Backbone modifications to modulate oligonucleotide targeting in vivo

Abstract

The present invention provides antisense compounds and methods for modulating the expression of target genes expressed in the kidney. In particular, this invention provides antisense oligonucleotide compounds optimized for targeting nucleic acid molecules expressed in the kidney. Such compounds are shown herein to efficiently modulate the expression of target genes PTEN, SGLT2 and connective tissue growth factor (CTGF) in the kidney.

1 . A method of modulating pharmacokinetic and pharmacodynamic properties of a gapmer antisense compound by modulating the number of phosphodiester linkages.  
     
     
         2 . The method of  claim 2 , wherein the gapmer comprises two wings and wherein the antisense compound has at least one phosphodiester linkage.  
     
     
         3 . The method of  claim 2 , wherein the at least one phosphodiester linkage is in a wing.  
     
     
         4 . The method of  claim 3 , wherein the wings have one or more regions of alternating phosphorothioate and phosphodiester linkages in the wings.  
     
     
         5 . The method of  claim 4 , wherein each region comprise up to 2 nucleobases.  
     
     
         6 . The method of  claim 4 , wherein each region comprises 1 nucleobase.  
     
     
         7 . The method of  claim 1 , wherein the gapmer antisense compound comprises a first central region consisting of at least 5 contiguous 2′-deoxy nucleosides flanked by a second 5′ region and a third 3′ region, each of said second and third regions independently consisting of at least one 2′-O-methoxyethyl nucleoside.  
     
     
         8 . The method of  claim 1 , wherein the pharmacodynamic property is tissue distribution.  
     
     
         9 . The method of  claim 1 , wherein the pharamacodynamic property is serum protein binding.  
     
     
         10 . A method of enhancing antisense inhibition of expression of a preselected cellular RNA target in a kidney cell or kidney tissue comprising contacting a kidney cell or tissue with an antisense compound 8 to 80 nucleobases in length which is substantially complementary to a preselected cellular RNA target, 
 wherein said antisense compound comprises a first central region consisting of at least 5 contiguous 2′-deoxy nucleosides flanked by a second 5′ region and a third 3′ region, each of said second and third regions independently consisting of at least one 2′-O-methoxyethyl nucleoside, and wherein the internucleoside linkages of the first region are phosphorothioate linkages and the internucleoside linkages of the second and third regions comprise at least one phosphodiester linkage, so that expression of said RNA target is inhibited.    
     
     
         11 . The method of  claim 10 , wherein the antisense compound is an antisense oligonucleotide.  
     
     
         12 . The method of  claim 10 , wherein the antisense compound comprises 10 to 50 nucleobases.  
     
     
         13 . The method of  claim 10 , wherein the antisense compound comprises 13 to 30 nucleobases.  
     
     
         14 . The method of  claim 10 , wherein the antisense compound comprises 15 to 25 nucleobases.  
     
     
         15 . The method of  claim 10 , wherein the antisense compound comprises 18 to 22 nucleobases.  
     
     
         16 . The method of  claim 10 , wherein the antisense compound has at least 70% complementarity with a nucleic acid molecule encoding said preselected cellular RNA target.  
     
     
         17 . The method of  claim 10 , wherein the antisense compound has at least 80% complementarity with a nucleic acid molecule encoding said preselected cellular RNA target.  
     
     
         18 . The method of  claim 10 , wherein the antisense compound has at least 90% complementarity with a nucleic acid molecule encoding said preselected cellular RNA target.  
     
     
         19 . The method of  claim 10 , wherein the antisense compound has at least 95% complementarity with a nucleic acid molecule encoding said preselected cellular RNA target.  
     
     
         20 . A gapped oligomeric compound comprising: 
 a contiguous sequence of nucleosides linked by internucleoside linking groups comprising an internal region of β-D-deoxyribonucleosides linked by phosphorothioate internucleoside linkages and flanked on each side by external regions of 2′-sugar modified nucleosides;    the oligomeric compound further comprising phosphorothioate internucleoside linkages at least between the two nucleosides located at the 5′ terminus, the two nucleosides located at the 3′ terminus and between the junctions located between each external region and the internal region; and    wherein each of the external regions independently comprises at least one phosphodiester internucleoside linkage.    
     
     
         21 . The gapped oligomeric compound of  claim 20  wherein each of the external regions comprises at least two phosphodiester internucleoside linkages.  
     
     
         22 . The gapped oligomeric compound of  claim 20  wherein each external region has alternating phosphorothioate and phosphodiester internucleoside linkages.  
     
     
         23 . The gapped oligomeric compound of  claim 20  wherein each of the 2′-sugar modified nucleosides comprises a 2′-substituent group, independently, selected from halo, amino, azido, O-allyl, O—C1-10 alkyl, OCF3, O—(CH2)2-O—CH3, O(CH2)2SCH3, O—(CH2)2-O—N(Rm)(Rn) or O—CH2-C(═O)—N(Rm)(Rn), wherein each Rm and Rn is, independently, H, an amino protecting group or substituted or unsubstituted C1-10 alkyl.  
     
     
         24 . The gapped oligomeric compound of  claim 23  wherein each 2′-substituent group is, independently, fluoro, O—CH3, OCF3 or O—(CH2)2-O—CH3.  
     
     
         25 . The gapped oligomeric compound of  claim 24  wherein each 2′-substituent group is O—(CH2)2-O—CH3.  
     
     
         26 . The gapped oligomeric compound of  claim 20  wherein each external region is, independently from 3 to about 6 nucleosides in length and the internal region is from about 7 to about 14 nucleosides in length.  
     
     
         27 . The gapped oligomeric compound of  claim 20  wherein each external region is, independently from 3 to about 5 nucleosides in length and the internal region is from about 7 to about 12 nucleosides in length.  
     
     
         28 . The gapped oligomeric compound of  claim 20  having from about 12 to about 30 nucleosides in length.  
     
     
         29 . The gapped oligomeric compound of  claim 20  having from about 12 to about 24 nucleosides in length.  
     
     
         30 . The gapped oligomeric compound of  claim 20  having the formula:  
         5′-M s (M j ) n M s -(N s ) r -(M j ) m M s M-3′ wherein:    each M is a is a 2′-sugar modified nucleoside;    each N is a β-D-deoxyribonucleoside;    each s is a phosphorothioate internucleoside linkage;    each j is, independently, a phosphorothioate or phosphodiester internucleoside linkage;    n and m are each, independently, from 1 to 4;    r is from about 6 to about 14; and    wherein at least one j in each external region is a phosphodiester internucleoside linkage.    
     
     
         31 . The gapped oligomeric compound of  claim 30  wherein n and m are each 1.  
     
     
         32 . The gapped oligomeric compound of  claim 31  wherein n and m are each, independently, from 2 to 4.  
     
     
         33 . The gapped oligomeric compound of  claim 30  wherein n and m are each, independently, 3 and each external region independently comprises two phosphodiester internucleoside linkages.  
     
     
         34 . A method of preventing or delaying the onset of a disease or condition in an animal, wherein said disease or condition is associated with expression of a preselected cellular RNA target expressed in the kidney, said method comprising administering to an animal an effective amount of the antisense compound of  claim 20 .  
     
     
         35 . The method of  claim 34 , wherein said disease or condition is a metabolic disease or condition.  
     
     
         36 . The method of  claim 34 , wherein said disease or condition is diabetes.  
     
     
         37 . The method of  claim 34 , wherein said disease or condition is type 2 diabetes.