Use of multivalent chimeric peptide-loaded, MHC/Ig molecules to detect, activate or suppress antigen-specific T cell-dependent immune responses

• US 20020006903A1
• Filed: 02/22/2001
• Published: 01/17/2002
• Est. Priority Date: 03/28/1997
• Status: Active Grant

• Current Assignee: Johns Hopkins University
• Original Assignee: Johns Hopkins University
• Inventors: Greten, Tim, Slansky, Jill, OʼHerrin, Sean, Pardoll, Drew, Schneck, Jonathan
• Granted Patent: US 6,734,013 B2
• Time in Patent Office: Days
• US Class Current: 514/12
• CPC Class Codes: A61K 2035/124 the cells being hematopoietic, bone marrow derived or blood cells
• CPC Class Codes: A61K 2039/5154 Antigen presenting cells [APCs], e.g. dendritic cells or macrophages
• CPC Class Codes: A61K 2039/5158 Antigen-pulsed cells, e.g. T-cells
• CPC Class Codes: A61K 2039/605 MHC molecules or ligands thereof
• CPC Class Codes: A61K 2039/6056 Antibodies
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61K 39/00 Medicinal preparations containing antigens or antibodies materials for immunoassay G01N33/53
• CPC Class Codes: A61K 39/385 Haptens or antigens, bound to carriers
• CPC Class Codes: A61K 47/6425 the peptide or protein in the drug conjugate being a receptor, e.g. CD4, a cell surface antigen, i.e. not a peptide ligand targeting the antigen, or a cell surface determinant, i.e. a part of the surface of a cell
• CPC Class Codes: A61K 47/6811 the drug being a protein or peptide, e.g. transferrin or bleomycin
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/02 Immunomodulators
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07K 14/005 from viruses
• CPC Class Codes: C07K 14/7051 T-cell receptor (TcR)-CD3 complex
• CPC Class Codes: C07K 14/70539 MHC-molecules, e.g. HLA-molecules
• CPC Class Codes: C07K 2319/00 Fusion polypeptide
• CPC Class Codes: C07K 2319/30 Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto
• CPC Class Codes: C12N 2710/14143 viral genome or elements thereof as genetic vector
• CPC Class Codes: C12N 2740/13022 : New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• CPC Class Codes: C12N 2740/14022 : New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• CPC Class Codes: C12N 2760/16122 : New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• CPC Class Codes: C12N 2760/16134 : Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein

Abstract

To increase the effective affinity of soluble analogs of peptide/MHC molecules for their cognate ligands, divalent peptide/MHC complexes were constructed. Using a recombinant DNA strategy, DNA encoding the MHC class I was ligated to DNA coding for murine Ig heavy chain. MHC/Ig complexes were exploited to homogeneously load with peptides of interest. The results of flow cytometry demonstrated that the ^pepMHC/Ig complexes bound specifically with high affinity to cells bearing their cognate receptors.

^pepMHC/Ig complexes are also useful in modulating effector functions of antigen-specific T cells. These ^pepMHC/Ig complexes are useful for studying TCR/MHC interactions and lymphocyte tracking and have uses as specific regulators of immune responses.

Claims

1. A composition comprising a chimeric protein which comprises an MHC molecule and an immunoglobulin chain, wherein the chimeric protein associates to form molecular complexes comprising at least two chimeric proteins per complex, wherein an antigenic peptide is bound to each MHC molecule, wherein each MHC molecule within the molecular complex is bound to an identical antigenic peptide.

2. The composition of claim 1 wherein each MHC molecule in the composition is bound to an identical antigenic peptide.

3. The composition of claim 1 wherein the antigenic peptide is passively bound.

4. The composition of claim 1 wherein the antigenic peptide is actively bound.

5. The composition of claim 1 wherein the antigenic peptide is covalently bound to the MHC molecule.

6. The composition of claim 1 wherein the antigenic peptide is bound to the amino-terminus of β

2 microglobulin.

7. The composition of claim 1 wherein the antigenic peptide is bound to the amino-terminus of an MHC class II β

chain.

8. The composition of claim 7 wherein the antigenic peptide is bound to the amino-terminus of the MHC class II β

chain by means of a peptide tether.

9. The composition of claim 1 wherein the molecular complex comprises two of said chimeric proteins.

10. The composition of claim 1 wherein the molecular complex is conjugated to a toxin.

11. The composition of claim 1 wherein the MHC molecule is an MHC class I molecule.

12. The composition of claim 11 wherein the MHC class 1 molecule is an HLA class I molecule.

13. The composition of claim 12 wherein the MHC molecule is an HLA-A2 molecule.

14. The composition of claim 1 wherein the antigenic peptide is selected from the group consisting of an HTLV-Tax11-19 peptide, a p17 Gag77-85 peptide, and an influenza virus A M158-66 peptide.

15. A vector encoding a chimeric protein comprising an immunoglobulin heavy chain and an MHC molecule, wherein the immunoglobulin is not IgG-1, wherein the immunoglobulin heavy chain is C-terminal to the MHC molecule.

16. The vector of claim 15 wherein the MHC molecule is an MHC class I molecule.

17. The vector of claim 16 wherein the MHC class I molecule is an HLA class I molecule.

18. The vector of claim 17 wherein the HLA class I molecule is an HLA-A2 molecule.

19. A composition comprising a chimeric protein which comprises an MHC molecule and an Ig chain, wherein the chimeric protein associates to form molecular complexes comprising at least two chimeric proteins per complex, wherein the immunoglobulin chain is not an IgG heavy chain.

