Identification of chemically modified peptide from poly(D,L-lactide-co-glycolide) microspheres under in vitro release conditions

Abstract

ABSTRACTmers within polymeric microspheres. The data suggest that due to steric hindrance factors, polymers with greater lactide content were less amenable to the formation of adduct impurities compared with PLGA 50:50 copolymers.The purpose of this research was to study the chemical reactivity of a somatostatin analogue, octreotide acetate, formulated in microspheres with polymers of varying molecular weight and co-monomer ratio under in vitro testing conditions. Poly(D,L-lactide-coglycolide) (PLGA) and poly(D,L-lactide) (PLA) microspheres were prepared by a solvent extraction/evaporation method. The microspheres were characterized for drug load, impurity content, and particle size. Further, the microspheres were subjected to in vitro release testing in acetate buffer (pH 4.0) and phosphate buffered saline (PBS) (pH 7.2). In acetate buffer, 3 microsphere batches composed of low molecular weight PLGA 50:50, PLGA 85:15, and PLA polymers (≤10 kDa) showed 100% release with minimal impurity formation (<10%). The high molecular weight PLGA 50:50 microspheres (28 kDa) displayed only 70% cumulative release in acetate buffer with significant impurity formation (~24%). In PBS (pH 7.4), on the other hand, only 50% release was observed with the same low molecular weight batches (PLGA 50:50, PLGA 85:15, and PLA) with higher percentages of hydrophobic impurity formation (ie, 40%, 26%, and 10%, respectively). In addition, in PBS, the high molecular weight PLGA 50:50 microspheres showed only 20% drug release with ~66% mean impurity content. The chemically modified peptide impurities inside microspheres were structurally confirmed through Fourier transform-mass spectrometry (FT-MS) and liquid chromatography/mass spectrometry (LC-MS/MS) analyses after extraction procedures. The adduct compounds were identified as covalently modified conjugates of octreotide with lactic and glycolic acid mono-

Synthesis and evaluation of biodegradable PCL/PEG nanoparticles for neuroendocrine tumor targeted delivery of somatostatin analog

“…Likewise, when Octreotide microspheres were subjected to in vitro release in acetate buffers, '100% drug release with insignificant degradation products' while testing were observed. On the contrary, another research showed that at physiological pH (pH 7.4), no instability was shown by Octreotide (Murty et al, 2003). Therefore in this study two buffers were used to study drug release.…”

Effect of excipients on PLGA film degradation and the stability of an incorporated peptide

“…4e and Table 1). Preferential protonation of the proton sponge (pKa = 12.34), rather than protonation of the peptide primary amine (pKa = 9.72 [27]), is a likely cause, since the peptide N-terminus is expected to be a better nucleophile in the unprotonated form [16,28,29]. The reduction in deamidation and cleavage products (~ 7% each) in comparison to the unbuffered control can also be attributed to preferential protonation of the proton sponge rather than the Asn side chain or backbone amide nitrogen, respectively, as these are acid catalyzed reactions [16].…”

Reversible Blocking of Amino Groups of Octreotide for the Inhibition of Formation of Acylated Peptide Impurities in Poly(Lactide-co-Glycolide) Delivery Systems

“…3a, various acylated octreotides were found during incubation with PLGA in 0.1 M phosphate buffer (pH 7.4) at 37°C. Among the acylated octreotides, three distinct peaks (peaks 2, 3, and 4) were observed and they were identified based on the previous study (9,15,25) as follows: peak 2, mono-glycoyl-conjugated octreotide at Lys residue; peak 3, mono-glycoyl-conjugated octreotide at N terminus; peak 4, di-glycoyl-conjugated octreotide at the N terminus and Lys residue. The most prevalent acylation product was the N-terminally glycoylconjugated octreotide (peak 3).…”

“…Currently, octreotide has been commercially formulated in PLGA microspheres (Sandostatin LAR depot, Novartis Pharma, Basel, Switzerland) as a monthly dosage form for the treatment of acromegaly (14). In a previous study, the octreotide in Sandostatin LAR has been also shown to form acylated peptide impurities after in vitro incubation in phosphate-buffered saline (15).…”