IP Library › Granted Patent US 10,487,313
Granted Patent B2
US 10,487,313 · App. 15/705,705 · Granted Nov 26, 2019

Efficient induction of definitive endoderm from pluripotent stem cells

Inventors: Nina Funa (Lund, SE); Katja Hess (Lund, SE); Jenny Ekberg (Malmoe, SE); Henrik Semb (Bjaerred, SE)
Assignees: Novo Nordisk A/S; Takara Bio Europe AB
C12N5/0678C12N5/0606C12N5/0676C12N5/0696C12N2501/115C12N2501/16C12N2501/40C12N2506/02
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Quick Facts
Patent No.
US 10,487,313
App. No.
15/705,705
Granted
Nov 26, 2019
Kind
B2
Abstract

The present invention relates to a method to differentiate pluripotent stem cells to a primitive streak cell population, in a stepwise manner for further maturation to definitive endoderm.

Claims (16)

1. A method for differentiating human embryonic stern cells into definitive endoderm cells, comprising:

(a) incubating human embryonic stem cells in a first medium comprising CHIR in a range of 2-7 μM, wherein activin A is not present in the medium; and

(b) subsequently incubating the stem cells in a second medium comprising activin A, wherein CHIR is not present in the second medium; and wherein human definitive endoderm cells are obtained.

2. The method according to claim 1 , wherein the concentration of CHIR is at least about 3 μM.

3. The method according to claim 1 , wherein the concentration of CHIR is about 4 μM.

4. The method according to claim 1 , wherein the incubating step with CHIR is performed for at least 24 hours.

5. The method according to claim 1 , wherein the incubating step with CHIR is performed for 24 hours.

6. The method according to claim 1 , wherein the incubating step with CHIR is performed for between 24 and 48 hours.

7. The method according to claim 1 , wherein the concentration of activin A is selected from the group consisting of about 80 ng/ml and about 100 ng/ml.

8. The method according to claim 1 , wherein the concentration of activin A is 100 ng/ml.

9. The method according to claim 1 , wherein the incubating step with activin A is performed for 48 to 72 hours.

10. The method according to claim 2 , wherein at least 91.5% of the stem cells after step (b) are positive for SOX17.

11. A method for differentiating human embryonic stem cells into definitive endoderm cells, comprising:

(a) incubating human embryonic stem cells in a first medium comprising about 2-7 μM CHIR for 24 hours, wherein activin A is not present in the first medium; and

(b) subsequently incubating the stem cells in a second medium comprising activin A in a concentration of about 100 ng/ml for 48 to 72 hours, wherein CHIR is not present in the second medium, wherein at least 91.5% of the stem cells after step (b) are positive for SOX17; and wherein human definitive endoderm cells are obtained.

12. The method according to claim 11 , wherein the concentration of CHIR is 4 μM and the concentration of activin A is 100 ng/ml.

Assignments (1)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Apr 1, 2026
From: NOVO NORDISK A/S
To: ASPECT BIOSYSTEMS LTD.
Reel/Frame 075363/0935 →
Priority Claims (1)
EP 11170713 · Jun 21, 2011 · regional
Continuity (3)
Continuation 14127296
Provisional Application 61501351 · Jun 27, 2011
Related Publication 20180002668A1 · Jan 4, 2018