Design of a stable cell line producing recombinant darbepoetin alpha based on CHO cells

Shukurov, R. R.; Lobanova, N. V.; Savinova, Iryna; Vorobyova, I. G.; Nurbakov, A. A.; Ermolina, L. V.; Орлова, Н. В.; Mosina, A. G.; Antonova, Liliya P.; Хамитов, Р. А.; Seryogin, Yu. A.

doi:10.1134/s0003683814090063

Cited by 3 publications

(1 citation statement)

References 19 publications

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“…[26] DPO is a highly glycosylated protein with over 20 isoforms. [27] However, only six to eight isoforms remain in the final pharmaceutical product, because of their desired high content of sialic acidic groups, which is an important quality criterion for DPO. Therefore, the goal was to use continuous glucose monitoring and supply to assess different cultivation conditions without compromising product quality.…”

Section: Introductionmentioning

confidence: 99%

Continuous optical in‐line glucose monitoring and control in CHO cultures contributes to enhanced metabolic efficiency while maintaining darbepoetin alfa product quality

Lederle

Tric

Roth

et al. 2021

Biotechnology Journal

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Great efforts are directed towards improving productivity, consistency and quality of biopharmaceutical processes and products. One particular area is the development of new sensors for continuous monitoring of critical bioprocess parameters by using online or in‐line monitoring systems. Recently, we developed a glucose biosensor applicable in single‐use, in‐line and long‐term glucose monitoring in mammalian cell bioreactors. Now, we integrated this sensor in an automated glucose monitoring and feeding system capable of maintaining stable glucose levels, even at very low concentrations. We compared this fed‐batch feedback system at both low (< 1 mM) and high (40 mM) glucose levels with traditional batch culture methods, focusing on glycosylation and glycation of the recombinant protein darbepoetin alfa (DPO) produced by a CHO cell line. We evaluated cell growth, metabolite and product concentration under different glucose feeding strategies and show that continuous feeding, even at low glucose levels, has no harmful effects on DPO quantity and quality. We conclude that our system is capable of tight glucose level control throughout extended bioprocesses and has the potential to improve performance where constant maintenance of glucose levels is critical.

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Section: Introductionmentioning

confidence: 99%

Continuous optical in‐line glucose monitoring and control in CHO cultures contributes to enhanced metabolic efficiency while maintaining darbepoetin alfa product quality

Lederle

Tric

Roth

et al. 2021

Biotechnology Journal

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Purification and Characterization of Recombinant Darbepoetin Alfa from Leishmania tarentolae

et al. 2016

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Darbepoetin alfa is a biopharmaceutical glycoprotein that stimulates erythropoiesis and is used to treat anemia, which associated with renal failure and cancer chemotherapy. We herein describe the structural characterization of recombinant darbepoetin alfa produced by Leishmania tarentolae T7-TR host. The DNA expression cassette was integrated into the L. tarentolae genome through homologous recombination. Transformed clones were selected by antibiotic resistance, diagnostic PCRs, and protein expression analysis. The structure of recombinant darbepoetin alfa was analyzed by isoelectric focusing, ultraviolet-visible spectrum, and circular dichroism (CD) spectroscopy. Expression analysis showed the presence of a protein band at 40 kDa, and its expression level was 51.2 mg/ml of culture medium. Darbepoetin alfa have 5 isoforms with varying degree of sialylation. The UV absorption and CD spectra were analogous to original drug (Aranesp), which confirmed that the produced protein was darbepoetin alfa. Potency test results revealed that the purified protein was biologically active. In brief, the structural and biological characteristics of expressed darbepoetin alfa were very similar to Aranesp which has been normally expressed in CHO. Our data also suggest that produced protein has potential to be developed for clinical use.

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Ubiquitous chromatin opening element enhance Darbepoetin alfa expression in CHO DG44 cell line

Hasheminejad,

Jazayeri,

Halfinezhad

et al. 2024

Biochemical Engineering Journal

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Design of a stable cell line producing recombinant darbepoetin alpha based on CHO cells

Cited by 3 publications

References 19 publications

Continuous optical in‐line glucose monitoring and control in CHO cultures contributes to enhanced metabolic efficiency while maintaining darbepoetin alfa product quality

Continuous optical in‐line glucose monitoring and control in CHO cultures contributes to enhanced metabolic efficiency while maintaining darbepoetin alfa product quality

Purification and Characterization of Recombinant Darbepoetin Alfa from Leishmania tarentolae

Ubiquitous chromatin opening element enhance Darbepoetin alfa expression in CHO DG44 cell line

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