title: Studies of protein oxidation as a product quality attribute on a scale-down model for cell culture process development.
abstract: Of importance to the biological properties of proteins produced in cell culture systems are the complex post-translational modifications that are affected by variations in process conditions. Protein oxidation, oxidative modification to intracellular proteins that involves cleavage of the polypeptide chain, and modifications of the amino acid side chains can be affected by such process variations. Dissolved oxygen is a parameter of increasing interest since studies have shown that despite the necessity of oxygen for respiration, there may also be some detrimental effects of oxygen to the cell. Production and accumulation of reactive oxygen species can cause damage to proteins as a result of oxidation of the cell and cellular components. Variation, or changes to cell culture products, can affect function, clearance rate, immunogenicity, and specific activity, which translates into clinical implications. The effect of increasing dissolved oxygen on protein oxidation in immunoglobulin G3-producing mouse hybridoma cells was studied using 50 mL high-throughput mini-bioreactors that employ non-invasive optical sensor technology for monitoring and closed feedback control of pH and dissolved oxygen. Relative protein carbonyl concentration of proteins produced under varying levels of dissolved oxygen was measured by enzyme-linked immunosorbent assay and used as an indicator of oxidative damage. A trend of increasing protein carbonyl content in response to increasing dissolved oxygen levels under controlled conditions was observed.
  
  
  
  Protein oxidation, oxidative modification to intracellular proteins that involves cleavage of the polypeptide chain, and modifications of the amino acid side chains can be affected by variations in dissolved oxygen levels in cell culture systems. Studies have shown that despite the necessity of oxygen for respiration, there may be detrimental effects of oxygen to the cell. Production and accumulation of reactive oxygen species can cause damage to proteins as a result of oxidation of the cell and cellular components, affecting function, clearance rate, immunogenicity, and specific activity, which translates into clinical implications. The effect of increasing dissolved oxygen on protein oxidation in immunoglobulin G3-producing mouse hybridoma cells was studied using 50 mL high-throughput mini-bioreactors that employ non-invasive optical sensor technology for monitoring and closed feedback control of pH and dissolved oxygen. Protein carbonyl concentration of proteins produced under varying levels of dissolved oxygen was measured by enzyme-linked immunosorbent assay and used as an indicator of oxidative damage. A trend of increasing protein carbonyl content in response to increasing dissolved oxygen levels under controlled conditions was observed.
authors: Department of Biomedical Sciences, Hood College, Frederick, MD; Currently at MedImmune, Inc., Gaithersburg, MD; LeeND@MedImmune.com.
authors: Nacole D
authors: ND
authors: Lee
authors: Center for Advanced Sensor Technology and Department of Chemical, Biochemical and Environmental Engineering; University of Maryland, Baltimore County; Baltimore, MD; Currently at Vaccine Research Center, NIH; and.
authors: Bhargavi
authors: B
authors: Kondragunta
authors: Center for Advanced Sensor Technology and Department of Chemical, Biochemical and Environmental Engineering; University of Maryland, Baltimore County; Baltimore, MD;
authors: Shaunak
authors: S
authors: Uplekar
authors: Center for Advanced Sensor Technology and Department of Chemical, Biochemical and Environmental Engineering; University of Maryland, Baltimore County; Baltimore, MD; Currently at MS&T-AstraZeneca, Frederick, MD.
authors: Jose
authors: J
authors: Vallejos
authors: Center for Advanced Sensor Technology and Department of Chemical, Biochemical and Environmental Engineering; University of Maryland, Baltimore County; Baltimore, MD;
authors: Antonio
authors: A
authors: Moreira
authors: Center for Advanced Sensor Technology and Department of Chemical, Biochemical and Environmental Engineering; University of Maryland, Baltimore County; Baltimore, MD;
authors: Govind
authors: G
authors: Rao
mesh_terms: D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D019149:Bioreactors; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D057166:High-Throughput Screening Assays; D006825:Hybridomas / Q000276:immunology*; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000276:immunology*; D051379:Mice; D010084:Oxidation-Reduction; D010100:Oxygen / Q000737:chemistry; D050050:Protein Carbonylation; D011506:Proteins / Q000737:chemistry*
chemical_list: D000596:Amino Acids; D007074:Immunoglobulin G; D011506:Proteins; D010100:Oxygen