Ligand Density and Linker Length are Critical Factors for Multivalent Nanoparticle–Receptor Interactions

Although there are a large number of studies available for the evaluation of the therapeutic efficacy of targeted polymeric nanoparticles, little is known about the critical attributes that can further influence their uptake into target cells. In this study, varying cRGD ligand densities (0–100% surface functionalization) were combined with different poly­(ethylene glycol) (PEG) spacer lengths (2/3.5/5 kDa), and the specific receptor binding of targeted core–shell structured poly­(lactic-co-glycolic acid)/poly­(lactic acid)–PEG nanoparticles was evaluated using αvβ3 integrin-overexpressing U87MG glioblastoma cells. Nanoparticles with 100% surface functionalization and short PEG2k linkers displayed a high propensity to form colloidal clusters, allowing for the cooperative binding to integrin receptors on the cellular membrane. In contrast, the high flexibility of longer PEG chains enhanced the chance of ligand entanglement and shrouding, decreasing the number of ligand–receptor binding events. As a result, the combination of short PEG2k linkers and a high cRGD surface modification synergistically increased the uptake of nanoparticles into target cells. Even though to date, the nanoparticle size and its degree of functionalization are considered to be the major determinants for controlling the uptake efficiency of targeted colloids, these results strongly suggest that the role of the linker length should be carefully taken into consideration for the design of targeted drug delivery formulations to maximize the therapeutic efficacy and minimize adverse side effects.