Precise deletion of chromosomal sequences in vivo and treatment of nucleotide repeat expansion disorders using engineered nucleases

A method for treating a subject having a nucleotide repeat expansion disorder, wherein said nucleotide repeat expansion disorder is characterized by expansion of a nucleotide repeat in a gene of interest, said method comprising delivering to target cells in said subject:

• at least a first nucleic acid encoding a first engineered nuclease and a second nucleic acid encoding a second engineered nuclease, wherein said first engineered nuclease and said second engineered nuclease are expressible in said target cells in vivo;
• wherein said first engineered nuclease recognizes and cleaves a first recognition sequence positioned 5′ upstream of said nucleotide repeat in said gene of interest;
• and wherein said second engineered nuclease recognizes and cleaves a second recognition sequence positioned 3′ downstream of said nucleotide repeat in said gene of interest;
• and wherein an intervening DNA fragment between said first recognition sequence and said second recognition sequence is excised and the number of said nucleotide repeat is reduced in said gene of interest;
• and wherein said first engineered nuclease and said second engineered nuclease generate complementary overhangs which promote direct re-ligation of said gene of interest.