Use of a TGF-βR-1 kinase to produce human pancreatic endocrine cells

• US 10,494,609 B2
• Filed: 05/14/2018
• Issued: 12/03/2019
• Est. Priority Date: 11/27/2007
• Status: Active Grant

• Current Assignee: Janssen Biotech, Inc. (Johnson & Johnson)
• Original Assignee: Janssen Biotech, Inc. (Johnson & Johnson)
• Inventors: Rezania, Alireza
• Primary Examiner(s): Ton, Thaian N
• Assistant Examiner(s): Montanari, David A.
• Application Number:
  US15/978,935
• Publication Number:
  US 20180258402A1
• Time in Patent Office: 568 Days
• CPC Class Codes: A61P 1/18 for pancreatic disorders, e.g. pancreatic enzymes
• CPC Class Codes: C12N 2501/105 Insulin-like growth factors [IGF]
• CPC Class Codes: C12N 2501/117 Keratinocyte growth factors (KGF-1, i.e. FGF-7; KGF-2, i.e. FGF-12)
• CPC Class Codes: C12N 2501/12 Hepatocyte growth factor [HGF]
• CPC Class Codes: C12N 2501/15 Transforming growth factor beta (TGF-β)
• CPC Class Codes: C12N 2501/155 Bone morphogenic proteins [BMP]; Osteogenins; Osteogenic factor; Bone inducing factor
• CPC Class Codes: C12N 2501/16 Activin; Inhibin; Mullerian inhibiting substance
• CPC Class Codes: C12N 2501/335 Glucagon; Glucagon-like peptide [GLP]; Exendin
• CPC Class Codes: C12N 2501/385 of the family of the retinoic acid recptor, e.g. RAR, RXR; Peroxisome proliferator-activated receptor [PPAR]
• CPC Class Codes: C12N 2501/40 Regulators of development
• CPC Class Codes: C12N 2501/41 Hedgehog proteins; Cyclopamine (inhibitor)
• CPC Class Codes: C12N 2501/415 Wnt; Frizzeled
• CPC Class Codes: C12N 2501/42 Notch; Delta; Jagged; Serrate
• CPC Class Codes: C12N 2501/70 Enzymes
• CPC Class Codes: C12N 2501/727 Kinases (EC 2.7.)
• CPC Class Codes: C12N 2501/998 Proteins not provided for elsewhere
• CPC Class Codes: C12N 2501/999 Small molecules not provided for elsewhere
• CPC Class Codes: C12N 2506/02 from embryonic cells
• CPC Class Codes: C12N 2533/90 Substrates of biological origin, e.g. extracellular matrix, decellularised tissue
• CPC Class Codes: C12N 5/0676 Pancreatic cells
• CPC Class Codes: C12N 5/0678 : Stem cells; Progenitor cells; Precursor cells

Abstract

The present invention provides methods to promote the differentiation of pluripotent stem cells and the products related to or resulting from such methods. In particular, the present invention provides an improved method for the formation of pancreatic hormone expressing cells and pancreatic hormone secreting cells. In addition, the present invention also provides methods to promote the differentiation of pluripotent stem cells without the use of a feeder cell layer and the products related to or resulting from such methods. The present invention also provides methods to promote glucose-stimulated insulin secretion in insulin-producing cells derived from pluripotent stem cells.

Claims

1. A method of differentiating human pancreatic endoderm cells into human pancreatic endocrine cells comprising

treating the human pancreatic endoderm cells with an inhibitor of TGF-β

R-1 kinase, thereby differentiating the human pancreatic endoderm cells into the human pancreatic endocrine cells.

2. The method of claim 1, wherein the method produces human pancreatic endocrine cells capable of glucose-stimulated insulin secretion.

3. The method of claim 1, wherein the method comprises treating the human pancreatic endoderm cells with about 0.1 μ

M to about 100 μ

M of the inhibitor of TGF-β

R-1 kinase.

4. The method of claim 1, wherein the method comprises treating the human pancreatic endoderm cells with about 1 μ

M to about 10 μ

M of the inhibitor of TGF-β

R-1 kinase.

5. The method of claim 1, wherein the inhibitor of TGF-f3R-1 kinase is 2-(3-(6- Methylpyridin-2-yl)-1H-pyrazol-4-yl)-1,5-naphthyridine.

6. The method of claim 1, the inhibitor of TGF-β

R-1 kinase is [3-(Pyridin-2-yl)-4-(4- quinonyl)]-1H-pyrazole.

7. The method of claim 1, wherein the method comprises treating the human pancreatic endoderm cells for about five to about twelve days.

8. The method of claim 1, wherein the method comprises treating the human pancreatic endoderm cells for about twelve days.

9. The method of claim 1, wherein the human pancreatic endoderm cells are derived from human pluripotent stem cells.

10. The method of claim 9, wherein the human pluripotent stem cells are human embryonic stem cells.

11. The method of claim 1, wherein the method further comprises obtaining the human pancreatic endoderm cells.

12. The method of claim 1, wherein the method further comprises culturing the human pancreatic endoderm cells.

13. The method of claim 1, wherein the method comprises treating the human pancreatic endoderm cells with an inhibitor of TGF-β

R-1 kinase and a factor that inhibits the Notch signaling pathway.

14. The method of claim 13, wherein the factor that inhibits the Notch signaling pathway is a γ

-secretase inhibitor.

15. The method of claim 14, wherein the γ

-secretase inhibitor is L-685,458.

16. The method of claim 1, wherein the method further comprises encapsulating the cells.