Differentiation of Human Embryonic Stem Cells
First Claim
Patent Images
1. A method for differentiating pluripotent stem cells into cells expressing markers characteristic of the pancreatic endocrine lineage, comprising the steps of:
- a. Culturing the pluripotent stem cells,b. Differentiating the pluripotent stem cells into cells expressing markers characteristics of the definitive endoderm lineage, by treating the pluripotent cells with activin A,c. Differentiating the cells expressing markers characteristics of the definitive endoderm lineage into cells expressing markers characteristics of the pancreatic endoderm lineage, by treating the cells expressing markers characteristics of the definitive endoderm lineage with at least one fibroblast growth factor, or with retinoic acid and at least one fibroblast growth factor, andd. Differentiating the cells expressing markers characteristics of the pancreatic endoderm lineage into cells expressing markers characteristics of the pancreatic endocrine lineage, by treating the cells expressing markers characteristics of the pancreatic endoderm lineage with a gamma secretase inhibitor.
2 Assignments
0 Petitions
Accused Products
Abstract
The present invention provides methods to promote the differentiation of pluripotent stem cells. In particular, the present invention provides an improved method for the formation of pancreatic endoderm, pancreatic hormone expressing cells and pancreatic hormone secreting cells. The present invention also provides methods to promote the differentiation of pluripotent stem cells without the use of a feeder cell layer.
-
Citations
308 Claims
-
1. A method for differentiating pluripotent stem cells into cells expressing markers characteristic of the pancreatic endocrine lineage, comprising the steps of:
-
a. Culturing the pluripotent stem cells, b. Differentiating the pluripotent stem cells into cells expressing markers characteristics of the definitive endoderm lineage, by treating the pluripotent cells with activin A, c. Differentiating the cells expressing markers characteristics of the definitive endoderm lineage into cells expressing markers characteristics of the pancreatic endoderm lineage, by treating the cells expressing markers characteristics of the definitive endoderm lineage with at least one fibroblast growth factor, or with retinoic acid and at least one fibroblast growth factor, and d. Differentiating the cells expressing markers characteristics of the pancreatic endoderm lineage into cells expressing markers characteristics of the pancreatic endocrine lineage, by treating the cells expressing markers characteristics of the pancreatic endoderm lineage with a gamma secretase inhibitor. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 26, 27, 28, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 63, 64, 65, 66, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 248, 249, 250, 251, 252, 253, 254, 279, 280, 281, 282, 283, 284)
-
- 24. The method of claim 24, wherein the GSK-3B inhibitor is selected from the group consisting of GSK-3B inhibitor IX and GSK-3B inhibitor XI.
- 29. The method of claim 29, wherein the extracellular matrix is MATRIGEL.
-
30. The method of claim 30, wherein the MATRIGEL is used at a dilution from about 1:
- 60 to about 1;
10.
- 60 to about 1;
-
62. The method of claim 62, wherein the GSK-3B inhibitor is GSK-3B inhibitor IX.
-
67. The method of claim 67, wherein the MATRIGEL is used at a dilution from about 1:
- 60 to about 1;
10.
- 60 to about 1;
-
99. A method for differentiating cells expressing markers characteristic of the definitive endoderm lineage into cells expressing markers characteristic of the pancreatic endocrine lineage, comprising the steps of:
-
a. Treating the cells expressing markers characteristics of the definitive endoderm lineage into cells expressing markers characteristics of the pancreatic endoderm lineage with at least one fibroblast growth factor, or with retinoic acid and at least one fibroblast growth factor, and b. Differentiating the cells expressing markers characteristics of the pancreatic endoderm lineage into cells expressing markers characteristics of the pancreatic endocrine lineage, by treating the cells expressing markers characteristics of the pancreatic endoderm lineage with a gamma secretase inhibitor. - View Dependent Claims (100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 255, 256, 257, 258, 259, 260, 261)
-
- 129. A method for differentiating cells expressing markers characteristic of the pancreatic endoderm lineage into cells expressing markers characteristic of the pancreatic endocrine lineage, comprising the step of treating the cells expressing markers characteristic of the pancreatic endoderm lineage with a factor that inhibits the Notch signaling pathway.
-
144. A method for differentiating pluripotent stem cells into cells expressing markers characteristic of the pancreatic endoderm lineage, comprising the steps of:
-
a. Culturing the pluripotent stem cells, b. Differentiating the pluripotent stem cells into cells expressing markers characteristics of the definitive endoderm lineage, by treating the pluripotent cells with activin A, and c. Differentiating the cells expressing markers characteristics of the definitive endoderm lineage into cells expressing markers characteristics of the pancreatic endoderm lineage, by treating the cells expressing markers characteristics of the definitive endoderm lineage with at least one fibroblast growth factor, or with retinoic acid and at least one fibroblast growth factor. - View Dependent Claims (145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 235, 236, 237, 238, 239, 268, 269, 285, 286, 287, 288, 289, 290)
-
-
164. A method for differentiating pluripotent stem cells into cells expressing markers that are characteristic of the definitive endoderm lineage, comprising the steps of:
-
a. Plating the pluripotent stem cells on a tissue culture substrate coated with an extracellular matrix, and b. Culturing the pluripotent stem cells with activin A and a Wnt ligand. - View Dependent Claims (165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 270, 274, 275, 291, 292, 293, 294, 295, 296)
-
-
192. A method for differentiating pluripotent stem cells into cells expressing markers that are characteristic of the definitive endoderm lineage, comprising the steps of:
-
a. Plating the pluripotent stem cells on a tissue culture substrate coated with an extracellular matrix, b. Culturing the pluripotent stem cells with activin A and a Wnt ligand in a first culture medium, and c. Culturing the pluripotent stem cells with activin A in a second culture medium. - View Dependent Claims (193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 271, 276, 277, 297, 298, 299, 300, 301, 302)
-
- 231. A method for differentiating cells expressing markers characteristic of the definitive endoderm into cells expressing markers characteristic of the pancreatic endoderm lineage, comprising the steps of treating the cells expressing markers characteristic of the definitive endoderm with at least one fibroblast growth factor, or with retinoic acid and at least one fibroblast growth factor.
-
278. A method for treating a patient suffering from diabetes, comprising the steps of:
-
a. Culturing pluripotent stem cells, b. Differentiating the pluripotent stem cells into cells expressing markers characteristics of the definitive endoderm lineage, c. Differentiating the cells expressing markers characteristics of the definitive endoderm lineage into cells expressing markers characteristics of the pancreatic endoderm lineage, d. Differentiating the cells expressing markers characteristics of the pancreatic endoderm lineage into cells of a β
-cell lineage, ande. Implanting the cells of a β
-cell lineage into the patient. - View Dependent Claims (303, 304, 305, 306, 307, 308)
-
Specification