Impact of Structurally Active Lipids on Lipid Nanoparticle Delivery Processes

RNA therapeutics have the potential to treat many diseases that are not targetable by small molecules. One of the biggest challenges in RNA therapy is RNA delivery. Lipid nanoparticles(LNPs) is one of the most successful RNA delivery systems to date, however the delivery efficiency of LNP systems is still hindered by inefficient endosomal escape. How LNPs impact the endosomal escape process is still poorly understood. In this work, we explore how structurally active lipids that can form bicontinuous cubic and inverse hexagonal phases impact the properties of active model endosomal membranes, to better understand the change of endosomes when lipids from LNPs diffuse into the endosomal membrane. We formed giant unilamellar vesicles (GUVs) with different molar percentages of structurally active lipids, and with fluctuation analysis showed that the bending modulus of the membrane reduces with the increase in structurally active lipid content. The presence of membrane protein can greatly impact endosomal membrane properties, so GUVs with V-ATPase, a membrane protein that is essential to the endosomal escape process, were also formed to model active endosomal membranes.