AMNIOTIC FLUID DERIVED CELLS

• US 20070122903A1
• Filed: 05/30/2006
• Published: 05/31/2007
• Est. Priority Date: 05/27/2005
• Status: Abandoned Application

• Current Assignee: Lifescan Incorporated (Duv Holding Corporation)
• Original Assignee: Lifescan Incorporated (Duv Holding Corporation)
• Inventors: Xu, Jean, Rezania, Alireza
• Application Number:
  US11/420,895
• Publication Number:
  US 20070122903A1
• Time in Patent Office: Days
• US Class Current: 435/325
• CPC Class Codes: A61P 1/18 for pancreatic disorders, e.g. pancreatic enzymes
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 5/50 for increasing or potentiating the activity of insulin
• CPC Class Codes: C12N 2500/02 Atmosphere, e.g. low oxygen conditions
• CPC Class Codes: C12N 2533/52 Fibronectin; Laminin
• CPC Class Codes: C12N 2533/54 Collagen; Gelatin
• CPC Class Codes: C12N 5/0605 Cells from extra-embryonic tissues, e.g. placenta, amnion, yolk sac, Wharton's jelly

Abstract

This invention relates to an expandable population of amniotic fluid-derived cells that can be differentiated into a β-cell lineage. This invention also provides methods for isolating and expanding such amniotic fluid-derived cells, as well as related methods and compositions for utilizing such cells in the therapeutic treatment of diabetes.

Claims

1. A substantially pure population of amniotic fluid-derived cells.

2. The population of amniotic fluid-derived cells according to claim 1, wherein the cells of said population are substantially negative in the expression of at least one protein marker selected from the group consisting of:

CD117, Oct4, and Tra2-54.

3. The population of amniotic fluid-derived cells according to claim 2, wherein the cells of said population are substantially positive for the expression of at least one protein marker selected from the group consisting of:

HNF-1 beta, GATA-6, and Sox-17.

4. The population of amniotic fluid-derived cells according to claim 2, wherein the cells of said population also express at least one gene selected from the group consisting of:

HNF-3 beta, Hes-1, GATA-4 and Musashi-1.

5. The population of amniotic fluid-derived cells according to claim 2, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, AXL, bFGF, EGF receptor, Fas/TNFRSF6, GRO, GRO-alpha, ICAM-1, IL-1 alpha, Il-3, Il-6, Il-8, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

6. The population of amniotic fluid-derived cells according to claim 2, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17.

7. The population of amniotic fluid-derived cells according to claim 6, wherein the cells of said population are substantially negative in the expression of cytokeratin protein.

8. The population of amniotic fluid-derived cells according to claim 6, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, bFGF, EGF receptor, GRO, GRO-alpha, ICAM-1, Il-3, Il-6, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

9. The population of amniotic fluid-derived cells according to claim 1, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17, and also express at least one gene selected from the group consisting of:

Hes-1 and Musashi-1.

10. The population of amniotic fluid-derived cells according to claim 1, wherein the cells of said population are substantially negative in the expression of CD 117, Oct4, Sox-17, and Tra2-54.

11. The population of amniotic fluid-derived cells according to claim 10, wherein the cells of said population are substantially negative in the expression of cytokeratin protein.

12. The population of amniotic fluid-derived cells according to claim 10, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, bFGF, EGF receptor, GRO, GRO-alpha, ICAM-1, Il-3, Il-6, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

13. The population of amniotic fluid-derived cells according to claim 10, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17, and also express at least one gene selected from the group consisting of:

Hes-1 and Musashi-1.

14. The population of pancreatic amniotic fluid-derived cells according to claim 1, capable of propagating in vitro.

15. The population of amniotic fluid-derived cells according to claim 1, capable of propagating in vitro under hypoxic conditions.

16. The population of amniotic fluid-derived cells according to claim 1, capable of differentiating into cells displaying the characteristics of the β

-cell lineage.

