Use of a Gsk-3 Inhibitor to Maintain Potency of Culture Cells

• US 20080194021A1
• Filed: 07/31/2006
• Published: 08/14/2008
• Est. Priority Date: 07/29/2005
• Status: Abandoned Application

• Current Assignee: ABT Holding Company (Athersys, Inc.)
• Original Assignee: ABT Holding Company (Athersys, Inc.)
• Inventors: Mays, Robert W.
• Application Number:
  US11/996,890
• Publication Number:
  US 20080194021A1
• Time in Patent Office: Days
• US Class Current: 435/375
• CPC Class Codes: C12N 2501/40 Regulators of development
• CPC Class Codes: C12N 2501/415 Wnt; Frizzeled
• CPC Class Codes: C12N 5/0607 Non-embryonic pluripotent stem cells, e.g. MASC induced pluripotent stem cells C12N5/0696

Abstract

The present invention is directed to the culture of non-embryonic cells, that can differentiate into cell types of more than one embryonic lineage, in culture under conditions that maintain differentiation capacity during expansion; more particularly, culturing non-embryonic cells in the presence of at least one GKS-3 inhibitor, such as BIO.

Claims

1. A culture method comprising culturing non-embryonic cells in the presence of at least one GSK-3 inhibitor, wherein said cells can differentiate into cell types of more than one embryonic lineage.

30. The culture method of claim 1, wherein said cells maintain or increase their capacity to differentiate to a greater extent than said cells cultured in the absence of a GSK-3 inhibitor.

31. The culture method of claim 1, wherein gene expression of Oct-3A, telomerase, or combination thereof is maintained or increased compared to said cells cultured in the absence of a GSK-3 inhibitor.

32. The culture method of claim 1, wherein the GSK-3 inhibitor is a compound of formula (I):

33. The method of claim 32, wherein one X is O and the other X is N-OH.

34. The method of claims 32 or 33, wherein one Y is Br.

35. The method of claim 34, wherein one Y is Br at the 6′

-position.

36. The method of claim 32, wherein one n is 0 and the other n is 1.

37. The method of claim 32, wherein each R is H.

38. The method claim 1, wherein the GSK-3 inhibitor comprises

39. The method of claim 1, wherein the GSK3 inhibitor comprises LiCl, hymenialdisine, flavopiridol, kenpaullone, alsterpaullone, azakenpaullone, Indirubin-3′

-oxime, 6-Bromoindirubin-3′

-oxime (BIO), 6-Bromoindirubin-3′

-acetoxime, Aloisine A, Aloisine B, TDZD8, Pyrazolopyridine 18, Pyrazolopyridine 9, Pyrazolopyridine 34, CHIR98014, CHIR99021, CHIR-637, CT20026, SU9516, ARA014418, Staurosporine, GF109203x (bisindolyl-maleimide I), Ro318220 (bisindolyl-maleimide IX), SB216763, SB415286, CGP60474, TWS119, or a thiazolo 5,4-f quinazolin-9-one.

40. The method of claim 1, wherein the non-embryonic cells are non-embryonic, non-germ, non-embryonic germ cells.

41. The method of claim 1 further comprising removing or inactivating the GSK-3 inhibitor from culture and culturing said cells to allow differentiation.

2-29. -29. (canceled)

42. A composition comprising non-embryonic cells in combination with at least one GSK-3 inhibitor, wherein said cells can differentiate into cell types of more than one embryonic lineage.

43. The composition of claim 42, wherein the composition is in cell culture medium.

44. The composition of claim 42, wherein the non-embryonic cells are non-embryonic, non-germ, non-embryonic germ cells.

45. A method to prepare a composition comprising admixing non-embryonic cells and at least one GSK-3 inhibitor, wherein said cells can differentiate into cell types of more than one embryonic lineage.

46. The method claim 45, wherein the composition is in cell culture medium or a pharmaceutically acceptable carrier.

47. The method of claim 45, wherein the non-embryonic cells are non-embryonic, non-germ, non-embryonic germ cells.

48. A method of drug discovery comprising screening the composition of claim 47 with a compound and assaying for an effect on the differentiation phenotype of the non-embryonic cells.