RGD-derivatized PEI-PEG copolymers: Influence of the degree of substitution on the targeting behavior

PEGylation is widely used to improve the stability of gene delivery vectors through the decrease of non-specific binding to serum proteins. In order to provide “stealth” vectors with targeting properties, the grafting of specific ligands is mandatory. For instance, the tripeptide arginine-glycine-aspartic acid (RGD) has been shown to confer selectivity towards some integrin-overexpressing tumor cells. Unfortunately, owing to the different RGD grafting degrees, some difference in starting materials and experimental conditions, the literature displays contradictory findings about the efficacy of this strategy. Starting from branched poly(ethylene imine) (bPEI) transfectant and the heterobifunctional linker N-hydroxy-succinimide-poly(ethylene glycol)-maleimide, a series of variably substituted PEI-PEGs were synthesized and functionalized with linear and cyclic RGDs. bPEI-PEG-RGDs were as effective as bPEI in complexing DNA while derivatization did affect the physicochemical properties of polyplexes. A degree of substitution of 1.31% led to enhanced targeting of cognate receptor-expressing cells without impairing the transfection efficiency of bPEI, even in the presence of serum. Of note, both RGDs were equally effective to confer selectivity to polyplexes. These results highlight the key role of the degree of substitution on the effectiveness and selectivity of bPEI-PEG-RGDs, suggesting that a systematic approach is needed for the development of more effective targeted transfectants.