Methods for the purification of polypeptide conjugates

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, said method comprising:

• (a) contacting a mixture comprising said first polypeptide conjugate and a hydrophobic interaction chromatography (HIC) medium;
• and
• (b) eluting said first polypeptide conjugate from said hydrophobic interaction chromatography medium,
• thereby making said composition comprising said first polypeptide conjugate, wherein said first polypeptide is a member selected from bone morphogenetic protein 2 (BMP-2), bone morphogenetic protein 7 (BMP-7), bone morphogenetic protein 15 (BMP-15), neurotrophin-3 (NT-3), granulocyte colony stimulating factor (G-CSF), granulocyte-macrophage colony stimulating factor (GM-CSF), erythropoietin (EPO), interferon alpha, interferon beta, interferon gamma, α1-antitrypsin (α-1 protease inhibitor), glucocerebrosidase, tissue-type plasminogen activator (TPA), interleukin-2 (IL-2), leptin, hirudin, urokinase, human DNase, insulin, hepatitis B surface protein (HbsAg), chimeric diphtheria toxin-IL-2, human growth hormone (hGH), human chorionic gonadotropin (hCG), thyroid peroxidase (TPO), alpha-galactosidase, alpha-L-iduronidase, beta-glucosidase, alpha-galactosidase A, acid α-glucosidase (acid maltase), anti-thrombin III (AT III), follicle stimulating hormone (FSH), glucagon-like peptide-1 (GLP-1), glucagon-like peptide-2 (GLP-2), fibroblast growth factor 7 (FGF-7), fibroblast growth factor 21 (FGF-21), fibroblast growth factor 23 (FGF-23), prokinetisin, extendin-4, CD4, tumor necrosis factor receptor (TNF-R), α-CD20, P-selectin glycoprotein ligand-1 (PSGL-1), complement, transferrin, glycosylation-dependent cell adhesion molecule (GlyCAM), neural-cell adhesion molecule (N-CAM), TNF receptor-IgG Fc region fusion protein, anti-HER2 monoclonal antibody, monoclonal antibody to respiratory syncytial virus, monoclonal antibody to protein F of respiratory syncytial virus, monoclonal antibody to TNF-α, monoclonal antibody to glycoprotein IIb/IIIa, monoclonal antibody to CD20, monoclonal antibody to VEGF-A, monoclonal antibody to PSGL-1, monoclonal antibody to CD4, monoclonal antibody to a CD3, monoclonal antibody to EGF, monoclonal antibody to carcinoembryonic antigen (CEA) and monoclonal antibody to IL-2 receptor.

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, 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.