Mixed Tricyclo-DNA, 2'-Modified RNA Oligonucleotide Compositions and Uses Thereof

Abstract

In some embodiments, disclosed herein are oligomeric compounds which include one or more tricyclo-deoxyribonucleic acid (tc-DNA) nucleosides and one or more 2′-modified ribonucleic acid (2′-modified-RNA) nucleosides, and which optionally also include one or more non-nucleotides, each of which is joined by a plurality of internucleoside linkages, including pharmaceutical compositions and methods of using the pharmaceutical compositions for the treatment of diseases including Duchenne muscular dystrophy treatment of familial dysautonomia, spinal muscular atrophy, ataxia telangiectasia, congenital disorder of glycosylation, fronto-temporal dementia, Parkinsonism linked to chromosome 17, Niemann-Pick disease type C. neurofibromatosis type 1, neurofibromatosis type 2, megalencephalic leukoencephalopathy with subcortical cysts type 1. Pelizaeus-Merzbacher disease, Pompe disease, myotonic dystrophy type 2 (DM2 or proximal myotonic myopathy), and myotonic dystrophy type 1 (DM1 or Steinert disease).

We claim: 
     
         1 . An oligomeric compound comprising:
 one or more tricyclo-deoxyribonucleic acid (tc-DNA) nucleosides and one or more 2′-modified ribonucleic acid (2′-modified-RNA) nucleosides, and optionally including one or more non-nucleosides;   which are joined by a plurality of internucleoside linkages.   
     
     
         2 . The oligomeric compound of  claim 1 , comprising from about 10 to about 20 nucleotides. 
     
     
         3 . The oligomeric compound of any one of  claim 1  to  claim 2 , wherein the melting temperature (T m ) of the oligomeric compound is at least 50° C. 
     
     
         4 . The oligomeric compound of any one of  claim 1  to  claim 3 , wherein the one or more 2′-modified-RNA nucleosides are incorporated in positions such that self-complementary binding under physiological conditions is prevented. 
     
     
         5 . The oligomeric compound of any one of  claim 1  to  claim 4 , wherein the oligomeric compound is a monomer as determined by non-denaturing gel electrophoresis. 
     
     
         6 . The oligomeric compound of any one of  claim 1  to  claim 5 , wherein the oligomeric compound exhibits a monomer to multimer ratio of at least 100 to 1 as determined by size-exclusion chromatography. 
     
     
         7 . The oligomeric compound of any one of  claim 1  to  claim 6 , wherein the C 3  percentage determined in a complement assay in an in vitro serum is at least 55% (relative to non-activated complement at 100%), wherein the in vitro serum is selected from the group consisting of human, monkey, dog, mouse, and rat serum. 
     
     
         8 . The oligomeric compound of any one of  claim 1  to  claim 7 , wherein the oligomeric compound is not toxic in an animal model at a dose that would be equivalent to a therapeutically effective dose in a human, wherein the animal model is a mouse model or a primate model. 
     
     
         9 . The oligomeric compound of any one of  claim 1  to  claim 8 , wherein the one or more 2′-modified-RNA nucleosides are incorporated in at least one position that forms self-complementary Watson-Crick base pairs. 
     
     
         10 . The oligomeric compound of any one of  claim 1  to  claim 9 , wherein the one or more 2′-modified-RNA nucleosides are incorporated in at least two adjacent positions that form self-complementary Watson-Crick base pairs. 
     
     
         11 . The oligomeric compound of any one of  claim 1  to  claim 10 , wherein the one or more 2′-modified-RNA nucleosides are incorporated at three or more adjacent positions that form self-complementary Watson-Crick base pairs. 
     
     
         12 . The oligomeric compound of any one of  claim 1  to  claim 11 , wherein the oligomeric compound does not contain a direct tc-DNA to tc-DNA phosphorothioate internucleoside linkage. 
     
     
         13 . The oligomeric compound of any one of  claim 1  to  claim 12 , which is complementary to a target sequence. 
     
     
         14 . The oligomeric compound of any one of  claim 1  to  claim 13 , wherein the one or more 2′-modified-RNA nucleosides are 2′-O-methyl-RNA nucleosides. 
     
     
         15 . The oligomeric compound of any one of  claim 1  to  claim 13 , wherein the one or more 2′-modified-RNA nucleosides are 2′-fluoro-RNA nucleosides. 
     
     
         16 . The oligomeric compound of any one of  claim 1  to  claim 13 , wherein the one or more 2′-modified-RNA nucleosides are locked nucleic acid RNA nucleosides. 
     
