title: Differences in N-glycosylation of recombinant human coagulation factor VII derived from BHK, CHO, and HEK293 cells.
abstract: BACKGROUND &
  
  
  METHODS
  Recombinant factor VII (rFVII), the precursor molecule for recombinant activated FVII (rFVIIa), is, due to its need for complex post translational modifications, produced in mammalian cells. To evaluate the suitability of a human cell line in order to produce rFVII with post-translational modifications as close as possible to pdFVII, we compared the biochemical properties of rFVII synthesized in human embryonic kidney-derived (HEK)293 cells (HEK293rFVII) with those of rFVII expressed in Chinese hamster ovary (CHO, CHOrFVII) and baby hamster kidney (BHK, BHKrFVII) cells, and also with those of plasma derived FVII (pdFVII), using various analytical methods. rFVII was purified from selected production clones derived from BHK, CHO, and HEK293 cells after stable transfection, and rFVII isolates were analyzed for protein activity, impurities and post-translational modifications. RESULTS &
  
  
  CONCLUSIONS
  The analytical results showed no apparent gross differences between the various FVII proteins, except in their N-linked glycosylation pattern. Most N-glycans found on rFVII produced in HEK293 cells were not detected on rFVII from CHO and BHK cells, or, somewhat unexpectedly, on pdFVII; all other protein features were similar. HEK293rFVII glycans were mainly characterized by a higher structural variety and a lower degree of terminal sialylation, and a high amount of terminal N-acetyl galactosamines (GalNAc). All HEK293rFVII oligosaccharides contained one or more fucoses (Fuc), as well as hybrid and high mannose (Man) structures.
  
  
  CONCLUSIONS
  From all rFVII isolates investigated, CHOrFVII contained the highest degree of sialylation and no terminal GalNAc, and CHO cells were therefore assumed to be the best option for the production of rFVII.
authors.affiliation: Baxalta Innovations GmbH, Uferstraße 15, A-2304, Orth/Donau, Austria. ernst.boehm@baxalta.com.
authors.forename: Ernst
authors.initials: E
authors.lastname: Böhm
authors.affiliation: Baxalta Innovations GmbH, Uferstraße 15, A-2304, Orth/Donau, Austria. birgit.seyfried@baxalta.com.
authors.forename: Birgit K
authors.initials: BK
authors.lastname: Seyfried
authors.affiliation: Baxalta Innovations GmbH, Uferstraße 15, A-2304, Orth/Donau, Austria. michael.dockal@baxalta.com.
authors.forename: Michael
authors.initials: M
authors.lastname: Dockal
authors.affiliation: Baxalta Innovations GmbH, Uferstraße 15, A-2304, Orth/Donau, Austria. michael.graninger@baxalta.com.
authors.forename: Michael
authors.initials: M
authors.lastname: Graninger
authors.affiliation: Baxalta Innovations GmbH, Uferstraße 15, A-2304, Orth/Donau, Austria. meinhard.hasslacher@baxalta.com.
authors.forename: Meinhard
authors.initials: M
authors.lastname: Hasslacher
authors.affiliation: Baxalta Innovations GmbH, Uferstraße 15, A-2304, Orth/Donau, Austria. marianne.neurath@baxalta.com.
authors.forename: Marianne
authors.initials: M
authors.lastname: Neurath
authors.affiliation: Baxalta Innovations GmbH, Uferstraße 15, A-2304, Orth/Donau, Austria. christian.konetschny@baxalta.com.
authors.forename: Christian
authors.initials: C
authors.lastname: Konetschny
authors.affiliation: BaxaltaInnovations GmbH, Industriestraße 72, A-1220, Vienna, Austria. peter.matthiessen@baxalta.com.
authors.forename: Peter
authors.initials: P
authors.lastname: Matthiessen
authors.affiliation: Baxalta Innovations GmbH, Uferstraße 15, A-2304, Orth/Donau, Austria. artur.mitterer@baxalta.com.
authors.forename: Artur
authors.initials: A
authors.lastname: Mitterer
authors.affiliation: Baxalta Innovations GmbH, Donau-City Straße 7, 1220, Vienna, Austria. fritz.scheiflinger@baxalta.com.
authors.forename: Friedrich
authors.initials: F
authors.lastname: Scheiflinger
mesh_terms: D000818:Animals; D016466:CHO Cells; D002240:Carbohydrate Sequence; D006224:Cricetinae; D003412:Cricetulus; D015942:Factor VIIa / Q000737:chemistry* / Q000378:metabolism*; D006031:Glycosylation; D057809:HEK293 Cells; D006801:Humans; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; C103587:recombinant FVIIa; D015942:Factor VIIa