Chirally-enriched double-stranded rna agents

Abstract

The present invention relates to a chirally-modified dsRNA agent capable of inhibiting the expression of a target gene. The sense and antisense strands of chirally-modified dsRNA agent independently or in combination comprises one or more site specificsite specific/position specific, chirally-modified internucleotide linkages.

We claim: 
     
         1 . A chirally-modified double-stranded RNA (dsRNA) agent capable of inhibiting the expression of a target gene, said chirally-modified dsRNA agent comprising a sense strand and an antisense strand, each strand having 14 to 40 nucleotides,
 wherein:   the antisense strand comprises one or more terminal, chirally-modified internucleotide linkages at the 5′ end and one or more terminal, chirally-modified internucleotide linkages at the 3′ end;   each terminal, chirally-modified internucleotide linkage comprises a chiral linkage phosphorus atom; and   a terminal, chiral modification occurs at the first internucleotide linkage at the 5′ end of the antisense strand, and the chiral linkage phosphorus atom is in Rp configuration.   
     
     
         2 . The chirally-modified dsRNA agent of  claim 1 , wherein the chiral purity with respect to the chiral linkage phosphorus atom for each terminal, chirally-modified internucleotide linkage is at least 50%. 
     
     
         3 . The chirally-modified dsRNA agent of  claim 1 , wherein the chirally-modified dsRNA agent comprises:
 a terminal, chiral modification occurring at the first internucleotide linkage at the 3′ end of the antisense strand, having the linkage phosphorus atom in Sp configuration, and   a terminal, chiral modification occurring at the first internucleotide linkage at the 5′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration.   
     
     
         4 . The chirally-modified dsRNA agent of  claim 1 , wherein the chirally-modified dsRNA agent comprises three or more terminal, chirally-modified internucleotide linkages at the antisense strand. 
     
     
         5 . The chirally-modified dsRNA agent of  claim 4 , wherein the chirally-modified dsRNA agent comprises:
 a terminal, chiral modification occurring at the first and second internucleotide linkages at the 3′ end of the antisense strand, having the linkage phosphorus atom in Sp configuration, and   a terminal, chiral modification occurring at the first internucleotide linkage at the 5′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration.   
     
     
         6 . The chirally-modified dsRNA agent of  claim 4 , wherein the chirally-modified dsRNA agent comprises:
 a terminal, chiral modification occurring at the first internucleotide linkages at the 3′ end of the antisense strand, having the linkage phosphorus atom in Sp configuration,   a terminal, chiral modification occurring at the second internucleotide linkages at the 3′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration, and   a terminal, chiral modification occurring at the first internucleotide linkage at the 5′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration.   
     
     
         7 . The chirally-modified dsRNA agent of  claim 4 , wherein the chirally-modified dsRNA agent comprises:
 a terminal, chiral modification occurring at the first internucleotide linkages at the 3′ end of the antisense strand, having the linkage phosphorus atom in Sp configuration, and   a terminal, chiral modification occurring at the first and second internucleotide linkage at the 5′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration.   
     
     
         8 . The chirally-modified dsRNA agent of  claim 4 , wherein the chirally-modified dsRNA agent comprises:
 a terminal, chiral modification occurring at the first internucleotide linkages at the 3′ end of the antisense strand, having the linkage phosphorus atom in Sp configuration,   a terminal, chiral modification occurring at the first internucleotide linkage at the 5′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration, and   a terminal, chiral modification occurring at the second internucleotide linkage at the 5′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration.   
     
     
         9 . The chirally-modified dsRNA agent of  claim 1 , wherein the chirally-modified dsRNA agent comprises four or more terminal, chirally-modified internucleotide linkages at the antisense strand. 
     
     
         10 . The chirally-modified dsRNA agent of  claim 9 , wherein the chirally-modified dsRNA agent comprises:
 a terminal, chiral modification occurring at the first, second, and third internucleotide linkages at the 3′ end of the antisense strand, having the linkage phosphorus atom in Sp configuration, and   a terminal, chiral modification occurring at the first internucleotide linkage at the 5′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration.   
     
     
         11 . The chirally-modified dsRNA agent of  claim 9 , wherein the chirally-modified dsRNA agent comprises:
 a terminal, chiral modification occurring at the first and second internucleotide linkages at the 3′ end of the antisense strand, having the linkage phosphorus atom in Sp configuration,   a terminal, chiral modification occurring at the third internucleotide linkages at the 3′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration, and   a terminal, chiral modification occurring at the first internucleotide linkage at the 5′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration.   
     
     
         12 . The chirally-modified dsRNA agent of  claim 9 , wherein the chirally-modified dsRNA agent comprises:
 a terminal, chiral modification occurring at the first internucleotide linkages at the 3′ end of the antisense strand, having the linkage phosphorus atom in Sp configuration,   a terminal, chiral modification occurring at the first and second internucleotide linkages at the 3′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration, and   a terminal, chiral modification occurring at the first internucleotide linkage at the 5′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration.   
     
     
         13 . The chirally-modified dsRNA agent of  claim 9 , wherein the chirally-modified dsRNA agent comprises:
 a terminal, chiral modification occurring at the first and second internucleotide linkages at the 3′ end of the antisense strand, having the linkage phosphorus atom in Sp configuration, and   a terminal, chiral modification occurring at the first and second internucleotide linkages at the 5′ end of the antisense strand, having the linkage phosphorus atom in Rp configuration.   
     
