GALNAC COMPOSITIONS FOR IMPROVING SIRNA BIOAVAILABILITY

Abstract

Provided herein is a composition comprising GalNAc moiety that can be conjugated to oligonucleotide.The oligonucleotide can be small interfering RNA or antisense oligonucleotide.Also provided herein is a method of treatment comprising administering the composition to a subject.

Description

[Technical field]

[0001] cross reference This application claims the benefit of U.S. Provisional Patent Application No. 63 / 320,431, filed March 16, 2022, U.S. Provisional Patent Application No. 63 / 354,359, filed June 22, 2022, and U.S. Provisional Patent Application No. 63 / 430,542, filed December 6, 2022, which applications are incorporated by reference herein.

[0002] Incorporation by reference of sequence listing This application is filed with a Sequence Listing in electronic format. The Sequence Listing is provided in a file entitled 54462-735601_PCT.xml, created on March 13, 2023, and is 922,453 bytes in size. The information in the electronic format of the Sequence Listing is incorporated by reference in its entirety. [Background technology]

[0003] Cardiovascular, metabolic, and liver-related disorders are abundant and can affect many different people, and improved therapeutic agents are needed to treat these disorders. Summary of the Invention

[0004] Formula (I) or (II)

[0005] [ka] or a salt thereof, J is an oligonucleotide; Each w is independently selected from any value of 1 to 20; Each v is independently selected from any value from 1 to 20; n is selected from any value between 1 and 20, m is selected from any value between 1 and 20, z is selected from any value between 1 and 3, when z is 3, Y is C; If z is 2, Y is CR 6 or When z is 1, Y is C(R 6 ) 2 and Q is halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , -S(O)R 7 , and C 1-6 C optionally substituted with one or more substituents independently selected from alkyl 3-10 Carbocyclic rings, C 1-6 Alkyl is a halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 and optionally substituted with one or more substituents independently selected from R 1 -O-, -S-, -N(R 7 )-, -C(O)-, -C(O)N(R 7 )-, -N(R 7 )C(O)-, -N(R 7 )C(O)N(R 7 )-, -OC(O)N(R 7 )-, -N(R 7 )C(O)O-, -C(O)O-, -OC(O)-, -S(O)-, -S(O) 2 -, -OS(O) 2 -,-OP(O)(OR 7 )O-, -SP(O)(OR7 )O-, -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, -OP(O)(OR 7 )S-, -OP(O)(O - )O-, -SP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, -OP(O)(O - )S-, -OP(O)(OR 7 )NR 7 -,-OP(O)(N(R 7 ) 2 )NR 7 -,-OP(OR 7 )O-, -OP(N(R 7 ) 2 )O-, -OP(OR 7 )N(R 7 )-, and -OPN(R 7 ) 2 NR 7 -, R 2 are each independently halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6 alkyl, R 3 and R 4 are each independently -OR 7, -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 is selected from R 5 are each independently -OC(O)R 7 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)R 7 , -C(O)OR 7 , and -C(O)N(R 7 ) 2 is selected from R 6 are each independently hydrogen, Halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 , and Halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6 alkyl, R 7 are each independently hydrogen, C 1-6 Alkyl, C 2-6 Alkenyl, and C 2-6 Alkynyl (each of which is halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 3-10 optionally substituted with one or more substituents independently selected from carbocycle, and 3- to 10-membered heterocycle; and C 3-10 Carbocycles and 3- to 10-membered heterocycles, each of which is independently selected from the group consisting of halogen, -CN, -OH, -SH, -NO 2 , -NH2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-10 Carbocyclic rings, 3- to 10-membered heterocyclic rings, and C 1-6 haloalkyl). In some embodiments, each w is independently selected from any value from 1 to 10. In some embodiments, each w is independently selected from any value from 1 to 5. In some embodiments, each w is 1. In some embodiments, each v is independently selected from any value from 1 to 10. In some embodiments, each v is independently selected from any value from 1 to 5. In some embodiments, each v is 1. In some embodiments, n is selected from any value from 1 to 10. In some embodiments, n is selected from any value from 1 to 5. In some embodiments, n is 2. In some embodiments, m is selected from any value from 1 to 10. In some embodiments, m is selected from any value from 1 to 5. In some embodiments, m is selected from 1 and 2. In some embodiments, z is 3 and Y is C. In some embodiments, Q is selected from halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2, -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 5-6 In some embodiments, Q is selected from halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 C optionally substituted with one or more substituents independently selected from 5-6 In some embodiments, Q is selected from phenyl and cyclohexyl, each of which is selected from halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 In some embodiments, Q is selected from phenyl. In some embodiments, Q is selected from cyclohexyl. In some embodiments, R 1 -OP(O)(OR 7 )O-, -SP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, -OP(O)(OR 7 )S-, -OP(O)(O - )O-, -SP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, -OP(O)(O - )S-, -OP(O)(OR 7 )NR 7 -,-OP(O)(N(R 7 ) 2 )NR 7 -,-OP(OR 7 )O-, -OP(N(R 7 ) 2 )O-, -OP(OR 7 )N(R 7 )-, and -OPN(R 7 ) 2 NR 7In some embodiments, R 1 -OP(O)(OR 7 )O-, -SP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, -OP(O)(OR 7 )S-, -OP(O)(O - )O-, -SP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, -OP(O)(O - )S-, and -OP(OR 7 In some embodiments, R 1 -OP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, and -OP(OR 7 In some embodiments, R 1 -OP(O)(OR 7 )O- and -OP(OR 7 In some embodiments, R 2 is halogen, -OR 7 , -OC(O)R 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , and -S(O)R 7 C substituted with one or more substituents independently selected from 1-3 In some embodiments, R 2 -OR 7 , -OC(O)R 7 , -SR 7 , and -N(R 7 ) 2 C substituted with one or more substituents independently selected from 1-3In some embodiments, R 2 -OR 7 and -OC(O)R 7 C substituted with one or more substituents independently selected from 1-3 In some embodiments, R 3 is halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -OC(O)R 7 , and -S(O)R 7 In some embodiments, R 3 -OR 7 , -SR 7 , -OC(O)R 7 , and -N(R 7 ) 2 In some embodiments, R 3 -OR 7 - and -OC(O)R 7 In some embodiments, R 4 is halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -OC(O)R 7 , and -S(O)R 7 In some embodiments, R 4 -OR 7 , -SR 7 , -OC(O)R 7 , and -N(R 7 ) 2 In some embodiments, R 4 -OR 7 - and -OC(O)R 7 In some embodiments, R 5 is -OC(O)R 7 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)R7 , -N(R 7 )C(O)N(R 7 ) 2 , and -N(R 7 )C(O)OR 7 In some embodiments, R 5 is -OC(O)R 7 and -N(R 7 )C(O)R 7 In some embodiments, R 7 are each independently hydrogen and the halogens -CN, -OH, -SH, and -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 3-10 C optionally substituted with one or more substituents independently selected from a carbocycle or a 3- to 10-membered heterocycle 1-6 In some embodiments, R 7 are each independently halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , and -NH(C 1-6 C optionally substituted with one or more substituents independently selected from 1-6 In some embodiments, R 7 each independently represents C optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, and -SH; 1-6 In some embodiments, w is 1, v is 1, n is 2, m is 1 or 2, z is 3, and Y is C and Q is phenyl or cyclohexyl, each of which is selected from halogen, -CN, -OH, -SH, -NO 2 , -NH2 , and C 1-3 optionally substituted with one or more substituents independently selected from alkyl, R 1 -OP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, and -OP(OR 7 )O-, R 2 is -OH or -OC(O)CH 3 C replaced with 1 is alkyl, R 3 is -OH or -OC(O)CH 3 and R 4 is -OH or -OC(O)CH 3 and R 5 -NH(O)CH 3 In some embodiments, the compound is

[0006] [ka]

[0007] [ka]

[0008] [ka]

[0009] [ka]

[0010] [ka] In some embodiments, the oligonucleotide (J) is attached at the 5' or 3' end of the oligonucleotide. In some embodiments, the oligonucleotide comprises DNA. In some embodiments, the oligonucleotide comprises RNA. In some embodiments, the oligonucleotide comprises one or more modified internucleoside linkages. In some embodiments, the one or more modified internucleoside linkages comprise alkyl phosphonates, phosphorothioates, methyl phosphonates, phosphorodithioates, alkyl phosphonothioates, phosphoramidates, carbamates, carbonates, phosphotriesters, acetamidates, or carboxymethyl esters, or combinations thereof. In some embodiments, the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 modified internucleoside linkages. In some embodiments, the oligonucleotide comprises one or more modified nucleosides. In some embodiments, the one or more modified nucleosides include locked nucleic acid (LNA), hexitol nucleic acid (HLA), cyclohexene nucleic acid (CeNA), 2'-methoxyethyl, 2'-O-alkyl, 2'-O-allyl, 2'-O-allyl, 2'-fluoro, or 2'-deoxy, or a combination thereof. In some embodiments, the one or more modified nucleosides include 2',4' constrained ethyl nucleosides, 2'-O-methyl nucleosides, 2'-deoxyfluoro nucleosides, 2'-ON-methylacetamide (2'-O-NMA), 2'-O-dimethylaminoethoxyethyl (2'-O-DMAEOE) nucleosides, 2'-O-aminopropyl (2'-O-AP) nucleosides, 2'-ara-F, 2'fluoro, or 2'O-alkyl, or a combination thereof. In some embodiments, the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, or more modified nucleosides. In some embodiments, the oligonucleotide comprises a lipid attached at the 3' or 5' end of the oligonucleotide.In some embodiments, the lipid comprises cholesterol, myristoyl, palmitoyl, stearoyl, lithocholoyl, docosanoyl, docosahexaenoyl, myristyl, palmitylstearyl, or alpha-tocopherol, or a combination thereof. In some embodiments, the oligonucleotide comprises an arginine-glycine-aspartic acid (RGD) peptide attached at the 3' or 5' end of the oligonucleotide. In some embodiments, the RGD peptide comprises cyclo(-Arg-Gly-Asp-D-Phe-Cys), cyclo(-Arg-Gly-Asp-D-Phe-Lys), cyclo(-Arg-Gly-Asp-D-Phe-Azide), RGD derived from aminobenzoic acid, or a combination thereof. In some embodiments, the oligonucleotide comprises a small interfering RNA (siRNA) comprising a sense strand and an antisense strand. In some embodiments, the sense strand is 12-30 nucleosides in length. In some embodiments, the antisense strand is 12-30 nucleosides in length. In some embodiments, the sense strand and the antisense strand form a double-stranded RNA duplex. In some embodiments, the first base pair of the double-stranded RNA duplex is an AU base pair. In some embodiments, the sense strand or the antisense strand comprises a 3' overhang. In some embodiments, the 3' overhang comprises one, two, or more nucleosides. In some embodiments, the sense strand comprises any one of the 1S to 6S or 1S#2 to 6S#2 modification patterns. In some embodiments, the antisense strand comprises any one of the 1AS to 9AS modification patterns. In some embodiments, the oligonucleotide comprises an antisense oligonucleotide (ASO). In some embodiments, the ASO is 12-30 nucleosides in length. In some embodiments, the ASO comprises the modification pattern ASO1. In some embodiments, the compound binds to an asialoglycoprotein receptor. In some embodiments, the compound targets hepatocytes.