17. The population of amniotic fluid-derived cells according to claim 1, capable of differentiating into a gut hormone-producing cell.

18. A method of obtaining a population of cells from amniotic fluid, comprising:

a. Isolating amniotic fluid,

b. Isolating the cells from the amniotic fluid,

c. Placing the cells in culture medium,

d. Plating the cells in a culture vessel, and

e. Allowing the cells to grow in said medium for at least about five days, thereby obtaining a population of amniotic fluid-derived cells.

19. The method according to claim 18, wherein the cells of said population are substantially negative in the expression of at least one protein marker selected from the group consisting of:

CD117, Oct4, and Tra2-54.

20. The method according to claim 19, wherein the cells of said population are substantially positive for the expression of at least one protein marker selected from the group consisting of:

HNF-1 beta, GATA-6, and Sox-17.

21. The method according to claim 19, wherein the cells of said population also express at least one gene selected from the group consisting of:

HNF-3 beta, Hes-1, GATA-4 and Musashi-1.

22. The method according to claim 19, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, AXL, bFGF, EGF receptor, Fas/TNFRSF6, GRO, GRO-alpha, ICAM-1, IL-1 alpha, Il-3, Il-6, Il-8, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

23. The method according to claim 19, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17.

24. The method according to claim 23, wherein the cells of said population are substantially negative in the expression of cytokeratin protein.

25. The method according to claim 23, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, bFGF, EGF receptor, GRO, GRO-alpha, ICAM-1, Il-3, Il-6, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

26. The method according to claim 18, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17, and also express at least one gene selected from the group consisting of:

Hes-1 and Musashi-1.

27. The method according to claim 18, wherein the cells of said population are substantially negative in the expression of CD117, Oct4, Sox-17, and Tra2-54.

28. The method according to claim 27, wherein the cells of said population are substantially negative in the expression of cytokeratin protein.

29. The method according to claim 27, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, bFGF, EGF receptor, GRO, GRO-alpha, ICAM-1, Il-3, Il-6, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

30. The method according to claim 27, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17, and also express at least one gene selected from the group consisting of:

Hes-1 and Musashi-1.

31. The method according to claim 18, wherein the cells of said population are capable of propagating in vitro.

32. The method according to claim 18, wherein the cells of said population are capable of propagating in vitro under hypoxic conditions.

33. The method according to claim 18, wherein the cells of said population are capable of differentiating into cells displaying the characteristics of the β

-cell lineage.

34. The method according to claim 18, wherein the cells of said population are capable of differentiating into a gut hormone-producing cell.

35. A method of obtaining a population of cells from amniotic fluid, comprising:

a. Isolating amniotic fluid,

b. Isolating the cells from the amniotic fluid,

c. Selecting cells that express at least one of the markers selected from the group consisting of SSEA-4, SSEA-3, TRA1-60 and TRA1-81,

d. Placing the cells in culture medium,

e. Plating the cells in a culture vessel, and

f. Allowing the cells to grow in said medium for at least about five days, thereby obtaining a population of amniotic fluid-derived cells.

36. The method according to claim 35, wherein the cells of said population are substantially negative in the expression of at least one protein marker selected from the group consisting of:

CD117, Oct4, and Tra2-54.

37. The method according to claim 36, wherein the cells of said population are substantially positive for the expression of at least one protein marker selected from the group consisting of:

HNF-1 beta, GATA-6, and Sox-17.

38. The method according to claim 36, wherein the cells of said population also express at least one gene selected from the group consisting of:

HNF-3 beta, Hes-1, GATA-4 and Musashi-1.

39. The method according to claim 36, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, AXL, bFGF, EGF receptor, Fas/TNFRSF6, GRO, GRO-alpha, ICAM-1, IL-1 alpha, Il-3, Il-6, Il-8, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

40. The method according to claim 36, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17.

41. The method according to claim 40, wherein the cells of said population are substantially negative in the expression of cytokeratin protein.

