Host cell protein profiling and immunogenicity assessment in E. coli-produced interferon beta

Area of Science:

• Biopharmaceutical Manufacturing
• Protein Chemistry
• Immunology

Background:

• Host cell proteins (HCPs) are critical impurities in biologic drugs.
• Assessing immunogenicity risk of bacterial HCPs is less understood than for CHO-derived platforms.

Purpose of the Study:

• To profile and assess the immunogenicity risk of HCPs in a recombinant human interferon beta-1b (rhIFNβ-1b) produced in E. coli.
• To establish a framework for bacterial HCP risk assessment.

Main Methods:

• Liquid chromatography-tandem mass spectrometry (LC-MS/MS) for HCP identification.
• Recombinant expression and orthogonal immunogenicity assays (THP-1 monocytes, RAW-Blue™ reporter cells, ex vivo human T-cell assays).

Main Results:

• Nine residual HCPs were identified, mainly cytoplasmic enzymes.
• Two HCPs (W7, E6) showed pro-inflammatory gene expression at high concentrations but no NF-κB activation or T-cell responses.
• One HCP (42) exhibited no immunostimulatory activity.

Conclusions:

• The bacterial HCPs evaluated pose a low immunogenicity risk at relevant concentrations in the purified drug product.
• This study addresses knowledge gaps for non-CHO expression systems and provides a risk assessment framework.