[0011] Disclosed herein is a pharmaceutical composition comprising any one of the compounds described herein and a pharma- ceutically acceptable carrier, excipient, or diluent. In some embodiments, the pharmaceutical composition is sterile. In some embodiments, the pharmaceutical composition comprises a pharma- ceutically acceptable carrier. In some embodiments, the pharma- ceutical composition comprises water, buffer, or saline. In some embodiments, the oligonucleotide targets a target mRNA and reduces target mRNA or target protein by at least 10% when administered to a subject in an effective amount.

[0012] Disclosed herein is a method of decreasing target mRNA or target protein in a subject, comprising administering an effective amount of any one of the pharmaceutical compositions of the compounds described herein. In some embodiments, the effective amount decreases the measurement of target mRNA or target protein in the subject compared to a baseline measurement of target mRNA or target protein. In some embodiments, the effective amount treats a disorder in the subject. In some embodiments, the effective amount decreases the measurement of a symptom or parameter associated with the disorder in the subject compared to a baseline measurement of the symptom or parameter. In some embodiments, the measurement of a symptom or parameter associated with the disorder in the subject is decreased for at least 10 days. In some embodiments, the measurement of a symptom or parameter associated with the disorder in the subject is decreased for at least 100 days. In some embodiments, the disorder comprises a metabolic disorder. In some embodiments, the disorder comprises a liver disorder.

[0013] Formula (A) or (B)

[0014] [ka] or a salt thereof, wherein Each w is independently selected from any value of 1 to 20; Each v is independently selected from any value from 1 to 20; n is selected from any value between 1 and 20, m is selected from any value between 1 and 20, z is selected from any value between 1 and 3, when z is 3, Y is C; If z is 2, Y is CR 6 or When z is 1, Y is C(R 6 ) 2 and Q is halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , -S(O)R 7 , and C 1-6 C optionally substituted with one or more substituents independently selected from alkyl 3-10 Carbocyclic rings, C 1-6 Alkyl is a halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 and optionally substituted with one or more substituents independently selected from R 1 -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7)C(O)R 7 、-N(R 7 )C(O)N(R 7 ) 2 、-OC(O)N(R 7 ) 2 、-N(R 7 )C(O)OR 7 、-C(O)OR 7 、-OC(O)R 7 、-S(O)R 7 、-S(O) 2 R 7 、-OS(O) 2 R 7 、-OP(O)(OR 7 ) 2 、-OP(S)(OR 7 ) 2 、-SP(O)(OR 7 ) 2 、-OP(O)(SR 7 )(OR 7 )、-OP(O)(OR 7 )N(R 7 ) 2 、-OP(S)(OR 7 )N(R 7 ) 2 、-SP(O)(OR 7 )N(R 7 ) 2 、-OP(O)(SR 7 )N(R 7 ) 2 、-OP(O)(N(R 7 ) 2 ) 2 、-OP(S)(N(R 7 ) 2 ) 2 、-SP(O)(N(R 7 ) 2 ) 2 、-OP(OR 7 ) 2 、-SP(OR 7 ) 2 、-OP(OR 7 )(SR 7 )、-OP(OR 7 )N(R 7 ) 2 、-OP(SR 7 )N(R 7 )2 , -SP(OR 7 )N(R 7 ) 2 , -OP(N(R 7 ) 2 ) 2 , and -SP(N(R 7 ) 2 ) 2 is selected from R 2 are each independently halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6 alkyl, R 3 and R 4 are each independently -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7, and -S(O)R 7 is selected from R 5 are each independently -OC(O)R 7 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)R 7 , -C(O)OR 7 , and -C(O)N(R 7 ) 2 is selected from R 6 are each independently hydrogen, Halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 , and Halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7)C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6 alkyl, R 7 are each independently hydrogen, C 1-6 Alkyl, C 2-6 Alkenyl, and C 2-6 Alkynyl (each of which is halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 3-10 optionally substituted with one or more substituents independently selected from carbocycle, and 3- to 10-membered heterocycle; and C 3-10 Carbocycles and 3- to 10-membered heterocycles, each of which is independently selected from the group consisting of halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-10 Carbocyclic rings, 3- to 10-membered heterocyclic rings, and C 1-6 haloalkyl). In some embodiments, each w is independently selected from any value between 1 and 10. In some embodiments, each w is independently selected from any value between 1 and 5. In some embodiments, each w is 1. In some embodiments, each v is independently selected from any value between 1 and 10. In some embodiments, each v is independently selected from any value between 1 and 5. In some embodiments, each v is 1. In some embodiments, n is selected from any value between 1 and 10. In some embodiments, n is selected from any value between 1 and 5. In some embodiments, n is 2. In some embodiments, m is selected from any value between 1 and 10. In some embodiments, m is selected from any value between 1 and 5. In some embodiments, m is selected from 1 and 2. In some embodiments, z is 3 and Y is C. In some embodiments, Q is selected from halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 5-6 In some embodiments, Q is selected from halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 C optionally substituted with one or more substituents independently selected from 5-6In some embodiments, Q is selected from phenyl and cyclohexyl, each of which is selected from halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 In some embodiments, Q is selected from phenyl. In some embodiments, Q is selected from cyclohexyl. In some embodiments, R 1 -OP(O)(OR 7 ) 2 , -OP(O)(OR 7 )N(R 7 ) 2 , -OP(O)(N(R 7 ) 2 ) 2 , -OP(OR 7 ) 2 , -OP(OR 7 )N(R 7 ) 2 , and -OP((NR 7 ) 2 ) 2 In some embodiments, R 1 -OP(O)(OR 7 ) 2 and -OP(OR 7 )N(R 7 ) 2 In some embodiments, R 1 -OP(O)(OCH 2 CH 3 )OH and -OP(OCH 2 CH 2 CN)N(CH(CH 3 ) 2 ) 2 In some embodiments, R 1 -OP(OCH 2 CH 2 CN)N(CH(CH 3 ) 2 ) 2 In some embodiments, R 2 is halogen, -OR 7 , -OC(O)R 7, -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , and -S(O)R 7 C substituted with one or more substituents independently selected from 1-3 In some embodiments, R 2 -OR 7 , -OC(O)R 7 , -SR 7 , and -N(R 7 ) 2 C substituted with one or more substituents independently selected from 1-3 In some embodiments, R 2 -OR 7 and -OC(O)R 7 C substituted with one or more substituents independently selected from 1-3 In some embodiments, R 3 is halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -OC(O)R 7 , and -S(O)R 7 In some embodiments, R 3 -OR 7 , -SR 7 , -OC(O)R 7 , and -N(R 7 ) 2 In some embodiments, R 3 -OR 7 - and -OC(O)R 7 In some embodiments, R 4 is halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -OC(O)R 7 , and -S(O)R 7 In some embodiments, R4 -OR 7 , -SR 7 , -OC(O)R 7 , and -N(R 7 ) 2 In some embodiments, R 4 -OR 7 - and -OC(O)R 7 In some embodiments, R 5 is -OC(O)R 7 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , and -N(R 7 )C(O)OR 7 In some embodiments, R 5 is -OC(O)R 7 and -N(R 7 )C(O)R 7 In some embodiments, R 7 are each independently hydrogen and the halogens -CN, -OH, -SH, and -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 3-10 C optionally substituted with one or more substituents independently selected from a carbocycle or a 3- to 10-membered heterocycle 1-6 In some embodiments, R 7 are each independently halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , and -NH(C 1-6C optionally substituted with one or more substituents independently selected from 1-6 In some embodiments, R 7 each independently represents C optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, and -SH; 1-6 In some embodiments, w is 1, v is 1, n is 2, m is 1 or 2, z is 3, and Y is C and Q is phenyl or cyclohexyl, each of which is selected from halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , and C 1-3 optionally substituted with one or more substituents independently selected from alkyl, R 1 -OP(O)(OR 7 ) 2 and -OP(OR 7 )N(R 7 ) 2 Selected from R 2 is -OH or -OC(O)CH 3 C replaced with 1 is alkyl, R 3 is -OH or -OC(O)CH 3 and R 4 is -OH or -OC(O)CH 3 and R 5 -NH(O)CH 3 In some embodiments, the compound is

[0015] [ka]

[0016] [ka]

[0017] [ka] Includes. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0018] N-acetylgalactosamine (GalNAc) is an amino sugar derivative of galactose. GalNAc and GalNAc-containing moieties can bind to lectins such as asialoglycoprotein receptors. These receptors can be contained on hepatocytes. Thus, GalNAc can target oligonucleotides to hepatocytes or the liver.

[0019] Provided herein is GalNAc moiety.These GalNAc moieties can be conjugated to oligonucleotides, such as small interfering RNA (siRNA) or antisense oligonucleotide (ASO).The oligonucleotides conjugated to GalNAc moiety can be administered to subjects, targeted to liver or liver cells, or used to treat liver-related disorders in subjects.

[0020] I. Composition In some embodiments, a composition is provided herein that includes an oligonucleotide and an N-acetylgalactosamine (GalNAc) moiety. In some embodiments, the composition includes an oligonucleotide. The oligonucleotide can inhibit a target gene or oligonucleotide. The oligonucleotide can bind to a target oligonucleotide. In some embodiments, the composition is used in the methods described herein.

[0021] In some embodiments, the present invention provides a compound comprising an oligonucleotide and a GalNAc moiety. In some embodiments, the compound comprises an oligonucleotide. The oligonucleotide can bind to a target oligonucleotide. In some embodiments, the compound is used in the method described herein. In some embodiments, the compound is included in the composition described herein.

[0022] The oligonucleotides of the compounds or compositions described herein may comprise small interfering RNA (siRNA) or antisense oligonucleotides (ASO).

[0023] Some embodiments include compositions comprising a GalNAc moiety and an oligonucleotide that, when administered to a subject in an effective amount, reduces target mRNA or protein levels in a cell, body fluid, or tissue of the subject. Some embodiments include compositions comprising a GalNAc moiety and an oligonucleotide that, when administered to a subject in an effective amount, reduces target mRNA or protein levels in liver tissue or liver cells. In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide that, when administered to a subject in an effective amount, reduces the level of a target, e.g., mRNA, in a cell or tissue. In some embodiments, the cell is a liver cell. In some embodiments, the tissue is a liver tissue. Some embodiments include compositions comprising a GalNAc moiety and an oligonucleotide that, when administered to a subject in an effective amount, reduces target mRNA levels in liver tissue. Some embodiments include compositions comprising a GalNAc moiety and an oligonucleotide that, when administered to a subject in an effective amount, reduces target mRNA levels in liver cells.

