase (RVG-RTU-LNP). Furthermore, we also demonstrated that RVG-RTU-LNP can be produced via a high-throughput 96-well plate format, yielding LNPs with consistent size, polydispersity index (PDI) and EE. These results indicate that ligand-functionalized, RTU LNPs provide a practical and effective platform for mRNA delivery to RPE cells, offering potential for nonviral therapeutic interventions in retinal diseases.