ANTISENSE OLIGONUCLEOTIDES FOR INDUCING EXON SKIPPING AND METHODS OF USE THEREOF
An antisense molecule capable of binding to a selected target site to induce exon skipping in the dystrophin gene, as set forth in SEQ ID NO: 1 to 202.
1 - 14 . (canceled)
15 . An isolated antisense oligonucleotide of 10 to 50 nucleotides in length comprising a sequence that is specifically hybridizable to an exon 45 target region of the Dystrophin gene designated as annealing site H45A(−6+20), H45A(+91+110), H45A(+125+151), H45D(+16−04), H45A(+71+90) or a combination thereof.
16 . The antisense oligonucleotide of claim 15 , comprising SEQ ID NO:207, SEQ ID NO:208, SEQ ID NO:209, SEQ ID NO:210 or SEQ ID NO:211 wherein the uracil bases are optionally thymine bases.
17 . The antisense oligonucleotide of claim 16 , wherein the uracil bases are thymine bases.
18 . The antisense oligonucleotide of claim 15 , which is 20 to 31 nucleotides in length.
19 . The antisense oligonucleotide of claim 15 , which is 25 to 31 nucleotides in length.
20 . The antisense oligonucleotide of claim 15 , wherein the oligonucleotide does not activate RNase H.
21 . The antisense oligonucleotide of claim 15 , comprising a non-natural backbone.
22 . The antisense oligonucleotide of claim 21 , wherein the sugar moieties of the oligonucleotide backbone are replaced with non-natural moieties.
23 . The antisense oligonucleotide of claim 22 , wherein the non-natural moieties are morpholinos.
24 . The antisense oligonucleotide of claim 15 , wherein the inter-nucleotide linkages of the oligonucleotide backbone are replaced with non-natural inter-nucleotide linkages.
25 . The antisense oligonucleotide of claim 24 , wherein the non-natural inter-nucleotide linkages are modified phosphates.
26 . The antisense oligonucleotide of claim 25 , wherein the modified phosphates are methyl phosphonates, methyl phosphorothioates, phosphoromorpholidates, phosphoropiperazidates or phosphoroamidates.
27 . The antisense oligonucleotide of claim 26 , wherein the modified phosphates are phosphoramidates.
28 . The antisense oligonucleotide of claim 26 , wherein the modified phosphates are phosphoromorpholidates.
29 . The antisense oligonucleotide of claim 15 , wherein the sugar moieties of the oligonucleotide backbone are replaced with non-natural moieties and the internucleotide linkages of the oligonucleotide backbone are replaced with non-natural inter-nucleotide linkages.
30 . The antisense oligonucleotide of claim 29 , wherein the non-natural moieties are morpholinos and the non-natural internucleotide linkages are modified phosphates.
31 . The antisense oligonucleotide of claim 30 , wherein the modified phosphates are methyl phosphonates, methyl phosphorothioates, phosphoromorpholidates, phosphoropiperazidates or phosphoroamidates.
32 . The antisense oligonucleotide of claim 31 , wherein the modified phosphates are phosphoramidates.
33 . The antisense oligonucleotide of claim 31 , wherein the modified phosphates are phosphoromorpholidates.
34 . The antisense oligonucleotide of claim 15 , wherein the oligonucleotide is a 2′-O-methyl-oligoribonucleotide.
35 . The antisense oligonucleotide claim 15 , wherein the oligonucleotide is a peptide nucleic acid.
36 . The antisense oligonucleotide of claim 15 , wherein the oligonucleotide is chemically linked to one or more moieties or conjugates that enhance the activity, cellular distribution, or cellular uptake of the antisense oligonucleotide.
37 . The antisense oligonucleotide of claim 36 , wherein the oligonucleotide is chemically linked to a polyethylene glycol molecule.
38 . The antisense oligonucleotide of claim 15 , wherein the antisense oligonucleotide consists of the sequence identified as SEQ ID NO:207, SEQ ID NO:208, SEQ ID NO:209, SEQ ID NO:210 or SEQ ID NO:211, wherein the uracil bases are optionally thymine bases.
39 . The antisense oligonucleotide of claim 38 , wherein the uracil bases are thymine bases.
40 . A pharmaceutical composition, comprising the antisense oligonucleotide of claim 15 and a pharmaceutically acceptable carrier.
41 . The pharmaceutical composition of claim 40 , wherein the pharmaceutically acceptable carrier is phosphate-buffered saline.
42 . A method of inducing exon-skipping of dystrophin exon 45 comprising delivering an antisense oligonucleotide of claim 15 to a cell, thereby inducing exon-skipping of the dystrophin exon.
43 . The method of claim 42 , wherein the cell is a human muscle cell.
44 . The method of claim 43 , wherein the human muscle cell is in a patient.
45 . The method of claim 44 , wherein the patient has muscular dystrophy.
46 . The method of claim 45 , wherein the muscular dystrophy is Duchenne Muscular Dystrophy.
47 . A pharmaceutically acceptable salt of the antisense oligonucleotide of claim 15 .