Methods for characterizing disulfide bonds
AssigneeRegeneron Pharmaceuticals, Inc.
InventorShunhai Wang
Life of this patent
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Abstract
Compositions and methods for analyzing disulfide bonds are provided. An exemplary method includes preparing peptide standards having no disulfide bonds, scrambled disulfide bond peptide standards, and native disulfide bond peptide standards according to the sequence of the region of the protein drug product that includes the disulfide bond, digesting a sample of protein drug product into peptides, separating the protein drug product peptides, analyzing the protein drug product peptides and the peptide standards, identifying scrambled and native disulfide bond peptides by retention time, and quantifying the level of scrambled disulfide bond peptides.
Claims18
A method of producing a protein drug product, comprising:
- producing the protein drug product in a cell culture;
- identifying scrambled disulfide bonds of the protein drug product, wherein identifying the scrambled disulfide bonds of the protein drug product comprises:
- preparing peptide standards comprising regions of the protein drug product containing one or more disulfide bonds, wherein the disulfide bonds in the peptide standards are formed through an oxidation reaction that comprises exposing the peptide standard to zinc;
- and
- modifying one or more cell culture, purification or excipient conditions to reduce the amount of crossed hinge disulfide bonds of the protein drug product to less than 1.0%.
Dependencies are inferred from the claim text.