Use of small molecules to enhance MAFA expression in pancreatic endocrine cells
AssigneeJanssen Biotech, Inc.
InventorAlireza Rezania
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Abstract
The present invention provides methods, cell cultures and differentiation media to promote differentiation of pluripotent stem cells to pancreatic endocrine cells of a mature phenotype. The resulting pancreatic endocrine cells express single hormonal insulin, PDX1, NKX6.1, and MAFA. In one or more differentiation stages, culturing may be carried out in a culture vessel at the air-liquid interface.
Claims18
An in vitro cell culture comprising (i) a population human pancreatic endocrine cells, in which at least 10% of the differentiated cells express insulin, PDX1, NKX6.1, and MAFA, and (ii) a differentiation medium supplemented with an antioxidant, L-alanyl-glutamine and an inhibitor selected from the group consisting of an aurora kinase inhibitor, an RSK inhibitor, an inhibitor of protein methyltransferase DOT1L, and a combination thereof, wherein the population is obtained by a step-wise differentiation protocol which comprises culturing human pancreatic endocrine cells in the differentiation medium, and wherein the antioxidant is vitamin C, vitamin E, or a derivative thereof.
An in vitro cell culture comprising:
- (i) a population of human pancreatic endocrine cells, in which at least 10% of the differentiated cells express insulin, PDX1, NKX6.1, and MAFA, and
- (ii) a differentiation medium supplemented with an antioxidant and an inhibitor of protein methyltransferase DOT1L, an aurora kinase inhibitor, or a RSK inhibitor, wherein the antioxidant is vitamin C, vitamin E, or a derivative thereof, and
- wherein the population is obtained by a step-wise differentiation protocol comprising:
- (a) culturing human pluripotent stem cells in a medium supplemented with activin A and wnt3A to obtain definitive endoderm cells;
- (b) differentiating the definitive endoderm cells into pancreatic endocrine cells;
- and
- (c) culturing the pancreatic endocrine cells in the differentiation medium supplemented with the antioxidant, L-alanyl-glutamine and an inhibitor selected from the group consisting of an aurora kinase inhibitor, an RSK inhibitor, an inhibitor of protein methyltransferase DOT1L, and combinations thereof.
Dependencies are inferred from the claim text.