mRNA-lipid nanoparticle surfaces, endocytosis, and endosomal escape: a structure-activity relationship

mRNA vaccines against infectious disease and cancer require encapsulation in lipid nanoparticles (LNPs) to improve stability and cell entry. Polyethylene glycol (PEG) lipids are used to stabilise the LNP surface. However PEG impairs endocytic cellular uptake and escape from endosomes, likely by reducing bilayer interactions. These are 2 key cellular barriers to mRNA-LNP efficacy. Moreover, increasing allergic response to PEG in patients who receive mRNA-LNP vaccines limits the safety and efficacy of mRNA-LNPs. I have synthesised alternative polymer-lipids that improve mRNA-LNP efficacy by 100-fold in vitro and 3-fold in vivo compared to formulations containing PEG. However, the mechanism behind this improved efficacy is unclear.
This project aims to uncover the mechanism by which LNP surface chemistry impacts cellular endocytosis and endosomal escape by using neutron reflectometry (NR) to resolve molecular interactions between model lipid bilayers and 4 LNPs with unique surfaces (no polymer, PEG, a commercial PEG alternative, and my leading novel PEG alternative) at physiological and endosomal pH (7.4, 5.5 respectively). NR will provide unreported biophysical insights unachievable via other techniques. These data will be complemented by in vitro microscopy and in silico molecular dynamic simulations to build a structure-activity relationship between LNP surface chemistry, endocytosis, and endosomal escape that will inform the design of future mRNA-LNPs with enhanced potency.