EphA2 T-cell epitopes and uses therefor
View Patent ↗EphA2 T-cell epitope are provided herein. The epitopes include peptides corresponding to specific fragments of human EphA2 protein containing one or more T-cell epitopes, and conservative derivatives thereof. The EphA2 T-cell epitopes are useful in an assay, such as an ELISPOT assay, that may be used to determine and/or quantify a patient's immune responsiveness to EphA2. The epitopes also are useful in methods of modulating a patient's immune reactivity to EphA2, which has substantial utility as a treatment for cancers that overexpress EphA2, such as renal cell carcinoma (RCC). The EphA2 epitopes also can be used to vaccinate a patient against EphA2, by in vivo or ex vivo methods.
1. A purified EphA2 T-cell epitope that binds to a major histocompatibility complex (“MHC”) Class II molecule, wherein the epitope is a peptide that consists of 9 to 35 contiguous amino acid residues of native human EphA2 (SEQ ID NO:2), and wherein the EphA2 T-cell epitope has a predicted binding score of greater than 39 for the MHC Class II molecule HLA-DRβ1*0401 as determined using the Southwood algorithm.
2. A purified polypeptide comprising two or more EphA2 T-cell epitopes of claim 1 , wherein the two or more EphA2 T-cell epitopes are not contiguous to each other in the amino acid sequence of SEQ ID NO:2.
3. A purified polypeptide comprising a first EphA2 T-cell epitope and a second EphA2 T-cell epitope, wherein the first EphA2 T-cell epitope is the purified EphA2 T-cell epitope of claim 1 , and wherein the first and second EphA2 T-cell epitopes are not contiguous to each other in the amino acid sequence of SEQ ID NO:2.
4. A purified chimeric polypeptide comprising the EphA2 T-cell epitope of claim 1 and a heterologous amino acid sequence.
5. The purified chimeric polypeptide of claim 4 , wherein the heterologous amino acid sequence permits affinity purification or facilitates delivery of the purified chimeric polypeptide.
6. The purified EphA2 T-cell epitope of claim 1 , wherein the EphA2 T-cell epitope consists of 9 to 20 amino acids.
7. The purified polypeptide of claim 2 or 4 , wherein the EphA2 T-cell epitope consists of 9 to 20 amino acids.
8. The purified polypeptide of claim 3 , wherein the first EphA2 T-cell epitope that binds to an MHC Class II molecule consists of 9 to 20 amino acids.
9. The purified polypeptide of claim 2 or 3 , wherein each EphA2 T-cell epitope is separated from the other by a peptide spacer or a protease cleavage site.
10. The purified chimeric polypeptide of claim 4 , wherein a protease cleavage site is included between the EphA2 T-cell epitope and the heterologous amino acid sequence.
11. The purified chimeric polypeptide of claim 5 , wherein the heterologous amino acid sequence is a poly(his) tag.
12. The purified chimeric polypeptide of claim 5 , wherein the heterologous amino acid sequence is a portion of lactadherin or other protein to facilitate presentation of the peptide to dendritic cells in membrane vesicles.
13. The purified polypeptide of claim 2 , wherein said two or more EphA2 T-cell epitopes are the same EphA2 T-cell epitope.
14. The purified EphA2 T-cell epitope of claim 1 , wherein the MHC Class II molecule to which the EphA2 T-cell epitope binds is selected from the list of MHC Class II molecules shown in FIG. 3 .
15. The purified polypeptide of claim 1 or 3 , wherein the MHC Class II molecule to which the EphA2 T-cell epitope binds is selected from the list of MHC Class II molecules shown in FIG. 3 .
16. The purified chimeric polypeptide of claim 4 , wherein the MHC Class II molecule to which the EphA2 T-cell epitope binds is selected from the list of MHC Class II molecules shown in FIG. 3 .
17. The purified EphA2 T-cell epitope of claim 1 , wherein the MHC Class II molecule to which the EphA2 T-cell epitope binds is HLA-DRβ1*0401.
18. The purified polypeptide of claim 2 or 3 , wherein the MHC Class II molecule to which the EphA2 T-cell epitope binds is HLA-DRβ1*0401.
19. The purified chimeric polypeptide of claim 4 , wherein the MHC Class II molecule to which the EphA2 T-cell epitope binds is HLA-DRβ1*0401.
20. The purified EphA2 T-cell epitope of claim 1 , wherein the epitope is capable of inducing a CD4 + T cell response as determined by IFN-γ or IL-5 ELISPOT assays, wherein the mean IFN-γ or IL-5 spots per 100,000 CD4 + T cells analyzed is at least 11, and wherein the CD4 + T cells are obtained from a donor with the same MHC Class II molecule as the MHC Class II molecule to which the EphA2 T-cell epitope binds.