42. The method according to claim 40, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, bFGF, EGF receptor, GRO, GRO-alpha, ICAM-1, Il-3, Il-6, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

43. The method according to claim 35, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17, and also express at least one gene selected from the group consisting of:

Hes-1 and Musashi-1.

44. The method according to claim 40, wherein the cells of said population are substantially negative in the expression of CD117, Oct4, Sox-17, and Tra2-54.

45. The method according to claim 44, wherein the cells of said population are substantially negative in the expression of cytokeratin protein.

46. The method according to claim 44, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, bFGF, EGF receptor, GRO, GRO-alpha, ICAM-1, Il-3, Il-6, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

47. The method according to claim 44, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17, and also express at least one gene selected from the group consisting of:

Hes-1 and Musashi-1.

48. The method according to claim 35, wherein the cells of said population are capable of propagating in vitro.

49. The method according to claim 35, wherein the cells of said population are capable of propagating in vitro under hypoxic conditions.

50. The method according to claim 35, wherein the cells of said population are capable of differentiating into cells displaying the characteristics of the β

-cell lineage.

51. The method according to claim 35, wherein the cells of said population are capable of differentiating into a gut hormone-producing cell.

52. A method of treating a patient with diabetes mellitus or at risk of developing diabetes, comprising:

a. Isolating a population of amniotic fluid-derived cells from a donor, and

b. Transferring the cells into the patient.

53. The method according to claim 52, wherein the cells of said population are substantially negative in the expression of at least one protein marker selected from the group consisting of:

CD117, Oct4, and Tra2-54.

54. The method according to claim 53, wherein the cells of said population are substantially positive for the expression of at least one protein marker selected from the group consisting of:

HNF-1 beta, GATA-6, and Sox-17.

55. The method according to claim 53, wherein the cells of said population also express at least one gene selected from the group consisting of:

HNF-3 beta, Hes-1, GATA-4 and Musashi-1.

56. The method according to claim 53, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, AXL, bFGF, EGF receptor, Fas/TNFRSF6, GRO, GRO-alpha, ICAM-1, IL-1 alpha, Il-3, Il-6, Il-8, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

57. The method according to claim 53, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17.

58. The method according to claim 57, wherein the cells of said population are substantially negative in the expression of cytokeratin protein.

59. The method according to claim 57, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, bFGF, EGF receptor, GRO, GRO-alpha, ICAM-1, Il-3, Il-6, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

60. The method according to claim 52, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17, and also express at least one gene selected from the group consisting of:

Hes-1 and Musashi-1.

61. The method according to claim 57, wherein the cells of said population are substantially negative in the expression of CD117, Oct4, Sox-17, and Tra2-54.

62. The method according to claim 61, wherein the cells of said population are substantially negative in the expression of cytokeratin protein.

63. The method according to claim 61, wherein the cells of said population are positive for the expression of at least one protein marker selected from the group consisting of:

Liver Activator Protein, PDGF receptor β

, bFGF, EGF receptor, GRO, GRO-alpha, ICAM-1, Il-3, Il-6, MIF, Osteoprotegerin, TIMP-2, and TRAIL R3.

64. The method according to claim 61, wherein the cells of said population are substantially negative in the expression of the protein marker Sox-17, and also express at least one gene selected from the group consisting of:

Hes-1 and Musashi-1.

65. The method according to claim 52, wherein the cells of said population are differentiated into pancreatic hormone producing cells prior to the step of transferring into the patient.

66. The method according to claim 52, wherein the cells of said population differentiate into pancreatic hormone producing cells after the step of transferring into the patient.

67. The method according to claim 52, wherein the cells of said population are capable of propagating in vitro.

68. The method according to claim 52, wherein the cells of said population are capable of propagating in vitro under hypoxic conditions.

69. The method according to claim 52, wherein the cells of said population are capable of differentiating into cells displaying the characteristics of the β

-cell lineage.

70. The method according to claim 52, wherein the cells of said population are capable of differentiating into a gut hormone-producing cell.