PLURIPOTENT STEM CELL-DERIVED BROWN ADIPOCYTES, PLURIPOTENT STEM CELL-DERIVED CELL AGGREGATE, METHOD FOR PRODUCING SAME, AND CELL THERAPY AND MEDICAL THERAPY THEREFOR

• US 20140140967A1
• Filed: 04/26/2012
• Published: 05/22/2014
• Est. Priority Date: 04/27/2011
• Status: Active Grant

• Current Assignee: Id Pharma Co., Ltd. (I'rom Group Co., Ltd.), National Center for Global Health and Medicine
• Original Assignee: National Center for Global Health and Medicine
• Inventors: Yuo, Akira, Saeki, Koichi, Hasegawa, Mamoru, Saeki, Kumiko, Nishio, Miwako, Kawasaki, Masako
• Granted Patent: US 9,492,485 B2
• Time in Patent Office: Days
• US Class Current: 424/93.7
• 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: A61K 35/35 Fat tissue; Adipocytes; Stromal cells; Connective tissues adipose-derived stem cells A61K35/28; collagen A61K38/39
• CPC Class Codes: A61P 3/06 Antihyperlipidemics
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 7/06 Antianaemics
• CPC Class Codes: A61P 9/10 for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: C12N 2501/105 Insulin-like growth factors [IGF]
• CPC Class Codes: C12N 2501/155 Bone morphogenic proteins [BMP]; Osteogenins; Osteogenic factor; Bone inducing factor
• CPC Class Codes: C12N 2501/165 Vascular endothelial growth factor [VEGF]
• CPC Class Codes: C12N 2501/2306 Interleukin-6 (IL-6)
• CPC Class Codes: C12N 2501/26 Flt-3 ligand (CD135L, flk-2 ligand)
• CPC Class Codes: C12N 2506/45 from artificially induced pluripotent stem cells
• CPC Class Codes: C12N 5/0653 Adipocytes; Adipose tissue

Abstract

Provided are a method of producing brown adipocytes from pluripotent stem cells, a method of producing cell aggregates as an intermediate product thereof, pluripotent stem cell-derived cell aggregates and pluripotent stem cell-derived brown adipocytes produced by these methods, and cell therapy using the pluripotent stem cell-derived brown adipocytes. In the method of producing brown adipocytes from pluripotent stem cells, cell aggregates are produced from pluripotent stem cells by a method including the step (A), and brown adipocytes are prepared from the cell aggregates by a method including the step (B). The step (A) is a step of producing cell aggregates by non-adhesive culture of pluripotent stem cells in serum-free environment in the presence of a hematopoietic cytokine, and the step (B) is a step of producing brown adipocytes by adhesion culture of the cell aggregates in the presence of a hematopoietic cytokine.

Claims

1. A method of producing brown adipocytes from pluripotent stem cells, the method comprising:

forming cell aggregates from pluripotent stem cells by a method including the step (A); and

preparing brown adipocytes from the cell aggregates by a method including the step (B);

(A) production of cell aggregates by non-adhesive culture of pluripotent stem cells in serum-free environment in the presence of one or more hematopoietic cytokines; and

(B) production of brown adipocytes by adhesion culture of the cell aggregates in the presence of one or more hematopoietic cytokines.

4. The method according to claim 1 or 2, wherein

the hematopoietic cytokine used in the step (A) is any one of BMP4, VEGF, SCF, Flt3L, IL6, and IGF2.

5. The method according to claim 1 or 3, wherein

the hematopoietic cytokine used in the step (B) is any one of BMP7, VEGF, SCF, Flt3L, IL6, and IGF2.

6. The method according to any one of claims 1 to 5, wherein

the pluripotent stem cells are ES cells or iPS cells.

7. The method of according to claim 6, wherein

the iPS cells are those established using a Sendai virus vector.

8. The method according to any one of claims 1 to 7, wherein

the pluripotent stem cells are human pluripotent stem cells.

2. A method of producing cell aggregates from pluripotent stem cells, the method comprising:

forming cell aggregates from pluripotent stem cells by a method including the step (A);

(A) production of cell aggregates by non-adhesive culture of pluripotent stem cells in serum-free environment in the presence of one or more hematopoietic cytokines.

3. A method of producing brown adipocytes from pluripotent stem cell-derived cell aggregate, the method comprising:

preparing brown adipocytes from pluripotent stem cell-derived cell aggregates by a method including the step (B);

(B) production of brown adipocytes by adhesion culture of cell aggregates in the presence of one or more hematopoietic cytokine.

9. A brown adipocyte produced using a pluripotent stem cell, wherein the brown adipocyte is produced from the pluripotent stem cell by a method including the steps (A) and (B):

(A) production of cell aggregates by non-adhesive culture of pluripotent stem cells in serum-free environment in the presence of a hematopoietic cytokine; and

(B) production of brown adipocytes by adhesion culture of the cell aggregates in the presence of a hematopoietic cytokine.

10. A cell aggregate produced using a pluripotent stem cell, wherein the cell aggregate is produced from the pluripotent stem cell by a method including the step (A):

(A) production of cell aggregates by non-adhesive culture of pluripotent stem cells in serum-free environment in the presence of a hematopoietic cytokine.

13. The cell aggregate derived from a pluripotent stem cell or the brown adipocyte derived from a pluripotent stem cell according to any one of claims 9 to 12, wherein

the pluripotent stem cell is an ES cell or an iPS cell.

14. The cell aggregate derived from a pluripotent stem cell or the brown adipocyte derived from a pluripotent stem cell according to any one of claims 9 to 13, wherein

the pluripotent stem cell is a human pluripotent stem cell.

11. A brown adipocyte produced using a pluripotent stem cell-derived cell aggregate, wherein the brown adipocyte is produced from the cell aggregate by a method including the step (B):

(B) production of brown adipocytes by adhesion culture of cell aggregates in the presence of a hematopoietic cytokine.

15. Cell therapy for preventing or treating obesity, the therapy comprising:

transplantation of the brown adipocyte derived from a pluripotent stem cell according to any one of claims 9 and 11 to 14.

16. Cell therapy for improving insulin resistance or preventing or treating diabetes mellitus, the therapy comprising:

transplantation of the brown adipocyte derived from a pluripotent stem cell according to any one of claims 9 and 11 to 14.

17. Cell therapy for preventing or treating hyperlipidemia, the therapy comprising:

transplantation of the brown adipocyte derived from a pluripotent stem cell according to any one of claims 9 and 11 to 14.

18. Cell therapy for surgery of producing a coronary artery bypass, the therapy comprising:

transplantation of the brown adipocyte derived from a pluripotent stem cell according to any one of claims 9 and 11 to 14.

19. Cell therapy for facilitating hematogenesis, the therapy comprising:

transplantation of the brown adipocyte derived from a pluripotent stem cell according to any one of claims 9 and 11 to 14.

20. Internal therapy being performed together with any of prevention or treatment of obesity, improvement of insulin resistance, prevention or treatment of diabetes mellitus, prevention or treatment of hyperlipidemia, treatment for facilitating hematogenesis, and surgery of producing a coronary artery bypass, the therapy comprising:

administration of a material secreted by the brown adipocyte derived from a pluripotent stem cell according to any one of claims 9 and 11 to 14.

12. A brown adipocyte produced using a pluripotent stem cell, wherein

the brown adipocyte derived from the pluripotent stem cell induces the expressions of at least UCP1, PRDM16, PGC1α

, Cyt-c, CIDE-A, ELOVL3, PPARα

, EVA1, and NTRK3 genes and has multilocular lipid droplets.