Targeted capture of Chinese hamster ovary host cell proteins: Peptide ligand binding by proteomic analysis

2019 ◽  
Vol 117 (2) ◽  
pp. 438-452 ◽  
Author(s):  
R. Ashton Lavoie ◽  
Alice Fazio ◽  
Taufika Islam Williams ◽  
Ruben Carbonell ◽  
Stefano Menegatti
Keyword(s):  
Ligand Binding ◽  
Host Cell ◽  
Chinese Hamster Ovary ◽  
Proteomic Analysis ◽  
Chinese Hamster ◽  
Peptide Ligand ◽  
Host Cell Proteins ◽  
Targeted Capture

scholarly journals Targeted Capture of Chinese Hamster Ovary Host Cell Proteins: Peptide Ligand Discovery

2019 ◽  
Vol 20 (7) ◽  
pp. 1729 ◽  
Author(s):  
R. Lavoie ◽  
Alice di Fazio ◽  
R. Blackburn ◽  
Michael Goshe ◽  
Ruben Carbonell ◽  
...  
Keyword(s):  
Host Cell ◽  
Cell Line ◽  
Chinese Hamster Ovary ◽  
Mixed Mode ◽  
Solid Phase ◽  
Chinese Hamster ◽  
Peptide Ligand ◽  
Host Cell Proteins ◽  
Ligand Discovery ◽  
Exchange Resins

The growing integration of quality-by-design (QbD) concepts in biomanufacturing calls for a detailed and quantitative knowledge of the profile of impurities and their impact on the product safety and efficacy. Particularly valuable is the determination of the residual level of host cell proteins (HCPs) secreted, together with the product of interest, by the recombinant cells utilized for production. Though often referred to as a single impurity, HCPs comprise a variety of species with diverse abundance, size, function, and composition. The clearance of these impurities is a complex issue due to their cell line to cell line, product-to-product, and batch-to-batch variations. Improvements in HCP monitoring through proteomic-based methods have led to identification of a subset of “problematic” HCPs that are particularly challenging to remove, both at the product capture and product polishing steps, and compromise product stability and safety even at trace concentrations. This paper describes the development of synthetic peptide ligands capable of capturing a broad spectrum of Chinese hamster ovary (CHO) HCPs with a combination of peptide species that allow for advanced mixed-mode binding. Solid phase peptide libraries were screened for identification and characterization of peptides that capture CHO HCPs while showing minimal binding of human IgG, utilized here as a model product. Tetrameric and hexameric ligands featuring either multipolar or hydrophobic/positive amino acid compositions were found to be the most effective. Tetrameric multipolar ligands exhibited the highest targeted binding ratio (ratio of HCP clearance over IgG loss), more than double that of commercial mixed-mode and anion exchange resins utilized by industry for IgG polishing. All peptide resins tested showed preferential binding to HCPs compared to IgG, indicating potential uses in flow-through mode or weak-partitioning-mode chromatography.

scholarly journals Recovery of Chinese hamster ovary host cell proteins for proteomic analysis

2013 ◽  
Vol 9 (1) ◽  
pp. 87-99 ◽  
Author(s):  
Kristin N. Valente ◽  
Amy K. Schaefer ◽  
Hannah R. Kempton ◽  
Abraham M. Lenhoff ◽  
Kelvin H. Lee
Keyword(s):  
Host Cell ◽  
Chinese Hamster Ovary ◽  
Proteomic Analysis ◽  
Chinese Hamster ◽  
Host Cell Proteins

scholarly journals CRISPR/Cas9-Multiplexed Editing of Chinese Hamster Ovary B4Gal-T1, 2, 3, and 4 TailorsN-Glycan Profiles of Therapeutics and Secreted Host Cell Proteins

2018 ◽  
Vol 13 (10) ◽  
pp. 1800111 ◽  
Author(s):  
Thomas Amann ◽  
Anders Holmgaard Hansen ◽  
Stefan Kol ◽  
Gyun Min Lee ◽  
Mikael Rørdam Andersen ◽  
...  
Keyword(s):  
Host Cell ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster ◽  
Host Cell Proteins

scholarly journals Improving Chinese hamster ovary host cell protein ELISA using Ella®: an automated microfluidic platform

