Activatable interleukin-2 polypeptides and methods of use thereof
The disclosure features fusion proteins that are conditionally active variants of IL-2. 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.
1. A cytokine comprising:
a) a first half-life extension domain, wherein the first half-life extension domain is polyethylene glycol, human serum albumin or a fragment thereof that binds neonatal Fc receptor (FcRn), an immunoglobulin Fc, or an antigen-binding portion of an antibody that binds to FcRn or to human serum albumin;
b) an IL-2 polypeptide; and
c) an IL-2 blocking moiety, wherein the IL-2 blocking moiety comprises a ligand-binding domain or fragment of a cognate receptor for the IL-2 polypeptide or an antibody or antigen-binding fragment of an antibody that binds the IL-2 polypeptide;
wherein the IL-2 blocking moiety is operably linked to the first half-life extension domain via a first linker, and wherein the IL-2 polypeptide is operably linked to the IL-2 blocking moiety via a second linker comprising a cleavable peptide.
2. The cytokine of claim 1 , wherein the first half-life extension domain is human serum albumin or a fragment thereof that binds neonatal Fc receptor (FcRn), an immunoglobulin Fc, or an antigen-binding portion of an antibody that binds to FcRn directly or to human serum albumin.
3. The cytokine of claim 1 , wherein the first half-life extension domain is an immunoglobulin Fc.
4. The cytokine of claim 1 , wherein the IL-2 polypeptide binds to a IL-2 receptor.
5. The cytokine of claim 1 , wherein the IL-2 blocking moiety inhibits activation of IL-2 receptor alpha/beta/gamma by the IL-2 polypeptide.
6. The cytokine of claim 1 , wherein the IL-2 blocking moiety inhibits activation of IL-2 receptor beta/gamma by the IL-2 polypeptide.
7. The cytokine of claim 1 , wherein the serum half-life of the IL-2 polypeptide that is produced by cleavage of the second linker is comparable to the half-life of naturally occurring IL-2.
8. The cytokine of claim 1 , wherein the IL-2 blocking moiety is an antigen-binding fragment of an antibody that binds the IL-2 polypeptide and is selected from the group consisting of a single domain antibody, Fab, and scFv.
9. The cytokine of claim 1 , wherein the IL-2 polypeptide comprises a naturally-occurring IL-2 polypeptide or a functional fragment thereof.
10. The cytokine of claim 1 , wherein the IL-2 polypeptide has attenuated IL-2-receptor activating activity at least 10 fold less than the IL-2 receptor activating activity of the polypeptide that comprises the IL-2 polypeptide that is produced by cleavage of the second linker comprising a cleavable linker.
11. The cytokine of claim 1 , wherein the IL-2 receptor activating activity is assessed by a CTLL-2 proliferation assay, a phosphor STAT ELISA, or HEK Blue receptor cell assay and by equal amounts, on a mole basis, of the IL-2 polypeptide and the cytokine.
12. A cytokine comprising:
a) a first half-life extension domain, wherein the first half-life extension domain is an immunoglobulin Fc;
b) an IL-2 polypeptide; and
c) an IL-2 blocking moiety, wherein the IL-2 blocking moiety comprises a fragment of a cognate receptor for the IL-2 polypeptide;
wherein the IL-2 blocking moiety is operably linked to the first half-life extension domain via a first linker, and wherein the IL-2 polypeptide is operably linked to the IL-2 blocking moiety via a second linker comprising a cleavable peptide.
13. The cytokine of claim 12 , wherein the cognate receptor for the IL-2 polypeptide is IL-2RBeta (CD122).
14. The cytokine of claim 12 , wherein the cognate receptor for the IL-2 polypeptide is IL-2Ralpha (CD25).
15. The cytokine of claim 12 , wherein the cognate receptor for the IL-2 polypeptide is IL-2Rgamma.
16. The cytokine of claim 12 , wherein the IL-2 polypeptide binds to a IL-2 receptor.
17. The cytokine of claim 12 , wherein the IL-2 blocking moiety inhibits activation of IL-2 receptor alpha/beta/gamma by the IL-2 polypeptide.
18. The cytokine of claim 12 , wherein the IL-2 blocking moiety inhibits activation of IL-2 receptor beta/gamma by the IL-2 polypeptide.
19. The cytokine of claim 12 , wherein the IL-2 polypeptide comprises a naturally-occurring IL-2 polypeptide or a functional fragment thereof.
20. The cytokine of claim 12 , wherein the IL-2 polypeptide has attenuated IL-2-receptor activating activity at least 10 fold less than the IL-2 receptor activating activity of the polypeptide that comprises the IL-2 polypeptide that is produced by cleavage of the second linker comprising a cleavable linker.
21. The cytokine of claim 12 , wherein the IL-2 receptor activating activity is assessed by a CTLL-2 proliferation assay, a phosphor STAT ELISA, or HEK Blue receptor cell assay and by equal amounts, on a mole basis, of the IL-2 polypeptide and the cytokine.