Pegylation modulates ApoE association and hepatic accumulation of DC-Chol-based nanoparticles through steric exclusion.

Due in part to adsorption of apolipoprotein E (ApoE) and subsequent receptor-mediated uptake by hepatocytes, lipid nanoparticles (LNPs) have a propensity to accumulate within the liver. The mechanistic contribution of PEGylation to this phenomenon has yet to be fully elucidated. Using fluorescence quenching, circular dichroism spectroscopy, molecular dynamics simulations, and in vivo bioluminescence imaging, we investigated how PEGylation influences ApoE interactions with DC-Chol-based nanoparticles. Association studies revealed that non-PEGylated formulations bound ApoE ∼ 1.7-fold more strongly than particles with surface PEG moieties, quantified by Stern-Volmer constants of 0.083 vs. 0.048 μM-1, respectively. Circular dichroism measurements demonstrated a greater structural perturbation to ApoE upon binding to non-PEGylated particles (80% helical loss) versus PEGylated LNPs (24-48% helical loss). Molecular dynamics simulations showed that PEG-2000 creates a dynamic steric barrier that reduces direct protein-lipid contact formation by 2.9-fold and decreases the tendency of ApoE to remain closely associated with the nanoparticle surface. Correspondingly, biodistribution studies in mice showed rapid hepatic accumulation of non-PEGylated nanoparticles within 24 h, whereas PEGylated formulations exhibited delayed liver accumulation that became prominent at 48 h. Together, these findings indicate that PEGylation reduces ApoE association primarily through steric exclusion and highlight PEGylation as a useful strategy for modulating nanoparticle biodistribution.