Ionic Coating of siRNA Polyplexes with cRGD–PEG–Hyaluronic Acid To Modulate Serum Stability and In Vivo Performance

Efficient delivery of siRNA-based polyplexes to tumors remains a major challenge. Nonspecific interactions in the bloodstream, limited circulation time, and nontargeted biodistribution hamper sufficient tumor accumulation. To address these challenges, we developed an ionic hyaluronic acid (HA) coating to shield sequence-defined oligoaminoamide-based polyplexes. This coating should shield the positive polyplex surface charge, thus reducing nonspecific interactions and enhancing serum stability. Additionally, we modified the HA coating with the cyclic RGDfK (cRGD) peptide to specifically target tumor endothelial cells (TECs). Optionally, a polyethylene glycol (PEG) spacer was also introduced to improve ligand presentation on the polyplex surface. The HA-coated polyplexes exhibited favorable physicochemical properties, including a negative zeta potential and effective siRNA retention within the polyplex, which was not adversely affected by PEG or cRGD modification. In vitro analyses revealed that these polyplexes not only enhanced tumor cell association and preserved the high transfection efficiency of plain cationic polyplexes but also exhibited coating-dependent cellular internalization, as evidenced by a competitive inhibition experiment. Even in the presence of serum, the HA-coated polyplexes encapsulated siRNA effectively, exhibited suitable particle sizes, and maintained a high gene silencing efficiency. In vivo studies involving intravenous administration into Neuro2a tumor-bearing mice showed that the HA coating, particularly when modified with PEG and cRGD, significantly increased the tumor accumulation of polyplexes. HA–PEG–cRGD-shielded polyplexes exhibited significantly enhanced in vivo gene silencing in tumors compared with plain polyplexes. Collectively, our results indicate a superior performance of HA-coated polyplexes in terms of stability and cellular uptake, both in vitro and in vivo.