COMPOSITION FOR CULTURE OF PLURIPOTENT STEM CELLS

• US 20190330597A1
• Filed: 01/11/2018
• Published: 10/31/2019
• Est. Priority Date: 01/11/2017
• Status: Active Application

• Current Assignee: The Francis Crick Institute Limited
• Original Assignee: The Francis Crick Institute Limited
• Inventors: Niakan, Kathy, Wamaitha, Sissy
• Application Number:
  US16/476,796
• Publication Number:
  US 20190330597A1
• Time in Patent Office: Days
• CPC Class Codes: C12N 2500/32 Amino acids
• CPC Class Codes: C12N 2500/99 Serum-free medium
• CPC Class Codes: C12N 2501/105 Insulin-like growth factors [IGF]
• CPC Class Codes: C12N 2501/113 Acidic fibroblast growth factor (aFGF, FGF-1)
• CPC Class Codes: C12N 2501/15 Transforming growth factor beta (TGF-β)
• CPC Class Codes: C12N 2501/16 Activin; Inhibin; Mullerian inhibiting substance
• CPC Class Codes: C12N 2501/727 Kinases (EC 2.7.)
• CPC Class Codes: C12N 2503/02 Drug screening
• CPC Class Codes: C12N 2506/02 from embryonic cells
• CPC Class Codes: C12N 2533/52 Fibronectin; Laminin
• CPC Class Codes: C12N 5/0604 Whole embryos; Culture medium therefor
• CPC Class Codes: C12N 5/0606 Pluripotent embryonic cells, e.g. embryonic stem cells [ES] embryonic germ cells C12N5/0611, induced pluripotent stem cells C12N5/0696
• CPC Class Codes: C12N 5/067 Hepatocytes
• CPC Class Codes: C12N 5/0696 Artificially induced pluripotent stem cells, e.g. iPS

Abstract

The present invention relates to a chemically defined or minimal cell medium for culturing pluripotent stem cells, including embryonic stem cells and induced pluripotent stem cells. The said medium may consist of the following components: a base medium, IGF1, a TGF-β family member and a glutamine supplement. The invention also relates to methods and uses involving the cell medium.

Claims

1. A cell medium for culturing a pluripotent stem cell, wherein said medium is a chemically defined or minimal medium which comprises insulin-like growth factor 1 (IGF1) and which is free or substantially free from fibroblast growth factor (FGF) and activators of any FGF receptor, and which does not comprise an ErbB3 ligand.

2. The cell medium of claim 1 for culturing an embryonic stem cell or induced pluripotent cell.

3. The cell medium of either of claim 1 or 2 for culturing a mammalian cell.

4. The cell medium of claim 3 for culturing a human cell.

5. The medium of any one of claims 1 to 4 wherein said IGF1 comprises the sequence of SEQ ID NO:

1 or SEQ ID NO;

2.

6. The medium of any one of claims 1 to 5 further comprising a TGF-β

family member.

7. The medium of claim 6 wherein said TGF-β

family member is Activin.

11. An in vitro method for culturing a pluripotent stem cell, said method comprising culturing said cell in the medium according to any one of claims 1 to 10.

12. Use of the medium of any one of claims 1 to 10 in the culture of a pluripotent stem cell.

13. The method of claim 11 or the use of claim 12 wherein said cell is an embryonic stem cell or an induced pluripotent cell.

14. The method or use of claim 13 wherein said cell is a mammalian cell.

15. The method or use of claim 14 wherein said cell is a human cell.

16. A cell obtainable or obtained by the method of any one of claims 11 to 15.

17. A medium for culturing an embryo, wherein said medium is as defined in any one of claims 1 to 10.

18. An in vitro method for culturing an embryo, comprising culturing said embryo in the medium of claim 17.

19. The method of claim 18 wherein said embryo is a human embryo.

20. Use of the medium of claim 17 in the culture of an embryo.

21. The method or use of any one of claims 11 to 15 or 17 to 20 wherein said culture is carried out in conjunction with a basement membrane.

22. The method or use of claim 21 wherein said basement membrane comprises laminin.

8. A cell medium for culturing a human pluripotent stem cell, wherein said medium is a chemically defined or minimal medium and wherein said medium consists of a base medium, IGF1, a TGF-β

family member and a glutamine supplement.

9. The medium of claim 8 wherein said TGF-β

family member is Activin.

10. The medium of claim 8 or 9 wherein said base medium is advanced DMEM/F12.

23. A method of screening for factors that are essential for the culture of human embryos, wherein said method comprises the following steps:

(i) Preparing a medium by adding a factor to be analysed;

(ii) Culturing a human embryo in said medium;

(iii) Comparing said embryo to an embryo that has been cultured in a medium without said factor or in the presence of an activator or inhibitor of said factor; and

(iv) Determining whether said factor is essential for culture of said embryo.