Compounds and methods for modulating expression apob

Abstract

The present disclosure describes short antisense compounds, including such compounds comprising chemically-modified high-affinity monomers 8-16 monomers in length. Certain such short antisense compound are useful for the reduction of target nucleic acids and/or proteins in cells, tissues, and animals with increased potency and improved therapeutic index. Thus, provided herein are short antisense compounds comprising high-affinity nucleotide modifications useful for reducing a target RNA in vivo. Such short antisense compounds are effective at lower doses than previously described antisense compounds, allowing for a reduction in toxicity and cost of treatment. In addition, the described short antisense compounds have greater potential for oral dosing.

1 . A short antisense compound 8 to 16 monomers in length, comprising a 2′-deoxyribonucleotide gap region flanked on each side by a wing, wherein each wing independently comprises 1 to 3 high-affinity modified monomers and wherein the short antisense compound is targeted to a nucleotide encoding ApoB. 
     
     
         2 . The short antisense compound of  claim 1 , wherein said high-affinity modified monomers are sugar-modified nucleotides. 
     
     
         3 . The short antisense compound of  claim 2 , wherein at least one of the sugar-modified nucleotides comprises a bridge between the 4′ and the 2′ position of the sugar. 
     
     
         4 . The short antisense compound of  claim 2 , wherein each of said high-affinity modified nucleotides confers a ΔT m  of 1 to 4 degrees per nucleotide. 
     
     
         5 . The short antisense compound of  claim 2 , wherein each of said sugar-modified nucleotides comprises a 2′-substituent group that is other than H or OH. 
     
     
         6 . The short antisense compound of  claim 5 , wherein at least one of said sugar-modified nucleotides is a 4′ to 2′ bridged bicyclic nucleotide. 
     
     
         7 . The short antisense compound of  claim 5 , wherein each of the 2′-substituent groups is, independently, alkoxy, substituted alkoxy, or halogen. 
     
     
         8 . The short antisense compound of  claim 7 , wherein each of the 2′-substituent groups is OCH 2 CH 2 OCH 3 . 
     
     
         9 . The short antisense compound  claim 3 , wherein the conformation of each of said sugar-modified nucleotides is, independently, β-D or α-L. 
     
     
         10 . The short antisense compound  claim 5 , wherein each of said bridges independently comprises 1 or from 2 to 4 linked groups independently selected from —[C(R 1 )(R 2 )] n —, —C(R 1 )═C(R 2 )—, —C(R 1 )═N—, —C(═NR 1 )—, —C(═O)—, —C(═S)—, —O—, —Si(R 1 ) 2 —, —S(═O) x — and —N(R 1 )—;
 wherein
 x is 0, 1, or 2; 
 n is 1, 2, 3, or 4; 
 each R 1  and R 2  is, independently, H, a protecting group, hydroxyl, C 1 -C 12  alkyl, substituted C 1 -C 12  alkyl, C 2 -C 12  alkenyl, substituted C 2 -C 12  alkenyl, C 2 -C 12  alkynyl, substituted C 2 -C 12  alkynyl, C 5 -C 20  aryl, substituted C 5 -C 20  aryl, heterocycle radical, substituted heterocycle radical, heteroaryl, substituted heteroaryl, C 5 -C 7  alicyclic radical, substituted C 5 -C 7  alicyclic radical, halogen, OJ 1 , NJ 1 J 2 , SJ 1 , N 3 , COOJ 1 , acyl (C(═O)—H), substituted acyl, CN, sulfonyl (S(═O) 2 -J 1 ), or sulfoxyl (S(═O)-J 1 ); and 
 each J 1  and J 2  is, independently, H, C 1 -C 12  alkyl, substituted C 1 -C 12  alkyl, C 2 -C 12  alkenyl, substituted C 2 -C 12  alkenyl, C 2 -C 12  alkynyl, substituted C 2 -C 12  alkynyl, C 5 -C 20  aryl, substituted C 5 -C 20  aryl, acyl (C(═O)—H), substituted acyl, a heterocycle radical, a substituted heterocycle radical, C 1 -C 12  aminoalkyl, substituted C 1 -C 12  aminoalkyl or a protecting group. 
 
 
     
     
         11 . The short antisense compound of  claim 10 , wherein each of said bridges is, independently, 4′-CH 2 -2′, 4′-(CH 2 ) 2 -2′, 4′-CH 2 —O-2′, 4′-(CH 2 ) 2 —O-2′, 4′-CH 2 —O—N(R 1 )-2′ and 4′-CH 2 —N(R 1 )—O-2′- wherein each R 1  is, independently, H, a protecting group or C 1 -C 12  alkyl. 
     
