Patent ID: 7973015
Filing Date: 2011-07-05
Classification: A01K,A61K,A61P,C12N

Abstract:
1. A method for directing splicing of a dystrophin pre-mRNA in a first cell, the method comprising: contacting said dystrophin pre-mRNA in said first cell with a first antisense oligonucleotide having from 14 to 40 nucleotides, and being complementary to a portion of a first exon of the dystrophin pre-mRNA, said first exon selected from the group consisting of exon number 2, 8, 29, 43, 44, 45, 46, 50, 51, 52 and 53 of a dystrophin gene, and wherein said first antisense oligonucleotide is able to bind to said first exon and mask said first exon from said first cell's splicing apparatus, providing said first cell with a second antisense oligonucleotide, said second antisense oligonucleotide having from 14 to 40 nucleotides, and being complementary to a portion of a second exon of said dystrophin pre-mRNA wherein said second exon is different from said first exon, wherein said second antisense oligonucleotide is able to bind to said second exon and mask the second exon from the first cell's splicing apparatus, splicing said dystrophin pre-mRNA, and translating an mRNA produced from splicing of said dystrophin pre-mRNA to form a final mRNA excluding said first exon and said second exon, wherein said translation of said mRNA results in a mutant dystrophin protein having a Becker mutant's functionality, wherein said first antisense oligonucleotide is obtained by: providing a second cell with said first oligonucleotide, the cell having a pre-mRNA containing said first exon, culturing said second cell to form mRNA in said second cell from said pre-mRNA in said second cell, and determining whether said first exon is absent from the thus formed mRNA in said second cell, wherein said second antisense oligonucleotide is obtained by: providing a third cell or said second cell with said second oligonucleotide, said third cell or said second cell having a pre-mRNA containing said second exon, culturing said third cell or said second cell to form mRNA in said third cell or said second cell from said pre-mRNA in said third cell or said second cell, and determining whether said second exon is absent from the thus formed mRNA in said third cell or said second cell, and wherein the method of determining whether said first exon or said second exon is absent from the thus formed mRNA comprises RT-PCR and sequence analysis.