کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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3027616 | 1182981 | 2012 | 8 صفحه PDF | دانلود رایگان |
![عکس صفحه اول مقاله: Human hepatoma cell line HuH-7 is an effective cellular system to produce recombinant factor IX with improved post-translational modifications Human hepatoma cell line HuH-7 is an effective cellular system to produce recombinant factor IX with improved post-translational modifications](/preview/png/3027616.png)
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 rFIX exhibiting PTM similar to plasma-derived FIX.
Journal: Thrombosis Research - Volume 130, Issue 5, November 2012, Pages e266–e273