Identification and quantification of host cell protein impurities in biotherapeutics using mass spectrometry

Residual host cell proteins (HCPs) in biotherapeutics can present potential safety risks to patients or compromise product stability. As such, their levels are typically monitored using a multicomponent HCP enzyme-linked immunosorbent assay (ELISA) to ensure adequate removal. However, it is not poss...

Full description

Saved in:
Bibliographic Details
Published in:Analytical biochemistry 2012-09, Vol.428 (2), p.150-157
Main Authors: Schenauer, Matthew R., Flynn, Gregory C., Goetze, Andrew M.
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Biological Therapy
Chromatography, Liquid
detection limit
drugs
enzyme-linked immunosorbent assay
Escherichia coli
Escherichia coli - metabolism
Escherichia coli Proteins - metabolism
Host cell proteins
Humans
liquid chromatography
Mass spectrometry
Mass Spectrometry - methods
Molecular Sequence Data
patients
peptides
Peptides - analysis
Peptides - chemistry
proteins
Proteins - analysis
Proteins - chemistry
Quantification
Receptors, Fc - metabolism
Recombinant Fusion Proteins - metabolism
Reference Standards
risk
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Residual host cell proteins (HCPs) in biotherapeutics can present potential safety risks to patients or compromise product stability. As such, their levels are typically monitored using a multicomponent HCP enzyme-linked immunosorbent assay (ELISA) to ensure adequate removal. However, it is not possible to guarantee ELISA coverage of every possible HCP impurity, and the specific HCPs remaining following purification are rarely identified. In the current study, we characterized the ability of an advanced two-dimensional liquid chromatography/mass spectrometry platform (2D–LC/MSE) to identify and quantify known low-level spiked protein impurities in a therapeutic peptide Fc fusion protein. The label-free quantification procedure based on the “top 3” intensity tryptic peptides per protein was applied and improved on for this application. Limits of detection for unknown HCPs were approximated from the spiked protein data along with estimates for the quantitative accuracy of the method. In all, we established that most protein impurities present at 13±4ppm can be identified with a quantitative error of less than 2-fold using the more sensitive of two tested method formats. To conclude the study, we characterized all detectable Escherichia coli proteins present in this Fc fusion protein drug substance and discuss future applications of the method.
ISSN:0003-2697
1096-0309
DOI:10.1016/j.ab.2012.05.018