Interferon-alpha polypeptides and conjugates
View Patent ↗The present invention provides interferon-alpha polypeptides and conjugates, and nucleic acids encoding the polypeptides. The invention also includes compositions comprising these polypeptides, conjugates, and nucleic acids; cells containing or expressing the polypeptides, conjugates, and nucleic acids; methods of making the polypeptides, conjugates, and nucleic acids; and methods of using the polypeptides, conjugates, and nucleic acids.
1. An isolated or recombinant polypeptide comprising a sequence which
(a) differs in 0 to 8 amino acid positions from SEQ ID NO:38;
(b) comprises Ala or Leu at position 48; and
(c) optionally further comprises a methionine at the N-terminus;
which polypeptide exhibits antiviral activity.
2. The polypeptide of claim 1 , which sequence comprises Ala at position 48.
3. The polypeptide of claim 1 , wherein the antiviral activity of the polypeptide is equal to or greater than the antiviral activity of huIFN-alpha 2b or huIFN-alpha 2a.
4. The polypeptide of claim 3 , wherein the antiviral activity of the polypeptide is at least two-fold greater than the antiviral activity of huIFN-alpha 2b or huIFN-alpha 2a.
5. The polypeptide of claim 1 , wherein the polypeptide exhibits a ratio of antiviral activity/antiproliferative activity at least two-fold greater than the ratio of antiviral activity/antiproliferative activity exhibited by huIFN-alpha 2b or huIFN-alpha 2a.
6. A conjugate comprising
(a) the polypeptide of claim 1 , and
(b) a non-polypeptide moiety covalently attached to the polypeptide,
which conjugate exhibits antiviral activity.
7. The conjugate of claim 6 , comprising at least two non-polypeptide moieties covalently attached to the polypeptide.
8. The conjugate of claim 6 , comprising a non-polypeptide moiety covalently attached to a lysine residue or to the N-terminal amino group.
9. The conjugate of claim 6 , comprising a non-polypeptide moiety covalently attached to a lysine residue.
10. The conjugate of claim 6 , comprising a non-polypeptide moiety attached to the N-terminal amino group.
11. The conjugate of claim 6 , wherein the non-polypeptide moiety is a polymer.
12. The conjugate of claim 11 , wherein the polymer is a polyethylene glycol.
13. The conjugate of claim 12 , wherein the polyethylene glycol is a 40 kDa mPEG2 moiety covalently attached to a lysine residue.
14. A composition comprising the polypeptide of claim 1 , and a pharmaceutically acceptable excipient.
15. The polypeptide of claim 1 , comprising a sequence which differs from SEQ ID NO: 38 in 1 to 4 amino acid positions.
16. A composition comprising the conjugate of claim 6 , and a pharmaceutically acceptable excipient.
17. An isolated or recombinant polypeptide comprising the sequence SEQ ID NO:38 or the sequence of a variant thereof wherein the variant differs in 1 to 2 amino acid positions from SEQ ID NO:38, optionally further comprising a methionine at the N-terminus, which polypeptide exhibits antiviral activity.
18. A composition comprising the polypeptide of claim 17 , and a pharmaceutically acceptable excipient.
19. The polypeptide of claim 17 , comprising the sequence SEQ ID NO:38.
20. The polypeptide of claim 17 , wherein the antiviral activity of the polypeptide is equal to or greater than the antiviral activity of huIFN-alpha 2b or huIFN-alpha 2a.
21. The polypeptide of claim 20 , wherein the antiviral activity of the polypeptide is at least two-fold greater than the antiviral activity of huIFN-alpha 2b or huIFN-alpha 2a.
22. The polypeptide of claim 17 , wherein the polypeptide exhibits a ratio of antiviral activity/antiproliferative activity at least two-fold greater than the ratio of antiviral activity/antiproliferative activity exhibited by huIFN-alpha 2b or huIFN-alpha 2a.
23. A conjugate comprising
(a) the polypeptide of claim 17 , and
(b) a non-polypeptide moiety covalently attached to the polypeptide,
which conjugate exhibits antiviral activity.
24. The conjugate of claim 23 , comprisingat least two non-polypeptide moieties covalently attached to the polypeptide.
25. The conjugate of claim 23 , comprising a non-polypeptide moiety covalently attached to a lysine residue or to the N-terminal amino group.
26. The conjugate of claim 23 , comprising a non-polypeptide moiety covalently attached to a lysine residue.
27. The conjugate of claim 23 , comprising a non-polypeptide moiety attached to the N-terminal amino group.
28. The conjugate of claim 23 , wherein the non-polypeptide moiety is a polymer.
29. The conjugate of claim 28 , wherein the polymer is a polyethylene glycol.
30. The conjugate of claim 29 , wherein the polyethylene glycol is a 40 kDa mPEG2 moiety.
31. A composition comprising the conjugate of claim 23 , and a pharmaceutically acceptable excipient.
32. A method for preparing a conjugate, the method comprising
providing the polypeptide of claim 1 or 17 , and
attaching at least one non-polypeptide moiety to the polypeptide, wherein the resulting conjugate exibits antiviral activity.
33. The method of claim 32 , wherein the step of attaching the at least one non-polypeptide moiety to the polypeptide comprises:
reacting the polypeptide with 40 kDa mPEG2-NHS at a molar ratio of mPEG2-NHS:polypeptide of about 3:1 to 5:1 at pH 9 and 4° C. for 1 hour to overnight.
34. A conjugate comprising
(a) a polypeptide comprising the sequence SEQ ID NO:38 or the sequence of a variant thereof wherein the variant differs in 1 to 2 amino acid positions from SEQ ID NO:38, optionally further comprising a methionine at the N-terminus; and
(b) a PEG moiety covalently attached to a lysine residue of the polypeptide, which conjugate exhibits antiviral activity.
35. The conjugate of claim 34 , wherein the PEG moiety is a 40 kDa mPEG2 moiety.
36. The conjugate of claim 35 , wherein the 40 kDa mPEG2 moiety is covalently attached to a lysine residue selected from Lys31, Lys122, and Lys135.
37. A composition comprising the conjugate of claim 34 , and a pharmaceutically acceptable excipient.
38. The conjugate of claim 34 , wherein the polypeptide comprises the sequence SEQ ID NO:38.
39. A composition comprising
(a) a conjugate comprising a polypeptide comprising the sequence SEQ ID NO:38, optionally further comprising a methionine at the N-terminus, and a 40 kDa mPEG2 moiety covalently attached to Lys31;
(b) a conjugate comprising a polypeptide comprising the sequence SEQ ID NO:38, optionally further comprising a methionine at the N-terminus, and a 40 kDa mPEG2 moiety covalently attached to Lys122; and
(b) a conjugate comprising a polypeptide comprising the sequence SEQ ID NO:38, optionally further comprising a methionine at the N-terminus, and a 40 kDa mPEG2 moiety covalently attached to Lys135;
which composition exhibits antiviral activity.
40. The composition of claim 39 further comprising a pharmaceutically acceptable excipient.