Induction, propagation and isolation of liver progenitor cells

Abstract

The present invention relates to methods of induction and isolation of progenitor cells from stem cell cultures, specifically liver progenitor cells from human embryonic stem cell cultures. In one embodiment, the present invention provides a method of inducing hepatocyte-like progenitor cells by placing a quantity of human embryonic stem cells in a medium supplemented with an inhibitor of the MAPK/MEK/ERK signaling pathway, FGFR, GSK3 and/or BMP.

1 . A method of inducing and/or propagating a quantity of endoderm progenitor cells, comprising:
 a) providing a quantity of human pluripotent stem cells;   b) placing the quantity of human pluripotent stem cells in a medium supplemented with an inhibitor of MKK1/2, a fibroblast growth factor receptor (FGFR) inhibitor, and a glycogen synthase kinase 3 (GSK3) inhibitor; and   c) culturing the human pluripotent stem cells to form endoderm progenitor cells.   
     
     
         2 . The method of  claim 1 , wherein the human pluripotent stem cells are human embryonic stem cells. 
     
     
         3 . The method of  claim 1 , wherein the inhibitor of MKK1/2 comprises a compound of the formula: 
       
         
           
           
               
               
           
         
         or a salt thereof. 
       
     
     
         4 . The method of  claim 1 , wherein the inhibitor of MKK1/2 comprises a compound of the formula: 
       
         
           
           
               
               
           
         
         or a salt thereof. 
       
     
     
         5 . The method of  claim 1 , wherein the FGFR inhibitor comprises a compound of the formula: 
       
         
           
           
               
               
           
         
         or a salt thereof. 
       
     
     
         6 . The method of  claim 1 , wherein the GSK3 inhibitor comprises a compound of the formula: 
       
         
           
           
               
               
           
         
         or a salt thereof. 
       
     
     
         7 . The method of  claim 1 , wherein the endoderm progenitor cells are hepatocyte-like. 
     
     
         8 . The method of  claim 1 , wherein the human pluripotent stem cells comprises human embryonic stem cell 2 (HES2) and/or human embryonic stem cell 3 (HES3). 
     
     
         9 . The method of  claim 1 , wherein the medium comprises 0.2 to 3.0 μM of the inhibitor of MKK1/2. 
     
     
         10 . The method of  claim 1 , wherein the medium comprises 1.0 to 6.0 μM of the FGFR inhibitor. 
     
     
         11 . The method of  claim 1 , wherein the medium comprises 1.0 to 6.0 μM of the GSK3 inhibitor. 
     
     
         12 . The method of  claim 1 , wherein the medium further comprises a bone morphogenetic protein (BMP) inhibitor. 
     
     
         13 . The method of  claim 12 , wherein the medium comprises 1.0 to 6.0 μM of the BMP inhibitor. 
     
     
         14 . The method of  claim 12 , wherein the BMP inhibitor comprises dorsomorphin. 
     
     
         15 . A composition comprising an inhibitor of MKK1/2, an FGFR inhibitor, a GSK3 inhibitor, and a BMP inhibitor. 
     
     
         16 . A composition comprising activin, BMP, epidermal growth factor (EGF), heregulin, fibroblast growth factor 1 (FGF-1), insulin growth factor (IGF), vascular endothelial growth factor (VEGF), angiopoietin, and transforming growth factor beta (TGF beta). 
     
     
         17 . A composition, comprising a quantity of endoderm progenitor cells, produced by the process of  claim 1 .