[0024] In some embodiments, the reduction in target oligonucleotide levels is specific to hepatocytes with respect to other cell types. In some embodiments, the reduction in target RNA levels is specific to hepatocytes with respect to other cell types. In some embodiments, the reduction in target mRNA levels is specific to hepatocytes with respect to other cell types. In some embodiments, the reduction in target protein levels is specific to hepatocytes with respect to other cell types. In some embodiments, the reduction in target oligonucleotide levels is specific to liver tissue with respect to other tissue types. In some embodiments, the reduction in target RNA levels is specific to liver with respect to other tissue types. In some embodiments, the reduction in target mRNA levels is specific to liver with respect to other tissue types. In some embodiments, the reduction in target protein levels is specific to liver with respect to other tissue types.

[0025] In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide that binds to a target oligonucleotide, which when administered to a subject in an effective amount reduces the target oligonucleotide level in a cell or tissue. In some embodiments, the target oligonucleotide level is reduced by about 2.5% or more, about 5% or more, or about 7.5% or more compared to before administration. In some embodiments, the target oligonucleotide level is reduced by about 10% or more compared to before administration. In some embodiments, the target oligonucleotide level is reduced by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% compared to before administration. In some embodiments, the target oligonucleotide level is reduced by about 2.5% or less, about 5% or less, or about 7.5% or less compared to before administration. In some embodiments, the target oligonucleotide level is reduced by about 10% or less compared to before administration. In some embodiments, the target oligonucleotide level is reduced by about 20% or less, about 30% or less, about 40% or less, about 50% or less, about 60% or less, about 70% or less, about 80% or less, about 90% or less, or about 100% or less compared to before administration. In some embodiments, the target oligonucleotide level is reduced by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, or a range defined by any of the two aforementioned percentages.

[0026] In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide that binds to a target mRNA, and when administered to a subject in an effective amount, reduces the target mRNA level in a cell or tissue. In some embodiments, the target mRNA level is reduced by about 2.5% or more, about 5% or more, or about 7.5% or more compared to before administration. In some embodiments, the target mRNA level is reduced by about 10% or more compared to before administration. In some embodiments, the target mRNA level is reduced by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% compared to before administration. In some embodiments, the target mRNA level is reduced by about 2.5% or less, about 5% or less, or about 7.5% or less compared to before administration. In some embodiments, the target mRNA level is reduced by about 10% or less compared to before administration. In some embodiments, the target mRNA level is reduced by about 20% or less, about 30% or less, about 40% or less, about 50% or less, about 60% or less, about 70% or less, about 80% or less, about 90% or less, or about 100% or less compared to before administration. In some embodiments, the target mRNA level is reduced by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, or a range defined by any of the two aforementioned percentages.

[0027] In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide encoding a target protein that reduces target protein levels in a cell or tissue when the composition is administered to a subject in an effective amount. In some embodiments, the cell is a hepatocyte. In some embodiments, the tissue is a liver tissue. In some embodiments, the target protein level is reduced by about 2.5% or more, about 5% or more, or about 7.5% or more compared to before administration. In some embodiments, the target protein level is reduced by about 10% or more compared to before administration. In some embodiments, the target protein level is reduced by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% compared to before administration. In some embodiments, the target protein level is reduced by about 2.5% or less, about 5% or less, or about 7.5% or less compared to before administration. In some embodiments, the target protein level is reduced by about 10% or less compared to before administration. In some embodiments, the target protein level is reduced by about 20% or less, about 30% or less, about 40% or less, about 50% or less, about 60% or less, about 70% or less, about 80% or less, about 90% or less, or about 100% or less compared to before administration. In some embodiments, the target protein level is reduced by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, or a range defined by any of the two aforementioned percentages.

[0028] In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide that binds to a target oligonucleotide (e.g., mRNA) and reduces a deleterious phenotype (e.g., a symptom of a disorder associated with the target oligonucleotide) when administered to a subject in an effective amount. In some embodiments, the deleterious phenotype is reduced by about 2.5% or more, about 5% or more, or about 7.5% or more compared to before administration. In some embodiments, the deleterious phenotype is reduced by about 10% or more compared to before administration. In some embodiments, the deleterious phenotype is reduced by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% compared to before administration. In some embodiments, the deleterious phenotype is reduced by about 2.5% or less, about 5% or less, or about 7.5% or less compared to before administration. In some embodiments, the deleterious phenotype is reduced by about 10% or less compared to before administration. In some embodiments, the deleterious phenotype is reduced by about 20% or less, about 30% or less, about 40% or less, about 50% or less, about 60% or less, about 70% or less, about 80% or less, about 90% or less, or about 100% or less compared to before administration. In some embodiments, the deleterious phenotype is reduced by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, or a range defined by any of the two aforementioned percentages.

[0029] In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide that binds to a target oligonucleotide (e.g., mRNA) and increases a protective phenotype (e.g., prevents a disorder) when administered to a subject in an effective amount. In some embodiments, the protective phenotype is increased by about 2.5% or more, about 5% or more, or about 7.5% or more compared to before administration. In some embodiments, the protective phenotype is increased by about 10% or more compared to before administration. In some embodiments, the protective phenotype is increased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more compared to before administration. In some embodiments, the protective phenotype is increased by about 200% or more, about 300% or more, about 400% or more, about 500% or more, about 600% or more, about 700% or more, about 800% or more, about 900% or more, or about 1000% or more compared to before administration. In some embodiments, the protective phenotype is increased by about 2.5% or less, about 5% or less, or about 7.5% or less compared to before administration. In some embodiments, the protective phenotype is increased by about 10% or less compared to before administration. In some embodiments, the protective phenotype is increased by about 20% or less, about 30% or less, about 40% or less, about 50% or less, about 60% or less, about 70% or less, about 80% or less, about 90% or less, or about 100% or less compared to before administration. In some embodiments, the protective phenotype is increased by about 200% or less, about 300% or less, about 400% or less, about 500% or less, about 600% or less, about 700% or less, about 800% or less, about 900% or less, or about 1000% or less compared to before administration. In some embodiments, the protective phenotype is increased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 150%, 200%, 250%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, or 1000%, or a range defined by any of the two aforementioned percentages.

[0030] A. GalNAc Moieties and Compounds In some embodiments, a composition comprising a GalNAc moiety is provided herein. In some embodiments, a composition comprising a GalNAc moiety and an oligonucleotide is provided herein. In some embodiments, a composition comprising a GalNAc moiety and an oligonucleotide is described by a compound of formula (I) or formula (II). In some embodiments, the oligonucleotide of formula (I) or formula (II) is J. In some embodiments, the GalNAc moiety of formula (I) or formula (II) is a molecular moiety linked to J. The oligonucleotide may comprise a small interfering RNA (siRNA) or an antisense oligonucleotide (ASO).

[0031] In some embodiments, formula (I) or (II)

[0032] [ka] or a salt thereof, wherein J is an oligonucleotide; Each w is independently selected from any value of 1 to 20; Each v is independently selected from any value from 1 to 20; n is selected from any value between 1 and 20, m is selected from any value between 1 and 20, z is selected from any value between 1 and 3, when z is 3, Y is C; If z is 2, Y is CR 6 or When z is 1, Y is C(R 6 ) 2 and Q is halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 )2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , -S(O)R 7 , and C 1-6 C optionally substituted with one or more substituents independently selected from alkyl 3-10 Carbocyclic rings, C 1-6 Alkyl is a halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 and optionally substituted with one or more substituents independently selected from R 1 -O-, -S-, -N(R 7 )-, -C(O)-, -C(O)N(R 7 )-, -N(R 7 )C(O)-, -N(R 7 )C(O)N(R 7 )-, -OC(O)N(R 7 )-, -N(R 7 )C(O)O-, -C(O)O-, -OC(O)-, -S(O)-, -S(O) 2 -, -OS(O) 2 -,-OP(O)(OR 7 )O-, -SP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, -OP(O)(OR 7 )S-, -OP(O)(O - )O-, -SP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, -OP(O)(O - )S-, -OP(O)(OR 7 )NR 7 -,-OP(O)(N(R 7) 2 )NR 7 -,-OP(OR 7 )O-, -OP(N(R 7 ) 2 )O-, -OP(OR 7 )N(R 7 )-, and -OPN(R 7 ) 2 NR 7 -, R 2 are each independently halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6 alkyl, R 3 and R 4 are each independently -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR7 , -OC(O)R 7 , and -S(O)R 7 is selected from R 5 are each independently -OC(O)R 7 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)R 7 , -C(O)OR 7 , and -C(O)N(R 7 ) 2 is selected from R 6 are each independently hydrogen, Halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 , and Halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7)C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6 alkyl, R 7 are each independently hydrogen, C 1-6 Alkyl, C 2-6 Alkenyl, and C 2-6 Alkynyl (each of which is halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 3-10 optionally substituted with one or more substituents independently selected from carbocycle, and 3- to 10-membered heterocycle; and C 3-10 Carbocycles and 3- to 10-membered heterocycles, each of which is independently selected from the group consisting of halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-10 Carbocyclic rings, 3- to 10-membered heterocyclic rings, and C 1-6 haloalkyl).

[0033] In some embodiments, each w is independently selected from any value from 1 to 20. In some embodiments, each w is independently selected from any value from 1 to 15. In some embodiments, each w is independently selected from any value from 1 to 10. In some embodiments, each w is independently selected from any value from 1 to 5. In some embodiments, each w is independently selected from any value from 1 to 4. In some embodiments, each w is independently selected from any value from 1 to 3. In some embodiments, each w is independently selected from any value from 1 to 2. In some embodiments, each w is independently 1. In some embodiments, w is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20.

[0034] In some embodiments, each v is independently selected from any value from 1 to 20. In some embodiments, each v is independently selected from any value from 1 to 15. In some embodiments, each v is independently selected from any value from 1 to 10. In some embodiments, each v is independently selected from any value from 1 to 5. In some embodiments, each v is independently selected from any value from 1 to 4. In some embodiments, each v is independently selected from any value from 1 to 3. In some embodiments, each v is independently selected from any value from 1 to 2. In some embodiments, each v is independently 1. In some embodiments, v is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20.

[0035] In some embodiments, n is selected from any value between 1 and 20. In some embodiments, n is selected from any value between 1 and 15. In some embodiments, n is selected from any value between 1 and 10. In some embodiments, n is selected from any value between 1 and 9. In some embodiments, n is selected from any value between 1 and 8. In some embodiments, n is selected from any value between 1 and 7. In some embodiments, n is selected from any value between 1 and 6. In some embodiments, n is selected from any value between 1 and 5. In some embodiments, n is selected from any value between 1 and 4. In some embodiments, n is selected from any value between 2 and 4. In some embodiments, n is selected from any value between 1 and 3. In some embodiments, n is 2 or 3. In some embodiments, n is 3. In some embodiments, n is selected from any value between 1 and 2. In some embodiments, n is 2. In some embodiments, n is 1. In some embodiments, n is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20.

