DYSTROPHIN EXON SKIPPING OLIGONUCLEOTIDES
Provided herein are antisense oligonucleotides (AONs) that induce skipping of exon 51 of human dystrophin pre-mRNA, and pharmaceutically acceptable derivatives thereof. Also provided are pharmaceutical compositions containing the AONs and methods of using the AONs and compositions for treating a subject with Duchenne muscular dystrophy.
1 . An AON comprising or consisting of the sequence of any one of AON #1-93 which contains at least one modification.
2 . The AON of claim 1 wherein all nucleotides are RNA.
3 . The AON of claim 1 , wherein the modification is a chemical modification of the sugar moiety of all nucleotides in the AON.
4 . The AON of claim 3 , wherein the chemical modification of the sugar moiety of all nucleotides in the AON is 2′-MOE.
5 . The AON of claim 4 , wherein the chemical modification of the sugar moiety is 1, 2, 3 or 4 LNAs, and the remaining nucleotides are 2′-MOE.
6 . The AON of claim 5 , wherein the chemical modification of the sugar moiety of the two 5′ terminal nucleotide positions of the AON and the two 3′ terminal nucleotide positions of the AON are all LNA, and the remaining nucleotides are 2′-MOE.
7 . The AON of claim 3 , wherein the chemical modification of the sugar moiety of all nucleotides in the AON is 2′-OMe.
8 . The AON of claim 3 , wherein the chemical modification of the sugar moiety is 1, 2, 3 or 4 LNAs, and the remaining nucleotides are 2′-OMe.
9 . The AON of claim 8 , wherein the chemical modification of the sugar moiety of the two 5′ terminal nucleotide positions of the AON and the two 3′ terminal nucleotide positions of the AON are all LNA, and the remaining nucleotides are 2′-OMe.
10 . The AON of claim 3 , wherein the chemical modification of the sugar moiety of one or more nucleotides in the AON is a morpholine (PMO, PPMO, PMO-X), a peptide derivative (PNA), a boron-cluster modified PNA, a pyrrolidine-based oxy-peptide nucleic acid (POPNA), a glycol- or glycerol-based nucleic acid (GNA), a threose-based nucleic acid (TNA), an acyclic threoninol-based nucleic acid (aTNA), a cationic morpholino-based oligomer (PMOPlus), an oligonucleotide with integrated bases and backbones (ONIBs), a pyrrolidine-amide oligonucleotides (POMs); or a derivative thereof.
11 . The AON of claim 1 , wherein the modification is a chemical modification of the base moiety of 1, 2, 3, 4 or all nucleotides in the AON.
12 . The AON of claim 11 , wherein all cytosine bases are replaced with 5-methylcytosine.
13 . The AON of claim 11 , wherein all thymine bases are replaced with uracil.
14 . The AON of claim 1 , wherein the backbone is a fully phosphorothioate backbone linkage.
15 . The AON of claim 1 , wherein the AON comprises a hydroxyalkoxy group at the 5′ terminus of the AON, the 3′ terminus of the AON, or at both the 5′ and 3′ ends of the AON.
16 . The AON of claim 15 , wherein the hydroxyalkoxy group comprises or consists of an ethylene glycol monomer, ethylene glycol oligomer or ethylene glycol polymer (also known as polyethylene glycol, PEG).
17 . The AON of claim 15 , wherein the hydroxyalkoxy group is a TEG or a HEG.
18 . The AON of claim 1 , comprising or consisting of the sequence of AON #33, 34, 35, 36, 37, 38 or 39.
19 . The AON of claim 1 , comprising or consisting of the sequence of AON #33, 38 or 39.
20 . The AON of claim 1 , wherein one or two nucleotides are omitted from the 5′ terminus of the AON, or where one or two nucleotides are omitted from the 3′ terminus of the AON, or where one nucleotide is omitted from the 5′ terminus of the AON and one nucleotide is omitted from the 3′ terminus of the AON.
21 . The AON of claim 1 that is 16 to 30 nucleotides in length.
22 . The AON of claim 1 that is 16, 17, 18, 19 or 20 nucleotides in length.
23 . The AON of claim 1 that is 18 nucleotides in length.
24 . The AON of claim 1 , that is fully 2′-MOE RNA modified, wherein all cytosines are replaced with 5-methylcytosine, and wherein the backbone is a fully phosphorothioate backbone.
25 . The AON of claim 1 , wherein all cytosines are replaced with 5-methylcytosine, the two 5′ terminal nucleotides of the AON are LNA, the two 3′ terminal nucleotides of the AON are LNA, the remaining nucleotides are 2′-MOE RNA modified, and wherein the backbone is a fully phosphorothioate backbone.
26 . The AON of claim 1 , that is fully 2′-OMe RNA modified, wherein all cytosines are replaced with 5-methylcytosine, and wherein the backbone is a fully phosphorothioate backbone.
27 . The AON of claim 1 , wherein all cytosines are replaced with 5-methylcytosine, the two 5′ terminal nucleotides of the AON are LNA, the two 3′ terminal nucleotides of the AON are LNA, the remaining nucleotides are 2′-OMe RNA modified, and wherein the backbone is a fully phosphorothioate backbone.
28 . A pharmaceutical composition, comprising the AON of claim 1 and a pharmaceutically acceptable carrier.
29 . A method of treating a subject having DMD, comprising administering to the subject the AON of claim 1 .
30 . A method of delaying the onset of DMD in a subject, comprising administering to the subject the AON of claim 1 .
31 . A method of inducing skipping of exon 51 of human dystrophin pre-mRNA, comprising contacting human dystrophin pre-mRNA with the AON of claim 1 .