Mathematical Modeling, Numerical Simulation, and Optimization of Targeted Drug Delivery Systems Using Liposomal Surface Remodeling

Abstract

We present a reformulated liposomal drug delivery system in which conventional targeting ligands are replaced by a computationally engineered O-glycosyltransferase variant that site-specifically remodels threonine-rich peptide ligands on the liposome surface. This approach substitutes traditional stochastic conjugation with a precisely controlled enzymatic glycosylation step, thereby transforming the surface-conjugated moiety into a glycoengineered peptide–carbohydrate hybrid. The system is built upon a PEGylated liposomal carrier, into which threonine-rich peptide targeting ligands are pre-conjugated via thiol-maleimide chemistry. The key innovation is a variant of the O-glycosyltransferase from Pasteurella multocida, designated PmOGT-GalNAz, which was redesigned through a computational pipeline combining RosettaDesign and molecular dynamics simulations. This variant harbors three mutations that widen the nucleotide-sugar binding pocket and improve catalytic efficiency for the unnatural donor substrate UDP-GalNAz, achieving a catalytic rate constant of  and a Michaelis constant of . The enzyme catalyzes the site-specific addition of GalNAz to threonine residues on the peptide ligands, forming a stable alpha-O-glycosidic linkage. The resulting azido-functionalized sugar corona reduces serum protein adsorption from 45% to 12% and provides a bioorthogonal handle for subsequent strain-promoted azide-alkyne cycloaddition with dibenzocyclooctyne-functionalized prodrug esters. This reaction yields a stable triazole-linked prodrug–ligand conjugate without disturbing the liposomal bilayer. The degree of glycosylation is tunable by adjusting enzyme concentration, reaction time, and donor substrate concentration, enabling precise control over ligand display density and payload loading. Furthermore, the prodrug is designed to undergo intracellular esterase-mediated cleavage, releasing active doxorubicin specifically in acidic tumor microenvironments.