20. The composition of claim 19 wherein the chimeric protein is conjugated to a toxin.

21. A composition comprising a cell in which a chimeric protein is bound to the surface of the cell, wherein the chimeric protein comprises an MHC molecule and an immunoglobulin chain;

wherein the chimeric protein associates to form molecular complexes comprising at least two chimeric proteins per complex, wherein an antigenic peptide is bound to each MHC molecule, wherein each MHC molecule within the molecular complex is bound to an identical antigenic peptide.

22. The composition of claim 21 wherein the cell is a dendritic cell.

23. The composition of claim 21 wherein a population of said chimeric proteins is bound to the cell, wherein each MHC molecule in the population of said chimeric proteins is bound to an identical antigenic peptide.

24. A method for treating a patient suffering from an allergy, comprising:

administering to the patient at a dose sufficient to suppress a T cell response associated with an allergy of the patient a chimeric protein which comprises an MHC molecule and an immunoglobulin chain;

wherein the chimeric protein associates to form molecular complexes comprising at least two chimeric proteins per complex, wherein an antigenic peptide is bound to each MHC molecule, wherein each MHC molecule within the molecular complex is bound to an identical antigenic peptide, and wherein the antigenic peptide is an antigen to which the patient has an allergic response.

25. A method for treating a patient who has received or will receive an organ transplant, comprising:

administering to the patient at a dose sufficient to suppress an immune response to the transplanted organ a chimeric protein which comprises an MHC molecule and an immunoglobulin chain;

wherein the chimeric protein associates to form molecular complexes comprising at least two chimeric proteins per complex, wherein an antigenic peptide is bound to each MHC molecule, wherein each MHC molecule within the molecular complex is bound to an identical antigenic peptide, and wherein the antigenic peptide is an alloantigen.

26. A method for treating a patient suffering from an autoimmune disease, comprising:

administering to the patient at a dose sufficient to suppress an immune response associated with the autoimmune disease a chimeric protein which comprises an MHC molecule and an immunoglobulin chain;

wherein the chimeric protein associates to form molecular complexes comprising at least two chimeric proteins per complex, wherein an antigenic peptide is bound to each MHC molecule, wherein each MHC molecule within the molecular complex is bound to an identical antigenic peptide, and wherein the antigenic peptide is one to which the patient expresses an autoimmune response.

27. The method of claim 26 wherein the autoimmune disease is HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP).

28. The method of claim 26 wherein the MHC molecule is an MHC class I molecule.

29. The method of claim 28 wherein the MHC class I molecule is an HLA class I molecule.

30. The method of claim 29 wherein the HLA class I molecule is an HLA-A2 molecule.

31. The method of claim 27 wherein the antigenic peptide is HTLV-1 Tax11-19.

32. A method for treating a patient having a tumor, comprising:

administering to the patient at a dose sufficient to induce or enhance an immune response to the tumor a chimeric protein which comprises an MHC molecule and an immunoglobulin chain;

wherein the chimeric protein associates to form molecular complexes comprising at least two chimeric proteins per complex, wherein an antigenic peptide is bound to each MHC molecule, wherein each MHC molecule within the molecular complex is bound to an identical antigenic peptide, and wherein the antigenic peptide is a tumor-associated peptide.

33. A method for treating a patient having an infection caused by an infectious agent, comprising:

administering to the patient at a dose sufficient to induce or enhance an immune response to the infection a chimeric protein which comprises an MHC molecule and an immunoglobulin chain;

wherein the chimeric protein associates to form molecular complexes comprising at least two chimeric proteins per complex, wherein an antigenic peptide is bound to each MHC molecule, wherein each MHC molecule within the molecular complex is bound to an identical antigenic peptide, and wherein the antigenic peptide is a peptide of an infectious agent.

34. The method of claim 33 wherein the infection is an HIV infection.

35. The method of claim 34 wherein the antigenic peptide is p17 Gag77-85.

36. The method of claim 33 wherein the MHC molecule is an MHC class I molecule.

37. The method of claim 36 wherein the MHC class I molecule is an HLA class I molecule.

38. The method of claim 37 wherein the HLA class I molecule is an HLA-2 molecule.

39. The method of claim 33 wherein the infection is an influenza infection.

40. The method of claim 39 wherein the antigenic peptide is an influenza virus A M158-66 peptide.

41. A method of labeling antigen-specific T cells, comprising:

contacting a sample which comprises antigen-specific T cells with a chimeric protein;

wherein the chimeric protein comprises an MHC molecule and an immunoglobulin chain, wherein the chimeric protein associates to form molecular complexes comprising at least two chimeric proteins per complex, and wherein each MHC molecule within the molecular complex is bound to an identical antigenic peptide, whereby the antigenic peptide specifically binds to the antigen-specific T cells, thereby labeling the cells with the chimeric protein.

42. The method of claim 41 further comprising the step of:

separating the antigen-specific T cells which are bound to the antigenic peptides from cells which are not bound.

43. The method of claim 42 wherein the step of separating is performed using flow cytometry.

44. The method of claim 41 further comprising the step of:

counting the number of antigen-specific T cells which are bound to the antigenic peptides.

45. The method of claim 41 wherein said step of contacting is performed in vitro.

46. The method of claim 41 wherein said step of contacting is performed in vivo.

47. The method of claim 41 wherein the MHC molecule is an MHC class I molecule.

48. The method of claim 47 wherein the MHC molecule is an HLA class I molecule.

49. The method of claim 48 wherein the HLA class I molecule is an HLA-A2 molecule.

50. The method of claim 41 wherein the antigenic peptide is selected from the group consisting of an HTLV-1 Tax11-19 peptide, a p17 Gag77-85 peptide, and an influenza virus A M158-66 peptide.

51. The method of claim 41 further comprising the step of:

detecting a marker for activation of the antigen-specific T cells.

52. The method of claim 51 wherein the marker for activation of antigen-specific T cells is a secreted lymphokine.