     
         17 . The oligomeric compound of any one of  claim 1  to  claim 16 , wherein each of the plurality of internucloside linkages is independently selected from a phosphorothioate linkage, a phosphorodithioate linkage, and a phosphorodiester linkage, a phosphotriester linkage, an aminoalkylphosphotriester linkage, a methyl phosphonate linkage, an alkyl phosphonate linkage, a 5′-alkylene phosphonate linkage, a phosphonate linkage, a phosphinate linkage, a phosphoramidate linkage, an 3′-aminophosphoramidate linkage, an aminoalkyl phosphoramidate linkage, a thionophosphoramidate linkage, a thionoalkylphosphonate linkage, a thionoalkylphosphotriester linkage, a selenophosphate linkage, or a boranophosphate linkage. 
     
     
         18 . The oligomeric compound of any one of  claim 1  to  claim 16 , wherein the internucloside linkages comprise a plurality of phosphorothioate linkages and a plurality of phosphorodiester linkages. 
     
     
         19 . The oligomeric compound of  claim 18 , wherein no more than 50% of the plurality of internucleoside linkages are phosphorothioate linkages. 
     
     
         20 . The oligomeric compound of  claim 18 , wherein no more than 33% of the plurality of internucleoside linkages are phosphorothioate linkages. 
     
     
         21 . The oligomeric compound of  claim 18 , wherein no more than 25% of the plurality of internucleoside linkages are phosphorothioate linkages. 
     
     
         22 . The oligomeric compound of  claim 18 , wherein at least 50% of the plurality of internucleoside linkages are phosphorodiester linkages. 
     
     
         23 . The oligomeric compound of  claim 18 , wherein at least 66% of the plurality of internucleoside linkages are phosphorodiester linkages. 
     
     
         24 . The oligomeric compound of  claim 18 , wherein at least 75% of the plurality of internucleoside linkages are phosphorodiester linkages. 
     
     
         25 . The oligomeric compound of any one of  claim 1  to  claim 24  wherein the oligomeric compound comprises a sequence selected from the group consisting of: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO: 43) 
                 
                     
                   5′-A*a*G*a*u*g*GCATTTCTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 44) 
                 
                     
                   5′-AAGATGGCA*u*T*u*C*u*A-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 45) 
                 
                     
                   5′-GAAGATGGCA*u*u*u*c*T-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 46) 
                 
                     
                   5′-AGGAAGATGGCA*u*u*u*c*u*A-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 47) 
                 
                     
                   5′-AGGAAGATGG*c*a*u*u*u*CTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 48) 
                 
                     
                   5′-AGGAAGATGG*c*a*u*u*u*c*u*A-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 49) 
                 
                     
                   5′-A*a*G*a*u*g*GCATTTCTAGTT-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 50) 
                 
                     
                   5′-A*a*g*a*u*g*GCATTTCTAGTT-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 51) 
                 
                     
                   5′-AA*g*a*u*g*GCATTTCTAGTT-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 52) 
                 
                     
                   5′-AGGAAG*a*u*g*GCATTTCTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 53) 
                 
                     
                   5′-AGGA*a*g*a*u*g*GCATTTCTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 54) 
                 
                     
                   5′-GGA*a*g*a*u*g*GCATTTCTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 55) 
                 
                     
                   5′-GGA*a*g*a*u*g*GCATTTCT-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 56) 
                 
                     
                   5′-AGGAA*g*a*u*g*g*CATTTCTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 57) 
                 
                     
                   5′-AGGA*a*g*a*u*g*g*CATTTCTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 58) 
                 
                     
                   5′-AGGAAGATGG*c*a*u*u*u*c*u-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 59) 
                 
                     
                   5′-a*g*a*u*g*GCATTTCTAGTT-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 60) 
                 
                     
                   5′-AGGAAGATGG*c*a*u*u*u*c*u-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 61) 
                 
                     
                   5′-s*s*AAGAuGGCATTTCTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 62) 
                 
                     
                   5′-s*s*s*s*AAGAuGGCATTTCTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 63) 
                 
                     
                   5′-s*s*s*s*s*s*AAGAuGGCATTTCTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 64) 
                 
                     
                   5′-s*s*s*s*s*s*s*s*AAGAuGGCATTTCTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 65) 
                 
                     
                   5′-s*s*s*s*s*s*s*s*s*s*AAGAuGGCATTTCTA-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 66) 
                 
                     
                   5′-A*A*G*A*T*G*G*C*A*u*T*T*C*T*A-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 67) 
                 
                     
                   5′-A*A*G*A*T*G*G*C*a*T*T*T*C*T*A-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 88) 
                 
                     
                   5′-C*A*T*C*C*T*G*G*A*G*T*T*C*C*T-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 89) 
                 
                     
                   5′-C*A*T*C*C*T*G*g*A*G*T*T*C*C*T-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 90) 
                 
                     
                   5′-G*C*C*A*T*C*C*T*G*G*A*G*T*T*C-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 91) 
                 
                     
                   5′-G*C*C*A*T*C*C*T*G*g*A*G*T*T*C-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 92) 
                 
