title: Transcriptional profiling of gene expression changes in a PACE-transfected CHO DUKX cell line secreting high levels of rhBMP-2.
abstract: Chinese hamster ovary (CHO) cells are widely used in the biopharmaceutical industry for the production of recombinant human proteins including complex polypeptides such as recombinant human bone morphogenic protein 2 (rhBMP-2). Large-scale manufacture of rhBMP-2 has associated production difficulties resulting from incomplete processing of the recombinant human protein due to insufficient endogenous levels of the paired basic amino acid cleaving enzyme (PACE) in CHO. In order to resolve this issue, CHO DUKX cells expressing rhBMP-2 were transfected with the soluble version of human PACE (PACEsol) resulting in improved amino-terminal homogeneity and a fourfold increase in rhBMP-2 productivity. In this article, we present a microarray expression profile analysis comparing the parental lineage to the higher producing subclone co-expressing PACEsol using a proprietary CHO-specific microarray. Using this technology we observed 1,076 significantly different genes in the high-productivity cells co-expressing PACEsol. Following further analysis of the differentially expressed genes, the Unfolded Protein Response (UPR) component of the endoplasmic reticulum stress response pathway was identified as a key candidate for effecting increased productivity in this cell system. Several additional ER- and Golgi-localised proteins were identified which may also contribute to this effect. The results presented here support the use of large-scale microarray expression profiling as a viable and valuable route towards understanding the behaviour of bioprocess cultures in vitro.
authors.affiliation: National Institute for Cellular Biotechnology, Dublin City University, Glasnevin, Dublin 9, Ireland. padraig.doolan@dcu.ie
authors.forename: Padraig
authors.initials: P
authors.lastname: Doolan
authors.forename: Mark
authors.initials: M
authors.lastname: Melville
authors.forename: Patrick
authors.initials: P
authors.lastname: Gammell
authors.forename: Martin
authors.initials: M
authors.lastname: Sinacore
authors.forename: Paula
authors.initials: P
authors.lastname: Meleady
authors.forename: Kevin
authors.initials: K
authors.lastname: McCarthy
authors.forename: Linda
authors.initials: L
authors.lastname: Francullo
authors.forename: Mark
authors.initials: M
authors.lastname: Leonard
authors.forename: Timothy
authors.initials: T
authors.lastname: Charlebois
authors.forename: Martin
authors.initials: M
authors.lastname: Clynes
mesh_terms: D000818:Animals; D001483:Base Sequence; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000235:genetics*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D017931:DNA Primers; D045683:Furin / Q000235:genetics*; D020869:Gene Expression Profiling; D006801:Humans; D012333:RNA, Messenger / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics; D014158:Transcription, Genetic; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000235:genetics*
chemical_list: C525718:BMP2 protein, human; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D017931:DNA Primers; D012333:RNA, Messenger; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; D045683:Furin