Method of producing pancreatic hormone-producing cells
| Title: | Method of producing pancreatic hormone-producing cells |
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| Patent Number: | 9,157,069 |
| Publication Date: | October 13, 2015 |
| Appl. No: | 13/814878 |
| Application Filed: | August 08, 2011 |
| Abstract: | Disclosed is a production method of pancreatic hormone-producing cells in a form that mimicks the pancreatogenesis, the method comprising subjecting stem cells to the following steps: (1) cultivating stem cells in a medium containing a Rho kinase inhibitor, (2) cultivating the cells obtained in (1) in a medium containing a GSK3 inhibitor, (3) cultivating the cells obtained in (2) in a medium containing GSK3 inhibitor and an activator of activin receptor-like kinase-4,7, (4) forming a cell mass from the cells obtained in (3), and cultivating the cell mass in a suspension state in a medium, (5) cultivating the cells obtained in (4) in a medium containing a retinoic acid receptor agonist, an inhibitor of AMP-activated protein kinase and/or activin receptor-like kinase-2,3,6, an inhibitor of activin receptor-like kinase-4,5,7 and a cell growth factor, and (6) cultivating the cells obtained in (5). |
| Inventors: | Hosoya, Masaki (Fujisawa, JP); Shoji, Masanobu (Fujisawa, JP) |
| Assignees: | Takeda Pharmaceutical Company Limited (Osaka-Shi, JP) |
| Claim: | 1. A method of producing human pancreatic hormone-producing cells, comprising subjecting human pluripotent stem cells to the following steps (1)-(6): (1) a step of cultivating human pluripotent stem cells in a medium containing (+)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl)cyclohexanamide dihydrochloride, (2) a step of cultivating the cells obtained in the aforementioned step (1) in a medium containing a 6-[[2-[[4-(2,4-dichlorophenyl)-5-(4-methyl-1H-imidazol-2-yl)-2-pyrimidinyl]amino]ethyl]amino]nicotinonitrile or (2′Z,3′E)-6-bromoindirubin-3′-oxime, (3) a step of cultivating the cells obtained in the aforementioned step (2) in a medium containing 6-[[2-[[4-(2,4-dichlorophenyl)-5-(4-methyl-1H-imidazol-2-yl)-2-pyrimidinyl]amino]ethyl]amino]nicotinonitrile and activin (4) a step of forming a cell mass from the cells obtained in the aforementioned (3), and cultivating the cell mass in a suspension state in a medium (5) a step of cultivating the cells obtained in the aforementioned step (4) in a medium containing dorsomorphin, retinoic acid, 4-[4-(1,3-benzodioxol-5-yl)-5-(2-pyridinyl)-1H-imidazol-2-yl]-benzamide or hydrate thereof and basic fibroblast growth factor, (6) a step of cultivating the cells obtained in the aforementioned step (5) to produce human pancreatic hormone-producing cells. |
| Claim: | 2. The production method according to claim 1 , wherein the steps (1)-(6) do not substantially use a feeder cell. |
| Claim: | 3. The production method according to claim 1 , wherein the medium in steps (1)-(6) does not substantially contain a serum. |
| Claim: | 4. The production method according to claim 1 , wherein the pancreatic hormone-producing cells are any selected from the group consisting of insulin-producing cells, glucagon-producing cells, somatostatin-producing cells, pancreatic polypeptide (PP)-producing cells and ghrelin-producing cells. |
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| Primary Examiner: | Ton, Thaian N |
| Attorney, Agent or Firm: | Mintz Levin Cohn Ferris Glovsky and Popeo, P.C. Conlin, Esq., David G. |
| Accession Number: | edspgr.09157069 |
| Database: | USPTO Patent Grants |
| Language: | English |
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