Biased IL2 muteins methods and compositions

1. A method of treating a human subject suffering from a neoplastic disease, the method comprising administering to said subject a polypeptide comprising a sequence of formula 1:

wherein:

AA18 is selected from the group consisting of L, R, G, M, F, E, W, K, Q, S, V, I, Y, H, D and T;

AA22 is selected from the group consisting of Q, E, G, A, L, M, W, K, S, V, I, Y, H, R, N, D, T and F; and

AA126 is selected from the group consisting of H, M, K, C, D, E, G, I, R and S, wherein AA18 is not L when AA22 is Q, and wherein:

AA1 is A or deleted;

AA2 is P or deleted;

AA3 is T, C, A, G, Q, E, N, D, R, K, P or deleted;

AA4 is S or deleted;

AA5 is S or deleted;

AA6 is S or deleted;

AA7 is T or deleted;

AA8 is K or deleted;

AA9 is K or deleted;

AA35 is K or E;

AA38 is R, W or G;

AA39 is M, L or V;

AA55 is H or Y;

AA69 is V or A;

AA74 is Q, P, N, H or S;

AA80 is L, F or V;

AA81 is R, I, D or T;

AA85 is L or V;

AA86 is I or V;

AA89 is I or V;

AA91 is V, R or K

AA92 is I or F;

AA97 is K or Q;

AA104 is M or A;

AA109 is D or C;

AA113 is T or N;

AA125 is C, A or S;

AA130 is S, T, G or R; and

wherein (a) said polypeptide does not comprise the group of amino acid substitutions 80F, 81D, 85V, 86V and 92F; (b) the polypeptide exhibits diminished binding to CD132 relative to wild-type hIl-2 of SEQ ID NO:1 and (c) (i) the ratio of pSTAT5 induction in CD25 pos T cells relative to pSTAT5 induction in CD25 neg T cells upon contacting the CD25 pos and CD25 neg T cells with the polypeptide, is greater than (ii) the ratio of pSTAT5 induction in CD25 pos T cells relative to pSTAT5 induction in CD25 neg T cells upon contacting with the CD25 pos and CD25 neg T cells with wild-type hIL2 of SEQ ID NO:1.

2. The method of claim 1 , wherein AA1 is deleted.

3. The method of claim 1 , wherein the polypeptide comprises a set of mutations selected from the following sets of mutations:

18R, 22E, and 126H;

18R, 22E, and 126K;

18R, 22E and 126M;

18R and 126H;

22E and 126H;

18A, 22E and 126H;

18M, 22E and 126H;

18F, 22E and 126H;

18W, 22E and 126H;

18K, 22E and 126H;

18Q, 22E and 126H;

18E, 22E and 126H;

18S, 22E and 126H;

18V, 22E and 126H;

18I, 22E and 126H;

18Y, 22E and 126H;

18H, 22E and 126H;

18N, 22E and 126H;

18D, 22E and 126H;

18T, 22E and 126H;

18R, 22G and 126H;

18R, 22A and 126H;

18R, 22L and 126H;

18R, 22M and 126H;

18R, 22F and 126H;

18R, 22W and 126H;

18R, 22K and 126H;

18R, 22S and 126H;

18R, 22V and 126H;

18R, 22I and 126H;

18R, 22Y and 126H;

18R, 22H and 126H;

18R, 22R and 126H;

18R, 22N and 126H;

18R, 22D and 126H; and

18R, 22T and 126H.

4. The method of claim 1 , wherein the polypeptide is PEGylated.

5. The method of claim 4 , wherein the polypeptide is PEGylated with a PEG having a molecular weight of 10,000 to 50,000 Daltons.

6. The method of claim 1 , wherein the administering comprises administering a nucleic acid encoding the polypeptide.

7. The method of claim 6 , wherein the nucleic acid is DNA.

8. The method of claim 6 , wherein the nucleic acid is a recombinant expression vector.

9. The method of claim 8 , wherein said vector is a viral vector.

10. The method of claim 8 , wherein said vector is a non-viral vector.

11. The method of claim 1 , wherein said method further comprises administering a supplementary agent to said subject.

12. The method of claim 11 , wherein said supplementary agent is selected from the group consisting of a chemotherapeutic agent, an antibody, an immune checkpoint modulator, tumor infiltrating lymphocytes (TILs), a CAR-T cell, and a physical method.

13. The method of claim 12 , wherein the supplementary agent is an immune checkpoint modulator.

14. The method of claim 13 , wherein the immune checkpoint modulator is an anti-PD-1 antibody or an anti-PD-L1 antibody.

15. The method of claim 1 , wherein the polypeptide comprises SEQ ID NO:8.

16. The method of claim 15 , wherein the polypeptide comprises an amino-terminal proline linked to:

17. The method of claim 14 , wherein the anti-PD-1 antibody is nivolumab or pembrolizumab.

18. A method of treating a human subject suffering from a neoplastic disease, the method comprising adminisering to said subject a polypeptide comprising a sequence of formula 1:

AA18 is selected from the group consisting of L, R, G, M, F, E, W, K, Q, S, V, I, Y, H, D and T;

AA22 is selected from the group consisting of Q E, G, A, L, M, W, K, S, V, I, Y, H, R, N, D, T and F; and

AA126 is selected from the group consisting of H, M, K, C, D, E, G, I, R and S,

wherein AA18 is not L when AA22 is Q,

and wherein:

AA1 is A or deleted;

AA2 is P or deleted;

AA3 is T, C, A, G, Q, E, N, D, R, K, P or deleted;

AA4 is S or deleted;

AA5 is S or deleted;

AA6 is S or deleted;

AA7 is T or deleted;

AA8 is K or deleted;

AA9 is K or deleted;

AA35 is K;

AA38 is R, W or G;

AA39 is M, L or V;

AA55 is H or Y;

AA69 is V or A;

AA74 is Q, N, H or S;

AA80 is L, F or V;

AA81 is R, I, D or T;

AA85 is L or V;

AA86 is I or V;

AA89 is I or V;

AA91 is V, R or K AA92 is I or F;

AA97 is K or Q;

AA104 is M or A;

AA109 is D or C;

AA113 is T or N;

AA125 is C, A or S;

AA130 is S, T or R; and

wherein (a) said polypeptide does not comprise the group of amino acid substitutions 80F, 81D, 85V, 86V and 92F; (b) the polypeptide exhibits diminished binding to CD132 relative to wild-type hIL-2 of SEQ ID NO:1 and (c) (i) the ratio of pSTAT5 induction in CD25 pos T cells relative to pSTAT5 induction in CD25 neg T cells upon contacting the CD25 pos and CD25 neg T cells with the polypeptide, is greater than (ii) the ratio of pSTAT5 induction in CD25 pos T cells relative to pSTAT5 induction in CD25 neg T cells upon contacting with the CD25 pos and CD25 neg T cells with wild-type IL2 of SEQ ID NO:1.