EXTRA-HEPATIC DELIVERY IRNA COMPOSITIONS AND METHODS OF USE THEREOF
The present invention provides double stranded ribonucleic acid (dsRNA) agents for inhibiting expression of a target gene, comprising an antisense strand which is complementary to the target gene; a sense strand which is complementary to the antisense strand and forms a double stranded region with the antisense strand; and one or more C 22 hydrocarbon chains conjugated to one or more internal positions on at least one strand, compositions comprising such dsRNA agents, and methods of use thereof for treating a subject having a skeletal muscle disorder, a cardiac muscle disorder, or an adipose tissue disorder.
1 . A double stranded ribonucleic acid (dsRNA) agent for inhibiting expression of a target gene, comprising
an antisense strand which is complementary to the target gene;
a sense strand which is complementary to the antisense strand and forms a double stranded region with the antisense strand; and
one or more C 22 hydrocarbon chains conjugated to one or more internal positions on at least one strand.
2 . The dsRNA agent of claim 1 , wherein the C 22 hydrocarbon chain is saturated or unsaturated.
3 . The dsRNA agent of claim 1 , wherein the C 22 hydrocarbon chain is linear or branched
4 .- 8 . (canceled)
9 . The dsRNA agent of claim 1 , wherein the one or more C 22 hydrocarbon chains are conjugated to one or more of the following internal positions: positions 4-8 and 13-18 on the sense strand, and positions 6-10 and 15-18 on the antisense strand, counting from the 5′ end of each strand.
10 . (canceled)
11 . The dsRNA agent of claim 9 , wherein the one or more C 22 hydrocarbon chains are conjugated to position 6 on the sense strand, counting from the 5′-end of the sense strand.
12 . The dsRNA agent of claim 1 , wherein the sense the antisense strands are each independently 15-30 nucleotides in length; 19 to 25 nucleotides in length; or 21 to 23 nucleotides in length.
13 . The dsRNA agent of claim 1 , wherein the one or more C 22 hydrocarbon chains is an aliphatic, alicyclic, or polyalicyclic compound.
14 . (canceled)
15 . The dsRNA agent of claim 1 , wherein the one or more C 22 hydrocarbon chains is a C 22 acid, a C 22 alcohol, or a C 22 amide.
16 .- 20 . (canceled)
21 . The dsRNA agent of claim 1 , wherein the one or more C 22 hydrocarbon chains is conjugated via a carrier that replaces one or more nucleotide(s) in the internal position(s).
22 . (canceled)
23 . The dsRNA agent of claim 1 , wherein the one or more C 22 hydrocarbon chains is conjugated to the dsRNA agent via a linker containing an ether, thioether, urea, carbonate, amine, amide, maleimide-thioether, disulfide, phosphodiester, sulfonamide linkage, a product of a click reaction, or carbamate.
24 . The dsRNA agent of claim 1 , wherein the one or more C 22 hydrocarbon chains is conjugated to the dsRNA agent via a linker or a carrier or via internucleotide phosphate linkage.
25 . The dsRNA agent of claim 1 , wherein the dsRNA agent comprises at least one single-stranded overhang.
26 . (canceled)
27 . The dsRNA agent of claim 1 , wherein at least one end of the dsRNA agent is blunt-ended.
28 . The dsRNA agent of claim 1 , wherein the one or more C22 hydrocarbon chains is conjugated to a nucleobase, sugar moiety, or internucleosidic phosphate linkage.
29 . (canceled)
30 . (canceled)
31 . The dsRNA agent of claim 1 , further comprising a modified phosphate at the 5′-end of the antisense strand.
32 . The dsRNA agent of claim 1 , wherein the one or more C 22 hydrocarbon chains or targeting ligand is conjugated via a biocleavable linker selected from the group consisting of a DNA linker, an RNA linker, a disulfide linker, an amide linker, a protease cleavable peptide linker, a functionalized monosaccharide linker, a galactosamine oligosaccharide linker, a glucosamine linker, a glucose linker, a galactose linker, a mannose linker, and combinations thereof.
33 . The dsRNA agent of claim 1 , wherein the 3′ end of the sense strand is protected via an end cap which is a cyclic group having an amine, said cyclic group being selected from the group consisting of pyrrolidinyl, pyrazolinyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, piperidinyl, piperazinyl, [1,3]dioxolanyl, oxazolidinyl, isoxazolidinyl, morpholinyl, thiazolidinyl, isothiazolidinyl, quinoxalinyl, pyridazinonyl, tetrahydrofuranyl, and decalinyl.