[0036] In some embodiments, m is selected from any value between 1 and 20. In some embodiments, m is selected from any value between 1 and 15. In some embodiments, m is selected from any value between 1 and 10. In some embodiments, m is selected from any value between 1 and 9. In some embodiments, m is selected from any value between 1 and 8. In some embodiments, m is selected from any value between 1 and 7. In some embodiments, m is selected from any value between 3 and 7. In some embodiments, m is selected from any value between 1 and 6. In some embodiments, m is selected from any value between 2 and 6. In some embodiments, m is selected from any value between 3 and 6. In some embodiments, m is selected from any value between 4 and 6. In some embodiments, m is 6. In some embodiments, m is selected from any value between 1 and 5. In some embodiments, m is selected from any value between 3 and 5. In some embodiments, m is 5. In some embodiments, m is 4 or 5. In some embodiments, m is selected from any value between 1 and 4. In some embodiments, m is 4. In some embodiments, m is 3 or 4. In some embodiments, m is selected from any value from 2 to 4. In some embodiments, m is selected from any value from 1 to 3. In some embodiments, m is 3. In some embodiments, m is selected from any value from 1 to 2. In some embodiments, m is 2. In some embodiments, m is 1. In some embodiments, m is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20.

[0037] In some embodiments, z is selected from any value from 1 to 3. In some embodiments, z is 3 and Y is C. In some embodiments, z is 2 and Y is CR 6 In some embodiments, z is 1 and Y is C(R6 ) 2 It is.

[0038] In some embodiments, Q is halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , -S(O)R 7 , and C 1-6 C optionally substituted with one or more substituents independently selected from alkyl 3-10 Carbocyclic rings, C 1-6 Alkyl is a halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 In some embodiments, Q is optionally substituted with one or more substituents independently selected from halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R7 , -S(O)R 7 , and C 1-6 C optionally substituted with one or more substituents independently selected from alkyl 3-6 Carbocyclic rings, C 1-6 Alkyl is a halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 In some embodiments, Q is optionally substituted with one or more substituents independently selected from halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 5-6 In some embodiments, Q is selected from halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 6 In some embodiments, Q is selected from halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 5 In some embodiments, Q is selected from halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 C optionally substituted with one or more substituents independently selected from 5-6 In some embodiments, Q is selected from phenyl, cyclohexyl, cyclopentadiene, and cyclopentyl, each of which is selected from halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 In some embodiments, Q is selected from phenyl and cyclohexyl, each of which is halogen, -CN, -OH, -SH, -NO 2 , and -NH 2In some embodiments, Q is optionally substituted with one or more substituents independently selected from: In some embodiments, Q is selected from phenyl and cyclohexyl. In some embodiments, Q is phenyl. In some embodiments, Q is cyclohexyl.

[0039] In some embodiments, R 1 -O-, -S-, -N(R 7 )-, -C(O)-, -C(O)N(R 7 )-, -N(R 7 )C(O)-, -N(R 7 )C(O)N(R 7 )-, -OC(O)N(R 7 )-, -N(R 7 )C(O)O-, -C(O)O-, -OC(O)-, -S(O)-, -S(O) 2 -, -OS(O) 2 -,-OP(O)(OR 7 )O-, -SP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, -OP(O)(OR 7 )S-, -OP(O)(OR 7 )NR 7 -,-OP(O)(N(R 7 ) 2 )NR 7 -,-OP(OR 7 )O-, -OP(N(R 7 ) 2 )O-, -OP(OR 7 )N(R 7 )-, and -OPN(R 7 ) 2 NR 7 In some embodiments, R 1 -O-, -S-, -N(R 7 )-, -C(O)-, -C(O)N(R 7 )-, -N(R 7 )C(O)-, -N(R 7 )C(O)N(R 7 )-, -OC(O)N(R 7)-, -N(R 7 )C(O)O-, -C(O)O-, -OC(O)-, -S(O)-, -S(O) 2 -, -OS(O) 2 -,-OP(O)(OR 7 )O-, -SP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, -OP(O)(OR 7 )S-, -OP(O)(O - )O-, -SP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, -OP(O)(O - )S-, -OP(O)(OR 7 )NR 7 -,-OP(O)(N(R 7 ) 2 )NR 7 -,-OP(OR 7 )O-, -OP(N(R 7 ) 2 )O-, -OP(OR 7 )N(R 7 )-, and -OPN(R 7 ) 2 NR 7 In some embodiments, R 1 -OP(O)(OR 7 )O-, -SP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, -OP(O)(OR 7 )S-, -OP(O)(O - )O-, -SP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, -OP(O)(O - )S-, -OP(O)(OR 7 )NR 7 -,-OP(O)(N(R 7 ) 2 )NR 7 -,-OP(OR7 )O-, -OP(N(R 7 ) 2 )O-, -OP(OR 7 )N(R 7 )-, and -OPN(R 7 ) 2 NR 7 In some embodiments, R 1 -OP(O)(OR 7 )O-, -SP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, -OP(O)(OR 7 )S-, -OP(O)(O - )O-, -SP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, -OP(O)(O - )S-, and -OP(OR 7 In some embodiments, R 1 -OP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, and -OP(OR 7 In some embodiments, R 1 -OP(O)(OR 7 )O- and -OP(OR 7 In some embodiments, R 1 -OP(O)(OH)O-, -SP(O)(OH)O-, -OP(S)(OH)O-, -OP(O)(SH)O-, -OP(O)(OH)S-, -OP(O)(O - )O-, -SP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, and -OP(O)(O - )S-. In some embodiments, R1 -OP(O)(OR 7 )O-, -OP(O)(OR 7 )NR 7 -,-OP(O)(N(R 7 ) 2 )NR 7 -,-OP(OR 7 )O-, -OP(N(R 7 ) 2 )O-, -OP(OR 7 )N(R 7 )-, and -OPN(R 7 ) 2 NR 7 In some embodiments, R 1 -S-, -S(O)-, -S(O) 2 -, -OS(O) 2 , -SP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, -OP(O)(OR 7 )S-, -SP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, and -OP(O)(O - In some embodiments, R 1 -S-, -S(O)-, -S(O) 2 -, -OS(O) 2 , -SP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, and -OP(O)(OR 7 In some embodiments, R 1 -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, and -OP(O)(OR 7 In some embodiments, R 1 -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, -OP(S)(O- )O-, and -OP(O)(S - In some embodiments, R 1 -OP(S)(OR 7 )O-, and -OP(O)(SR 7 In some embodiments, R 1 -OP(O)(OR 7 )O-, -OP(OR 7 )N(R 7 )-, and -OPN(R 7 ) 2 In some embodiments, R 1 is -OP(O)(OH)O-, -OP(O)(OCH 2 CH 3 )O-, -OP(OCH 2 CH 2 CN)N(CH(CH 3 ) 2 )-, or -OPN(CH(CH 3 ) 2 ) 2 In some embodiments, R 1 is -OP(O)(OH)O-, and OP(O)(O - In some embodiments, R 1 In some embodiments, R 1 In some embodiments, R 1 In some embodiments, R 1 is a linker selected from -O- or -S-. In some embodiments, R 1 In some embodiments, R 1 is -S-.

[0040] In some embodiments, R 2 are each independently halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6 In some embodiments, R 2 are each independently halogen, -OR 7 , -OC(O)R 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , and -S(O)R 7 C substituted with one or more substituents independently selected from 1-3 In some embodiments, R 2 are each independently -OR 7 , -OC(O)R 7 , -SR 7 , and -N(R 7 ) 2 C substituted with one or more substituents independently selected from 1-3 In some embodiments, R 2 are each independently -OR 7 and -OC(O)R 7 C substituted with one or more substituents independently selected from 1 In some embodiments, R 2 are each independently -CH 2 OH and -CH 2 OC(O)CH 3 is selected from.

[0041] In some embodiments, R3 are each independently -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 In some embodiments, R 3 are each independently -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -OC(O)R 7 , and -S(O)R 7 In some embodiments, R 3 are each independently -OR 7 , -SR 7 , -OC(O)R 7 , -SR 7 , and -N(R 7 ) 2 In some embodiments, R 3 are each independently -OR 7 and -OC(O)R 7 In some embodiments, R 3 are each independently -OH and -OC(O)CH 3 is selected from.

[0042] In some embodiments, R 4 are each independently -OR 7 , -SR 7 , -N(R 7 )2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 In some embodiments, R 4 are each independently halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -OC(O)R 7 , and -S(O)R 7 In some embodiments, R 4 are each independently -OR 7 , -OC(O)R 7 , -SR 7 , and -N(R 7 ) 2 In some embodiments, R 4 are each independently -OR 7 and -OC(O)R 7 In some embodiments, R 4 are each independently -OH and -OC(O)CH 3 is selected from.

[0043] In some embodiments, R 5 are each independently -OC(O)R 7 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2, -N(R 7 )C(O)OR 7 , -C(O)R 7 , -C(O)OR 7 , and -C(O)N(R 7 ) 2 In some embodiments, R 5 are each independently -OC(O)R 7 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , and -N(R 7 )C(O)OR 7 In some embodiments, R 5 are each independently -OC(O)R 7 , and -N(R 7 )C(O)R 7 In some embodiments, R 5 are each independently -N(H)C(O)CH 3 is selected from.

[0044] In some embodiments, R 6 are each independently hydrogen, halogen, -CN, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 , and halogens, -CN, -OR7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6 In some embodiments, R 6 are each independently hydrogen, halogen, -CN, -OR 7 , -SR 7 , -N(R 7 ) 2 , and halogens, -OR 7 , -SR 7 , and -N(R 7 ) 2 C optionally substituted with one or more substituents independently selected from 1-6 In some embodiments, R 6 are each independently hydrogen, halogen, -CN, -OH, -SH, and -NH 2 are independently selected from

[0045] In some embodiments, R 7 are each independently hydrogen, C 1-6 Alkyl, C 2-6 Alkenyl, and C 2-6 Alkynyl (each of which is halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 3-10 Optionally substituted with one or more substituents independently selected from carbocycle, and 3- to 10-membered heterocycle, and 3-10 Carbocycles and 3- to 10-membered heterocycles, each of which is independently selected from the group consisting of halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-10 Carbocyclic rings, 3- to 10-membered heterocyclic rings, and C 1-6 haloalkyl). 7 are each independently hydrogen and the halogens -CN, -OH, -SH, and -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 3-10 C optionally substituted with one or more substituents independently selected from a carbocycle, a 3- to 10-membered heterocycle 1-6 In some embodiments, R 7 are each independently halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , and -NH(C 1-6 C optionally substituted with one or more substituents independently selected from1-6 In some embodiments, R 7 is independently selected from halogen, -CN, -OH, and -SH, 1-6 In some embodiments, R 7 Each is independently selected from hydrogen. 7 are each independently halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , and -NH(C 1-6 C optionally substituted with one or more substituents independently selected from 1-3 is selected from alkyl.

