Conversion of somatic cells to induced reprogrammed neural stem cells (irnscs)

Abstract

This application relates to a method for converting somatic cells to Neural Stem Cells (NSCs). Moreover this application relates to a method for converting human fibroblasts, keratinocytes or adipocytes to neural stem cells based on linked steps of genes transduction and chemically defined medium induction.

1 . A method of producing Neural Stem Cells (NCS), comprising:
 a) providing somatic cells,   b) reprogramming said somatic cells to NSCs by introducing at least two genes selected from the group consisting of Bmi 1, Sox2, Mash 1, Sox1 1, Emx2, Foxg1 and Pax6; and   c) inducing the reprogramming with growth factors and a small molecule.   
     
     
         2 . The method of  claim 1 , further comprising
 d) incubating the product of steps b) and c) under conditions suitable for proliferation of the NSCs.   
     
     
         3 . The method of  claim 1 , wherein the somatic cells of step a) are human cells. 
     
     
         4 . The method of  claim 3 , wherein the somatic cells of step a) are selected from the group consisting of fibroblasts, keratinocytes and adipocytes. 
     
     
         5 . The method of  claim 1 , wherein the growth factors and small molecule of step c) are supplements of a chemically defined medium. 
     
     
         6 . The method of  claim 5 , wherein the chemically defined medium is a serum free medium supplemented with insulin, transferrin and progesterone. 
     
     
         7 . The method of  claim 1 , wherein the at least two genes of step b) comprise Bmi 1 and Sox2. 
     
     
         8 . The method of  claim 7 , wherein the at least two genes of step b) additionally comprise at least one gene selected from the group of Mash 1, Sox1 1, Emx2, Foxg1 and Pax6. 
     
     
         9 . The method of  claim 7 , wherein the at least two genes of step b) comprise Bmi 1, Sox2 and Mash 1. 
     
     
         10 . The method of  claim 1 , wherein the growth factor of step c) is selected from the group consisting of FGF2, EGF and BDNF. 
     
     
         11 . The method of  claim 1 , wherein the small molecule of step c) comprises a ROCK inhibitor. 
     
     
         12 . The method of  claim 11 , wherein the ROCK inhibitor is selected from the group consisting of
 1-(5-Isoquinolinesulfonyl) homopiperazine,   N-Benzyl-2-(pyrimidin-4-ylamino)thiazole-4-carboxamide,   (+)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl) cyclo-hexanecarboxamide dihydrochloride) and   N-{(3R,4R)-4-[4-(2-Fluoro-6-hydroxy-3-methoxy-benzoyl)-benzoylamino]-azepan-3-yl}-4-hydroxy-3,5-dimethyl-benzamide.   
     
     
         13 . The method of  claim 1 , wherein the somatic cells are pretreated with a histone deacetylase (HDAC) inhibitor. 
     
     
         14 . The method of  claim 1 , wherein reprogramming of said somatic cells is achieved through delivery of a combination of at least two genes by a lentivirus. 
     
     
         15 . (canceled) 
     
     
         16 . (canceled) 
     
     
         17 . A therapeutic composition comprising the neural stem cells of  claim 15 . 
     
     
         18 . The therapeutic composition of  claim 17 , wherein the neural stem cells are differentiated into neurons or glia cells. 
     
     
         19 . (canceled) 
     
     
         20 . (canceled) 
     
     
         21 . A method of treating, preventing, or stabilizing a neurological disease in an individual in need of such treatment comprising administering a therapeutic composition of  claim 17  to the individual. 
     
     
         22 . The method of  claim 21 , wherein the neurological disease is one of Alzheimer's disease, Parkinson's disease, Huntington's disease, or ALS, lysosomal storage diseases, multiple sclerosis, and a spinal cord injury.