US2014234963A1PendingUtilityA1

Efficient induction of definitive endoderm from pluripotent stem cells

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Assignee: FUNA NINAPriority: Jun 21, 2011Filed: Jun 21, 2012Published: Aug 21, 2014
Est. expiryJun 21, 2031(~4.9 yrs left)· nominal 20-yr term from priority
Inventors:Nina FunaKatja HessJenny EkbergHenrik Semb
A61P 3/08C12N 2501/115C12N 2501/40C12N 2501/16C12N 5/0678C12N 5/0676C12N 5/0606C12N 5/0696C12N 2506/02
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Claims

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

exact text as granted — not AI-modified
1 - 15 . (canceled) 
     
     
         16 . A method for differentiation of stem cells into definitive endoderm comprising the steps of:
 a. incubating stem cells in a medium comprising at least 2 μM CHIR, wherein activin A is not present; and   b. subsequently incubating stem cells in a medium comprising activin A.   
     
     
         17 . The method according to  claim 16 , wherein said medium is RPMI-1640. 
     
     
         18 . The method according to  claim 16 , wherein said stem cells are embryonic stem cells or induced pluripotent stem cells. 
     
     
         19 . The method according to  claim 16 , wherein the concentration of CHIR is at least about 2.5 μM. 
     
     
         20 . The method according to  claim 16 , wherein the concentration of CHIR is at least about 3.1 μM. 
     
     
         21 . The method according to  claim 16 , wherein the concentration of CHIR is in a range selected from the group consisting of about 2.5-15 μM, about 3.1-15 μM, about 3.1-7 μM, 3.5-7 μM, about 3.5-6 μM, and about 3.5-5 μM. 
     
     
         22 . The method according to  claim 16 , wherein the concentration of CHIR is at least about 3.5 μM. 
     
     
         23 . The method according to  claim 16 , wherein said incubation with CHIR is at least 24 hours. 
     
     
         24 . The method according to  claim 16 , wherein said incubation with activin A is at least 24 hours. 
     
     
         25 . The method according to  claim 16 , wherein said incubation with activin A is 48 to 72 hours. 
     
     
         26 . The method according to  claim 16 , wherein endodermal cells are obtained from said definitive endoderm cells. 
     
     
         27 . The method according to  claim 26 , wherein said endoderm cells are pancreatic endoderm cells. 
     
     
         28 . Pancreatic endoderm cells obtainable by the method of  claim 27 . 
     
     
         29 . The method according to  claim 16 , wherein the concentration of CHIR in the culture medium is at least 3 μM. 
     
     
         30 . The method according to  claim 16 , wherein the concentration of CHIR in the culture medium is in a range selected from the group consisting of about 2-15 μM, about 3-15 μM, about 3.1-15 μM, about 3.1-7 μM, about 3.5-15 μM, and about 3.5-7 μM. 
     
     
         31 . A method for inducing primitive streak cells from embryonic stem cells by using CHIR in a concentration of at least 2 μM in the culture medium. 
     
     
         32 . A method for inducing definitive endoderm cells from embryonic stem cells by using CHIR in a concentration in the range of 3.5-7 μM.

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