Proteomics Reveals Distinctive Host Cell Protein Expression Patterns in Fed-Batch and Perfusion Cell Culture...

Area of Science:

• Biotechnology
• Proteomics
• Cell Culture Engineering

Background:

• Chinese hamster ovary (CHO) cells are key for recombinant protein production, particularly monoclonal antibodies (mAbs).
• Fed-batch (FB) processes are standard, but high-density perfusion processes offer advantages in productivity and cost.
• Controlling host cell proteins (HCPs) is critical for biologics safety and efficacy.

Purpose of the Study:

• To compare host cell protein (HCP) expression dynamics in Chinese hamster ovary (CHO) cell culture under fed-batch (FB) and perfusion processes.
• To identify differences in proteomic profiles and understand HCP abundance variations between process modes.
• To inform strategies for managing high-risk HCPs in biologics manufacturing.

Main Methods:

• Utilized data-independent acquisition (DIA) proteomics to analyze cell culture supernatants.
• Compared proteomic profiles from 14 FB clones and 3 perfusion clones producing the same mAb.
• Performed cluster analysis and differential abundance analysis to identify protein variations.

Main Results:

• Perfusion processes demonstrated enhanced cell growth and productivity compared to FB.
• Perfusion processes exhibited distinct proteomic profiles and more comparable HCP abundance across clones.
• Identified significant protein differences between FB and perfusion, including 46 problematic HCPs.

Conclusions:

• Perfusion processes offer a more consistent HCP profile, beneficial for biologics manufacturing.
• Understanding HCP expression dynamics under different process modes is essential for impurity control.
• This research provides a foundation for developing strategies to manage problematic HCPs.