De novo design of potent and selective interleukin mimetics
De novo designed polypeptides that bind to IL-2 receptor β c heterodimer (IL-2Rβ c ), IL-4 receptor α c heterodimer (IL-4Rα c ), or IL-13 receptor α subunit (IL-13Rα) are disclosed, as are methods for using and designing the polypeptides.
1. A non-naturally occurring polypeptide comprising domains X1, X2, X3, and X4, wherein:
(a) X1 is a peptide comprising an amino acid sequence at least 80% identical to the amino acid sequence PKKKIQLHAEHALYDALMILNI (SEQ ID NO: 4);
(b) X2 is a helical-peptide of at least 8 amino acids in length that has no binding site for the IL-2 receptor α subunit (IL-2Rα);
(c) X3 is a peptide comprising an amino acid sequence at least 80% identical to the amino acid sequence LEDYAFNFELILEEIARLFESG (SEQ ID NO:5); and
(d) X4 is a peptide comprising an amino acid sequence at least 80% identical to the amino acid sequence EDEQEEMANAIITILQSWIFS (SEQ ID NO:6);
wherein the domains are arranged N-terminal to C-terminal in an arrangement X1-X3-X2-X4;
wherein amino acid linkers may be present between any of the domains; and
wherein the polypeptide binds to the IL-2 receptor β c heterodimer (IL-2Rβ c ).
2. The polypeptide of claim 1 , wherein X1 comprises the amino acid sequence EHALYDAL (SEQ ID NO:1), X3 comprises the amino acid sequence YAFNFELI (SEQ ID NO:2), and X4 comprises the amino acid sequence ITILQSWIF (SEQ ID NO:3).
3. The polypeptide of claim 1 , wherein X2 comprises an amino acid sequence at least 80% identical to the amino acid sequence KDEAEKAKRMKEWMKRIKT (SEQ ID NO:7).
4. The polypeptide of claim 1 , wherein the polypeptide comprises an amino acid sequence set forth in SEQ ID NO:181, SEQ ID NO:182 or SEQ ID NO:246.
5. The polypeptide of claim 1 wherein:
(a) X1 comprises an amino acid sequence at least 85% identical to the amino acid sequence PKKKIQLHAEHALYDALMILNI (SEQ ID NO: 4);
(b) X2 comprises an amino acid sequence at least 85% identical to the amino acid sequence KDEAEKAKRMKEWMKRIKT (SEQ ID NO:7);
(c) X3 comprises an amino acid sequence at least 85% identical to the amino acid sequence LEDYAFNFELILEEIARLFESG (SEQ ID NO:5); and
(d) X4 comprises an amino acid sequence at least 85% identical to the amino acid sequence EDEQEEMANAIITILQSWIFS (SEQ ID NO:6).
6. The polypeptide of claim 1 wherein:
(a) X1 comprises an amino acid sequence at least 90% identical to the amino acid sequence PKKKIQLHAEHALYDALMILNI (SEQ ID NO: 4);
(b) X2 comprises an amino acid sequence at least 90% identical to the amino acid sequence KDEAEKAKRMKEWMKRIKT (SEQ ID NO:7);
(c) X3 comprises an amino acid sequence at least 90% identical to the amino acid sequence LEDYAFNFELILEEIARLFESG (SEQ ID NO:5); and
(d) X4 comprises an amino acid sequence at least 90% identical to the amino acid sequence EDEQEEMANAIITILQSWIFS (SEQ ID NO:6).
7. The polypeptide of claim 3 wherein:
(a) X1 comprises the amino acid sequence PKKKIQLHAEHALYDALMILNI (SEQ ID NO: 4);
(c) X3 comprises the amino acid sequence LEDYAFNFELILEEIARLFESG (SEQ ID NO:5); and
(d) X4 comprises the amino acid sequence EDEQEEMANAIITILQSWIFS (SEQ ID NO:6).
8. The polypeptide of claim 1 wherein the polypeptide activates the IL-2 receptor β c heterodimer (IL-2Rβ c ) as measured by the ability of the polypeptide to elicit phospho-STAT5 signaling on IL-2 responsive NK cells in an in vitro assay.
9. The polypeptide of claim 3 wherein the polypeptide activates the IL-2 receptor β c heterodimer (IL-2Rβ c ) as measured by the ability of the polypeptide to elicit phospho-STAT5 signaling on IL-2 responsive NK cells in an in vitro assay.
10. The polypeptide of claim 5 wherein the polypeptide activates the IL-2 receptor β c heterodimer (IL-2Rβ c ) as measured by the ability of the polypeptide to elicit phospho-STAT5 signaling on IL-2 responsive NK cells in an in vitro assay.
11. A pharmaceutical composition comprising a polypeptide that binds to the IL-2 receptor β c heterodimer (IL-2Rβ c ) and has no binding site for the IL-2 receptor α subunit wherein the polypeptide comprises an amino acid sequence at least 80% identical to the amino acid sequence set forth in SEQ ID NO: 181 and/or SEQ ID NO:182; and a pharmaceutically acceptable carrier.
12. The pharmaceutical composition of claim 11 , wherein the polypeptide comprises an amino acid sequence at least 85% identical to the amino acid sequence set forth in SEQ ID NO: 181.
13. The pharmaceutical composition of claim 11 , wherein the polypeptide comprises an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO: 181.
14. The pharmaceutical composition of claim 11 , wherein the polypeptide comprises an amino acid sequence at least 95% identical to the amino acid sequence set forth in SEQ ID NO: 181.
15. The pharmaceutical composition of claim 11 , wherein the polypeptide comprises an amino acid sequence at least 97% identical to the amino acid sequence set forth in SEQ ID NO: 181.
16. The pharmaceutical composition of claim 11 , wherein the polypeptide comprises the amino acid sequence set forth in SEQ ID NO: 181.
17. The pharmaceutical composition of claim 11 , wherein the polypeptide comprises the amino acid sequence set forth in SEQ ID NO: 181 except for a cysteine mutation at position 62 (E62C), wherein numbering is in accordance with SEQ ID NO: 181.
18. The pharmaceutical composition of claim 17 , wherein a polyethylene glycol containing moiety is linked via a maleimide to the cysteine residue at position 62.
19. The pharmaceutical composition of claim 11 , wherein the polypeptide activates the IL-2 receptor β c heterodimer (IL-2Rβ c ) as measured by the ability of the polypeptide to elicit phospho-STAT5 signaling on IL-2 responsive NK cells in an in vitro assay.
20. The pharmaceutical composition of claim 13 , wherein the polypeptide activates the IL-2 receptor β c heterodimer (IL-2Rβ c ) as measured by the ability of the polypeptide to elicit phospho-STAT5 signaling on IL-2 responsive NK cells in an in vitro assay.