Methods and compositions for treatment of a genetic condition

Abstract

Methods and compositions for a genetic disease are provided.

1 . An isolated genetically modified mammalian cell comprising an insertion and/or deletion within an endogenous BCL11A gene made by a nuclease comprising a DNA-binding domain that binds to a target site as shown in any of SEQ ID NOs:56, 63, 66, 71, 160, 170, 179, 183, 189, 193, 197, 200, 203, 207, 211 and 213, such that the BCL11A gene is inactivated and expression of at least one globin gene in the cell is altered. 
     
     
         2 . The genetically modified cell of  claim 1 , wherein the cell is red blood precursor cell. 
     
     
         3 . The genetically modified cell of  claim 1 , wherein the cell is a hematopoietic stem cell. 
     
     
         4 . The genetically modified cell of  claim 1 , wherein the genomic modification is within exon 2 of the BCL11A gene, exon 4 of BCL11A or the XL portion of a BCL11A-XL isoform. 
     
     
         5 . The genetically modified cell of  claim 1 , wherein the cell further comprises a transgene. 
     
     
         6 . The genetically modified cell of  claim 5 , wherein the transgene is integrated into the inactivated BCL11A gene. 
     
     
         7 . The genetically modified cell of  claim 1 , wherein the nuclease comprises a zinc finger nuclease (ZFN), a TAL-effector domain nuclease (TALEN) or a CRISPR/Cas nuclease. 
     
     
         8 . The genetically modified cell of  claim 7 , wherein the zinc finger nuclease comprises a zinc finger protein comprising 4, 5, or 6 zinc finger domains comprising a recognition helix and further wherein the zinc finger protein comprises the recognition helix regions of the proteins designated SBS#39172, SBS#43490, SBS#44642, SBS#45148, SBS#45147, SBS#39145, SBS#44490, SBS#44489, SBS#45081, SBS#44493, SBS#29527, SBS#29528, SBS#29525, SBS#29526, SBS#34678, SBS#34642, SBS#44889, SBS#44888, SBS#44905, SBS#44904, SBS#44911, SBS#44910, SBS#44945, SBS#44944, SBS#44947 or SBS#44946 of Table 1A. 
     
     
         9 . A pharmaceutical composition comprising the genetically modified cell of  claim 1 . 
     
     
         10 . A method of making a cell according to  claim 1 , the method comprising
 introducing, into the cell, one or more polynucleotides encoding a nuclease comprising a DNA-binding domain that binds to a target site as shown in any of SEQ ID NOs:56, 63, 66, 71, 160, 170, 179, 183, 189, 193, 197, 200, 203, 207, 211 and 213, such that the BCL11A gene is inactivated and expression of the globin gene is altered.   
     
     
         11 . The method of  claim 10 , wherein the proteins increase expression of a globin gene. 
     
     
         12 . The method of  claim 10 , wherein the globin gene is a gamma globin or beta globin gene. 
     
     
         13 . The method of  claim 10 , further comprising integrating a donor sequence into the genome of the cell. 
     
     
         14 . The method of  claim 13 , wherein the donor sequence is introduced to the cell using a viral vector, as an oligonucleotide or on a plasmid. 
     
     
         15 . The method of  claim 10 , wherein the cell is a red blood cell (RBC) precursor cell and a hematopoietic stem cell. 
     
     
         16 . The cell of  claim 15 , wherein the hematopoietic stem cell is a CD34+ hematopoietic stem cell. 
     
     
         17 . The method of  claim 13 , wherein the donor sequence comprises a transgene under the control of an endogenous promoter or an exogenous promoter. 
     
     
         18 . (canceled) 
     
     
         19 . A method of altering globin expression in a subject, the method comprising administering a cell according to  claim 1  to the subject. 
     
     
         20 . The method of  claim 19 , wherein the subject has a hemoglobinopathy. 
     
     
         21 . A method of altering globin expression in a subject, the method comprising administrating a pharmaceutical composition according to  claim 9  to the subject. 
     
     
         22 . (canceled)