Interleukin-9 and interleukin-4 chimeric antagonist muteins and methods of using same
Chimeric polypeptide antagonists that include an interleukin-4 (IL-4) mutein linked to an interleukin-9 (IL-9) mutein are provided, as are polynucleotides encoding the IL-4 and IL-9 chimeric mutein antagonists. Also provided are methods of using the chimeric mutein antagonists and encoding polypeptides to reduce or inhibit the responsiveness of a cell to a cytokine such as IL-4, IL-9 and/or interleukin-13. Methods using the compositions to treat disorders such as pulmonary disorders (e.g., asthma) also are provided.
1. A chimeric polypeptide, comprising an interleukin-4 (IL-4) mutein receptor antagonist operatively linked to an interleukin-9 (IL-9) mutein receptor antagonist, wherein the IL-4 mutein receptor antagonist comprises the amino acid sequence of wild-type human IL-4 with one or more of the amino acids at positions 121, 124 or 125 substituted with another natural amino acid, and the IL-9 mutein receptor antagonist comprises the amino acid sequence of wild-type human IL-9 with one or more of the amino acids at positions 126 or 133 substituted with another natural amino acid, wherein the IL-9 mutein receptor antagonist is positioned N-terminal to the IL-4 mutein receptor antagonist, and wherein the chimeric polypeptide reduces or inhibits the association of an interleukin with an IL-4 receptor, an IL-9 receptor, an interleukin-13 (IL-13) receptor, or a combination thereof.
2. The chimeric polypeptide of claim 1 , wherein the IL-4 mutein receptor antagonist comprises a R121D substitution.
3. The chimeric polypeptide of claim 1 , wherein the IL-4 mutein receptor antagonist comprises a Y124D substitution.
4. The chimeric polypeptide of claim 1 , wherein the IL-4 mutein receptor antagonist comprises a R121D substitution and a Y124D substitution.
5. The chimeric polypeptide of claim 1 , wherein the IL-9 mutein receptor antagonist comprises a K126E substitution.
6. The chimeric polypeptide of claim 1 , wherein the IL-9 mutein receptor antagonist comprises a Q133K substitution.
7. The chimeric polypeptide of claim 1 , wherein the IL-9 mutein receptor antagonist comprises a K126E and a Q133K substitution.
8. The chimeric polypeptide of claim 1 , which has the amino acid sequence as set forth in SEQ ID NO: 4, SEQ ID NO: 5, or SEQ ID NO: 7.
9. The chimeric polypeptide of claim 1 , further comprising a peptide tag selected from the group consisting of a polyhistidine tag, a myc tag, a FLAG epitope, a V5 tag, and a combination thereof.
10. The chimeric polypeptide of claim 1 , further comprising a detectable label.
11. The chimeric polypeptide of claim 10 , wherein the detectable label comprises a radionuclide, a fluorescent moiety, a luminescent moiety, a chemiluminescent moiety, a paramagnetic moiety, an enzyme, or a substrate.
12. The chimeric polypeptide of claim 1 , which specifically binds an IL-4 receptor α polypeptide, an IL-9 receptor α polypeptide, or a combination thereof, thereby reducing or inhibiting assembly of a functional interleukin receptor.
13. The chimeric polypeptide of claim 1 , which specifically binds an IL-4 receptor, an JL-9 receptor, or a combination thereof, thereby reducing or inhibiting specific binding of an interleukin to the IL-4 receptor, the IL-9 receptor, or the combination thereof.
14. A composition, comprising the chimeric polypeptide of claim 1 and a carrier.
15. The composition of claim 14 , wherein the carrier comprises a physiologically acceptable carrier, a pharmaceutically acceptable carrier, or a combination thereof.
16. A polynucleotide encoding the chimeric polypeptide of claim 1 .
17. The polynucleotide of claim 16 , which encodes the chimeric polypeptide as set forth in SEQ ID NO: 4, SEQ ID NO: 5, or SEQ ID NO: 7.
18. The polynucleotide of claim 16 , which comprises the nucleotide sequence as set forth in SEQ ID NO: 3 or SEQ ID NO: 6.
19. The polynucleotide of claim 16 , which is operatively linked to a heterologous nucleic acid molecule.
20. The polynucleotide of claim 19 , wherein the heterologous nucleic acid molecule comprises a transcriptional regulatory element, a translational regulatory element, or a combination thereof.
21. The polynucleotide of claim 19 , wherein the heterologous nucleic acid molecule encodes a peptide.
22. The polynucleotide of claim 21 , wherein the peptide comprises a tag, a cellular localization domain, or a combination thereof.
23. A vector, which comprises the polynucleotide of claim 16 .
24. The vector of claim 23 , which is an expression vector.
25. The vector of claim 24 , wherein the expression vector is a prokaryotic expression vector, a eukaryotic expression vector, or a shuttle vector.
26. The vector of claim 23 , which comprises a viral vector.
27. An isolated host cell, which comprises the polynucleotide of claim 16 .
28. The host cell of claim 27 , which comprises the vector of claim 23 .
29. The host cell of claim 28 , wherein the vector is an expression vector.
30. A method of making a chimeric polypeptide, which comprises an IL-4 mutein receptor antagonist operatively linked to an IL-9 mutein receptor antagonist, comprising expressing the polynucleotide of claim 16 , thereby making the chimeric polypeptide.
31. The method of claim 30 , which comprises culturing the host cell of claim 27 under conditions whereby the polynucleotide is expressed.
32. The method of claim 30 , which comprises culturing the host cell of claim 29 , under conditions suitable for expression of the polynucleotide.
33. A chimeric polypeptide, which comprises an IL-4 mutein receptor antagonist operatively linked to an IL-9 mutein receptor antagonist, wherein the IL-9 mutein receptor antagonist is positioned N-terminal to the IL-4 mutein receptor antagonist, produced by the method of claim 30 .
34. The method of claim 30 , further comprising purifying the chimeric polypeptide.
35. A purified chimeric polypeptide, which comprises an IL-4 mutein receptor antagonist operatively linked to an IL-9 mutein receptor antagonist, wherein the IL-9 mutein receptor antagonist is positioned N-terminal to the IL-4 mutein receptor antagonist, produced by the method of claim 34 .