Pegylated g-csf polypeptides and methods of producing same
AssigneeMaxygen Holdings Ltd.
InventorsCarsten GERMANSEN, Bobby SONI, Grethe RASMUSSEN
Life of this patent
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Abstract
A method for increasing the stability and uniformity of a PEGylated G-CSF polypeptide having at least one PEG moiety attached to the epsilon amino group of a lysine residue or the N-terminal amino group and at least one PEG moiety attached to a hydroxyl group, comprising subjecting the polypeptide to an elevated pH of above 8.0 for a period of time suitable to remove PEG moieties attached to a hydroxyl group, and reducing the pH to about 8.0 or lower; as well as PEGylated G-CSF polypeptides and compositions produced according to the method and methods for increasing neutrophil levels in a patient using the PEGylated G-CSF polypeptides and compositions.
Claims12
-39. (canceled)
A method for increasing the level of neutrophils in a patient suffering from or at risk of an insufficient neutrophil level, comprising administering to said patient an effective dose of a PEGylated G-CSF polypeptide produced by a method comprising:
- subjecting a G-CSF polypeptide to a PEGylation reaction with an amine-specific activated polyethylene glycol (PEG) to produce a PEGylated G-CSF polypeptide intermediate having at least one PEG moiety attached to the epsilon amino group of a lysine residue or the N-terminal amino group and at least one PEG moiety attached to a hydroxyl group, and
- subjecting the PEGylated G-CSF polypeptide intermediate to an elevated pH of above 8.0 for a period of time suitable to remove PEG moieties attached to a hydroxyl group, and
- reducing the pH to a reduced pH of about 8.0 or lower,
- thereby producing the PEGylated G-CSF polypeptide.
-47. (canceled)
A method for increasing the level of neutrophils in a patient suffering from or at risk of an insufficient neutrophil level, comprising administering to said patient an effective dose of a composition comprising a homogeneous mixture of positional PEG isomers of a PEGylated G-CSF polypeptide variant, wherein at least about 80% of the positional PEG isomers of the mixture consist of two positional PEG isomers each having PEG moieties consisting of two PEG moieties bound to epsilon amino groups of lysine and one PEG moiety bound to the N-terminal amino group.
(canceled)
A method for increasing the level of neutrophils in a patient suffering from or at risk of an insufficient neutrophil level, comprising administering to said patient an effective dose of a composition comprising a mixture of positional PEG isomers of a PEGylated G-CSF polypeptide, wherein the G-CSF polypeptide comprises the substitutions K16R, K34R, K40R, T105K and S159K relative to wild-type human G-CSF, and wherein at least about 80% of the positional PEG isomers of the polypeptide are lysine/N-terminal positional PEG isomers having three attached PEG moieties.
Dependencies are inferred from the claim text.