Activatable cytokine polypeptides and methods of use thereof
The disclosure features fusion proteins that are conditionally active variants of a cytokine of interest. In one aspect, the full-length polypeptides of the invention have reduced or minimal cytokine-receptor activating activity even though they contain a functional cytokine polypeptide. Upon activation, e.g., by cleavage of a linker that joins a blocking moiety, e.g., a steric blocking polypeptide, in sequence to the active cytokine, the cytokine can bind its receptor and effect signaling. Typically, the fusion proteins further comprise an in vivo half-life extension element, which may be cleaved from the cytokine in the tumor microenvironment.
1. A fusion polypeptide having the formula of: [D]-[L1]-[A]-[L2′]-[H], wherein,
[A] is an interferon alpha (IFNa) polypeptide, a mutein, or an active fragment thereof,
[D] is a blocking moiety,
[H] is a half-life extension moiety,
[L1] is a protease-cleavable polypeptide linker comprising the amino acid sequence of SEQ ID NO: 609, 612, or 615, and
[L2′] is a protease-cleavable polypeptide linker comprising the amino acid sequence of SEQ ID NO: 609, 612, or 615,
wherein the blocking moiety and the half-life extension moiety each independently comprise human serum albumin (HSA) or an antibody or antibody fragment that binds the HSA.
2. The fusion polypeptide of claim 1 , wherein the IFa polypeptide comprises a murine interferon alpha 1 (mIFNa1), murine interferon alpha 11 (mIFNa11), human interferon alpha 2b (IFNA2b), murine interferon alpha 11 (mIFNa11), interferon alpha 8 (IFNA8), interferon alpha 14 (IFNA14), interferon alpha 16 (IFNA16), or a mutein thereof.
3. The fusion polypeptide of claim 2 , wherein the IFNa polypeptide comprises the amino acid sequence of SEQ ID NO: 193, 548, 549 or 550.
4. The fusion polypeptide of claim 1 , wherein each of the blocking moiety and the half-life extension moiety comprises HSA.
5. The fusion polypeptide of claim 1 , wherein each of the blocking moiety and the half-life extension moiety comprises the antibody or antibody fragment that binds HSA.
6. The fusion polypeptide of claim 1 , wherein one of the blocking moiety and the half-life extension moiety comprises HSA and the other one of the blocking moiety and the half-life extension moiety comprises the antibody or antibody fragment that binds HSA.
7. The fusion polypeptide of claim 1 , wherein each of [L1] and [L2′] comprises SEQ ID NO: 609.
8. The fusion polypeptide of claim 7 , wherein at least one of the blocking moiety and the half-life extension moiety comprises the antibody or antibody fragment that binds HSA, and the antibody or antibody fragment has the amino acid sequence of residues 1 to 116 of SEQ ID NO: 427.
9. The fusion polypeptide of claim 8 , wherein the IFNa polypeptide comprises the amino acid sequence of SEQ ID NO: 193, 548, 549 or 550.
10. The fusion polypeptide of claim 7 , comprising the amino acid sequence of SEQ ID NO: 424, 427, 539, 541, 545, 546 or 547.
11. The fusion polypeptide of claim 1 , wherein each of [L1] and [L2′] comprises SEQ ID NO: 612.
12. The fusion polypeptide of claim 11 , wherein at least one of the blocking moiety and the half-life extension moiety comprises the antibody or antibody fragment that binds HSA, and the antibody or antibody fragment has the amino acid sequence of residues 1 to 116 of SEQ ID NO: 427.
13. The fusion polypeptide of claim 11 , wherein the IFNa polypeptide comprises an amino acid sequence of SEQ ID NO: 193, 548, 549 or 550.
14. The fusion polypeptide of claim 11 , comprising the amino acid sequence selected from the group consisting of SEQ ID NO: 425, 428, 540, 542, and 551-562.
15. The fusion polypeptide of claim 1 , wherein each of [L1] and [L2′] comprises SEQ ID NO: 615.
16. The fusion polypeptide of claim 15 , wherein at least one of the blocking moiety and the half-life extension moiety comprises the antibody or antibody fragment that binds HSA, and the antibody or antibody fragment has the amino acid sequence of residues 1 to 116 of SEQ ID NO: 427.
17. The fusion polypeptide of claim 15 , wherein the IFNa polypeptide comprises the amino acid sequence of SEQ ID NO: 193, 548, 549 or 550.
18. The fusion polypeptide of claim 15 , comprising the amino acid sequence SEQ ID NO: 426 or 429.
19. A fusion polypeptide comprising the amino acid sequence of SEQ ID NO: 427.
20. A fusion polypeptide comprising the amino acid sequence of SEQ ID NO: 428.
21. A fusion polypeptide comprising the amino acid sequence of SEQ ID NO: 429.
22. A nucleic acid encoding the fusion polypeptide of claim 1 .
23. A nucleic acid encoding the fusion polypeptide of claim 19 .
24. A nucleic acid encoding the fusion polypeptide of claim 20 .
25. A nucleic acid encoding the fusion polypeptide of claim 21 .
26. A pharmaceutical composition comprising the fusion polypeptide of claim 1 .
27. A pharmaceutical composition comprising the fusion polypeptide of claim 19 .
28. A pharmaceutical composition comprising the fusion polypeptide of claim 20 .
29. A pharmaceutical composition comprising the fusion polypeptide of claim 21 .