Compositions and Methods for Correcting Limb Girdle Muscular Dystrophy Type 2C Using Exon Skipping

Abstract

The invention is directed to one or more antisense polynucleotides and their use in pharmaceutical compositions in a strategy to induce exon skipping in the γ-sarcoglycan gene in patients suffering from Limb-Girdle Muscular Dystrophy-2C (LGMD2C) or in patients at risk of such a disease. The invention also provides methods of preventing or treating muscular dystrophy, e.g., LGMD2C, by exon skipping in the gamma sarcoglycan gene using antisense polynucleotides. Accordingly, in some aspects the invention provides an isolated antisense oligonucleotide, wherein the oligonucleotide specifically hybridizes to an exon target region of a γ-sarcoglycan RNA. In another aspect, the the invention provides a method of inducing exon-skipping of a gamma sarcoglycan RNA, comprising delivering an antisense oligonucleotide or a composition to a cell.

What is claimed is: 
     
         1 . An isolated antisense oligonucleotide (AON) selected from the group consisting of oligonucleotides listed in Table 2. 
     
     
         2 . The antisense oligonucleotide of  claim 1 , wherein the oligonucleotide cannot form an RNase H substrate. 
     
     
         3 . The antisense oligonucleotide of  claim 1  or  claim 2 , comprising a modified oligonucleotide backbone. 
     
     
         4 . The antisense oligonucleotide of  claim 3 , wherein the modified oligonucleotide backbone comprises a modified moiety substituted for the sugar of at least one of the oligonucleotides. 
     
     
         5 . The antisense oligonucleotide of  claim 4 , wherein the modified moiety is a Morpholino. 
     
     
         6 . The antisense oligonucleotide of any one of  claims 3 - 5 , wherein the modified oligonucleotide backbone of at least one of the oligonucleotides comprises at least one modified internucleotide linkage. 
     
     
         7 . The antisense oligonucleotide of  claim 6 , wherein the modified internucleotide linkage is a tricyclo-DNA (tc-DNA) modification. 
     
     
         8 . The antisense oligonucleotide of  claim 6 , wherein the modified internucleotide linkage comprises a modified phosphate. 
     
     
         9 . The antisense oligonucleotide of  claim 8 , wherein the modified phosphate is selected from the group consisting of a methyl phosphonate, a methyl phosphorothioate, a phosphoromorpholidate, a phosphoropiperazidate and a phosphoroamidate. 
     
     
         10 . The antisense oligonucleotide of any one of  claims 3 - 9 , wherein the oligonucleotide is a 2′-O-methyl-oligoribonucleotide. 
     
     
         11 . The antisense oligonucleotide of any one of  claims 1 - 10 , wherein the oligonucleotide comprises a peptide nucleic acid. 
     
     
         12 . The antisense oligonucleotide of any one of  claims 1 - 11 , wherein the oligonucleotide is chemically linked to one or more conjugates that enhance the activity, cellular distribution, or cellular uptake of the antisense oligonucleotide. 
     
     
         13 . The antisense oligonucleotide of  claim 12 , wherein the oligonucleotide is chemically linked to a polyethylene glycol molecule. 
     
     
         14 . The antisense oligonucleotide of  claim 12  or  claim 13  wherein the conjugate is a peptide that enhances cellular uptake. 
     
     
         15 . The antisense oligonucleotide of  claim 14  wherein the peptide is selected from the group consisting of a nuclear localization signal (NLS), HIV-1 TAT protein, a peptide comprising an integrin binding domain, oligolysine, adenovirus fiber protein and a peptide comprising a receptor-mediated endocytosis (RME) domain. 
     
     
         16 . A pharmaceutical composition, comprising the antisense oligonucleotide of any one of  claims 1 - 15  and a physiologically compatible buffer. 
     
     
         17 . A method of inducing exon-skipping of a gamma sarcoglycan RNA, comprising delivering to a cell the antisense oligonucleotide of any one of  claims 1 - 14  or the composition of  claim 15 , thereby inducing exon-skipping of the gamma sarcoglycan RNA. 
     
     
         18 . The method of  claim 17 , wherein the cell is a human muscle cell. 
     
     
         19 . The method of  claim 18 , wherein the human muscle cell is in a patient. 
     
     
         20 . The method of  claim 19 , wherein the patient has muscular dystrophy. 
     
     
         21 . The method of  claim 20 , wherein the muscular dystrophy is Limb Girdle Muscular Dystrophy type 2C (LGMD2C). 
     
     
         22 . A method of ameliorating Limb Girdle Muscular Dystrophy type 2C (LGMD2C) in a patient in need thereof comprising the step of administering to the patient a therapeutically effective amount of the composition of  claim 16 , thereby ameliorating LGMD2C. 
     
     
         23 . A method of inhibiting the progression of dystrophic pathology associated with LGMD2C in a patient in need thereof comprising the step of administering to the patient a therapeutically effective amount of the composition of  claim 16 , thereby inhibiting the progression of dystrophic pathology. 
     
     
         24 . A method of improving muscle function in a patient suffering from Limb Girdle Muscular Dystrophy type 2C (LGMD2C) comprising the step of administering to the patient a therapeutically effective amount of the composition of  claim 16 , thereby improving muscle function. 
     
     
         25 . The method of  claim 24  wherein the muscle is a cardiac muscle. 
     
     
         26 . The method of  claim 24  or  claim 25  wherein the improvement in muscle function is an improvement in muscle strength. 
     
     
         27 . The method of  claim 26  wherein the improvement in muscle strength is an improvement in respiratory muscle strength. 
     
     
         28 . The method of  claim 24  or  claim 25  wherein the improvement in muscle function is an improvement in motor stability, improved upper limb strength, or improved cardiac function. 
     
     
         29 . The method of  claim 28  wherein the improvement in motor stability results in an improved six-minute walk test by the patient relative to a previously measured six-minute walk test by that patient. 
     
     
         30 . A kit comprising the antisense oligonucleotide of any one of  claims 1 - 15 , optionally in a container, and a package insert, package label, instructions or other labeling. 
     
     
         31 . The kit of  claim 30 , further comprising an additional oligonucleotide, wherein the additional oligonucleotide specifically hybridizes to an exon in a gamma sarcoglycan RNA.