Differentiation of human embryonic stem cells into pancreatic endocrine cells using HB9 regulators
First Claim
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1. A method for producing pancreatic endocrine cells from human pluripotent stem cells, comprising the steps of:
- (a) differentiating human pluripotent stem cells into foregut endoderm cells; and
(b) differentiating the foregut endoderm cells into pancreatic endocrine cells by treatment with a medium supplemented with (i) a thyroid hormone selected from triiodothyronine, thyroxine, analogues of triiodothyronine, analogues of thyroxine or mixtures thereof, or (ii) both thyroid hormone and an ALK5 inhibitor.
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Abstract
The present invention provides methods to promote differentiation of pluripotent stem cells to pancreatic endoderm cells expressing PDX1, NKX6.1, and HB9. In particular, the methods encompass culturing Stage 4 to Stage 6 cells with a thyroid hormone (e.g. T3), an ALK5 inhibitor, or both.
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Citations
53 Claims
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1. A method for producing pancreatic endocrine cells from human pluripotent stem cells, comprising the steps of:
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(a) differentiating human pluripotent stem cells into foregut endoderm cells; and (b) differentiating the foregut endoderm cells into pancreatic endocrine cells by treatment with a medium supplemented with (i) a thyroid hormone selected from triiodothyronine, thyroxine, analogues of triiodothyronine, analogues of thyroxine or mixtures thereof, or (ii) both thyroid hormone and an ALK5 inhibitor. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18)
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19. A method of producing Stage 5 cells comprising culturing human foregut endoderm cells in a medium supplemented with (i) a thyroid hormone selected from triiodothyronine, thyroxine, analogues of triiodothyronine, analogues of thyroxine or mixtures thereof, or (ii) both thyroid hormone and an ALK5 inhibitor;
- wherein said culturing of foregut endoderm cells produces Stage 5 cells.
- View Dependent Claims (20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36)
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37. A method of (a) increasing HB9 expression and (b) suppressing SOX2 and albumin expression in human pancreatic foregut precursor cells, comprising culturing human pancreatic foregut precursor cells in a medium supplemented with triiodothyronine and an ALK5 inhibitor;
- wherein said culturing increases HB9 expression and suppresses SOX2 and albumin expression in human pancreatic foregut precursor cells as compared to human pancreatic foregut precursor cells not cultured in a medium supplemented with triiodothyronine and an ALK5 inhibitor.
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38. A method of down-regulating glucagon, somatostatin and ghrelin expression in human pancreatic foregut precursor cells, human endocrine precursor cells or human pancreatic endocrine cells, comprising culturing the cells in a medium supplemented with triiodothyronine and an ALK5 inhibitor;
- wherein said culturing down-regulates glucagon, somatostatin and ghrelin expression in human pancreatic foregut precursor cells, human endocrine precursor cells or human pancreatic endocrine cells as compared to human pancreatic foregut precursor cells, human endocrine precursor cells or human pancreatic endocrine cells not cultured in a medium supplemented with triiodothyronine and an ALK5 inhibitor.
- View Dependent Claims (39, 40)
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41. A method for generating human pancreatic endocrine cells, comprising culturing human foregut endoderm cells in a medium supplemented with (i) a thyroid hormone selected from triiodothyronine, thyroxine, analogues of triiodothyronine, analogues of thyroxine or mixtures thereof, or (ii) both thyroid hormone and an ALK5 inhibitor;
- wherein said culturing of human foregut endoderm cells produces pancreatic endocrine cells.
- View Dependent Claims (42, 43, 44, 45, 46, 47, 48, 49)
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50. A method for increasing the yield of human β
- cells via the differentiation of human pluripotent stem cells, comprising culturing said human pluripotent cells in a growth media supplemented with a thyroid hormone selected from triiodothyronine, thyroxine, analogues of triiodothyronine, analogues of thyroxine or mixtures thereof, or both thyroid hormone and ALK5 inhibitor;
wherein said culturing of human pluripotent stem cells increases the yield of human β
cells. - View Dependent Claims (51, 52, 53)
- cells via the differentiation of human pluripotent stem cells, comprising culturing said human pluripotent cells in a growth media supplemented with a thyroid hormone selected from triiodothyronine, thyroxine, analogues of triiodothyronine, analogues of thyroxine or mixtures thereof, or both thyroid hormone and ALK5 inhibitor;
Specification