Human hepatoma cell line HuH-7 is an effective cellular system to produce recombinant factor IX with improved post-translational modifications
Abstract
Recombinant factor IX (rFIX) is increasingly used to treat patients with hemophilia B. CHO (Chinese Hamster Ovary) cells are commonly used for the production of rFIX but they have a limited capacity for introducing post-translational modifications (PTM) leading to incomplete γ-carboxylation, low phosphorylation and sulfation profiles as compared with plasma-derived preparations. Imperfect PTM might have an impact on the activity of Factor IX molecule. Several studies in animal models as well as clinical trials have previously reported a lower recovery of rFIX compared to plasma-derived FIX concentrates. In the present study, we aimed to produce a rFIX having a profile of PTM similar to plasma-derived FIX, using human hepatoma cell line HuH-7. We showed that rFIX produced by HuH-7 cells followed the classical intracellular pathway before secretion. In addition, improved PTM were associated with fully active molecule compared to plasma-derived and recombinant control FIX molecules. Secreted rFIX presented as a single band at the correct molecular weight. HuH-7 cellular clones were obtained and they secreted a biologically active human FIX. FIX was then purified for a detailed evaluation of PTM. Glycosylation and sialylation profiles were similar to plasma-derived and rFIX and mass spectrometry analysis demonstrated the presence of phosphorylated and sulfated forms of rFIX. These data strongly support that HuH-7 cells may represent an effective cellular system for production of…
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CAP and HuH-7 cells display human-like glycosylation profiles.
“Two other cell lines, the CAP (CEVEC's Amniocyte Production) cells of human amniocytes origin and the HuH-7 cells of human hepatocellular carcinoma origin, are presently tested for recombinant glycoprotein production in preclinical phases and both display human-like glycosylation profiles (Schiedner et al., 2008;Wolfel et al., 2011;Enjolras et al., 2012;Swiech et al., 2012;Wissing et al., 2015).”
From Therapeutic glycoprotein production in mammalian cells (2017) · cites this paper as Enjolras et al., 2012
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Therapeutic glycoprotein production in mammalian cells
Lalonde, M.-E., & Durocher, Y. (2017) · Journal of Biotechnology · doi:10.1016/j.jbiotec.2017.04.028 · Cited by 296 · see citing works
- Authors
- Enjolras, N., Dargaud, Y., Pérot, E., Guillaume, F., Becchi, M., & Négrier, C.
- Affiliations
- Hospices Civils de Lyon (FR)
- Lyon 1 Université (FR)
- Corresponding author
- Enjolras, Nathalie
- Venue
- Thrombosis Research
- Publisher
- Elsevier BV
- Published
- 2012-11
- Volume
- 130
- Issue
- 5
- Pages
- e266-e273
- ISSN
- 0049-3848 (ISSN-L)
- OpenAlex
- W2070573402
- Cited by
- 26
- References
- 0
- Citations by year
- 2024: 2 · 2022: 1 · 2021: 1 · 2019: 6 · 2018: 5 · 2017: 3 · 2015: 5 · 2014: 2 · 2013: 1
- FWCI
- 1.70
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Thrombosis Research (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Health Sciences › Medicine › Hematology › Hemophilia Treatment and Research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Viral Infectious Diseases and Gene Expression in Insects
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein purification and stability
- Keywords
- recombinant factor IX, Huh7 cells, human hepatoma cell lines, post-translational modification, hemophilia B, glycosylation, sialylation, phosphorylation, sulfation, gamma-carboxylation, protein secretion, CHO cells, recombinant protein expression
- MeSH terms
- Animals, Factor IX, Glycosylation, Cricetinae, Humans, Liver Neoplasms, Protein Processing, Post-Translational, Recombinant Proteins, Transfection, CHO Cells, Cell Line, Tumor
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