[0046] In some embodiments, w is 1, v is 1, n is 2, m is 1 or 2, z is 3, and Y is C and Q is phenyl or cyclohexyl, each of which is selected from the group consisting of halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , and C 1-3 optionally substituted with one or more substituents independently selected from alkyl, R 1 -OP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, and -OP(OR 7 )O-, R 2 is -OH or -OC(O)CH 3 C replaced with 1 is alkyl, R 3 is -OH or -OC(O)CH 3 and R 4 is -OH or -OC(O)CH 3and R 5 -NH(O)CH 3 It is.

[0047] In some embodiments, w is 1, v is 1, n is 2, m is 1 or 2, z is 3, and Y is C and Q is phenyl or cyclohexyl, each of which is selected from the group consisting of halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , and C 1-3 optionally substituted with one or more substituents independently selected from alkyl, R 1 -OP(O)(OH)O-, -OP(S)(OH)O-, -OP(O)(O - )O-, -OP(S)(O - )O-, -OP(O)(S - )O-, and -OP(OH)O-; R 2 is -OH or -OC(O)CH 3 C replaced with 1 is alkyl, R 3 is -OH or -OC(O)CH 3 and R 4 is -OH or -OC(O)CH 3 and R 5 -NH(O)CH 3 In some embodiments, w is 1, v is 1, n is 2, m is 1 or 2, z is 3, and Y is C and Q is phenyl or cyclohexyl, each of which is selected from the group consisting of halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , and C 1-3 optionally substituted with one or more substituents independently selected from alkyl, R 1 is selected from -OP(O)(OH)O-, -OP(S)(OH)O-, and -OP(OH)O-; R 2 is -OH or -OC(O)CH 3 C replaced with 1 is alkyl, R 3 is -OH or -OC(O)CH 3 and R4 is -OH or -OC(O)CH 3 and R 5 -NH(O)CH 3 It is.

[0048] In some embodiments, the compound of formula (I) is

[0049] [ka]

[0050] [ka]

[0051] [ka] is selected from.

[0052] In some embodiments, the compound of formula (II) is

[0053] [ka]

[0054] [ka]

[0055] [ka]

[0056] [ka]

[0057] [ka] is selected from.

[0058] In some embodiments, the compound of formula (I), (II), or (III) binds to a lectin. In some embodiments, the compound binds to an asialoglycoprotein receptor. In some embodiments, the compound binds to a liver cell receptor. In some embodiments, the compound binds to a hepatocyte receptor. In some embodiments, the compound targets liver cells.

[0059] In some embodiments, the compositions described herein include a GalNAc compound. In some embodiments, the GalNAc compound is represented by formula (A) or formula (B):

[0060] [ka] or a salt thereof, wherein Each w is independently selected from any value of 1 to 20; Each v is independently selected from any value from 1 to 20; n is selected from any value between 1 and 20, m is selected from any value between 1 and 20, z is selected from any value between 1 and 3, when z is 3, Y is C; If z is 2, Y is CR 6 or When z is 1, Y is C(R 6 ) 2 and Q is halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2, -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , -S(O)R 7 , and C 1-6 C optionally substituted with one or more substituents independently selected from alkyl 3-10 Carbocyclic rings, C 1-6 Alkyl is a halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 and optionally substituted with one or more substituents independently selected from R 1 -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , -S(O)R 7 , -S(O) 2 R 7 , -OS(O) 2 R 7 , -OP(O)(OR 7 ) 2 , -OP(S)(OR 7 ) 2 , -SP(O)(OR 7 ) 2 , -OP(O)(SR 7 )(OR 7 ), -OP(O)(OR 7 )N(R 7 ) 2 , -OP(S)(OR 7 )N(R7 ) 2 , -SP(O)(OR 7 )N(R 7 ) 2 , -OP(O)(SR 7 )N(R 7 ) 2 , -OP(O)(N(R 7 ) 2 ) 2 , -OP(S)(N(R 7 ) 2 ) 2 , -SP(O)(N(R 7 ) 2 ) 2 , -OP(OR 7 ) 2 , -SP(OR 7 ) 2 , -OP(OR 7 )(SR 7 ), -OP(OR 7 )N(R 7 ) 2 , -OP(SR 7 )N(R 7 ) 2 , -SP(OR 7 )N(R 7 ) 2 , -OP(N(R 7 ) 2 ) 2 , and -SP(N(R 7 ) 2 ) 2 is selected from R 2 are each independently halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6 alkyl, R 3 and R 4 are each independently -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 is selected from R 5 are each independently -OC(O)R 7 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)R 7 , -C(O)OR 7 , and -C(O)N(R 7 ) 2 is selected from R 6 are each independently hydrogen, Halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 , and Halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6 alkyl, R 7 are each independently hydrogen, C 1-6 Alkyl, C 2-6 Alkenyl, and C 2-6 Alkynyl (each of which is halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2, -NH(C 1-6 Alkyl), C 3-10 optionally substituted with one or more substituents independently selected from carbocycle, and 3- to 10-membered heterocycle; and C 3-10 Carbocycles and 3- to 10-membered heterocycles, each of which is independently selected from the group consisting of halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-10 Carbocyclic rings, 3- to 10-membered heterocyclic rings, and C 1-6 haloalkyl).

[0061] In some embodiments, each w is independently selected from any value from 1 to 20. In some embodiments, each w is independently selected from any value from 1 to 15. In some embodiments, each w is independently selected from any value from 1 to 10. In some embodiments, each w is independently selected from any value from 1 to 5. In some embodiments, each w is independently selected from any value from 1 to 4. In some embodiments, each w is independently selected from any value from 1 to 3. In some embodiments, each w is independently selected from any value from 1 to 2. In some embodiments, W is 1. In some embodiments, w is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20.

[0062] In some embodiments, each v is independently selected from any value from 1 to 20. In some embodiments, each v is independently selected from any value from 1 to 15. In some embodiments, each v is independently selected from any value from 1 to 10. In some embodiments, each v is independently selected from any value from 1 to 5. In some embodiments, each v is independently selected from any value from 1 to 4. In some embodiments, each v is independently selected from any value from 1 to 3. In some embodiments, each v is independently selected from any value from 1 to 2. In some embodiments, each v is independently 1. In some embodiments, v is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20.

[0063] In some embodiments, n is selected from any value between 1 and 20. In some embodiments, n is selected from any value between 1 and 15. In some embodiments, n is selected from any value between 1 and 10. In some embodiments, n is selected from any value between 1 and 9. In some embodiments, n is selected from any value between 1 and 8. In some embodiments, n is selected from any value between 1 and 7. In some embodiments, n is selected from any value between 1 and 6. In some embodiments, n is selected from any value between 1 and 5. In some embodiments, n is selected from any value between 1 and 4. In some embodiments, n is selected from any value between 2 and 4. In some embodiments, n is selected from any value between 1 and 3. In some embodiments, n is 2 or 3. In some embodiments, n is 3. In some embodiments, n is selected from any value between 1 and 2. In some embodiments, n is 2. In some embodiments, n is 1. In some embodiments, n is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20.

[0064] In some embodiments, m is selected from any value between 1 and 20. In some embodiments, m is selected from any value between 1 and 15. In some embodiments, m is selected from any value between 1 and 10. In some embodiments, m is selected from any value between 1 and 9. In some embodiments, m is selected from any value between 1 and 8. In some embodiments, m is selected from any value between 1 and 7. In some embodiments, m is selected from any value between 3 and 7. In some embodiments, m is selected from any value between 1 and 6. In some embodiments, m is selected from any value between 2 and 6. In some embodiments, m is selected from any value between 3 and 6. In some embodiments, m is selected from any value between 4 and 6. In some embodiments, m is 6. In some embodiments, m is selected from any value between 1 and 5. In some embodiments, m is selected from any value between 3 and 5. In some embodiments, m is 5. In some embodiments, m is 4 or 5. In some embodiments, m is selected from any value between 1 and 4. In some embodiments, m is 4. In some embodiments, m is 3 or 4. In some embodiments, m is selected from any value from 2 to 4. In some embodiments, m is selected from any value from 1 to 3. In some embodiments, m is 3. In some embodiments, m is selected from any value from 1 to 2. In some embodiments, m is 2. In some embodiments, m is 1. In some embodiments, m is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20.

[0065] In some embodiments, z is selected from any value from 1 to 3. In some embodiments, z is 3 and Y is C. In some embodiments, z is 2 and Y is CR 6 In some embodiments, z is 1 and Y is C(R6 ) 2 It is.

[0066] In some embodiments, Q is halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , -S(O)R 7 , and C 1-6 C optionally substituted with one or more substituents independently selected from alkyl 3-10 Carbocyclic rings, C 1-6 Alkyl is a halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 In some embodiments, Q is optionally substituted with one or more substituents independently selected from halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R7 , -S(O)R 7 , and C 1-6 C optionally substituted with one or more substituents independently selected from alkyl 3-6 Carbocyclic rings, C 1-6 Alkyl is a halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 In some embodiments, Q is optionally substituted with one or more substituents independently selected from halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 5-6 In some embodiments, Q is selected from halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 6 In some embodiments, Q is selected from halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 5 In some embodiments, Q is selected from halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 C optionally substituted with one or more substituents independently selected from 5-6 In some embodiments, Q is selected from phenyl, cyclohexyl, cyclopentadiene, and cyclopentyl, each of which is selected from halogen, -CN, -OH, -SH, -NO 2 , and -NH 2 In some embodiments, Q is selected from phenyl and cyclohexyl, each of which is halogen, -CN, -OH, -SH, -NO 2 , and -NH 2In some embodiments, Q is optionally substituted with one or more substituents independently selected from: In some embodiments, Q is selected from phenyl and cyclohexyl. In some embodiments, Q is phenyl. In some embodiments, Q is cyclohexyl.

