Biphasic cultivation strategy to avoid Epo-Fc aggregation and optimize protein expression

Area of Science:

• Biotechnology
• Bioprocess Engineering
• Cell Culture Technology

Background:

• Biphasic cultivation strategies optimize cell growth and recombinant protein production.
• Chinese hamster ovary (CHO) cell lines are crucial for biopharmaceutical manufacturing.
• Protein aggregation is a critical quality attribute impacting therapeutic efficacy.

Purpose of the Study:

• To develop and optimize a biphasic cultivation strategy for a CHO cell line expressing an erythropoietin fusion protein (Epo-Fc).
• To investigate the influence of cultivation temperature and pH on cell growth, Epo-Fc production, and aggregation.
• To enhance the economic viability of the bioprocess through optimized culture conditions.

Main Methods:

• Utilized a biphasic batch cultivation strategy for CHO cells.
• Employed a Design of Experiments (DoE) approach with a fractional factorial design.
• Systematically evaluated the effects of temperature and pH shifts on cell growth, Epo-Fc production, and aggregation.

Main Results:

• Cultivation temperature significantly influenced cell growth, Epo-Fc production, and aggregation.
• A synergistic interaction between pH and temperature was identified, drastically reducing protein aggregation from 75% to <1% under optimized conditions (low temperature, acidic pH).
• Optimized conditions increased cell-specific Epo-Fc production rates and reduced nutrient consumption and waste production.

Conclusions:

• A biphasic cultivation strategy involving temperature and pH shifts is effective for controlling protein aggregation in CHO cell cultures.
• Optimized culture conditions enhance recombinant protein quality and production efficiency.
• This approach leads to a more economical and robust bioprocess for Epo-Fc production.