Apolipoprotein E Binding Drives Structural and Compositional Rearrangement of mRNA-Containing Lipid Nanoparticles

Abstract

Emerging therapeutic treatments based on the production of proteins by delivering mRNA have become increasingly important in recent times. While lipid nanoparticles (LNPs) are approved vehicles for small interfering RNA delivery, there are still challenges to use this formulation for mRNA delivery. LNPs are typically a mixture of a cationic lipid, distearoylphosphatidylcholine (DSPC), cholesterol, and a PEG-lipid. The structural characterization of mRNA-containing LNPs (mRNA-LNPs) is crucial for a full understanding of the way in which they function, but this information alone is not enough to predict their fate upon entering the bloodstream. The biodistribution and cellular uptake of LNPs are affected by their surface composition as well as by the extracellular proteins present at the site of LNP administration, e.g., apolipoproteinE (ApoE). ApoE, being responsible for fat transport in the body, plays a key role in the LNP’s plasma circulation time. In this work, we use small-angle neutron scattering, together with selective lipid, cholesterol, and solvent deuteration, to elucidate the structure of the LNP and the distribution of the lipid components in the absence and the presence of ApoE. While DSPC and cholesterol are found to be enriched at the surface of the LNPs in buffer, binding of ApoE induces a redistribution of the lipids at the shell and the core, which also impacts the LNP internal structure, causing release of mRNA. The rearrangement of LNP components upon ApoE incubation is discussed in terms of potential relevance to LNP endosomal escape.

Intracellular Routing and Recognition of Lipid-Based mRNA Nanoparticles

“…In the blood circulation and among different functions, ApoE is involved in the transportation of lipids to the liver for recycling. Recently, Sebastiani et al [ 117 ] showed that the binding of ApoE induce lipid rearrangement of the shell and the core of LNPs, potentially leading to a different endosomal escape. To study this effect, they produced LNPs made of deuterated lipids coupled with a small-angle neutron scattering (SANS) analysis, to decipher the composition and the global structure of the LNPs, but also more precisely the distribution of the lipids around the particle.…”

Lipid Bilayer Degradation Induced by SARS-CoV-2 Spike Protein as Revealed by Neutron Reflectometry

“…The deuterated cholesterol (d-45 cholesterol, 85% deuteration) was provided by the National Deuteration Facility at ANSTO (Sydney, Australia). The details of the biosynthetic production of d-25 Cholesterol are reported elsewhere 34 . The 18A peptide (sequence KAFYDKVAEKLKEAF acetylated and amidated at the N-terminal and C-terminal, respectively, >95% purity) was obtained from GenScript.…”

Immunomodulatory Nanomedicine for the Treatment of Atherosclerosis

“…Additionally, the clinically approved LNPs that are most famous at the moment are based on lipid encapsulation, i.e., the COVID-19 vaccines from BioNTech/Pfizer and Moderna, are lipid nanoparticles [ 35 ]. LNPs enter the cells by binding to the LDL-receptor aided by apolipoprotein E (ApoE), which stimulates its uptake significantly [ 36 ]. As the LDL-receptor is not only expressed on hepatocytes, but also on other cells like antigen-presenting cells, LNPs can target a variety of cell-types to exert its effects and modulate lipid and/or inflammatory mechanisms ( Figure 1 ).…”