[0067] In some embodiments, R 1 -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , -S(O)R 7 , -S(O) 2 R 7 , -OS(O) 2 R 7 , -OP(O)(OR 7 ) 2 , -OP(S)(OR 7 ) 2 , -SP(O)(OR 7 ) 2 , -OP(O)(SR 7 )(OR 7 ), -OP(O)(OR 7 )N(R 7 ) 2 , -OP(S)(OR 7 )N(R 7 ) 2 , -SP(O)(OR 7 )N(R 7 ) 2 , -OP(O)(SR 7 )N(R 7 ) 2 , -OP(O)(N(R 7 )2 ) 2 , -OP(S)(N(R 7 ) 2 ) 2 , -SP(O)(N(R 7 ) 2 ) 2 , -OP(OR 7 ) 2 , -SP(OR 7 ) 2 , -OP(OR 7 )(SR 7 ), -OP(OR 7 )N(R 7 ) 2 , -OP(SR 7 )N(R 7 ) 2 , -SP(OR 7 )N(R 7 ) 2 , -OP(N(R 7 ) 2 ) 2 , and -SP(N(R 7 ) 2 ) 2 In some embodiments, R 1 -OR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , -OP(O)(OR 7 ) 2 , -OP(O)(OR 7 )N(R 7 ) 2 , -OP(O)(N(R 7 ) 2 ) 2 , -OP(OR 7) 2 , OP(OR 7 )N(R 7 ) 2 , and -OP(N(R 7 ) 2 ) 2 In some embodiments, R 1 -SR 7 , -S(O)R 7 , -S(O) 2 R 7 , -OS(O) 2 R 7 , -OP(S)(OR 7 ) 2 , -SP(O)(OR 7 ) 2 , -OP(O)(SR 7 )(OR 7 ), -OP(S)(OR 7 )N(R 7 ) 2 , -SP(O)(OR 7 )N(R 7 ) 2 , -OP(O)(SR 7 )N(R 7 ) 2 , -OP(S)(N(R 7 ) 2 ) 2 , -SP(O)(N(R 7 ) 2 ) 2 , -SP(OR 7 ) 2 , -OP(OR 7 )(SR 7 ), -OP(SR 7 )N(R 7 ) 2 , -SP(OR 7 )N(R 7 ) 2 , and -SP(N(R 7 ) 2 ) 2 In some embodiments, R 1 -OP(O)(OR 7 ) 2 , -OP(S)(OR 7 ) 2 , -SP(O)(OR 7 )2 , -OP(O)(SR 7 )(OR 7 ), -OP(O)(OR 7 )N(R 7 ) 2 , -OP(S)(OR 7 )N(R 7 ) 2 , -SP(O)(OR 7 )N(R 7 ) 2 , -OP(O)(SR 7 )N(R 7 ) 2 , -OP(O)(N(R 7 ) 2 ) 2 , -OP(S)(N(R 7 ) 2 ) 2 , -SP(O)(N(R 7 ) 2 ) 2 , -OP(OR 7 ) 2 , -SP(OR 7 ) 2 , -OP(OR 7 )(SR 7 ), -OP(OR 7 )N(R 7 ) 2 , -OP(SR 7 )N(R 7 ) 2 , -SP(OR 7 )N(R 7 ) 2 , -OP(N(R 7 ) 2 ) 2 , and -SP(N(R 7 ) 2 ) 2 In some embodiments, R 1 -OP(O)(OR 7 ) 2 , -OP(O)(OR 7 )N(R 7 ) 2 , -OP(O)(N(R 7 ) 2 ) 2 , -OP(OR 7 ) 2, -OP(OR 7 )N(R 7 ) 2 , and -OP((NR 7 ) 2 ) 2 In some embodiments, R 1 -OP(O)(OR 7 ) 2 and -OP(OR 7 )N(R 7 ) 2 In some embodiments, R 1 -OP(O)(OCH 2 CH 3 )OH and -OP(OCH 2 CH 2 CN)N(CH(CH 3 ) 2 ) 2 In some embodiments, R 1 -OP(OCH 2 CH 2 CN)N(CH(CH 3 ) 2 ) 2 It is.

[0068] In some embodiments, R 2 are each independently halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6In some embodiments, R 2 are each independently halogen, -OR 7 , -OC(O)R 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , and -S(O)R 7 C substituted with one or more substituents independently selected from 1-3 In some embodiments, R 2 are each independently -OR 7 , -OC(O)R 7 , -SR 7 , and -N(R 7 ) 2 C substituted with one or more substituents independently selected from 1-3 In some embodiments, R 2 are each independently -OR 7 and -OC(O)R 7 C substituted with one or more substituents independently selected from 1 In some embodiments, R 2 are each independently -CH 2 OH and -CH 2 OC(O)CH 3 is selected from.

[0069] In some embodiments, R 3 are each independently -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 In some embodiments, R 3 are each independently -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -OC(O)R 7 , and -S(O)R 7 In some embodiments, R 3 are each independently -OR 7 , -SR 7 , -OC(O)R 7 , -SR 7 , and -N(R 7 ) 2 In some embodiments, R 3 are each independently -OR 7 and -OC(O)R 7 In some embodiments, R 3 are each independently -OH and -OC(O)CH 3 is selected from.

[0070] In some embodiments, R 4 are each independently -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7In some embodiments, R 4 are each independently halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -OC(O)R 7 , and -S(O)R 7 In some embodiments, R 4 are each independently -OR 7 , -OC(O)R 7 , -SR 7 , and -N(R 7 ) 2 In some embodiments, R 4 are each independently -OR 7 and -OC(O)R 7 In some embodiments, R 4 are each independently -OH and -OC(O)CH 3 is selected from.

[0071] In some embodiments, R 5 are each independently -OC(O)R 7 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)R 7 , -C(O)OR 7 , and -C(O)N(R 7 ) 2 In some embodiments, R 5 are each independently -OC(O)R 7 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7) 2 , and -N(R 7 )C(O)OR 7 In some embodiments, R 5 are each independently -OC(O)R 7 , and -N(R 7 )C(O)R 7 In some embodiments, R 5 are each independently -N(H)C(O)CH 3 is selected from.

[0072] In some embodiments, R 6 are each independently hydrogen, halogen, -CN, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 and halogen, -CN, -OR 7 , -SR 7 , -N(R 7 ) 2 , C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)OR 7 , C(O)OR7 , -OC(O)R 7 , and -S(O)R 7 C optionally substituted with one or more substituents independently selected from 1-6 In some embodiments, R 6 are each independently hydrogen, halogen, -CN, -OR 7 , -SR 7 , -N(R 7 ) 2 , and halogens, -OR 7 , -SR 7 , and -N(R 7 ) 2 C optionally substituted with one or more substituents independently selected from 1-6 In some embodiments, R 6 are each independently hydrogen, halogen, -CN, -OH, -SH, and -NH 2 are independently selected from

[0073] In some embodiments, R 7 are each independently hydrogen, C 1-6 Alkyl, C 2-6 Alkenyl, and C 2-6 Alkynyl (each of which is halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 3-10 Optionally substituted with one or more substituents independently selected from carbocycle, and 3- to 10-membered heterocycle, and 3-10 Carbocycles and 3- to 10-membered heterocycles, each of which is independently selected from the group consisting of halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2, -NH(C 1-6 Alkyl), C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-10 Carbocyclic rings, 3- to 10-membered heterocyclic rings, and C 1-6 haloalkyl). 7 are each independently hydrogen and the halogens -CN, -OH, -SH, and -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , -NH(C 1-6 Alkyl), C 3-10 C optionally substituted with one or more substituents independently selected from a carbocycle, a 3- to 10-membered heterocycle 1-6 In some embodiments, R 7 are each independently halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , and -NH(C 1-6 C optionally substituted with one or more substituents independently selected from 1-6 In some embodiments, R 7 Each is independently selected from hydrogen. 7 are each independently halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl) 2 , and -NH(C 1-6 C optionally substituted with one or more substituents independently selected from1-3 In some embodiments, R 7 each independently represents C optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, and -SH; 1-6 is selected from alkyl.

[0074] In some embodiments, the compound of formula (A) is

[0075] [ka]

[0076] [ka]

[0077] [ka] is selected from.

[0078] In some embodiments, the present invention relates to a compound of formula (B):

[0079] [ka]

[0080] [ka]

[0081] [ka] A compound represented by the formula:

[0082] In some embodiments, the salt of formula (I), (II), (A), or (B) is formed by deprotonating the phosphate. In some embodiments, the cation is a metal ion, such as a metal cation. Non-limiting examples of metal cations include Na + , K + , Mg 2+ , and Ca 2+ In some embodiments, the metal cation is Na + In some embodiments, the metal cation is K + In some embodiments, the metal cation is Mg 2+ In some embodiments, the metal cation is Ca 2+ In some embodiments, the cation is an organic or inorganic small molecule. In some embodiments, the compound is in the deprotonated form of Formula (I) or (II), the cation is a positively charged nucleic acid present in an oligonucleotide.

[0083] Some embodiments include the following, where J is an oligonucleotide.

[0084] [ka] J may include one or more additional phosphates or one or more phosphorothioates attached to the oligonucleotide. J may include one or more additional phosphates attached to the oligonucleotide. J may include one or more phosphorothioates attached to the oligonucleotide.

[0085] Some embodiments include the following, where J is an oligonucleotide.

[0086] [ka] J may include one or more additional phosphates or one or more phosphorothioates attached to the oligonucleotide. J may include one or more additional phosphates attached to the oligonucleotide. J may include one or more phosphorothioates attached to the oligonucleotide.

[0087] Some embodiments include the following, where J is an oligonucleotide.

[0088] [ka] J may comprise one or more phosphates or phosphorothioates attached to the oligonucleotide. J may comprise one or more phosphates attached to the oligonucleotide. J may comprise a phosphate attached to the oligonucleotide. J may comprise one or more phosphorothioates attached to the oligonucleotide. J may comprise a phosphorothioate attached to the oligonucleotide.

[0089] Some embodiments include the following, where J is an oligonucleotide.

[0090] [ka] The structure in this compound attached to an oligonucleotide (J) is an example of a GalNAc moiety. J may include one or more phosphates or phosphorothioates attached to the oligonucleotide. J may include one or more phosphates attached to the oligonucleotide. J may include a phosphate attached to the oligonucleotide. J may include one or more phosphorothioates attached to the oligonucleotide. J may include a phosphorothioate attached to the oligonucleotide.

[0091] Some embodiments include the following, where J is an oligonucleotide.

[0092] [ka] J may include one or more additional phosphates or one or more phosphorothioates attached to the oligonucleotide. J may include one or more additional phosphates attached to the oligonucleotide. J may include one or more phosphorothioates attached to the oligonucleotide.

[0093] Some embodiments include the following, where J is an oligonucleotide.

[0094] [ka] J may include one or more additional phosphates or one or more phosphorothioates attached to the oligonucleotide. J may include one or more additional phosphates attached to the oligonucleotide. J may include one or more phosphorothioates attached to the oligonucleotide.

[0095] Some embodiments include the following, where J is an oligonucleotide.

[0096] [ka] J may comprise one or more phosphates or phosphorothioates attached to the oligonucleotide. J may comprise one or more phosphates attached to the oligonucleotide. J may comprise a phosphate attached to the oligonucleotide. J may comprise one or more phosphorothioates attached to the oligonucleotide. J may comprise a phosphorothioate attached to the oligonucleotide.

[0097] Some embodiments include the following, where J is an oligonucleotide.

[0098] [ka] The structure in this compound bound to an oligonucleotide (J) may be referred to as "ETL17" and is an example of a GalNAc moiety. J may include one or more phosphates or phosphorothioates bound to the oligonucleotide. J may include one or more phosphates bound to the oligonucleotide. J may include a phosphate bound to the oligonucleotide. J may include one or more phosphorothioates bound to the oligonucleotide. J may include a phosphorothioate bound to the oligonucleotide.

