Small molecule based conversion of somatic cells into neural crest cells

Patent metadata

Field	Value
Publication number	US-2016369233-A1
Application number	US-201615004745-A
Country	US
Kind code	A1
Filing date	Jan 22, 2016
Priority date	Jul 23, 2013
Publication date	Dec 22, 2016
Grant date	—

Abstract

This application relates to a method for differentiating somatic cells into multi-competent neural crest cells based on linked steps of chemically defined medium inductions. Neural crest cells are able to differentiate into numerous cell types like Schwann cells, chondrocytes, smooth muscle cells or adipocytes.

First claim

1 . A method of producing Neural Crest cells from somatic cells, comprising:
a) culturing somatic cells in medium supplemented with valproic acid, b) culturing the cells obtained in step (a) in a serum-free medium supplemented with N-{(3R,4R)-4-[4-(2-Fluoro-6-hydroxy-3-methoxy-benzoyl)-benzoylamino]-azepan-3-yl}-4-hydroxy-3,5-dimethyl-benzamide.

2 . The method of claim 1 , wherein the method does not comprise genetically modifying the somatic cells or the cells obtained in step (a) by the introduction of genes.

3 . The method of claim 1 , wherein step b) comprises culturing the cells in suspension culture.

4 . The method of claim 1 , wherein the serum-free medium of step b) is supplemented with an inhibitor of bone morphogenetic protein (BMP).

5 . The method of claim 4 , wherein the inhibitor of BMP is noggin.

6 . The method of claim 1 , wherein the serum-free medium of step b) is supplemented with a small molecule inhibitor of Transforming growth factor beta (TGF β).

7 . The method of claim 6 , wherein the small molecule inhibitor of TGIF β is SB431542.

8 . The method of claim 1 , wherein the serum-free medium of step b) is supplemented with a small molecule inhibitor of glycogen synthase kinase 3 (GSK3β).

9 . The method of claim 8 , wherein the inhibitor of GSK3β is 3-Amino-phenyl)-4-(1-methyl-1H-indol-3-yl)-pyrrole-2,5-dione.

10 . The method of claim 1 , wherein step a) comprises culturing the cells for 2 days.

11 . The method of claim 1 , wherein step b) comprises culturing the cells for 7 days.

12 . The method of claim 1 , wherein the somatic cells are fibroblasts.

13 . The method of claim 1 , wherein the somatic cells are human cells.

14 . The method of claim 1 , wherein the somatic cells are obtained from a subject suffering from a neurological disease.

15 . (canceled)

16 . The method of claim 1 , further comprising c) incubating the product of steps b) under conditions suitable for differentiation of the neural crest cells into a differentiated cell selected from the group of Schwann Cell, chondrocyte, smooth muscle cell or adipocyte.

17 . (canceled)

18 . (canceled)

19 . A therapeutic composition comprising cells selected from Neural Crest cells, differentiated Schwann Cells, chondrocytes, smooth muscle cells, and adipocytes.

20 . A method of treating, preventing, or stabilizing a neurological disease in a patient, comprising:
a) obtaining somatic cells from a healthy subject not suffering from a neurological disease, b) culturing the somatic cells in a medium supplemented with valproic acid, c) culturing the cells obtained in step (b) in a serum-free medium supplemented with N-{(3R,4R)-4-[4-(2-Fluoro-6-hydroxy-3-methoxy-benzoyl)-benzoylamino]-azepan-3yl}-4-hydroxy-3,5-dimethyl-benzamide, to obtain Neural Crest cells, d) transplanting the Neural Crest cells to an individual in need of such treatment.

21 . The method of claim 20 , wherein the neurological disease is one of a demyelinating disease, Multiple sclerosis, Myelopathies, Experimental allergic encephalomyelitis (EAE), acute disseminated encephalomyelitis (ADEM), postinfectious or postvaccinal encephalomyelitis, peripheral neuropathies, Schwannomatosis, Charcot-MarieTooth disease, Guillain-Barre Syndrome, Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP).

Assignees

• Hoffmann La Roche

Inventors

• Graf Martin
• Iacone Roberto
• Thoma Eva Christina

Classifications

• A61P21/00

  Drugs for disorders of the muscular or neuromuscular system · CPC title

• A61P25/00

  Drugs for disorders of the nervous system · CPC title

• A61P19/00

  Drugs for skeletal disorders · CPC title

• A61K35/30

  Nerves; Brain; Eyes; Corneal cells; Cerebrospinal fluid; Neuronal stem cells; Neuronal precursor cells; Glial cells; Oligodendrocytes; Schwann cells; Astroglia; Astrocytes; Choroid plexus; Spinal cord tissue · CPC title

• C12N2501/727

  Kinases (EC 2.7.) · CPC title