Methods for selectively stimulating proliferation of T cells

• US 7,175,843 B2
• Filed: 03/02/2006
• Issued: 02/13/2007
• Est. Priority Date: 06/03/1994
• Status: Expired due to Fees

• Current Assignee: Genetics Institute LLC (Pfizer Inc.), the united states of america as represented by the secretary of the navy, Board of Regents of the University of Michigan (University of Michigan)
• Original Assignee: Genetics Institute LLC (Pfizer Inc.), the united states of america as represented by the secretary of the navy, Board of Regents of the University of Michigan (University of Michigan)
• Inventors: Nabel, Gary J., June, Carl H., Gray, Gary S., Rennert, Paul D., Thompson, Craig B.
• Primary Examiner(s): Gambel; Phillip
• Application Number:
  US11/366,331
• Publication Number:
  US 20060140919A1
• Time in Patent Office: 348 Days
• Field of Search: None
• US Class Current: 424/93.71
• CPC Class Codes: A61K 2039/5158 Antigen-pulsed cells, e.g. T-cells
• CPC Class Codes: A61K 39/4611 T-cells, e.g. tumor infiltrating lymphocytes [TIL], lymphokine-activated killer cells [LAK] or regulatory T cells [Treg]
• CPC Class Codes: A61K 39/464838 Viral antigens
• CPC Class Codes: C07K 14/705 Receptors; Cell surface antigens; Cell surface determinants tumour specific antigens C07K14/4748
• CPC Class Codes: C07K 16/2806 against CD2
• CPC Class Codes: C07K 16/2809 against the T-cell receptor (TcR)-CD3 complex
• CPC Class Codes: C07K 16/2818 against CD28 or CD152
• CPC Class Codes: C07K 16/2896 against molecules with a "CD"-designation, not provided for elsewhere
• CPC Class Codes: C07K 2317/34 Identification of a linear epitope shorter than 20 amino acid residues or of a conformational epitope defined by amino acid residues
• CPC Class Codes: C12N 2501/51 B7 molecules, e.g. CD80, CD86, CD28 (ligand), CD152 (ligand)
• CPC Class Codes: C12N 2501/515 CD3, T-cell receptor complex
• CPC Class Codes: C12N 2501/599 with CD designations not provided for elsewhere
• CPC Class Codes: C12N 5/0636 T lymphocytes

Abstract

Methods for inducing a population of T cells to proliferate by activating the population of T cells and stimulating an accessory molecule on the surface of the T cells with a ligand which binds the accessory molecule are described. T cell proliferation occurs in the absence of exogenous growth factors or accessory cells. T cell activation is accomplished by stimulating the T cell receptor (TCR)/CD3 complex or the CD2 surface protein. To induce proliferation of an activated population T cells, an accessory molecule on the surface of the T cells, such as CD28, is stimulated with a ligand which binds the accessory molecule. The T cell population expanded by the method of the invention can be genetically transduced and used for immunotherapy or can be used in methods of diagnosis.

Claims

1. A method for inducing a population of T cells to proliferate to sufficient numbers for use in therapy, comprising:

(a) activating a population of T cells in vitro with an agent which stimulates a TCR/CD3 complex-associated signal in the T cells, wherein the agent is selected from the group consisting of an anti-CD3 antibody or a CD3-binding fragment thereof, an anti-CD2 antibody or a CD2-binding fragment thereof, and an antigen in a form suitable to trigger a primary activation signal in the T cell when complexed with the TCR/CD3 complex, wherein said agent is attached on a surface; and

(b) stimulating a CD28 accessory molecule on the surface of the T cells in vitro with a ligand that binds the CD28 accessory molecule on the surface of the T cells, wherein the ligand is selected from the group consisting of an anti-CD28 antibody or a CD28-binding fragment thereof, B7-1 or a CD28-binding fragment thereof, and B7-2 or a CD28-binding fragment thereof, wherein said ligand is attached on the same surface as said agent;

the activating and stimulating steps thereby inducing proliferation of the T cells to sufficient numbers for use in therapy.

2. The method of claim 1, wherein the agent which stimulates a TCR/CD3 complex-associated signal in the T cells is an anti-CD3 antibody or a CD3-binding fragment thereof.

3. The method of claim 2, wherein the anti-CD3 antibody or the CD3-binding fragment thereof is an anti-human CD3 monoclonal antibody or a CD3-binding fragment thereof.

4. The method of claim 3, wherein the anti-CD3 antibody is OKT3.

5. The method of claim 1, wherein the T cells are activated by contact with an antigen or portion thereof.

6. The method of claim 1, wherein the ligand is an anti-CD28 antibody or a CD28-binding fragment thereof.

7. The method of claim 6, wherein said anti-CD28 antibody is a whole antibody.

8. The method of claim 6, wherein the anti-CD28 antibody or the CD28-binding fragment thereof is an anti-human CD28 monoclonal antibody or a CD28-binding fragment thereof.