21. The purified polypeptide of claim 2 or 3 , wherein the purified polypeptide is capable of inducing a CD4 + T cell response as determined by IFN-γ or IL-5 ELISPOT assays, wherein the mean IFN-γ or IL-5 spots per 100,000 CD4 + T cells analyzed is at least 11, and wherein the CD4 + T cells are obtained from a donor with the same MHC Class II molecule as the MHC Class II molecule to which the EphA2 T-cell epitope binds.
22. The purified chimeric polypeptide of claim 4 , wherein the purified chimeric peptide is capable of inducing a CD4 + T cell response as determined by IFN-γ or IL-5 ELISPOT assays, wherein the mean IFN-γ or IL-5 spots per 100,000 CD4 + T cells analyzed is at least 11, and wherein the CD4 + T cells are obtained from a donor with the same MHC Class II molecule as the MHC Class II molecule to which the EphA2 T-cell epitope binds.
23. The purified EphA2 T-cell epitope of claim 1 , wherein the purified EphA2 T-cell epitope has an amino acid sequence selected from the group consisting of: IMGQFSHHN (SEQ ID NO: 2, residues 666-674); YSVCNVMSG (SEQ ID NO: 2, residues 67-75); EAGIMGQFSHHNIIR (SEQ ID NO:2, residues 663-677); and PIYMYSVCNVMSG (SEQ ID NO:2, residues 63-75).
24. The purified polypeptide of claim 2 , wherein at least one of the EphA2 T-cell epitopes has an amino acid sequence selected from the group consisting of: IMGQFSHHN (SEQ ID NO: 2, residues 666-674); YSVCNVMSG (SEQ ID NO: 2, residues 67-75); EAGIMGQFSHHNIIR (SEQ ID NO:2, residues 663-677); and PIYMYSVCNVMSG (SEQ ID NO:2, residues 63-75).
25. The purified polypeptide of claim 3 , wherein the first EphA2 T-cell epitope that binds to an MHC Class II molecule has an amino acid sequence selected from the group consisting of: IMGQFSHHN (SEQ ID NO: 2, residues 666-674); YSVCNVMSG (SEQ ID NO: 2, residues 67-75); EAGIMGQFSHHNIIR (SEQ ID NO:2, residues 663-677); and PIYMYSVCNVMSG (SEQ ID NO:2, residues 63-75).
26. The purified chimeric polypeptide of claim 4 , wherein the EphA2 T-cell epitope has an amino acid sequence selected from the group consisting of: IMGQFSHHN (SEQ ID NO: 2, residues 666-674); YSVCNVMSG (SEQ ID NO: 2, residues 67-75); EAGIMGQFSHHNIIR (SEQ ID NO:2, residues 663-677); and PIYMYSVCNVMSG (SEQ ID NO:2, residues 63-75).
27. The purified EphA2 T-cell epitope of claim 1 , wherein the purified EphA2 T-cell epitope has the amino acid sequence EAGIMGQFSHHNIIR (SEQ ID NO:2, residues 663-677).
28. The purified polypeptide of claim 2 , wherein at least one of the purified EphA2 T-cell epitopes has the amino acid sequence EAGIMGQFSHHNIIR (SEQ ID NO:2, residues 663-677).
29. The purified polypeptide of claim 3 , wherein the first EphA2 T-cell epitope that binds to an MHC Class II molecule has the amino acid sequence EAGIMGQFSHHNIIR (SEQ ID NO:2, residues 663-677).
30. The purified chimeric polypeptide of claim 4 , wherein the EphA2 T-cell epitope has the amino acid sequence EAGIMGQFSHHNIIR (SEQ ID NO:2, residues 663-677).
31. A composition comprising the purified EphA2 T-cell epitope of claim 1 and a pharmaceutically acceptable excipient, carrier, diluent, or vehicle.
32. A composition comprising the purified polypeptide of claim 2 , 3 , or 4 and a pharmaceutically acceptable excipient, carrier, diluent, or vehicle.
33. A composition comprising the purified EphA2 T-cell epitope of claim 23 and a pharmaceutically acceptable excipient, carrier, diluent, or vehicle.