     
         14 . The chirally-modified dsRNA agent of  claim 1 , wherein the chirally-modified dsRNA agent comprises six or more terminal, chirally-modified internucleotide linkages. 
     
     
         15 . The chirally-modified dsRNA agent of  claim 1 , wherein the chirally-modified dsRNA agent comprises eight or more terminal, chirally-modified internucleotide linkages. 
     
     
         16 . The chirally-modified dsRNA agent of  claim 1 , wherein each nucleotide of a dinucleotide connected by the terminal, chirally-modified internucleotide linkage is independently modified with a modification selected from the group consisting of acyclic nucleotides, LNA, HNA, CeNA, 2′-O-methoxyalkyl, 2′ O-methyl, 2′-O-allyl, 2′-C-allyl, 2′-deoxy, 2′-fluoro, 2′-O—N-methylacetamido (2′ O NMA), 2′-O-dimethylaminoethoxyethyl (2′-O-DMAEOE), 2′-O-aminopropyl (2′ O AP), and 2′-ara F. 
     
     
         17 . The chirally-modified dsRNA agent of  claim 1 , wherein each of the sense and antisense strands is independently modified with a modification selected from the group consisting of acyclic nucleotides, LNA, HNA, CeNA, 2′ O-methoxyalkyl, 2′ O-methyl, 2′-O-allyl, 2′-C-allyl, 2′-deoxy, 2′-fluoro, 2′-O—N-methylacetamido (2′-O—NMA), 2′-O-dimethylaminoethoxyethyl (2′-O-DMAEOE), 2′ O aminopropyl (2′-O-AP), and 2′-ara-F. 
     
     
         18 . The chirally-modified dsRNA agent of  claim 1 , wherein each of the sense and antisense strands has 15-30 nucleotides. 
     
     
         19 . The chirally-modified dsRNA agent of  claim 18 , wherein the sense strand has 19-22 nucleotides, and the antisense strand has 19-25 nucleotides. 
     
     
         20 . The chirally-modified dsRNA agent of  claim 1 , further comprising one or more ligands. 
     
     
         21 . The chirally-modified dsRNA agent of  claim 20 , wherein at least one ligand is attached to the 3′ end of the sense strand or antisense strand. 
     
     
         22 . The chirally-modified dsRNA agent of  claim 20 , wherein at least one ligand is attached through one or more cleavable linkers and/or a cyclic or acyclic carrier. 
     
     
         23 . The chirally-modified dsRNA agent of  claim 20 , wherein at least one ligand is an ASGPR ligand. 
     
     
         24 . The chirally-modified dsRNA agent of  claim 23 , wherein the ASGPR ligand is one or more GalNAc derivatives attached through a bivalent or trivalent branched linker. 
     
     
         25 . The chirally-modified dsRNA agent of  claim 24 , wherein the ASGPR ligand is: 
       
         
           
           
               
               
           
         
       
     
     
         26 . The chirally-modified dsRNA agent of  claim 20 , wherein at least one ligand is a lipophilic moiety. 
     
     
         27 . The chirally-modified dsRNA agent of  claim 26 , wherein the lipophilic moiety is selected from the group consisting of a lipid, cholesterol, retinoic acid, cholic acid, adamantane acetic acid, 1-pyrene butyric acid, dihydrotestosterone, 1,3-bis-O(hexadecyl)glycerol, geranyloxyhexyanol, hexadecylglycerol, borneol, menthol, 1,3-propanediol, heptadecyl group, palmitic acid, myristic acid, O3-(oleoyl)lithocholic acid, O3-(oleoyl)cholenic acid, dimethoxytrityl, and phenoxazine. 
     
     
         28 . The chirally-modified dsRNA agent of  claim 26 , wherein the lipophilic moiety contains a saturated or unsaturated C 4 -C 3 o hydrocarbon chain, and an optional functional group selected from the group consisting of hydroxyl, amine, carboxylic acid, sulfonate, phosphate, thiol, azide, and alkyne. 
     
     
         29 . The chirally-modified dsRNA agent of  claim 26 , wherein the lipophilic moiety contains a C 6 -C 30  acid or a C 6 -C 30  alcohol. 
     
     
         30 . The chirally-modified dsRNA agent of  claim 29 , wherein the lipophilic moiety contains a C 6 -C 30  acid selected from the group consisting of hexanoic acid, heptanoic acid, octanoic acid, nonanoic acid, decanoic acid, undecanoic acid, dodcanoic acid, tridecanoic acid, tetradecanoic acid, pentadecanoic acid, hexadecanoic acid, heptadecanoic acid, octadecanoic acid, oleic acid, linoleic acid, arachidonic acid, and cis-4,7,10,13,16,19-docosahexanoic acid; or a C 6 -C 30  alcohol selected from the group consisting of hexanol, heptanol, octanol, nonanol, decanol, undecanol, dodcanol, tridecanol, tetradecanol, pentadecanol, hexadecanol, heptadecanol, octadecanol, oleyl alcohol, linoleyl alcohol, arachidonic alcohol, cis-4,7,10,13,16,19-docosahexanol, retinol, vitamin E, and cholesterol. 
     
     
         31 . A pharmaceutical composition comprising the chirally-modified dsRNA agent of  claim 1  and a pharmaceutically acceptable carrier or excipient. 
     
     
         32 . A method for inhibiting the expression of a target gene comprising the step of administering the chirally-modified dsRNA agent of  claim 1  in an amount sufficient to inhibit expression of the target gene. 
     
     
         33 . The method of  claim 32 , wherein the chirally-modified dsRNA agent is administered through subcutaneous or intravenous administration.