[0099] Some embodiments include the following, where J is an oligonucleotide.

[0100] [ka] The structure in this compound linked to oligonucleotide (J) is an example of a GalNAc moiety. J can include one or more phosphates or phosphorothioates linked to the oligonucleotide. J can include one or more phosphates linked to the oligonucleotide. J can include a phosphate linked to the oligonucleotide. J can include one or more phosphorothioates linked to the oligonucleotide. J can include a phosphorothioate linked to the oligonucleotide. In any embodiment, J can include an additional linker.

[0101] 1. Analog Chemical entities having carbon-carbon or carbon-nitrogen double bonds can exist in Z or E forms (or cis or trans forms). Additionally, some chemical entities can exist in various tautomeric forms. Unless otherwise specified, the compounds described herein are intended to include all Z, E, and tautomeric forms as well.

[0102] "Tautomer" refers to a molecule or moiety capable of proton transfer from one atom of a molecule to another atom of the same molecule. The compounds presented herein exist as tautomers in certain embodiments. In situations where tautomerization is possible, a chemical equilibrium of tautomers exists. The exact ratio of tautomers depends on a variety of factors, including physical conditions, temperature, solvent, and pH. Some examples of tautomeric equilibrium are:

[0103] [ka] Includes.

[0104] In some embodiments, the compounds and moieties disclosed herein are used in various isotopically enriched forms, e.g., 2 H, 3 H, 11 C. 13 C, and / or 14 C content. In one particular embodiment, the compound is deuterated at at least one position. Such deuterated forms can be made by the procedures described in U.S. Patent Nos. 5,846,514 and 6,334,997. As described in U.S. Patent Nos. 5,846,514 and 6,334,997, deuteration can improve metabolic stability or efficacy, thus increasing the duration of action of pharmaceutical agents.

[0105] Unless otherwise specified, the compounds and moieties described herein are intended to include compounds and moieties that differ only in the presence of one or more isotopically enriched atoms. For example, the replacement of hydrogen with deuterium or tritium, or 13 C or 14 Compounds having this structure, except for the replacement of a carbon with a C-rich carbon, are within the scope of this disclosure.

[0106] The compounds and moieties of the present disclosure optionally contain unnatural proportions of atomic isotopes at one or more of the atoms that constitute such compounds. For example, the compounds may contain deuterium ( 2H), tritium ( 3 H), iodine-125( 125 I), or carbon-14 ( 14 C). 2 H, 11 C. 13 C. 14 C. 15 C. 12 N, 13 N, 15 N, 16 N, 16 O. 17 O. 14 F, 15 F, 16 F, 17 F, 18 F, 33 S, 34 S, 35 S, 36 S, 35 Cl, 37 Cl, 79 Br, 81 Br, and 125 All isotopic substitutions with I are contemplated. All isotopic variations of the compounds and moieties of the present invention, whether radioactive or not, are encompassed within the scope of the present invention.

[0107] In certain embodiments, the compounds and moieties disclosed herein are 1 Some or all of the H atoms 2 It is substituted with an H atom. Methods for the synthesis of compounds containing deuterium are known in the art and include, by way of non-limiting example only, the following synthetic methods.

[0108] Deuterium substituted compounds are synthesized using a variety of methods such as those described in Dean, Dennis C.; Editor. Recent Advances in the Synthesis and Applications of Radiolabeled Compounds for Drug Discovery and Development. [In: Curr., Pharm. Des., 2000; 6(10)] 2000, 110pp; George W.; Varma, Rajender S. The Synthesis of Radiolabeled Compounds via Organometallic Intermediates, Tetrahedron, 1989, 45(21), 6601-21; and Evans, E. Anthony. Synthesis of radiolabeled compounds, J. Radioanal. Chem., 1981, 64(1-2), 9-32.

[0109] Deuterated starting materials are readily available and can be subjected to the synthetic methods described herein to effect the synthesis of compounds containing deuterium. Many deuterium-containing reagents and building blocks are commercially available from chemical commodity companies such as Aldrich Chemical Co.

[0110] The salts of the compounds described herein, particularly pharma-ceutically acceptable salts, are included in the present disclosure.The compounds of the present disclosure that have sufficiently acidic, sufficiently basic, or both functional groups can react with many inorganic bases, and inorganic and organic acids to form salts.Alternatively, compounds that are inherently charged, such as those that have quaternary nitrogen, can form salts with suitable counterions, for example, halides such as bromide, chloride, or fluoride, especially bromide.

[0111] The compounds described herein may exist as diastereomers, enantiomers, or other stereoisomeric forms in some cases. The compounds presented herein include all diastereomeric, enantiomeric, and epimeric forms, and the appropriate mixtures thereof. Separation of stereoisomers can be carried out by chromatography, or diastereomeric formation and recrystallization or chromatographic separation, or any combination thereof (Jean Jacques, Andre Collet, Samuel H. Wilen, "Enantiomers, Racemates and Resolutions", John Wiley And Sons, Inc., 1981, incorporated by reference for this disclosure). Stereoisomers can also be obtained by stereoselective synthesis.

[0112] The methods and compositions described herein include the use of amorphous forms as well as crystalline forms (also known as polymorphs). The compounds described herein may be in the form of pharmaceutically acceptable salts. Similarly, in some embodiments, active metabolites of such compounds having the same type of activity are included within the scope of this disclosure. In addition, the compounds described herein can exist in unsolvated form or in solvated form with pharmaceutically acceptable solvents such as water, ethanol, etc. The solvated forms of the compounds presented herein are also considered to be disclosed herein.

[0113] B. Oligonucleotides In some embodiments, the present invention provides a composition or compound that comprises oligonucleotide.The oligonucleotide can be conjugated to GalNAc moiety.The oligonucleotide can be directly bound to the linker that is bound to GalNAc moiety.The oligonucleotide can be used in the method described herein.

[0114] In some embodiments, the oligonucleotide binds to a target oligonucleotide. Examples of target oligonucleotides include target RNA or target DNA. In some embodiments, the oligonucleotide binds to a target DNA. In some embodiments, the oligonucleotide binds to a target DNA and inhibits RNA (e.g., mRNA) expression from the target DNA. In some embodiments, the oligonucleotide binds to a target RNA. The target RNA may include a target mRNA. In some embodiments, the oligonucleotide binds to a target mRNA. In some embodiments, the oligonucleotide inhibits a target mRNA, such as by reducing the amount of the target mRNA, causing degradation of the target mRNA, or reducing or preventing translation of the target mRNA. In some embodiments, the oligonucleotide reduces the amount of a target protein produced from a target mRNA, for example, by inhibiting the target mRNA. The oligonucleotide may include a small interfering RNA (siRNA). The oligonucleotide may include an antisense oligonucleotide (ASO).

[0115] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide and inhibits expression of a target protein encoded by the target oligonucleotide. In some embodiments, the composition comprises an oligonucleotide that binds to a target RNA and inhibits expression of a target protein encoded by the target RNA. In some embodiments, the composition comprises an oligonucleotide that binds to a target mRNA and inhibits expression of a target protein encoded by the target mRNA.

[0116] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide and inhibits expression of a second oligonucleotide encoded by the target oligonucleotide. In some embodiments, the composition comprises an oligonucleotide that binds to a target DNA and inhibits expression of a target RNA encoded by the target DNA. In some embodiments, the composition comprises an oligonucleotide that binds to a target DNA and inhibits expression of a target mRNA encoded by the target DNA.

[0117] Target oligonucleotides can be identified in a variety of ways. In some examples, the target oligonucleotide includes an mRNA that has an expression level associated with the occurrence of a disorder (e.g., liver disorder). In some examples, the target oligonucleotide includes an mRNA encoded by a gene that has a particular genotype associated with a disorder. Large-scale human genetic data can improve the success rate of drug discovery and development. Genome-wide association studies (GWAS) can detect associations between genetic variants and traits in population samples. GWAS can improve the biological understanding of disease and inform appropriate treatments. GWAS can utilize genotyping and / or sequencing data and often involve evaluating millions of genetic variants that are relatively evenly distributed across the genome. The most common GWAS design is a case-control study, which involves comparing variant frequencies in cases and controls. If a variant has a significantly different frequency in cases and controls, the variant is said to be associated with the disease. Potential association statistics used in GWAS are p-values ​​as an indicator of statistical significance, odds ratios (ORs) as an indicator of effect size, or beta coefficients (β) as an indicator of effect size. Researchers often assume an additive genetic model and calculate allelic odds ratios, which indicate the increased (or decreased) risk of a given disease for each additional copy of an allele (compared to having no copies of that allele). An additional concept in the design and interpretation of GWAS is that of linkage disequilibrium, which is the non-random association of alleles. The presence of linkage disequilibrium can obscure which variants are "causative."

[0118] Functional annotation of variants and / or wet lab experiments can identify causative gene variants identified through GWAS, often leading to the identification of disease-causing genes. Notably, understanding the functional effect of a causative gene variant (e.g., loss of protein function, gain of protein function, increased gene expression, or decreased gene expression) can allow the variant to be used as a surrogate for therapeutic modulation of a target gene, providing insight into the potential therapeutic efficacy and safety of therapeutics that modulate that target.

[0119] Identification of such gene-disease associations can provide insight into disease biology and be used to identify novel therapeutic targets for the pharmaceutical industry. To translate therapeutic insights gained from human genetics, disease biology in patients may be exogenously "programmed" to recapitulate observations from human genetics. There are several options for therapeutic modalities that can be added when applying therapeutic targets identified by human genetics to new drugs. These may include well-established therapeutic modalities such as small molecule compounds and monoclonal antibodies, modalities in the maturation stage such as oligonucleotides, as well as new modalities such as gene therapy and gene editing. The choice of therapeutic modality may depend on several factors, including the location of the target (e.g., intracellular, extracellular, or secreted), the relevant tissue (e.g., liver), and the relevant indication. Such studies may be performed to identify specific liver injury-associated targets for siRNA or ASO inhibition by the compositions or compounds described herein.

[0120] Some embodiments include the method of making oligonucleotide or siRNA using the method disclosed herein.For example, any aspect of the examples herein that includes the step for synthesis can be used.Some embodiments include making GalNAc moiety or making the oligonucleotide with GalNAc moiety.

[0121] 1. siRNA In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide (e.g., mRNA), and the oligonucleotide comprises a small interfering RNA (siRNA). In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide (e.g., mRNA), and the oligonucleotide comprises a siRNA comprising a sense strand and an antisense strand. In some embodiments, the sense strand comprises RNA. In some embodiments, the antisense strand comprises RNA.