34 . The dsRNA agent of claim 1 , wherein the target gene is selected from the group consisting of adrenoceptor beta 1 (ADRB1); calcium voltage-gated channel subunit alpha1 C (CACNA1C); calcium voltage-gated channel subunit alpha1 G (CACNA1G) (T type calcium channel); angiotensin II receptor type 1(AGTR1); Sodium Voltage-Gated Channel Alpha Subunit 2 (SCN2A); Hyperpolarization Activated Cyclic Nucleotide Gated Potassium Channel 1 (HCN1); Hyperpolarization Activated Cyclic Nucleotide Gated Potassium Channel 4 (HCN4); Hyperpolarization Activated Cyclic Nucleotide Gated Potassium Channel 3 (HCN3); Potassium Voltage-Gated Channel Subfamily A Member 5 (KCNA5); Potassium Inwardly Rectifying Channel Subfamily J Member 3 (KCNJ3); Potassium Inwardly Rectifying Channel Subfamily J Member 4 (KCNJ4); phospholamban (PLN); calcium/calmodulin dependent protein kinase II delta (CAMK2D); Phosphodiesterase 1 (PDE1), Delta 4-Desaturase, Sphingolipid 1 (DEGS1): leptin (LEP); folliculin (FLCN): Zinc Finger Protein 423 (ZFP423); Cyclin Dependent Kinase 6 (CDK6); Regulatory Associated Protein Of MTOR Complex 1 (RPTOR); Mechanistic Target Of Rapamycin Kinase, (mTOR): Forkhead Box P1 (FOXP1); Phosphodiesterase 3B (PDE3B); Activin A Receptor Type 1C (ACVR1C), myostatin (MSTN); Cholinergic Receptor Nicotinic Alpha 1 Subunit (CHRNA1): Cholinergic Receptor Nicotinic Beta 1 Subunit (CHRNB11): Cholinergic Receptor Nicotinic Delta Subunit (CHRND): Cholinergic Receptor Nicotinic Epsilon Subunit (CHRNE); Cholinergic Receptor Nicotinic Gamma Subunit (CHRNG): Collagen Type XIII Alpha 1 Chain (COL13A1): Docking Protein 7 (DOK7): LDL Receptor Related Protein 4 (LRP4): Muscle Associated Receptor Tyrosine Kinase (MUSK): Receptor Associated Protein Of The Synapse (RAPSN): Sodium Voltage-Gated Channel Alpha Subunit 4 (SCN4A); and Double Homeobox 4 (DUX4).
35 . (canceled)
36 . (canceled)
37 . A cell containing the dsRNA agent of claim 1 .
38 . A pharmaceutical composition for inhibiting expression of the target gene, comprising the dsRNA agent of claim 1 .
39 . A method of inhibiting expression of a target gene in a skeletal muscle cell, a cardiac muscle cell, or an adipocyte, comprising contacting the cell with a dsRNA agent that inhibits expression of a target gene, wherein the dsRNA agent comprises
an antisense strand which is complementary to the target gene;
a sense strand which is complementary to the antisense strand and forms a double stranded region with the antisense strand; and
one or more C 22 hydrocarbon chains conjugated to one or more internal positions on at least one strand.
40 .- 75 . (canceled)
76 . A method of treating a subject having a skeletal muscle disorder, a cardiac muscle disorder, or an adipose tissue disorder, comprising:
administering to the subject a therapeutically effective amount of the dsRNA agent of claim 1 , thereby treating the subject.
77 .- 84 . (canceled)
85 . The method of claim 76 , further comprising administering to the subject an additional agent or a therapy suitable for treatment or prevention of a cardiac muscle disorder, or an adipose tissue disorder.
86 . (canceled)
87 . (canceled)
88 . A method of synthesizing a nucleoside monomer having the structure of Formula (I):
wherein:
B is a modified or unmodified nucleobase;
R 1 is a hydroxyl protecting group;
R 2 is H or phosphoramidite;
R 3 is C 22 H 45
wherein the compound of formula (I) is free or substantially free of a compound of Formula (II)
wherein:
B is a modified or unmodified nucleobase;
R 1 is a hydroxyl protecting group;
R 2 is C 22 H 45
R 3 is H
89 . (canceled)