title: New expression method and characterization of recombinant human granulocyte colony stimulating factor in a stable protein formulation.
abstract: Human recombinant granulocyte colony stimulating factor (rhG-CSF) is widely used in hematology and oncology for the treatment of neutropenia, for the restoration of neutrophil production after bone marrow transplantation, for myelodysplastic syndromes, and aplastic anemia. The E. coli  expression system is commonly used for fast recombinant production of rhG-CSF at a large scale. We have applied a novel autoinduction method for the batch expression of rhG-CSF to study whether this new system would increase cell mass and target-protein yield compared to conventional E. coli  cell culture and induction with isopropyl β-D-thiogalactopyranoside (IPTG). We could demonstrate 3-fold higher culture densities and a 5-fold higher protein yield compared to IPTG induction without the need to monitor cell growth in a shortened 24 h expression procedure. rhG-CSF expressed  in autoinduction media was successfully extracted from E. coli inclusion bodies and refolded by dialysis. After size exclusion chromatography (SEC) purification, rhG-CSF showed similar conformation, biological activity and aggregation profile compared to the commercially available biosimilar  TEVAgrastim(®) (TEVA Pharma AG). Expression by autoinduction is suggested as a cost- and time-effective method for rhG-CSF production.
authors.affiliation: School of Pharmaceutical Sciences University of Geneva, University of Lausanne 30 Quai Ernest-Ansermet, CH-1211 Geneva 4, Switzerland.
authors.forename: Ralitza
authors.initials: R
authors.lastname: Boubeva
authors.forename: Christian
authors.initials: C
authors.lastname: Reichert
authors.forename: René
authors.initials: R
authors.lastname: Handrick
authors.forename: Claudia
authors.initials: C
authors.lastname: Müller
authors.forename: Jürgen
authors.initials: J
authors.lastname: Hannemann
authors.forename: Gerrit
authors.initials: G
authors.lastname: Borcharda
mesh_terms: D000818:Animals; D049109:Cell Proliferation / Q000187:drug effects; D002626:Chemistry, Pharmaceutical; D002942:Circular Dichroism; D003001:Cloning, Molecular; D003470:Culture Media; D004345:Drug Industry; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D016179:Granulocyte Colony-Stimulating Factor / Q000096:biosynthesis* / Q000737:chemistry / Q000494:pharmacology; D006801:Humans; D053613:Janus Kinase 1 / Q000502:physiology; D051379:Mice; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D050796:STAT3 Transcription Factor / Q000502:physiology; D015398:Signal Transduction / Q000187:drug effects; D013050:Spectrometry, Fluorescence; D013862:Thiogalactosides / Q000494:pharmacology
chemical_list: D003470:Culture Media; D011994:Recombinant Proteins; D050796:STAT3 Transcription Factor; D013862:Thiogalactosides; D016179:Granulocyte Colony-Stimulating Factor; D053613:Janus Kinase 1