Methods and compositions for modification of the human glucocorticoid receptor locus
AssigneeSangamo BioSciences, Inc.
InventorsAndreas Reik, Michael Jensen, Michael C. Holmes, Philip D. Gregory, Dale Ando
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Abstract
Disclosed herein are methods and compositions for inactivation of the human glucocorticoid receptor (GR) gene by targeted cleavage of genomic DNA encoding the GR. Such methods and compositions are useful, for example, in therapeutic applications which require retention of immune function during glucocorticoid treatment.
Claims19
A method for inactivating glucocorticoid receptor (GR) function in a cell, the method comprising:
- expressing in the cell a pair of fusion proteins, wherein each fusion protein comprises:
- (i) a zinc finger DNA-binding domain that has been engineered to bind a target sequence in a GR gene, and
- (ii) a cleavage half-domain;
- such that the fusion proteins catalyze a double-strand break in the GR gene.
A fusion protein comprising:
- (i) a zinc finger DNA-binding domain that has been engineered to bind a target sequence in a GR gene, and
- (ii) a cleavage domain or a cleavage half-domain.
A method of selecting cells comprising an exogenous sequence that has been integrated into a GR gene, the method comprising
- expressing a pair of fusion proteins in the cell, wherein each fusion protein comprises:
- (i) a zinc finger DNA-binding domain that has been engineered to bind a target sequence in a GR gene, and
- (ii) a cleavage half-domain;
- such that the fusion proteins catalyze a double-strand break in the GR gene;
- introducing a polynucleotide into the cell, wherein the polynucleotide comprises a first region of homology to sequences upstream of the double-strand break, a second region of homology to sequences downstream of the double-strand break and the exogenous sequence;
- and
- treating the cells with a natural or synthetic corticosteroid under conditions such that cells not comprising the integrated exogenous sequence are killed, thereby selecting cells into which an exogenous sequence has been introduced into a GR gene.
Dependencies are inferred from the claim text.