Comparison of the rates of deamidation, diketopiperazine formation, and oxidation in recombinant human vascular endothelial growth factor and model peptides
Journal Title: The AAPS Journal - Year 2000, Vol 2, Issue 1
Abstract
In this work, we examine the way in which stability information obtained from studies on small model peptides correlates with similar information acquired from a protein. The rates of deamidation, oxidation, and diketopiperazine reactions in model peptide systems were compared to those of recombinant human vascular endothelial growth factor (rhVEGF). The N-terminal residues of rhVEGF, a potent mitogen in angiogenesis, are susceptible to the aforementioned reactions. The degradation of the peptides L-Ala-L-Pro-L-Met (APM) and Gly-L-Gsn-L-His-L-His (GQNHH), residues 1–3 and 8–12 of rh VEGF, respectively, and rhVEGF were examined at pH 5 and 8 at 37°C. Capillary electrophoresis and high-performance liquid chromatography (HPLC) stability-indicating assays were developed to monitor the degradation of the penta- and tripeptides, respectively. The degradation of rhVEGF was determined by tryptic mapping and quantified by RP-HPLC. The rates of degradation of both peptides and the protein followed apparent first-order kinetics and increased with increasing pH. The tripeptide APM underwent diketopiperazine formation (Ala-Prodiketopiperazine) and oxidation of the Met residue, whereas the pentapeptide GQNHH degraded via the deamidation pathway. The results indicate that the rates of deamidation and oxidation of the protein are comparable to those observed in the model peptides at both pH values. However, the rate of the diketo-piperazine reaction was slower in the protein than in the model peptide, which may be the result of differences in the cis-trans equilibrium of the X-Pro peptide bonds in the 2 molecules.
Authors and Affiliations
Chimanlall Goolcharran, Andrew J. S. Jones, Ronald T. Borchardt
Keywords
Related Articles
- EP ID EP682080
- DOI 10.1208/ps020105
- Views 85
- Downloads 0
Multidimensional Atomic Force Microscopy: A Versatile Novel Technology for Nanopharmacology Research
The online version of this article (doi:10.1208/s12248-010-9232-y) contains supplementary material, which is available to authorized users.
Structure Activity Relationships in Alkylammonium C12-Gemini Surfactants Used as Dermal Permeation Enhancers
The purpose of this study was to determine the ability and the safety of a series of alkylammonium C12-gemini surfactants to act as permeation enhancers for three model drugs, namely lidocaine HCl, caffeine, and ketoprof...
Detergent-resistant membrane microdomains in the disposition of the lipid signaling molecule anandamide
The endogenous cannabinoid anandamide is an agonist of the cannabinoid receptors CB1 and CB2, as well as transient receptor potential vanilloid type 1 and type 4 ion channels. In recent years, there has been a great deal...
Ligand Binding Assays in the 21st Century Laboratory: Recommendations for Characterization and Supply of Critical Reagents
The online version of this article (doi:10.1208/s12248-012-9334-9) contains supplementary material, which is available to authorized users.
In Silico Evaluation of the Potential Impact of Bioanalytical Bias Difference between Two Therapeutic Protein Formulations for Pharmacokinetic Assessment in a Biocomparability Study
Formulation changes at later stages of biotherapeutics development require biocomparability (BC) assessment. Using simulation, this study aims to determine the potential effect of bias difference observed between the two...