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Tytuł:
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Novel peptide ligands for antibody purification provide superior clearance of host cell protein impurities.
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Autorzy:
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Reese HR; Department of Chemical and Biomolecular Engineering, North Carolina State University, Campus Box 7905, Raleigh, NC, 27695-7905, USA.
Xiao X; Department of Chemical and Biomolecular Engineering, North Carolina State University, Campus Box 7905, Raleigh, NC, 27695-7905, USA.
Shanahan CC; Department of Chemical and Biomolecular Engineering, North Carolina State University, Campus Box 7905, Raleigh, NC, 27695-7905, USA.
Chu W; Department of Chemical and Biomolecular Engineering, North Carolina State University, Campus Box 7905, Raleigh, NC, 27695-7905, USA.
Van Den Driessche GA; Department of Chemistry, North Carolina State University, Raleigh, NC, 27695-8204, USA.
Fourches D; Department of Chemistry, North Carolina State University, Raleigh, NC, 27695-8204, USA.
Carbonell RG; Department of Chemical and Biomolecular Engineering, North Carolina State University, Campus Box 7905, Raleigh, NC, 27695-7905, USA; Biomanufacturing Training and Education Center (BTEC), North Carolina State University, Raleigh, NC, 27695-7928, USA.
Hall CK; Department of Chemical and Biomolecular Engineering, North Carolina State University, Campus Box 7905, Raleigh, NC, 27695-7905, USA.
Menegatti S; Department of Chemical and Biomolecular Engineering, North Carolina State University, Campus Box 7905, Raleigh, NC, 27695-7905, USA; Biomanufacturing Training and Education Center (BTEC), North Carolina State University, Raleigh, NC, 27695-7928, USA. Electronic address: .
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Źródło:
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Journal of chromatography. A [J Chromatogr A] 2020 Aug 16; Vol. 1625, pp. 461237. Date of Electronic Publication: 2020 May 23.
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Typ publikacji:
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Journal Article
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Język:
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English
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Imprint Name(s):
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Original Publication: Amsterdam ; New York : Elsevier, 1993-
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MeSH Terms:
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Antibodies, Monoclonal/*isolation & purification
Chromatography, Affinity/*methods
Peptides/*chemistry
Amino Acid Sequence ; Animals ; Antibodies, Monoclonal/metabolism ; CHO Cells ; Cricetinae ; Cricetulus ; Immunoglobulin G/isolation & purification ; Immunoglobulin G/metabolism ; Ligands ; Peptides/chemical synthesis ; Peptides/metabolism ; Protein Binding ; Staphylococcal Protein A/chemistry ; Staphylococcal Protein A/metabolism
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Contributed Indexing:
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Keywords: Affinity chromatography; Peptide affinity ligands; Peptide design; Protein A; mAb purification
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Substance Nomenclature:
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0 (Antibodies, Monoclonal)
0 (Immunoglobulin G)
0 (Ligands)
0 (Peptides)
0 (Staphylococcal Protein A)
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Entry Date(s):
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Date Created: 20200726 Date Completed: 20200908 Latest Revision: 20200908
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Update Code:
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20240104
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DOI:
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10.1016/j.chroma.2020.461237
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PMID:
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32709313
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The quest for ligands alternative to Protein A for the purification of monoclonal antibodies (mAbs) has been pursued for almost three decades. Yet, the IgG-binding peptides known to date still fall short of the host cell protein (HCP) logarithmic removal value (LRV) set by Protein A media (2.5-3.1). In this study, we present an integrated computational-experimental approach leading to the discovery of peptide ligands that provide HCP LRVs on par with Protein A. First, the screening of 60,000 peptide variants was performed using a high-throughput search algorithm to identify sequences that ensure IgG affinity binding. Select sequences WQRHGI, MWRGWQ, RHLGWF, and GWLHQR were then negatively screened in silico against a panel of model HCPs to ensure the selection of peptides with high binding selectivity. Candidate ligands WQRHGI and MWRGWQ were conjugated to chromatographic resins and characterized by isothermal binding and breakthrough assays to quantify static and dynamic binding capacity (Q max and DBC 10% ), respectively. The resulting Q max were 52.6 mg of IgG per mL of adsorbent for WQRHGI and 57.48 mg/mL for MWRGWQ, while the DBC 10% (2 minutes residence time) were 30.1 mg/mL for WQRHGI and 36.4 mg/mL for MWRGWQ. Evaluation of the peptides by isothermal titration calorimetry (ITC) confirmed the binding energy predicted in silico, and an amino acid scanning study corroborated the affinity-like binding activity of the peptides. WQRHGI-WorkBeads resin was finally characterized by purification of a monoclonal antibody from a Chinese Hamster Ovary (CHO) cell culture harvest, affording a remarkable HCP LRV of 2.7, and consistent product yield and purity over 100 chromatographic cycles. These results demonstrate the potential of WQRHGI as an effective alternative to Protein A for antibody purification.
Competing Interests: Declaration of Competing Interest None.
(Copyright © 2020. Published by Elsevier B.V.)