9. The method of claim 8, wherein the anti-CD28 antibody is EX5.3D10, produced by hybridoma cell line ATCC No. HB11373.

10. The method of claim 1, wherein the agent which stimulates a TCR/CD3 complex-associated signal in the T cells and the ligand that binds the CD28 accessory molecule on the surface of the T cells are attached on the same surface via a covalent modification.

11. The method of claim 1, wherein the agent which stimulates a TCR/CD3 complex-associated signal in the T cells and the ligand that binds the CD28 accessory molecule on the surface of the T cells are attached on the same surface via an avidin-biotin complex.

12. The method of claim 1, wherein the agent which stimulates a TCR/CD3 complex-associated signal in the T cells and the ligand that binds the CD28 accessory molecule on the surface of the T cells are attached on the same surface via a streptavidin-biotin complex.

13. The method of claim 1, wherein the agent which stimulates a TCR/CD3 complex-associated signal in the T cells and the ligand that binds the CD28 accessory molecule on the surface of the T cells are directly attached on the same surface.

14. The method of claim 1, wherein the surface is a bead.

15. The method of claim 14, wherein the bead is a magnetic immunobead.

16. The method of claim 1, wherein the surface is a tissue culture dish.

17. The method of claim 1, wherein the T cells are induced to proliferate for at least 3 days.

18. The method of claim 1, wherein the T cells are induced to proliferate for at least 7 days.

19. The method of claim 1, wherein the T cells are induced to proliferate to about 10-fold the original T cell population.

20. The method of claim 1, wherein the T cells are induced to proliferate to about 100-fold the original T cell population.

21. The method of claim 1, wherein the T cells are induced to proliferate to about 1000-fold the original T cell population.

22. The method of claim 1, wherein the population of T cells is increased in number from about 10- to about 1000-fold the original T cell population.

23. The method of claim 1, wherein the T cells are induced to proliferate to about 100,000-fold the original T cell population.

24. The method of claim 1, further comprising reactivating and re-stimulating the T cells with the agent and the ligand when the rate of T cell proliferation has decreased to induce further proliferation of the T cells to produce a population of T cells increased in number of from about 100- to about 100,000-fold the original T cell population.

25. The method of claim 24, wherein the rate of T cell proliferation is monitored by examining cell size.

26. The method of claim 1, wherein the population of T cells comprises CD4⁺ T cells.

27. The method of claim 1, wherein the population of T cells is obtained from an individual infected with HIV.

28. The method of claim 27, wherein the method further comprises rendering the T cells resistant to HIV infection and restoring the T cells to the individual.

29. The method of claim 28, wherein the T cells are rendered resistant to HIV infection by contacting the T cells with at least one anti-retroviral agent which inhibits HIV replication or viral production.

30. The method of claim 28, wherein the T cells are rendered resistant to HIV infection by genetically transducing the T cells to produce molecules which inhibit HIV infection or replication.

58. The method of claim 1, wherein T cell proliferation is monitored by examining cell size.

60. The method of claim 1, wherein T cell proliferation is monitored by determining the level of expression of a cell surface molecule.

64. The method of claim 8, wherein the anti-CD28 antibody is 9.3, produced by hybridoma cell line ATCC No. HB10271.

31. An ex vivo method for inducing a population of CD4⁺ T cells to proliferate to sufficient numbers for use in therapy, comprising:

(a) activating a population of CD4⁺ T cells with an agent which stimulates a TCR/CD3 complex-associated signal in the CD4⁺ T cells, wherein the agent is selected from the group consisting of an anti-CD3 antibody or a CD3-binding fragment thereof, an anti-CD2 antibody or a CD2-binding fragment thereof, and an antigen in a form suitable to trigger a primary activation signal in the T cell when complexed with the TCR/CD3 complex, wherein the agent is attached on a surface; and

(b) stimulating a CD28 accessory molecule on the surface of the T cells with a ligand that binds the CD28 accessory molecule on the surface of the T cells, wherein the ligand is selected from the group consisting of an anti-CD28 antibody or a CD28-binding fragment thereof, B7-1 or a CD28-binding fragment thereof, and B7-2 or a CD28-binding fragment thereof, wherein the ligand is attached on the same surface as said agent,

the activating and stimulating steps thereby inducing proliferation of the CD4⁺ T cells to sufficient numbers for use in therapy, and resulting in a population of CD4⁺ T cells that are polyclonal with respect to antigen reactivity.

32. The method of claim 31, wherein the agent which stimulates a TCR/CD3 complex-associated signal in the T cells is an anti-CD3 antibody or a CD3-binding fragment thereof.

33. The method of claim 32, wherein the anti-CD3 antibody or CD3-binding fragment thereof is an anti-human CD3 monoclonal antibody or a CD3-binding fragment thereof.

34. The method of claim 33, wherein the anti-CD3 antibody is OKT3.

35. The method of claim 31, wherein the T cells are activated by contact with an antigen or portion thereof.

36. The method of claim 31, wherein the ligand is an anti-CD28 antibody or a CD28-binding fragment thereof.

37. The method of claim 31, wherein the anti-CD28 antibody is an anti-human CD28 monoclonal antibody or a CD28-binding fragment thereof.

