Inhibition of Octreotide Acylation Inside PLGA Microspheres by Derivatization of the Amines of the Peptide with a Self-Immolative Protecting Group

Acylation of biopharmaceuticals such as peptides has been identified as a major obstacle for the successful development of PLGA controlled release formulations. The purpose of this study was to develop a method to inhibit peptide acylation in poly(d,l-lactide-co-glycolide) (PLGA) formulations by reversibly and temporarily blocking the amine groups of a model peptide (octreotide) with a self-immolative protecting group (SIP), O-4-nitrophenyl-O'-4-acetoxybenzyl carbonate. The octreotide with two self-immolative protecting groups (OctdiSIP) on the N-terminus and lysine side chain was synthesized by reaction of the peptide with O-4-nitrophenyl-O'-4-acetoxybenzyl carbonate, purified by preparative RP-HPLC and characterized by mass spectrometry. Degradation studies of OctdiSIP in aqueous solutions of different pH values showed that protected octreotide was stable at low pH (pH 5) whereas the protecting group was eliminated at physiological pH, especially in the presence of an esterase, to generate native octreotide. OctdiSIP encapsulated in PLGA microspheres, prepared using a double emulsion solvent evaporation method, showed substantial inhibition of acylation as compared to the unprotected octreotide: 52.5% of unprotected octreotide was acylated after 50 days incubation of microspheres in PBS pH 7.4 at 37 °C, whereas OctdiSIP showed only 5.0% acylation in the same time frame. In conclusion, the incorporation of self-immolative protection groups provides a viable approach for inhibition of acylation of peptides in PLGA delivery systems.