     
         12 . The short antisense compound of  claim 1 , wherein each of the high-affinity modified monomer is independently selected from bicyclic nucleotides or other 2′-modified nucleotides. 
     
     
         13 . The short antisense compound of  claim 12 , wherein the 2′-modified nucleotides are selected from halogen, allyl, amino, azido, thio, O-allyl, O—C 1 -C 10  alkyl, —OCF 3 , O—(CH 2 ) 2 —O—CH 3 , 2′-O(CH 2 ) 2 SCH 3 , O—(CH 2 ) 2 —O—N(R m )(R n ) or O—CH 2 —C(═O)—N(R m )(R n ), where each R m  and R n  is, independently, H or substituted or unsubstituted C 1 -C 10  alkyl. 
     
     
         14 . The short antisense compound of  claim 13 , wherein the 2′-modified nucleotide is a 2′-OCH 2 CH 2 OCH 3  nucleotide. 
     
     
         15 . The short antisense compound of  claim 1 , wherein at least one monomeric linkage is a modified monomeric linkage. 
     
     
         16 . The antisense compound of  claim 15 , wherein the modified monomeric linkage is a phosphorothioate linkage. 
     
     
         17 . The short antisense compound of  claim 1 , wherein each monomeric linkage is a phosphorothioate internucleoside linkage. 
     
     
         18 . The short antisense compound of  claim 1 , that is 8-15 monomers in length. 
     
     
         19 - 37 . (canceled) 
     
     
         38 . The short antisense compound of  claim 1 , having a motif selected from 1-12-1; 3-10-3; 2-10-3; 2-10-2; 1-10-1; 1-10-2; 3-8-3; 2-8-2; 1-8-1; 3-6-3; and 1-6-11 wherein, the first number represents the number of monomers in the 5′-wing, the second number represents the number of monomers in the gap, and the third number represents the number of monomers in the 3′ wing. 
     
     
         39 . (canceled) 
     
     
         40 . The short antisense compound of  claim 1  having:
 (a) a motif selected from 1-1-10-2, 1-1-8-2, 1-1-6-3, and 1-2-8-2, wherein the first number represents the number of monomers in a first 5′ wing, the second number represents the number of monomers in a second 5′ wing, the third number represents the number of monomers in the gap, and the fourth number represents the number of monomers in the 3′ wing; 
 (b) a motif selected from 2-10-1-1, 2-8-1-1, 3-6-1-1, and 2-8-2-1, wherein the first number represents the number of monomers in the 5′ wing, the second number represents the number of monomers in the gap, the third number represents the number of monomers in a first 3′ wing, and the fourth number represents the number of monomers in a second 3′ wing; or 
 (c) a motif selected from 1-2-10-1-1; 1-1-8-1-1; 2-1-6-1-1; and 1-2-8-2-1, wherein the first number represents the number of monomers in a first 5′ wing, the second number represents the number of monomers in a second 5′ wing, the third number represents the number of monomers in the gap, the fourth number represents the number of monomers in a first 3′ wing and the fifth number represents the number of monomers in a second 3′wing. 
 
     
     
         41 . (canceled) 
     
     
         42 . (canceled) 
     
     
         43 . A method of modulating expression of a ApoB by contacting a nucleic acid encoding ApoB with a short antisense compound. 
     
     
         44 . (canceled) 
     
     
         45 . (canceled) 
     
     
         46 . (canceled) 
     
     
         47 . The method of  claim 1 , wherein the short antisense compound is a short antisense compound 8 to 16 monomers in length, comprising a 2′-deoxyribonucleotide gap region flanked on each side by a wing, wherein each wing independently comprises 1 to 3 high-affinity modified monomers and wherein the short antisense compound is targeted to a nucleotide encoding ApoB. 
     
     
         48 . Use of the short antisense compound of  claim 1  for the preparation of a medicament for reducing the expression of ApoB RNA in an animal. 
     
     
         49 . The use of  claim 48 , wherein the medicament decreases total serum cholesterol, serum LDL, serum VLDL, serum HDL, serum triglycerides, serum apolipoprotein(a) and/or free fatty acids in an animal. 
     
     
         50 . A method of inhibiting expression of ApoB RNA in an animal, comprising administering to said animal the short antisense compound of  claim 1 . 
     
     
         51 . A method of treating a cardiovascular disorder in an animal, comprising administering to an animal in need of such therapy the short antisense compound of  claim 1 .