38. The method of claim 37, wherein the anti-CD28 antibody is EX5.3D10, produced by hybridoma cell line ATCC No. HB11373.

39. The method of claim 31, wherein the agent which stimulates a TCR/CD3 complex-associated signal in the T cells and the ligand that binds the CD28 accessory molecule on the surface of the CD4⁺ T cells are attached on the same surface via a covalent modification.

40. The method of claim 31, wherein the agent which stimulates a TCR/CD3 complex-associated signal in the CD4⁺ T cells and the ligand that binds the CD28 accessory molecule on the surface of the CD4⁺ T cells are attached on the same surface via an avidin-biotin complex.

41. The method of claim 31, wherein the agent which stimulates a TCR/CD3 complex-associated signal in the CD4⁺ T cells and the ligand that binds the CD28 accessory molecule on the surface of the CD4⁺ T cells are attached on the same surface via a streptavidin-biotin complex.

42. The method of claim 31, wherein the agent which stimulates a TCR/CD3 complex-associated signal in the CD4⁺ T cells and the ligand that binds the CD28 accessory molecule on the surface of the CD4⁺ T cells are directly attached on the same surface.

43. The method of claim 31, wherein the surface is a bead.

44. The method of claim 43, wherein the bead is a magnetic immunobead.

45. The method of claim 31, wherein the surface is a tissue culture dish.

46. The method of claim 31, wherein the CD4⁺ T cells are induced to proliferate for at least 3 days.

47. The method of claim 31, wherein the CD4⁺ T cells are induced to proliferate for at least 7 days.

48. The method of claim 31, wherein the CD4⁺ T cells are induced to proliferate to about 10-fold the original CD4⁺ T cell population.

49. The method of claim 31, wherein the CD4⁺ T cells are induced to proliferate to about 100-fold the original CD4⁺ T cell population.

50. The method of claim 31, wherein the CD4⁺ T cells are induced to proliferate to about 1000-fold the original CD4⁺ T cell population.

51. The method of claim 31, wherein the population of CD4⁺ T cells is increased in number from about 10- to about 1000-fold the original CD4⁺ T cell population.

52. The method of claim 31, wherein the CD4⁺ T cells are induced to proliferate to about 100,000-fold the original CD4⁺ T cell population.

53. The method of claim 31, further comprising reactivating and re-stimulating the CD4⁺ T cells with the agent and the ligand when the rate of CD4⁺ T cell proliferation has decreased to induce further proliferation of the CD4⁺ T cells to produce a population of CD4⁺ T cells increased in number of from about 100- to about 100,000-fold the original CD4⁺ T cell population.

54. The method of claim 31, wherein the population of CD4⁺ T cells is obtained from an individual infected with HIV.

55. The method of claim 31, wherein the method further comprises rendering the CD4⁺ T cells resistant to HIV infection and restoring the CD4⁺ T cells to the individual.

56. The method of claim 55, wherein the CD4⁺ T cells are rendered resistant to HIV infection by contacting the CD4⁺ T cells with at least one anti-retroviral agent which inhibits HIV replication or viral production.

57. The method of claim 55, wherein the CD4⁺ T cells are rendered resistant to HIV infection by genetically transducing the CD4⁺ T cells to produce molecules which inhibit HIV infection or replication.

59. The method of claim 31, wherein T cell proliferation is monitored by examining cell size.

61. The method of claim 31, wherein T cell proliferation is monitored by determining the level of expression of a cell surface molecule.

65. The method of claim 37, wherein the anti-CD28 antibody is 9.3, produced by hybridoma cell line ATCC No. HB10271.

62. A method for expanding a population of T cells to sufficient numbers for use in therapy, comprising:

a) activating the population of T cells by contacting the T cells in vitro with an anti-CD3 antibody or a CD3-binding fragment thereof, which is attached on a surface; and

b) stimulating a CD28 accessory molecule on the surface of the T cells in vitro with an anti-CD28 antibody or a CD28-binding fragment thereof, wherein said anti-CD28 antibody or CD28-binding fragment thereof is attached on the same surface as the anti-CD3 antibody or CD3-binding fragment thereof,

the activating and stimulating steps thereby expanding the population of the T cells to sufficient numbers for use in therapy.

63. A method for producing a population of T cells suitable for augmenting or supporting T cell growth in vivo, comprising:

a) activating the population of T cells by contacting the T cells in vitro with an anti-CD3 antibody or a CD3-binding fragment thereof, which is attached on a surface; and

b) stimulating a CD28 accessory molecule on the surface of the T cells in vitro with an anti-CD28 antibody or a CD28-binding fragment thereof, wherein said anti-CD28 antibody or CD28-binding fragment thereof is attached on the same surface as the anti-CD3 antibody or CD3-binding fragment thereof,

the activating and stimulating steps thereby producing a population of T cells suitable for augmenting or supporting T cell growth in vivo.