Methods for increasing definitive endoderm differentiation of pluripotent human embryonic stem cells with PI-3 kinase inhibitors

A method of differentiating a pluripotent human embryonic stem cell culture, the method consisting essentially of the steps:

• (a) providing the pluripotent human embryonic stem cell culture, and
• (b) contacting the pluripotent human embryonic stem cell culture with an effective amount of an inhibitor of a PI-3-kinase signaling pathway and a member of the TGFβ family to differentiate pluripotent human embryonic stem cells to definitive endoderm cells,
• wherein the inhibitor of a PI-3-kinase signaling pathway and the member of the TGFβ family are added exogenously to the pluripotent human embryonic stem cell culture at the same time,
• wherein the member of the TGFβ family is selected from the group consisting of Nodal, Activin A, Activin B, TGF-β, BMP2, BMP4, and a mixture thereof,
• thereby producing a definitive endoderm cell culture, wherein the percentage of definitive endoderm cells in the definitive endoderm cell culture is greater than 50%.