Methods of culturing and expanding mesenchymal stem cells

• US 9,567,569 B2
• Filed: 07/19/2013
• Issued: 02/14/2017
• Est. Priority Date: 07/23/2012
• Status: Expired due to Fees

• Current Assignee: Gamida Cell, Ltd.
• Original Assignee: Gamida Cell, Ltd.
• Inventors: Peled, Tony, Steinhardt, Yair
• Primary Examiner(s): Lankford, Blaine
• Application Number:
  US13/946,272
• Publication Number:
  US 20140023623A1
• Time in Patent Office: 1,306 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: A61K 35/28 Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
• CPC Class Codes: C12N 2501/38 with nuclear receptors
• CPC Class Codes: C12N 2501/60 Transcription factors
• CPC Class Codes: C12N 5/0663 Bone marrow mesenchymal stem cells (BM-MSC)

Abstract

A method of culturing mesenchymal stem cells (MSCs) is provided. The method comprising culturing a population of the MSCs in a medium comprising an aryl hydrocarbon receptor antagonist, thereby culturing MSCs.

Claims

1. A method of culturing mesenchymal stem cells (MSCs) comprising culturing a population of the MSCs in a medium comprising an aryl hydrocarbon receptor antagonist, thereby culturing MSCs, wherein said aryl hydrocarbon receptor inhibitor comprises 4-(2-(2-(benzo[b]thiophen-3-yl)-9-isopropyl-9H-purin-6-ylamino) ethyl) phenol (StemReginin 1, SR1).

3. A method of generating cells useful for transplantation into a subject, the method comprising:

(a) culturing mesenchymal stem cells according to the method of claim 1 to generate a population of cultured mesenchymal stem cells;

(b) contacting said population of cultured mesenchymal stem cells with a differentiating agent, thereby generating cells useful for transplantation into a subject.

7. The method of claim 1, wherein the mesenchymal stem cells are derived from a tissue selected from the group consisting of bone marrow, adipose tissue, placenta and umbilical cord blood.

8. The method of claim 1, wherein the concentration of said SR1 in said medium is at a range of 100-1000 nM.

10. The method of claim 1, wherein said medium further comprises nicotinamide at a range of 1-20 mM.

11. The method of claim 10, wherein said medium further comprises FGF-4.

12. The method of claim 1, wherein a calcium concentration of said medium is greater than 1.8 mM.

13. The method of claim 1, wherein said culturing is effected on a plastic surface.

14. The method of claim 1, wherein said medium is devoid of platelet derived growth factor (PDGF).

18. The method of claim 3, wherein said differentiation agent comprises a growth factor.

2. A method of expanding a population of mesenchymal stem cells, the method comprising culturing a seeded population of mesenchymal stem cells for a period of time sufficient for cell expansion, wherein said culturing is in a medium comprising an aryl hydrocarbon receptor antagonist, thereby generating an expanded population of mesenchymal stem cells, and wherein said aryl hydrocarbon receptor inhibitor comprises 4-(2-(2-(benzo[b]thiophen-3-yl)-9-isopropyl-9H-purin-6-ylamino) ethyl) phenol (StemReginin 1, SR1).

4. A method of generating cells useful for transplantation, the method comprising:

(a) expanding mesenchymal stem cells according to the method of claim 2; and

(b) contacting the mesenchymal stem cells with a differentiating agent, thereby generating cells useful for transplantation.

9. The method of claim 2, wherein the concentration of said SR1 in said medium is at a range of 100-1000 nM.

15. The method of claim 2, wherein said expanding is effected under conditions that do not induce differentiation of said mesenchymal stem cells.

16. The method of claim 2, wherein said seeded population of mesenchymal stem cells were seeded in an absence of nicotinamide.

17. The method of claim 2, wherein said seeded population of mesenchymal stem cells were seeded in a presence of nicotinamide.

5. A cell culture, comprising mesenchymal stem cells and a medium which comprises an aryl hydrocarbon receptor antagonist, wherein said aryl hydrocarbon receptor inhibitor comprises 4-(2-(2-(benzo[b]thiophen-3-yl)-9-isopropyl-9H-purin-6-ylamino) ethyl) phenol (StemReginin 1, SR1).

6. The cell culture of claim 5, wherein said medium further comprises at least one of nicotinamide and a growth factor.

19. The cell culture of claim 5, wherein the concentration of said SR1 in said medium is at a range of 100-1000 nM.