Modified uricase and uses thereof
A modified uricase is described herein, as well as a method of reducing a level of uric acid by contacting a medium with the modified uricase. The modified uricase comprises a uricase polypeptide crosslinked by at least one bifunctional linking moiety that comprises a poly(alkylene glycol) moiety. A molecular weight of the bifunctional linking moiety is from about 1.5 kDa to about 4 kDa, and/or the modified uricase comprises a plurality of polypeptides having the amino acid sequence SEQ ID NO: 2. Further described is a polypeptide having the amino acid sequence SEQ ID NO: 2. A process of preparing the modified uricase is also described, comprising contacting the polypeptide with a crosslinking agent that comprises a poly(alkylene glycol) moiety and at least two aldehyde groups, to obtain a conjugate; and contacting the conjugate with a reducing agent.
1 . A modified uricase comprising a plurality of uricase polypeptides having the amino acid sequence SEQ ID NO: 2, crosslinked by at least one bifunctional linking moiety that comprises a poly(alkylene glycol) moiety, wherein a molecular weight of said bifunctional linking moiety is in a range of from about 1.5 kDa to about 4 kDa, and wherein said at least one bifunctional linking moiety is covalently attached to a lysine residue of a uricase polypeptide having the amino acid sequence SEQ ID NO: 2.
2 . The modified uricase of claim 1 , wherein said molecular weight of said bifunctional linking moiety is in a range of from about 2 kDa to about 4 kDa.
3 . The modified uricase of claim 1 , wherein said at least one bifunctional linking moiety comprises an alkylene group covalently attached to a nitrogen atom of an amine group comprised by a side chain of said lysine residue of the uricase polypeptide.
4 . The modified uricase of claim 1 , wherein said uricase polypeptide is attached to an average of at least 8 of said bifunctional linking moiety and/or wherein at least 30% of lysine residue side chains in the uricase polypeptide having the amino acid sequence SEQ ID NO: 2 are covalently attached to said at least one bifunctional linking moiety.
5 . The modified uricase of claim 1 , being in a form of a crosslinked tetramer.
6 . The modified uricase of claim 1 , wherein said uricase polypeptide is a plant recombinant polypeptide.
7 . The modified uricase of claim 1 , being characterized by a plasma half-life in rats of at least 50 hours.
8 . A method of treating a disease or disorder in which uricase activity is beneficial and/or a disease or disorder associated with excessive uric acid levels, in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of the modified uricase of claim 1 , thereby treating the disease or disorder.
9 . The method of claim 8 , said administering the modified uricase is at an interval of at least one week.
10 . The method of claim 9 , wherein administering the modified uricase is at a dosage of no more than 8 mg per month.
11 . A process of preparing the modified uricase of claim 1 , the process comprising:
(a) contacting said plurality of uricase polypeptides having the amino acid sequence SEQ ID NO: 2 with a crosslinking agent that comprises said poly(alkylene glycol) moiety, said crosslinking agent comprising at least two aldehyde groups, to thereby conjugate said plurality of uricase polypeptides having the amino acid sequence SEQ ID NO: 2 and said crosslinking agent; and
(b) contacting the plurality of uricase polypeptides having the amino acid sequence SEQ ID NO: 2 conjugated to said crosslinking agent with a reducing agent.
12 . A method of reducing a level of uric acid in a medium, the method comprising contacting the medium with the modified uricase of claim 1 .