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 one or more amino acid replacements in an unmodified PH20 polypeptide, wherein:
the unmodified PH20 polypeptide consists of the amino acid sequence selected from the group consisting of SEQ ID NOs: 3, 7 and 32-66;
the modified PH20 polypeptide exhibits increased hyaluronidase activity compared to the unmodified PH20 polypeptide not containing the amino acid replacement(s);
the amino acid replacement(s) confer(s) the increased hyaluronidase activity;
the amino acid replacement(s) is at a position corresponding to 313 with reference to amino acid positions of SEQ ID NO:3, which confers increased hyaluronidase activity;
corresponding amino acid positions are identified by alignment of the PH20 polypeptide with the polypeptide having the sequence of amino acids set forth in SEQ ID NO:3; and
the modified PH20 polypeptide has at least 91% sequence identity to the amino acid sequence selected from the group consisting of SEQ ID NOs: 3, 7 and 32-66.
2. The modified PH20 polypeptide of claim 1 , wherein the replacement is selected from among A at a position corresponding to position 313; G at a position corresponding to position 313; H at a position corresponding to position 313; K at a position corresponding to position 313; P at a position corresponding to position 313; R at a position corresponding to position 313; T at a position corresponding to position 313; and Y at a position corresponding to position 313.
3. The modified PH20 polypeptide of claim 2 , wherein the replacement at the position corresponding to 313 is K.
4. The modified PH20 polypeptide of claim 3 , wherein the unmodified PH20 polypeptide consists of the amino acid sequence of sequence of SEQ ID NO:35.
5. The modified PH20 polypeptide of claim 2 , wherein the unmodified PH20 polypeptide consists of the amino acid sequence selected from the group consisting of SEQ ID NOs: 3 and 32-66.
6. The modified PH20 polypeptide of claim 1 , wherein the unmodified PH20 polypeptide consists of the amino acid sequence selected from the group consisting of SEQ ID NOs: 3 and 32-66.
7. The modified PH20 polypeptide of claim 1 , wherein the modified PH20 polypeptide has at least 95% sequence identity to the amino acid sequence selected from the group consisting of SEQ ID NOs: 3, 7 and 32-66.
8. The modified PH20 polypeptide of claim 1 , wherein the modified PH20 polypeptide has at least 95% sequence identity to SEQ ID NO:35.
9. The modified PH20 polypeptide of claim 1 , wherein the modified PH20 polypeptide exhibits increased hyaluronidase activity that is at least 120% of the hyaluronidase activity compared to the unmodified PH20 polypeptide not containing the amino acid replacement or replacements.
10. The modified PH20 polypeptide of claim 1 that comprises one or more of a modification selected from among glycosylation, sialation, albumination, farnesylation, carboxylation, hydroxylation, and phosphorylation.
11. The modified PH20 polypeptide of claim 1 that comprises an N-acetylglucosamine moiety linked to each of at least three asparagine (N) residues.
12. The modified PH20 polypeptide of claim 1 that is conjugated to a polymer.
13. The modified PH20 polypeptide of claim 12 , wherein the polymer is dextran or polyethylene glycol (PEG).
14. The modified PH20 polypeptide of claim 1 , further comprising a heterologous signal sequence.
15. A chimeric polypeptide comprising the modified PH20 polypeptide of claim 1 .
16. A pharmaceutical composition, comprising theft modified PH20 polypeptide of claim 1 .
17. A modified PH20 polypeptide, comprising the amino acid sequence selected from the group consisting of SEQ ID NOs: 591-598, or a sequence of amino acids that exhibits at least 91% sequence identity to a sequence of amino acids set forth in any of SEQ ID NOs: 591-598 and that contains the amino acid replacement at residue 313 compared to corresponding positions in the amino acid sequence selected from the group consisting of SEQ ID NOs: 3 and 32-66.