BioTechniques ◽  
2020 ◽  
Vol 69 (3) ◽  
pp. 186-192
Author(s):  
Kathleen Van Manen-Brush ◽  
Jacob Zeitler ◽  
John R White ◽  
Paul Younge ◽  
Samantha Willis ◽  
...  
Keyword(s):  
Host Cell ◽  
Chinese Hamster Ovary ◽  
Expression System ◽  
Chinese Hamster ◽  
Cell Protein ◽  
Enzyme Linked Immunosorbent Assay ◽  
Host Cell Proteins ◽  
Common Technique ◽  
Automated Instrument ◽  
Cell Expression

Chinese hamster ovary (CHO) cells are a mammalian cell line used in the production of therapeutic proteins. Host cell proteins (HCPs) are process-related impurities that are derived from the host cell expression system. During biopharmaceutical drug development, removal of HCPs is required. Enzyme-linked immunosorbent assay (ELISA) is a common technique to quantitate HCPs, but is a labor-intensive process that takes up to 7 h. Ella® is an automated instrument that utilizes microfluidics and glass nanoreactors to quantitate HCPs in 75 min using similar ELISA reagents. The antibodies and antigens are captured on three distinct glass nanoreactors, resulting in sensitive reproducible data. Our results indicate that Ella quantitates CHO HCPs with precision, accuracy, sensitivity and trends comparable with our traditional CHO HCP ELISA.

scholarly journals Survival, mutagenesis, and host cell reactivation in a Chinese hamster ovary cell ERCC1 knock-out mutant

Mutagenesis ◽  
1997 ◽  
Vol 12 (4) ◽  
pp. 277-283 ◽  
Author(s):  
Rhonda L. Rolig ◽  
Susan K. Layher ◽  
Barbara Santi ◽  
Gerald M. Adair ◽  
Feng Gu ◽  
...  
Keyword(s):  
Host Cell ◽  
Chinese Hamster Ovary Cell ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster ◽  
Ovary Cell ◽  
Knock Out ◽  
Cell Reactivation ◽  
Host Cell Reactivation

scholarly journals Optimization of Sequence, Display, and Mode of Operation of IgG-Binding Peptide Ligands to Develop Robust, High-Capacity Affinity Adsorbents That Afford High IgG Product Quality

2019 ◽  
Vol 20 (1) ◽  
pp. 161 ◽  
Author(s):  
Tuhidul Islam ◽  
Amith D. Naik ◽  
Yasuhiro Hashimoto ◽  
Stefano Menegatti ◽  
Ruben G. Carbonell
Keyword(s):  
Binding Capacity ◽  
Protein A ◽  
Chinese Hamster ◽  
High Capacity ◽  
Alkaline Solutions ◽  
Peptide Ligand ◽  
Dynamic Binding ◽  
Ligand Density ◽  
Host Cell Proteins ◽  
Igg Binding

This work presents the use of peptide ligand HWRGWV and its cognate sequences to develop affinity adsorbents that compete with Protein A in terms of binding capacity and quality of the eluted product. First, the peptide ligand was conjugated to crosslinked agarose resins (WorkBeads) at different densities and using different spacer arms. The optimization of ligand density and display resulted in values of static and dynamic binding capacity of 85 mg/mL and 65 mg/mL, respectively. A selected peptide-WorkBeads adsorbent was utilized for purifying Mabs from Chinese Hamster Ovary (CHO) cell culture supernatants. The peptide-WorkBeads adsorbent was found able to withstand sanitization with strong alkaline solutions (0.5 M NaOH). The purity of the eluted product was consistently higher than 95%, with logarithmic removal value (LRV) of 1.5 for host cell proteins (HCPs) and 4.0 for DNA. HCP clearance was significantly improved by adding a post-load washing step with either 0.1 M Tris HCl pH 9 or 1 M NaCl. The cognate peptide of HWRGWV, constructed by replacing arginine (R) with citrulline, further increased the HCP LRV to 2.15. The peptide-based adsorbent also showed a remarkable performance in terms of removal of Mab aggregates; unlike Protein A, in fact, HWRGWV was found to bind only monomeric IgG. Collectively, these results demonstrate the potential of peptide-based adsorbents as alternative to Protein A for the purification of therapeutic antibodies.

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