title: Production, purification, and chemical stability of recombinant human interferon-γ in low oxygen tension condition: a formulation approach.
abstract: The low stability of recombinant human interferon-γ (rhIFN-γ) therapeutic protein imposes some restrictions in its medical applications. In the current study, the effect of oxygen tension on the stability of purified rhIFN-γ was investigated. The rhIFN-γ was purified (>99%) by a two-step chromatographic process. Storage vials were filled by purified formulated product under normal atmospheric oxygen and low oxygen tension conditions. At different time intervals, the amounts of rhIFN-γ covalent dimers and deamidated forms were analyzed using analytical high-performance liquid chromatography (HPLC; size exclusion and cation exchange) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) methods. To determine the biological activity of purified rhIFN-γ, an antiviral activity assay against vesicular stomatitis virus (VSV) was performed. Upon rhIFN-γ long-term storage in a low oxygen tension condition, the amounts of rhIFN-γ covalent dimers and deamidated forms and also the biological activity of rhIFN-γ changed a little. In contrast, by 9 months of storage of rhIFN-γ preparations under normal atmospheric condition, the amount of covalent dimers and deamidated forms increased with time and reached to approximately 3.5% and 11.5% of the initial amount, respectively. The antiviral specific activity of 9-month-old rhIFN-γ preparations decreased to 41% of the initial amount at normal storage condition, while no significant reduction was seen at the low oxygen tension condition. In conclusion, oxygen tension during storage could have a significant impact on rhIFN-γ stability and finally on the quality of pharmaceutical rhIFN-γ product.
authors.affiliation: Malek-e Ashtar University of Technology, Tehran, Iran.
authors.forename: Narges
authors.initials: N
authors.lastname: Malek-Sabet
authors.forename: Mohammad Reza
authors.initials: MR
authors.lastname: Masoumian
authors.forename: Majid
authors.initials: M
authors.lastname: Zeinali
authors.forename: Rasoul
authors.initials: R
authors.lastname: Khalilzadeh
authors.forename: Jafar Mohamadian
authors.initials: JM
authors.lastname: Mousaabadi
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D004355:Drug Stability; D004356:Drug Storage / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005285:Fermentation; D006801:Humans; D007371:Interferon-gamma / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology; D010100:Oxygen / Q000737:chemistry / Q000378:metabolism; D055503:Protein Multimerization; D055550:Protein Stability; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology; D013997:Time Factors; D018116:Vesiculovirus / Q000187:drug effects
chemical_list: D000998:Antiviral Agents; D011994:Recombinant Proteins; D007371:Interferon-gamma; D010100:Oxygen