Different oligoarginine modifications alter endocytic pathways and subcellular trafficking of polymeric nanoparticles

Oligoarginine is a class of cell-penetrating peptides known for their ability to enhance cellular uptake of different cargoes. Here, we aim to understand how differences in oligoarginine modifications affect the cellular internalization and subcellular trafficking of polymeric nanoparticles. We found that the length of oligoarginine not only influenced the rate of cellular uptake, but also directed the mechanism of endocytosis, endosomal escape and subcellular destinations. Confocal microscopy and flow cytometry analysis were conducted using submicron particles of poly(ethylene glycol)-b-poly(ε-caprolactone) (PEG-b-PCL) with the surface modified by 1-, 4- and 8-residue-long oligoarginines, designated as R1PECL, R4PECL and R8PECL respectively. R8PECL and R4PECL effectively increased cellular uptake by 12-fold and 5-fold, respectively, while the effect of R1 modification on the cellular uptake rate was negligible. Nanoparticles without oligoarginine and R1PECL particles entered cells via clathrin-mediated endocytosis and were both trapped in lysosomes. R4PECL particles were internalized via lipid-raft dependent endocytosis, but failed to escape from endosomes. R8PECL particles were taken up by both lipid-raft dependent endocytosis and macropinocytosis, and successfully escaped from endosomes to enter cytosol, ER and mitochondria. On the other hand, decreasing the degree of modification on the particle surface while keeping the length of oligoarginine only lowered the amount of uptake and endosomal escape, but did not alter the endocytic pathways and intracellular trafficking. In short, this study illustrates the effect of different surface modifications on the subcellular fate of polymeric nanoparticles, providing useful insights into the design of nanocarriers for subcellular targeting.