کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
5832173 | 1122593 | 2015 | 12 صفحه PDF | دانلود رایگان |

- Successfully expressed and purified the mammalian rhIGFBP7, an IGF-1 targeted inhibition protein, using transient gene expression and conventional non-affinity column chromatography purification method.
- High quality material IGFBP7 was generated for further pre-clinical drug development studies.
- Transient gene expression technology through optimization of vectors is practical to be used for production of recombinant non-antibody protein products for biopharmaceutical development.
Targeted inhibiting insulin-like growth factor 1 is an effective approach for cancer therapy. Insulin-like growth factor binding protein 7 (IGFBP7) is considered as a potential therapeutic protein. However, producing high quality of such non-IgG proteins in mammalian cells is still a challenge in biopharmaceutical development. Here, we report a rapid production process by using transient gene transfection in HEK 293E cells. A set of constructs combining several expression promoters, leader sequences, and 5â² un-translated regions were generated and optimized, from which the best vector with expression level at ~Â 50Â mg/L was selected for production at 2Â L cell culture scale. Comparison study in downstream purification methods led to development of a scalable, non-affinity chromatography strategy through Super Q, Fast Flow Q, and Heparin columns. The product was characterized in purity (99%), isoelectric point, molecule weight, glycosylation, and stability by using SEC-HPLC, SDS-PAGE, isoelectric focusing and mass spectrometry. The highly purified product shows IGF-1 binding activity and inhibits IGF-1-induced cell proliferation. This process not only provides a remarkable high expression at ~Â 50Â mg/L and pure glycosylated mammalian rhIGFBP7, also highlights that transient gene expression technology is practical to be used for production and early development of recombinant non-IgG therapeutic proteins.
Journal: International Immunopharmacology - Volume 29, Issue 2, December 2015, Pages 476-487