Compositions and methods for modulating dysferlin expression
Aspects of the disclosure relate to methods of altering RNA splicing in a subject. In some embodiments, methods are provided for correcting splicing in a cell that contains a DYSF gene having a mutation that results in defective splicing.
1. A method of modulating splicing in a cell that contains a DYSF gene comprising a c.4886+1249 (G>T) mutation, the method comprising:
delivering to the cell an antisense nucleic acid that targets a pre-messenger RNA expressed from the DYSF gene and alters splicing of the pre-messenger RNA such that exons 44 and 45 of the pre-messenger RNA are spliced together without an intervening pseudoexon, wherein the antisense nucleic acid is complementary to the entirety of SEQ ID NO: 56 or SEQ ID NO: 58.
2. The method of claim 1 , wherein the cell is heterozygous for the c.4886+1249 (G>T) point mutation.
3. The method of claim 1 , wherein the cell is homozygous for the c.4886+1249 (G>T) point mutation.
4. The method of claim 2 , wherein the cell contains a second DYSF gene encoding a wild-type DYSF protein.
5. The method of claim 2 , wherein the cell contains a second DYSF gene comprising a mutation that causes a premature stop codon.
6. The method of claim 1 , wherein the cell is a human cell.
7. The method of claim 6 , wherein the human cell is from a subject having a muscular dystrophy that is associated at least in part with the DYSF gene comprising the c.4886+1249 (G>T) mutation.
8. The method of claim 7 , wherein the muscular dystrophy is of the Miyoshi Myopathy-type, or other muscular dystrophy caused by abnormal expression of the dysferlin gene product.
9. The method of claim 1 , wherein the antisense nucleic acid comprises at least one modified nucleotide or at least one modified internucleotide linkage.
10. The method of claim 1 , wherein the antisense nucleic acid is expressed from a transgene.
11. The method of claim 10 , wherein the transgene is delivered to the cell using a viral vector.
12. The method of claim 11 , wherein the viral vector is a recombinant AAV vector.
13. A oligonucleotide of comprising a region of complementarity that is complementary with the entirety of SEQ ID NO: 56 or SEQ ID NO: 58, wherein the oligonucleotide comprises at least one modified nucleotide or at least one modified internucleotide linkage.
14. A composition comprising the oligonucleotide of claim 13 further comprising a carrier.
15. A pharmaceutical composition comprising the oligonucleotide of claim 13 further comprising a pharmaceutically-acceptable carrier.
16. The method of claim 1 , wherein the antisense nucleic acid comprises SEQ ID NO: 55.
17. The method of claim 1 , wherein the antisense nucleic acid comprises SEQ ID NO: 57.