PCL-PEG-based nanoparticles grafted with GRGDS peptide: preparation and surface analysis by XPS.

Nanoscopic particles self-assembled from amphiphilic block copolymers have been developed over the past years for the preparation of various drug delivery systems. The biodegradable carriers made of polyethylene glycol (PEG)-polyester diblock copolymers emerged as the most adequate devices due to their prolonged circulation half-life in blood, their ability to bypass natural barriers, their reduced rate of uptake by liver, and their controllable delivery potientiality. However, the coupling of ligands to achieve targeting toward particular cell types remains a challenge. Few methods of functionalization of polymeric micelles have been reported: they are based on the synthesis of maleimide, acetal, or biotin-terminated PEG blocks before block copolymerization and self-assembling. The first material is thiol-reactive and can be used for the direct covalent coupling of ligands such as Arg-Gly-Asp (RGD) containing molecules. The second one is used, after deprotection of the acetal, for the coupling of amine-terminated GRGDS (Gly-Arg-Gly-Asp-Ser) pentapeptide via a reductive amination standard protocol. The third one is able to fix avidin in a noncovalent, but highly strong, interaction; this enables the subsequent fixation of biotinylated ligands because several biotin binding sites remain available on the nanoparticles. In this note, we propose an alternative strategy inspired from the photoaffinity labeling, a technique well-known in biochemistry. Our method relies upon the implantation of activated ester functions in the copolymer matrix by UV irradiation of O-succinimidyl 4-(p-azido-phenyl)butanoate, intimately mixed with the material. Further reaction with NH2-terminated molecules allows the covalent coupling of ligands or molecular probes via an amide linkage resulting from NHS (N-hydroxysuccinimide) displacement. This has been illustrated by the functionalization of polycaprolactone(PCL)-polyethylene glycol(PEG) diblock copolymers with 3,5-bis-trifluoromethyl benzyl amine (Tag F6) and with Gly-Arg-Gly-Asp-Ser (GRGDS) pentapeptide. The Tag F6 is a fluorinated probe easily detectable and quantifiable by X-ray photoelectron spectroscopy (XPS) and GRGDS is a biologically active ligand aiming at the binding of the cell receptors called integrins. Those grafted PCL-PEGs were added in the formulation of PEGylated PLGA-based nanoparticles prepared by a double emulsion technique and designed for oral vaccination. Our aim was to develop a convenient method to introduce any desired compounds (ligands or molecular probes) in selfassembled, tailor-made nanoparticulate systems, constituted of an hydrophobic core of associated PLGA and PCL blocks surrounded by an hydrophilic shell of PEG chains exposing the targeting signals. By modifying the quantities and lengths of the different polymers (PLGA, PLGA-PEG, and PCL-PEG), the characteristics of the nanoparticles could be adjusted in terms of surface properties, drug encapsulation efficiency, and rate of degradation.