METHOD FOR PREPARING PLURIPOTENT STEM CELLS

• US 20160215258A1
• Filed: 09/03/2014
• Published: 07/28/2016
• Est. Priority Date: 09/04/2013
• Status: Active Grant

• Current Assignee: Otsuka Pharmaceutical Factory Incorporated (Otsuka Holdings Company Limited)
• Original Assignee: Otsuka Pharmaceutical Factory Incorporated (Otsuka Holdings Company Limited)
• Inventors: OKAIRI, Risa, NISHIMURA, Masuhiro, WADA, Tamaki
• Granted Patent: US 9,765,296 B2
• Time in Patent Office: Days
• 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: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C12N 2506/1353 from bone marrow mesenchymal stem cells (BM-MSC)
• CPC Class Codes: C12N 2506/1384 from adipose-derived stem cells [ADSC], from adipose stromal stem cells
• CPC Class Codes: C12N 5/0607 Non-embryonic pluripotent stem cells, e.g. MASC induced pluripotent stem cells C12N5/0696
• CPC Class Codes: C12N 5/0663 Bone marrow mesenchymal stem cells (BM-MSC)

Abstract

An object of the present invention is to provide a method capable of inexpensively and conveniently preparing cells having pluripotency and a very low risk of tumorigenic transformation. The cells having pluripotency and a very low risk of tumorigenic transformation can be prepared by suspension-culturing mammalian mesenchymal stem cells such as human mesenchymal stem cells from bone marrow (hMSC-BM) and human adipose tissue-derived mesenchymal stem cells (hAT-MSC) (also referred to as “human adipose-derived stem cells [hADSC]”), 7 types of human adherent mature cells (human hepatocyte cells [hHEP cells], human umbilical vein endothelial cells [HUVEC cells], human dermal lymphatic microvascular endothelial cells [HMVEC cells], human epidermal keratinocyte cells [NHEK cells], human bronchial epithelial cells [NHBE cells], human melanocyte cells [NHEM cells], and human smooth muscle cells [UASMC cells]), and 3 types of human adherent precursor cells (human dermal fibroblast cells [NHDF cells], human skeletal muscle myoblast cells [HSMM cells], and human osteoblast cells [NHOst cells]) to form cell masses (spheroids).

Claims

1. A method for preparing a pluripotent stem cell, comprising the step of suspension-culturing mammalian mesenchymal stem cells to form a cell mass of pluripotent stem cells.

2. The method according to claim 1, wherein the mammalian mesenchymal stem cells are human mesenchymal stem cells from bone marrow or human adipose tissue-derived mesenchymal stem cells.

3. The method according to claim 1, wherein the pluripotent stem cell expresses Nanog, Oct3/4, or Sox2.

4. The method according to claim 1, wherein the suspension-culturing is performed in a solution containing the following (A) or (B):

(A) gellan gum or a derivative thereof or a salt of these;

or

(B) dextran or a derivative thereof or a salt of these.

5. The method according to claim 1, wherein the suspension-culturing is performed in a physiological aqueous solution free from serum or a serum substitute.

6. A pluripotent stem cell obtainable by the method according to claim 1.

14. The method according to claim 2, wherein the pluripotent stem cell expresses Nanog, Oct3/4, or Sox2.

15. The method according to claim 2, wherein the suspension-culturing is performed in a solution containing the following (A) or (B):

(A) gellan gum or a derivative thereof or a salt of these;

or

(B) dextran or a derivative thereof or a salt of these.

16. The method according to claim 3, wherein the suspension-culturing is performed in a solution containing the following (A) or (B):

(A) gellan gum or a derivative thereof or a salt of these;

or

(B) dextran or a derivative thereof or a salt of these.

17. The method according to claim 14, wherein the suspension-culturing is performed in a solution containing the following (A) or (B):

(A) gellan gum or a derivative thereof or a salt of these;

or

(B) dextran or a derivative thereof or a salt of these.

18. The method according to claim 2, wherein the suspension-culturing is performed in a physiological aqueous solution free from serum or a serum substitute.

19. The method according to claim 3, wherein the suspension-culturing is performed in a physiological aqueous solution free from serum or a serum substitute.

20. The method according to claim 4, wherein the suspension-culturing is performed in a physiological aqueous solution free from serum or a serum substitute.

21. The method according to claim 14, wherein the suspension-culturing is performed in a physiological aqueous solution free from serum or a serum substitute.

22. The method according to claim 15, wherein the suspension-culturing is performed in a physiological aqueous solution free from serum or a serum substitute.

7. A pluripotent stem cell obtained by suspension-culturing mammalian mesenchymal stem cells.

8. The pluripotent stem cell according to claim 7, wherein the mammalian mesenchymal stem cells are human mesenchymal stem cells from bone marrow or human adipose tissue-derived mesenchymal stem cells.

9. The pluripotent stem cell according to claim 7, wherein the pluripotent stem cell expresses Nanog, Oct3/4, or Sox2.

10. The pluripotent stem cell according to claim 7, wherein the suspension-culturing is performed in a solution containing the following (A) or (B):

(A) gellan gum or a derivative thereof or a salt of these;

or

(B) dextran or a derivative thereof or a salt of these.

11. The pluripotent stem cell according to claim 7, wherein the suspension-culturing is performed in a physiological aqueous solution free from serum or a serum substitute.

12. (canceled)

13. (canceled)