34. A purified EphA2 T-cell epitope that binds at an MHC Class II molecule, wherein the EphA2 T-cell epitope has an amino acid sequence selected from the group consisting of: WLVPIGQCL (SEQ ID NO:2, residues 253-261); LLWGCALAA (SEQ ID NO:2, residues 12-20); GLTRTSVTV (SEQ ID NO:2, residues 391-399); NLYYAESDL (SEQ ID NO:2, residues 120-128); KLNVEERSV (SEQ ID NO:2, residues 162-170); MQNIMNDMP (SEQ ID NO:2, residues 55-63); DLMQNIMNDMPIYMYS (SEQ ID NO:2, residues 53-68); IMGQFSHHN (SEQ ID NO:2, residues 666-674); IVMWEVMTY (SEQ ID NO:2, residues 805-813); IVTSVTCEQ (SEQ ID NO:2, residues 360-368); IRLPSTSGS (SEQ ID NO: 2, residues 893-901; VELRWTPPQ (SEQ ID NO:2, residues 344-352); LRWTPPQDS (SEQ ID NO:2, residues 346-354); VVLLLVLAG (SEQ ID NO: 2, residues 545-553); ILVNSNLVC (SEQ ID NO:2, residues 745-753); and MNYTFTVEA (SEQ ID NO:2, residues 406-414).
35. A purified polypeptide comprising two or more EphA2 T-cell epitopes of claim 34 , wherein the two or more EphA2 T-cell epitopes are not contiguous to each other in the amino acid sequence of SEQ ID NO:2.
36. A purified polypeptide comprising a first EphA T-cell epitope and a second EphA2 T-cell epitope, wherein the first EphA2 T-cell epitope is the purified EphA2 T-cell epitope of claim 34 , and wherein the first and second EphA2 T-cell epitopes are not contiguous to each other in the amino acid sequence of SEQ ID NO:2.
37. A purified chimeric polypeptide comprising the EphA2 T-cell epitope of claim 34 and a heterologous amino acid sequence.
38. The purified EphA2 T-cell epitope of claim 34 , wherein the MHC Class II molecule to which the EphA2 T-cell epitope binds is selected from the list of MHC Class II molecules shown in FIG. 3 .
39. The purified polypeptide of claim 35 or 36 , wherein the MHC Class II molecule to which the EphA2 T-cell epitope binds is selected from the list of MHC Class II molecules shown in FIG. 3 .
40. The purified chimeric polypeptide of claim 37 , wherein the MHC Class II molecule to which the EphA2 T-cell epitope binds is selected from the list of MHC Class II molecules shown in FIG. 3 .
41. A composition comprising the purified EphA2 T-cell epitope of claim 34 and a pharmaceutically acceptable excipient, carrier, diluent, or vehicle.
42. A composition comprising the purified polypeptide of claim 35 , 36 , or 37 and a pharmaceutically acceptable excipient, carrier, diluent, or vehicle.
43. The purified polypeptide of claim 3 or 36 , wherein the second EphA2 T-cell epitope binds to an MHC Class I molecule.
44. The purified EphA2 T-cell epitope of claim 34 , wherein one of the sequences identified using ProPred to predict peptides for the MHC Class II molecule HLA-DRβ1*0402 has the amino acid sequence: IMGQFSHHN (SEQ ID NO: 2, residues 666-674); IVMWEVMTY (SEQ ID NO: 2, residues 805-813); IVYSVTCEQ (SEQ ID NO: 2, residues 360-368); IRLPSTSGS (SEQ ID NO: 2, residues 893-901; VELRWTPPQ (SEQ ID NO: 2, residues 344-352); LRWTPPQDS (SEQ ID NO: 2, residues 346-354); VVLLLVLAG (SEQ ID NO: 2, residues 545-553); ILVNSNLVC (SEQ ID NO: 2, residues 745-753); or MNYTFTVEA (SEQ ID NO: 2, residues 406-414).
45. The purified polypeptide of claim 3 , wherein the second EphA2 T-cell epitope binds to an MHC Class II molecule, and wherein the second EphA2 T-cell epitope has an amino acid sequence selected from the group consisting of: WLVPIGQCL (SEQ ID NO: 2, residues 253-261); LLWGCALAA (SEQ ID NO: 2, residues 12-20); GLTRTSVTV (SEQ ID NO: 2, residues 391-399); NLYYAESDL (SEQ ID NO: 2, residues 120-128); KLNVEERSV (SEQ ID NO: 2, residues 162-170); MQNIMNDMP (SEQ ID NO: 2, residues 55-63); DLMQNIMNDMPIYMYS (SEQ ID NO:2, residues 53-68); IMGQFSHHN (SEQ ID NO: 2, residues 666-674); IVMWEVMTY (SEQ ID NO: 2, residues 805-813); IVYSVTCEQ (SEQ ID NO: 2, residues 360-368); IRLPSTSGS (SEQ ID NO: 2, residues 893-901; VELRWTPPQ (SEQ ID NO: 2, residues 344-352); LRWTPPQDS (SEQ ID NO: 2, residues 346-354); VVLLLVLAG (SEQ ID NO: 2, residues 545-553); ILVNSNLVC (SEQ ID NO: 2, residues 745-753); and MNYTFTVEA (SEQ ID NO: 2, residues 406-414).