                     
                   5′-C*C*G*C*T*G*C*C*C*A*A*T*G*C*C-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 93) 
                 
                     
                   5′-T*G*C*C*G*C*T*G*C*C*C*A*A*T*G-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 94) 
                 
                     
                   5′-C*T*G*G*A*G*T*T*C*C*T*G*T*A*A-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 95) 
                 
                     
                   5′-C*T*G*g*A*G*T*T*C*C*T*G*T*A*A-3′ 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 96) 
                 
                     
                   5′-T*C*C*T*G*G*A*G*T*T*C*C*T*G*T-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 97) 
                 
                     
                   5′-T*C*C*T*G*g*A*G*T*T*C*C*T*G*T-3′ 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 98) 
                 
                     
                   5′-A*C*T*T*C*A*T*C*C*C*A*C*T*G*A-3′ 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 99) 
                 
                     
                   5′-A*C*T*T*c*A*T*C*C*C*A*C*T*G*A-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 100) 
                 
                     
                   5′-A*T*T*T*C*A*T*T*C*A*A*C*T*G*T-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 101) 
                 
                     
                   5′-G*T*G*T*T*C*T*T*G*T*A*C*T*T*C-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 102) 
                 
                     
                   5′-G*T*G*T*T*C*T*T*G*T*a*C*T*T*C-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 103) 
                 
                     
                   5′-C*T*G*A*A*G*G*T*G*T*T*C*T*T*G-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 104) 
                 
                     
                   5′-C*T*G*A*a*G*G*T*G*T*T*C*T*T*G-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 105) 
                 
                     
                   5′-C*T*C*C*G*G*T*T*C*T*G*A*A*G*G-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 106) 
                 
                     
                   5′-C*T*C*C*G*G*T*T*c*T*G*A*A*G*G-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 107) 
                 
                     
                   5′-T*T*G*A*A*T*C*C*T*T*T*A*A*C*A-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 108) 
                 
                     
                   5′-T*T*G*A*A*T*C*C*T*u*T*A*A*C*A-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 109) 
                 
                     
                   5′-C*T*T*T*C*A*T*A*A*T*G*C*T*G*G-3′, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 110) 
                 
                     
                   5′-C*T*T*T*C*a*T*A*A*T*G*C*T*G*G-3′, 
                 
                     
                   and 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 111) 
                 
                     
                   5′-C*T*T*T*C*A*T*A*A*u*G*C*T*G*G-3′, 
                 
             
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
               
            
           
         
       
       wherein an * between two nucleosides indicates a phosphorothioate, the absence of an * between two nucleosides indicates a phosphorodiester, the capitalized letters A, C, G, and T indicate tc-DNA nucleosides; the lowercase letters a, u, g, and t indicate 2′-O-methyl-RNA nucleosides, the nucleobase at all C positions is 5-methylcytosine, the nucleobase at all c positions is cytosine, and s represents a —O—CH 2 —CH 2 —CH 2 —O— (1,3-propanediol) non-nucleoside. 
     
     
         26 . A pharmaceutical composition comprising an oligonucleotide molecule according to any one of  claim 1  to  claim 25 . 
     
     
         27 . The pharmaceutical composition of  claim 26 , for use in the treatment of a neuromuscular or musculoskeletal disease. 
     
     
         28 . The use of  claim 27 , wherein the neuromuscular or musculoskeletal disease is selected from the group consisting of Duchenne muscular dystrophy, familial dysautonomia, spinal muscular atrophy, ataxia telangiectasia, congenital disorder of glycosylation, fronto-temporal dementia, Parkinsonism linked to chromosome 17, Niemann-Pick disease type C, neurofibromatosis type 1, neurofibromatosis type 2, megalencephalic leukoencephalopathy with subcortical cysts type 1, Pelizaeus-Merzbacher disease, Pompe disease, myotonic dystrophy type 2 (DM2), and myotonic dystrophy type 1 (DM1). 
     
     
         29 . A method for treating a neuromuscular or musculoskeletal disease comprising the step of administering to a patient a therapeutically effective dose of the pharmaceutical composition of  claim 26 . 
     
     
         30 . The method of  claim 29 , wherein the neuromuscular or musculoskeletal disease is selected from the group consisting of Duchenne muscular dystrophy, familial dysautonomia, spinal muscular atrophy, ataxia telangiectasia, congenital disorder of glycosylation, fronto-temporal dementia, Parkinsonism linked to chromosome 17, Niemann-Pick disease type C, neurofibromatosis type 1, neurofibromatosis type 2, megalencephalic leukoencephalopathy with subcortical cysts type 1, Pelizaeus-Merzbacher disease, Pompe disease, myotonic dystrophy type 2 (DM2), and myotonic dystrophy type 1 (DM1).