Fig. 1: Morphological and in vitro transfection properties of LNP mRNA systems formulated using various bilayer lipid to ionizable lipid molar ratios (RB/I).
From: Liposomal lipid nanoparticles for extrahepatic delivery of mRNA
LNP mRNA systems were formulated as described in Methods containing NanoLuc mRNA (808 nt; N/P = 6) with lipid compositions nor-MC3/ ESM: cholesterol/ PEG-DMG at varying ratios of bilayer lipid (ESM and equimolar cholesterol) to ionizable lipid (RB/I). These formulations correspond to RB/I of 9, 4, 2.3, 1.5, 1, 0.67 and 0.43. The Onpattro-like formulation consists of MC3/DSPC/cholesterol/PEG-DMG (50/10/38.5/1.5 mol/mol). If it is assumed that the cholesterol and DSPC reside exclusively in the monolayer or bilayer environments, the RB/I for this formulation is 1.03. A Lipids in ethanol are mixed with mRNA in 25 mM NaOAc (pH 4) to give rise to LNP mRNA systems that are either liposomal LNP systems or solid core systems where at least part of the exterior surface consists of a lipid monolayer. B Encapsulation efficiencies of LNP NanoLuc mRNA systems with varying RB/I values (mean ± S.D., n = 3). C Hydrodynamic diameter (DLS measurement, number mean) for various RB/I ratios (mean ± S.D., n = 3). D The classification of LNP morphologies within the formulated population. LNP morphology was defined by its oil core or lamellar/multilamellar liposomal features as determined from cryo-TEM micrographs. E Cryo-TEM micrographs of LNP mRNA systems prepared using various RB/I values, micrograph has been reproduced twice. F Luminescence of Huh7 cells incubated for 24 h with LNP NanoLuc mRNA systems (0.1–1 μg mRNA/mL), decrease in RB/I indicated from beige to dark red, with Onpattro-like formulation (indicated as gray) over a range of RB/I values (mean ± S.E.M., n = 6). Source data are provided as a Source Data file. Figure 1A was partially created in BioRender. Cheng, M. (2025) https://BioRender.com/o07c413. All LNP illustrations are original and were created using Inkscape.
