Targeting ligand-functionalized and redox-sensitive heparin-Pluronic nanogels for intracellular protein delivery.

The heparin-Pluronic (HP) conjugate was coupled via redox-sensitive disulfide bond and contains a vinyl sulfone (VS) group with high reactivity to some functional groups such as thiol group. Heparin was conjugated with cystamine and the terminal hydroxyl groups of Pluronic were activated with the VS group, followed by coupling of VS groups of Pluronic with cystamine of heparin. The chemical structure, heparin content and VS group content of the resulting product were determined by (1)H NMR, FT-IR, toluidine blue assay and Ellman's method. The HP conjugate formed a type of nanogel in an aqueous medium, showing a critical micelle concentration of approximately 129.35 mg L(-1), a spherical shape and the mean diameter of 115.7 nm, which were measured by AFM and DLS. The release test demonstrated that HP nanogel was rapidly degraded when treated with glutathione. Cytotoxicity results showed a higher viability of drug-free HP nanogel than that of drug-loaded one. Cyclo(Arg-Gly-Asp-D-Phe-Cys) (cRGDfC) peptide was efficiently conjugated to VS groups of HP nanogel and exhibited higher cellular uptake than unmodified nanogels. All results suggest a novel multi-functional nanocarrier delivery and effective release of proteins to the intracellular region in a redox-sensitive manner.