title: Novel human renal proximal tubular cell line for the production of complex proteins.
abstract: Human host cell lines for the production of biopharmaceutical proteins are of interest due to differences in the glycosylation patterns of human and animal cell lines. Specifically, sialylation, which has a major impact on half-life and immunogenicity of recombinant biopharmaceuticals, differs markedly. Here, we established and characterized an immortalized well documented and serum-free host cell line, RS, from primary human renal proximal tubular epithelial cells (RPTEC). In order to test its capacity to produce complex glycosylated proteins, stable recombinant human erythropoietin (rhEpo) producing clones were generated. The clone with highest productivity, RS-1C9 was further characterized and showed stable productivity. Biological activity was observed in in vitro assays and 28% of rhEpo glyco-isoforms produced by RS-1C9 were in range and distribution of the biological reference standard (BRP) isoform, as compared to 11.5% of a CHO based rhEpo. Additionally, cellular α-2,6 sialylation, Galactose-alpha-1,3-galactose (alpha-Gal) and N-glycolylneuraminic acid (NeuGc) patterns compare favourably to CHO cells. While productivity of RS still needs optimization, its amenability to upscaling in bioreactors, its production of glyco-isoforms that will increase yields after down-stream processing of about 2.5 fold, presence of sialylation and lack of Neu5Gc recommend RS as alternative human host cell line for production of biopharmaceuticals.
authors.affiliation: ACIB, Muthgasse 11, A-1190 Vienna, Austria.
authors.forename: Lukas
authors.initials: L
authors.lastname: Fliedl
authors.affiliation: ACIB, Muthgasse 11, A-1190 Vienna, Austria.
authors.forename: Gabriele
authors.initials: G
authors.lastname: Manhart
authors.affiliation: ACIB, Muthgasse 11, A-1190 Vienna, Austria.
authors.forename: Florian
authors.initials: F
authors.lastname: Kast
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences Vienna, Muthgasse 18, A-1190 Vienna, Austria.
authors.forename: Hermann
authors.initials: H
authors.lastname: Katinger
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences Vienna, Muthgasse 18, A-1190 Vienna, Austria.
authors.forename: Renate
authors.initials: R
authors.lastname: Kunert
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences Vienna, Muthgasse 18, A-1190 Vienna, Austria.
authors.forename: Johannes
authors.initials: J
authors.lastname: Grillari
authors.affiliation: ACIB, Muthgasse 11, A-1190 Vienna, Austria.
authors.forename: Matthias
authors.initials: M
authors.lastname: Wieser
authors.affiliation: ACIB, Muthgasse 11, A-1190 Vienna, Austria; Department of Biotechnology, University of Natural Resources and Life Sciences Vienna, Muthgasse 18, A-1190 Vienna, Austria. Electronic address: regina.grillari@boku.ac.at.
authors.forename: Regina
authors.initials: R
authors.lastname: Grillari-Voglauer
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000032:analysis; D016466:CHO Cells; D060846:Cell Engineering / Q000379:methods*; D002460:Cell Line; D003412:Cricetulus; D004847:Epithelial Cells / Q000378:metabolism*; D004921:Erythropoietin / Q000235:genetics / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D007687:Kidney Tubules, Proximal / Q000166:cytology*; D020033:Protein Isoforms / Q000378:metabolism; D014162:Transfection
chemical_list: D015415:Biomarkers; C554270:EPO protein, human; D020033:Protein Isoforms; D004921:Erythropoietin
keywords: Erythropoietin; Glycosylation; Isoelectric focussing; RPTEC; Sialylation