Polysorbates 20 and 80 Degradation by Group XV Lysosomal Phospholipase A2 Isomer X1 in Monoclonal Antibody Formulations
Autor: | Stephanie L. Sandefur, Troii Hall, Christopher C. Frye, Lihua Huang, Tammy L. Tuley |
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Rok vydání: | 2015 |
Předmět: |
0301 basic medicine
medicine.drug_class Drug Compounding Pharmaceutical Science Polysorbates CHO Cells Monoclonal antibody 030226 pharmacology & pharmacy Chinese hamster law.invention 03 medical and health sciences chemistry.chemical_compound Hydrolysis 0302 clinical medicine Cricetulus law Tandem Mass Spectrometry Phosphatidylcholine Cricetinae medicine Animals Polysorbate Chromatography biology Chemistry Chinese hamster ovary cell Antibodies Monoclonal biology.organism_classification Phospholipases A2 030104 developmental biology Biochemistry Recombinant DNA Polysorbate 20 Lysosomes Acyltransferases |
Zdroj: | Journal of pharmaceutical sciences. 105(5) |
ISSN: | 1520-6017 |
Popis: | Decreases in polysorbate (PS80) content were observed while evaluating the long-term storage stability of Chinese hamster ovary-derived, purified monoclonal antibodies. It was determined that polysorbate had been enzymatically degraded; therefore, studies were performed to identify and characterize the protein(s) responsible. Polysorbate degrading activity was enriched from Chinese hamster ovary media leading to the identification of group XV lysosomal phospholipase A2 isomer X1 (LPLA2) by shotgun proteomics. Recombinant LPLA2 was over expressed, purified, and functional integrity confirmed against a diheptanoyl phosphatidylcholine substrate. Incubation of recombinantly produced LPLA2 with PS20 and PS80 resulted in hydrolysis of PS20 and PS80 monoester but a much slower rate was observed for higher-order PS80. Endogenous LPLA2 was detected and quantitated at less than 1 ppm in 3 formulated antibodies while LPLA2 was not detected (or less than 0.1 ppm) in a fourth formulated antibody. Furthermore, antibodies with detectable quantities of endogenous LPLA2 demonstrated polysorbate hydrolysis while in contrast the antibody without detectable LPLA2 did not show polysorbate hydrolysis. Comparison of polysorbate degradation products generated from the formulated antibody and samples of polysorbate incubated with recombinant LPLA2 resulted in similar elution profiles by liquid chromatography-mass spectrometry. These results suggest that LPLA2 may play a key role in polysorbate degradation in some antibody preparations. |
Databáze: | OpenAIRE |
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