Methods for the purification of polypeptide conjugates comprising polyalkylene oxide using hydrophobic interaction chromatography
First Claim
1. A method of making a composition comprising a first polypeptide conjugate, said first polypeptide conjugate comprising a first number of poly(alkylene oxide) moieties covalently linked to said first polypeptide, wherein at least one of said poly(alkylene oxide) moieties is covalently linked to said first polypeptide via a glycosyl linking group, wherein said glycosyl linking group is either covalently linked to an amino acid residue of said first polypeptide or is covalently linked to a glycosyl moiety of said first polypeptide, said method comprising:
- (a) contacting a mixture comprising said first polypeptide conjugate with an anion exchange medium;
(b) eluting said first polypeptide conjugate from said anion exchange medium, forming a first eluate comprising said first polypeptide conjugate;
(c) contacting said first eluate with a hydrophobic interaction chromatography (HIC) medium; and
(d) eluting said first polypeptide conjugate from said hydrophobic interaction chromatography medium,thereby making said composition comprising said first polypeptide conjugate.
2 Assignments
0 Petitions
Accused Products
Abstract
The present invention provides processes for the manufacturing of polypeptide conjugates. In particular, the invention provides methods for the purification of polypeptide conjugates, which include at least one polymeric modifying groups, such as a poly(alkylene oxide) moiety. Exemplary poly(alkylene oxide) moieties include poly(ethylene glycol) (PEG) and poly(propylene glycol). In an exemplary process, hydrophobic interaction chromatography (HIC) is used to resolve different glycoforms of glycoPEGylated polypeptides.
-
Citations
65 Claims
-
1. A method of making a composition comprising a first polypeptide conjugate, said first polypeptide conjugate comprising a first number of poly(alkylene oxide) moieties covalently linked to said first polypeptide, wherein at least one of said poly(alkylene oxide) moieties is covalently linked to said first polypeptide via a glycosyl linking group, wherein said glycosyl linking group is either covalently linked to an amino acid residue of said first polypeptide or is covalently linked to a glycosyl moiety of said first polypeptide, said method comprising:
-
(a) contacting a mixture comprising said first polypeptide conjugate with an anion exchange medium; (b) eluting said first polypeptide conjugate from said anion exchange medium, forming a first eluate comprising said first polypeptide conjugate; (c) contacting said first eluate with a hydrophobic interaction chromatography (HIC) medium; and (d) eluting said first polypeptide conjugate from said hydrophobic interaction chromatography medium, thereby making said composition comprising said first polypeptide conjugate. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25)
-
-
26. A method of isolating a first polypeptide conjugate comprising a first number of poly(alkylene oxide) moieties covalently linked to said first polypeptide, wherein at least one of said poly(alkylene oxide) moieties is attached to said first polypeptide via a glycosyl linking group, wherein said glycosyl linking group is covalently linked to an amino acid residue of said first polypeptide or is covalently linked to a glycosyl moiety of said first polypeptide, from a second polypeptide conjugate comprising a second number of poly(alkylene oxide) moieties covalently linked to said second polypeptide, wherein said first number is selected from 1 to 20 and said second number is selected from 0-20, said first number and said second number being different, said method comprising:
-
(a) contacting a mixture comprising said first polypeptide conjugate and said second polypeptide conjugate with an anion exchange medium; (b) eluting said first polypeptide conjugate and said second polypeptide conjugate from said anion exchange medium, forming a first eluate comprising said first polypeptide conjugate and said second polypeptide conjugate; (c) contacting said first eluate with a hydrophobic interaction chromatography (HIC) medium; and (d) eluting said first polypeptide conjugate from said hydrophobic interaction chromatography medium, thereby isolating said first polypeptide conjugate from said second polypeptide conjugate. - View Dependent Claims (27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45)
-
-
46. A method of forming a composition comprising a first erythropoietin (EPO) conjugate, said first EPO conjugate comprising a first number of poly(alkylene oxide) moieties covalently linked to said EPO, wherein said EPO conjugate comprises at least one poly(alkylene oxide) moiety that is covalently linked to said EPO via a glycosyl linking group, wherein said glycosyl linking group is covalently linked to an amino acid residue of said EPO or is covalently linked to a glycosyl moiety of said EPO, said method comprising:
-
(a) contacting a mixture comprising said first EPO conjugate with an anion exchange medium; (b) eluting said first EPO conjugate from said anion exchange medium, forming a first eluate comprising said first EPO conjugate; (c) contacting said first eluate with a hydrophobic interaction chromatography (HIC) medium; and (d) eluting said first EPO conjugate from said hydrophobic interaction chromatography medium, thereby forming a composition comprising said first EPO conjugate. - View Dependent Claims (47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65)
-
Specification