Mono- and Di-Peg IL-10 Production; and Uses
AssigneeMerck Sharp & Dohme Corp.
InventorsSteven J. Blaisdell, Collette M. Cutler, Brittany C. Paporello, Alexandre Ambrogelly
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Abstract
Provided are methods of producing mono- and di-pegylated IL-10.
Claims20
A method of producing a mixture of mono- and di-pegylated IL-10, wherein at least one PEG molecule is covalently attached to at least one amino acid residue of at least one subunit of IL-10, comprising:
- a) reacting 1 mg/ml to 12 mg/ml of IL-10 protein with an activated PEG-linker such that the IL-10 to PEG-linker ratio is 1:1 to 1:7.7, in the presence of 0.75 mM to 35 mM of a reducing agent, at a pH of about 5.0 to 7.4 and a temperature of 5° C. to 30° C., for 12-15 hours;
- and
- b) purifying the mixture of mono- and di-pegylated IL-10.
The method of claim 9, wherein the mixture of mono- and di-PEG-IL-10 is purified by size exclusion chromatography.
A method of producing a mixture of mono- and di-pegylated IL-10, wherein at least one PEG molecule is covalently attached to at least one amino acid residue of at least one subunit of IL-10, comprising
- a) reacting 7.5 mg/ml of IL-10 with an activated PEG-linker such that the IL-10 to PEG-linker ratio is 1:3.5, in the presence of 25 mM of a reducing agent, at a pH of 6.3 and a temperature of 15° C., for 15 hours;
- and
- b) purifying the mixture of mono- and di-pegylated IL-10.
Dependencies are inferred from the claim text.