Tailored Monoacyl Poly(2-oxazoline)- and Poly(2-oxazine)-Lipids as PEG-Lipid Alternatives for Stabilization and Delivery of mRNA-Lipid Nanoparticles.

The successful use of lipid nanoparticles (LNPs) for clinical development of the COVID-19 mRNA vaccines marked a breakthrough in mRNA-LNP therapeutics. As one of the vital components of LNPs, poly(ethylene glycol)-lipid conjugates (PEG-lipids) influence particle biophysical properties and stability, as well as interactions within biological environments. Reports suggesting that anti-PEG antibodies can be detected quite commonly within the human population raise concerns that PEG content in commercial LNP products could further stimulate immune responses to PEG. The presence of anti-PEG antibodies has been linked to accelerated clearance of LNPs, potentially a source of variability in the biological response to mRNA-LNP products. This motivated us to explore potential PEG alternatives. Herein, we report physicochemical and biological properties of mRNA-LNPs assembled using poly(2-oxazoline) (POx)- and poly(2-oxazine) (POz)-based polymer-lipid conjugates. Notably, we investigated monoacyl lipids as alternatives to diacyl lipids. mRNA-LNPs produced using monoacyl POx/POz-lipids displayed comparable biophysical characteristics and cytocompatibility. Delivery of reporter mRNA resulted in similar transfection efficiencies, in both adherent and suspension cells, and in mice, compared to PEG-lipid equivalents. Our results suggest that monoacyl POx/POz-lipid-containing LNPs are promising candidates for the development of PEG-free LNP-based therapeutic products.