PH20 polypeptide variants, formulations and uses thereof
Modified PH20 hyaluronidase polypeptides, including modified polypeptides that exhibit increased stability and/or increased activity, are provided. Also provided are compositions and formulations and uses thereof.
1. A modified PH20 polypeptide comprising an amino acid sequence, wherein:
(a) at least 95% of the residues of the amino acid sequence of the modified PH20 polypeptide are identical to the residues in an amino acid sequence selected from the group consisting of SEQ ID NO: 3 and 32-66 when the sequence of the modified PH20 polypeptide is aligned at positions corresponding to the sequence selected from the group consisting of SEQ ID NO: 3 and 32-66 to maximize identical residues, and wherein terminal gaps are treated as non-identical; and
(b) the amino acid sequence of the modified PH20 polypeptide comprises an amino acid modification at a position corresponding to position 313 with reference to amino acid positions set forth in SEQ ID NO: 3; and
(c) the modification at position 313 is a replacement selected from among A, H, K, L, P, R, and Y.
2. The modified PH20 polypeptide of claim 1 , wherein the amino acid modification is at a position corresponding to position 313 with reference to amino acid positions set forth in SEQ ID NO: 3 is K.
3. The modified PH20 polypeptide of claim 2 , wherein at least 96% of the residues of the amino acid sequence of the modified PH20 polypeptide are identical to the residues in an amino acid sequence set forth in SEQ ID NO:35.
4. The modified PH20 polypeptide of claim 2 , wherein at least 95% of the residues of the amino acid sequence of the modified PH20 polypeptide are identical to the residues in an amino acid sequence set forth in SEQ ID NO:32.
5. The modified PH20 polypeptide of claim 1 , wherein the modified PH20 polypeptide exhibits increased hyaluronidase activity compared to the hyaluronidase activity of the polypeptide set forth in SEQ ID NO: 3, measured under identical conditions.
6. The modified PH20 polypeptide of claim 1 , wherein the hyaluronidase activity of the modified PH20 polypeptide is at least 120% of the hyaluronidase activity of the PH20 polypeptide of SEQ ID NO: 3, measured under identical conditions.
7. The modified PH20 polypeptide of claim 1 that comprises one or more modifications of the polypeptide selected from among glycosylation, sialylation, albumination, farnesylation, carboxylation, hydroxylation, and phosphorylation.
8. The modified PH20 polypeptide of claim 7 that is glycosylated.
9. The modified PH20 polypeptide of claim 8 , wherein the polypeptide is a glycoprotein that comprises an N-acetylglucosamine moiety linked to each of at least three asparagine (N) residues.
10. A nucleic acid encoding the modified PH20 polypeptide of claim 1 .
11. A recombinant expression vector, comprising the nucleic acid of claim 10 .
12. A host cell, comprising the vector of claim 11 .
13. A pharmaceutical composition, comprising the modified PH20 polypeptide of claim 1 .
14. The pharmaceutical composition of claim 13 , further comprising a therapeutically active agent.
15. A pharmaceutical composition of claim 14 , wherein the therapeutically active agent is a polypeptide, a protein, a nucleic acid, a drug, a small molecule, or an organic molecule.
16. The pharmaceutical composition of claim 15 , wherein the therapeutically active agent is an antibody.
17. The pharmaceutical composition of claim 15 , wherein the therapeutically active agent is a small molecule drug.
18. A method for administering a therapeutically active agent, comprising providing a pharmaceutical composition of claim 14 , and parenterally administering the composition.
19. The method of claim 18 , wherein said parenterally administering is subcutaneously administering.
20. A method for treating cancer, comprising administering a pharmaceutical composition of claim 14 to a subject in need thereof.
21. The method of claim 20 , wherein said administering is to a subject receiving treatment with an anticancer drug.
22. A method for manufacture of
the modified PH20 polypeptide of claim 1 , the method comprising: transfecting a plasmid DNA containing a cDNA encoding the modified PH20 polypeptide into a host cell; culturing the cell under conditions for expression of the cDNA as the modified PH20 polypeptide; and harvesting the modified PH20 polypeptide from the culture.