[0122] In some embodiments, the sense strand is 12-30 nucleosides in length. In some embodiments, the sense strand is 14-30 nucleosides in length. In some embodiments, the sense strand is at least about 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleosides in length. In some embodiments, the sense strand is at least 12 nucleotides in length. In some embodiments, the sense strand is at least 14 nucleotides in length. In some embodiments, the sense strand is at least 16 nucleotides in length. In some embodiments, the sense strand is at least 18 nucleotides in length. In some embodiments, the sense strand is at least 20 nucleotides in length. In some embodiments, the sense strand is at least 22 nucleotides in length. In some embodiments, the sense strand is at least 24 nucleotides in length. In some embodiments, the sense strand is at least 26 nucleotides in length. In some embodiments, the sense strand is at least 28 nucleotides in length. In some embodiments, the sense strand is at least 30 nucleotides in length. In some embodiments, the sense strand is about 10 or less, about 11 or less, about 12 or less, about 13 or less, about 14 or less, about 15 or less, about 16 or less, about 17 or less, about 18 or less, about 19 or less, about 20 or less, about 21 or less, about 22 or less, about 23 or less, about 24 or less, about 25 or less, about 26 or less, about 27 or less, about 28 or less, about 29 or less, or about 30 or less nucleosides in length. In some embodiments, the sense strand is 12 or less nucleotides in length. In some embodiments, the sense strand is 14 or less nucleotides in length. In some embodiments, the sense strand is 16 or less nucleotides in length. In some embodiments, the sense strand is 18 or less nucleotides in length. In some embodiments, the sense strand is 20 or less nucleotides in length. In some embodiments, the sense strand is 22 or less nucleotides in length. In some embodiments, the sense strand is 24 or less nucleotides in length. In some embodiments, the sense strand is 26 or less nucleotides in length.In some embodiments, the sense strand is 28 or fewer nucleotides in length. In some embodiments, the sense strand is 30 or fewer nucleotides in length.

[0123] In some embodiments, the antisense strand is 12-30 nucleosides in length. In some embodiments, the antisense strand is 14-30 nucleosides in length. In some embodiments, the antisense strand is at least about 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleosides in length. In some embodiments, the antisense strand is at least 12 nucleotides in length. In some embodiments, the antisense strand is at least 14 nucleotides in length. In some embodiments, the antisense strand is at least 16 nucleotides in length. In some embodiments, the antisense strand is at least 18 nucleotides in length. In some embodiments, the antisense strand is at least 20 nucleotides in length. In some embodiments, the antisense strand is at least 22 nucleotides in length. In some embodiments, the antisense strand is at least 24 nucleotides in length. In some embodiments, the antisense strand is at least 26 nucleotides in length. In some embodiments, the antisense strand is at least 28 nucleotides in length. In some embodiments, the antisense strand is at least 30 nucleotides in length. In some embodiments, the antisense strand is about 10 or less, about 11 or less, about 12 or less, about 13 or less, about 14 or less, about 15 or less, about 16 or less, about 17 or less, about 18 or less, about 19 or less, about 20 or less, about 21 or less, about 22 or less, about 23 or less, about 24 or less, about 25 or less, about 26 or less, about 27 or less, about 28 or less, about 29 or less, or about 30 or less nucleosides in length. In some embodiments, the antisense strand is 12 or less nucleotides in length. In some embodiments, the antisense strand is 14 or less nucleotides in length. In some embodiments, the antisense strand is 16 or less nucleotides in length. In some embodiments, the antisense strand is 18 or less nucleotides in length. In some embodiments, the antisense strand is 20 or less nucleotides in length. In some embodiments, the antisense strand is 22 or less nucleotides in length.In some embodiments, the antisense strand is 24 or less nucleotides in length. In some embodiments, the antisense strand is 26 or less nucleotides in length. In some embodiments, the antisense strand is 28 or less nucleotides in length. In some embodiments, the antisense strand is 30 or less nucleotides in length. In some embodiments, the antisense strand is the same length as the sense strand.

[0124] In some embodiments, the composition comprises an oligonucleotide that inhibits expression of a target oligonucleotide (e.g., mRNA), the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, the sense strand being 12-30 nucleosides in length. In some embodiments, the composition comprises a sense strand that is 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleosides in length, or in a range defined by either of the two preceding numbers. In some embodiments, the composition comprises an antisense strand that is 12-30 nucleosides in length. In some embodiments, the composition comprises an antisense strand that is 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleosides in length, or in a range defined by any of the foregoing two numbers.

[0125] In some embodiments, the compositions include oligonucleotides that inhibit expression of a target oligonucleotide (e.g., mRNA), the oligonucleotides include siRNAs that include a sense strand and an antisense strand, each strand being independently about 14-30 nucleosides in length, and at least one of the sense strand and the antisense strand includes a nucleoside sequence that includes about 12-30 contiguous nucleosides of a full-length human target mRNA sequence. In some embodiments, at least one of the sense strand and the antisense strand includes a nucleoside sequence that includes at least about 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or more contiguous nucleosides of one of the full-length human target mRNA sequences.

[0126] In some embodiments, the composition comprises the oligonucleotide that inhibits the expression of target protein, the oligonucleotide comprises siRNA that comprises sense strand and antisense strand, and the sense strand and the antisense strand form double-stranded RNA duplex.In some embodiments, the first base pair of the double-stranded RNA duplex is AU base pair.

[0127] In some embodiments, the sense strand further comprises a 3' overhang. In some embodiments, the 3' overhang comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleosides, or a range of nucleotides defined by any two of the foregoing numbers. In some embodiments, the 3' overhang comprises 1, 2, or more nucleosides. In some embodiments, the 3' overhang comprises 2 nucleosides. In some embodiments, the sense strand further comprises a 5' overhang. In some embodiments, the 5' overhang comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleosides, or a range of nucleotides defined by any two of the foregoing numbers. In some embodiments, the 5' overhang comprises 1, 2, or more nucleosides. In some embodiments, the 5' overhang comprises 2 nucleosides.

[0128] In some embodiments, the antisense strand further comprises a 3' overhang. In some embodiments, the 3' overhang comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleosides, or a range of nucleotides defined by any two of the aforementioned numbers. In some embodiments, the 3' overhang comprises 1, 2, or more nucleosides. In some embodiments, the 3' overhang comprises 2 nucleosides. In some embodiments, the antisense strand further comprises a 5' overhang. In some embodiments, the 5' overhang comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleosides, or a range of nucleotides defined by any two of the aforementioned numbers. In some embodiments, the 5' overhang comprises 1, 2, or more nucleosides. In some embodiments, the 5' overhang comprises 2 nucleosides.

[0129] In some embodiments, the composition comprises the oligonucleotide that inhibits the expression of target protein, the oligonucleotide comprises siRNA that comprises sense strand and antisense strand, and siRNA binds to 19-mer in human target mRNA that encodes target protein.In some embodiments, siRNA binds to 12-mer, 13-mer, 14-mer, 15-mer, 16-mer, 17-mer, 18-mer, 19-mer, 20-mer, 21-mer, 22-mer, 23-mer, 24-mer or 25-mer in human target mRNA.

[0130] In some embodiments, the composition comprises the oligonucleotide that inhibits the expression of target protein, the oligonucleotide comprises siRNA that comprises sense strand and antisense strand, and the siRNA binds to 17-mer in non-human primate target mRNA that encodes target protein.In some embodiments, the siRNA binds to 12-mer, 13-mer, 14-mer, 15-mer, 16-mer, 17-mer, 18-mer, 19-mer, 20-mer, 21-mer, 22-mer, 23-mer, 24-mer or 25-mer in non-human primate target mRNA.

[0131] In some embodiments, the composition comprises the oligonucleotide that inhibits the expression of target protein, the oligonucleotide comprises siRNA that comprises sense strand and antisense strand, and siRNA binds to 19-mer in human target mRNA that codes for target protein or combinations thereof.In some embodiments, siRNA binds to 12-mer, 13-mer, 14-mer, 15-mer, 16-mer, 17-mer, and 18-mer, 19-mer, 20-mer, 21-mer, 22-mer, 23-mer, 24-mer, or 25-mer in human target mRNA.

[0132] 2. ASO In some embodiments, the composition comprises an oligonucleotide that inhibits expression of a target oligonucleotide (e.g., mRNA), and the oligonucleotide comprises an antisense oligonucleotide (ASO). In some embodiments, the ASO is 12-30 nucleosides in length. In some embodiments, the ASO is 14-30 nucleosides in length. In some embodiments, the ASO is 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleosides in length, or a range defined by either of the two preceding numbers. In some embodiments, the ASO is 15-25 nucleosides in length. In some embodiments, the ASO is 20 nucleosides in length. In some embodiments, the ASO comprises DNA.

[0133] In some embodiments, the ASO is 12-30 nucleosides in length. In some embodiments, the ASO is 14-30 nucleosides in length. In some embodiments, the ASO is at least about 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleosides in length. In some embodiments, the ASO is at least 12 nucleotides in length. In some embodiments, the ASO is at least 14 nucleotides in length. In some embodiments, the ASO is at least 16 nucleotides in length. In some embodiments, the ASO is at least 18 nucleotides in length. In some embodiments, the ASO is at least 20 nucleotides in length. In some embodiments, the ASO is at least 22 nucleotides in length. In some embodiments, the ASO is at least 24 nucleotides in length. In some embodiments, the ASO is at least 26 nucleotides in length. In some embodiments, the ASO is at least 28 nucleotides in length. In some embodiments, the ASO is at least 30 nucleotides in length. In some embodiments, the ASO is about 10 or less, about 11 or less, about 12 or less, about 13 or less, about 14 or less, about 15 or less, about 16 or less, about 17 or less, about 18 or less, about 19 or less, about 20 or less, about 21 or less, about 22 or less, about 23 or less, about 24 or less, about 25 or less, about 26 or less, about 27 or less, about 28 or less, about 29 or less, or about 30 or less nucleosides in length. In some embodiments, the ASO is 12 or less nucleotides in length. In some embodiments, the ASO is 14 or less nucleotides in length. In some embodiments, the ASO is 16 or less nucleotides in length. In some embodiments, the ASO is 18 or less nucleotides in length. In some embodiments, the ASO is 20 or less nucleotides in length. In some embodiments, the ASO is 22 or less nucleotides in length. In some embodiments, the ASO is 24 or less nucleotides in length. In some embodiments, the ASO is 26 or less nucleotides in length. In some embodiments, the ASO is 28 or fewer nucleotides in length.In some embodiments, the ASO is 30 or fewer nucleotides in length.

[0134] In some embodiments, the composition comprises an oligonucleotide that inhibits expression of a target protein, the oligonucleotide comprises an ASO that is about 12-30 nucleosides in length and comprises a nucleoside sequence that is complementary to about 12-30 consecutive nucleosides of a full-length human pre-mRNA target sequence encoding the target protein, (i) the oligonucleotide comprises a modification that includes a modified nucleoside and / or a modified internucleoside linkage, and / or (ii) the composition comprises a pharma- ceutically acceptable carrier.

[0135] In some embodiments, the composition comprises an oligonucleotide that inhibits expression of a target protein, the oligonucleotide comprises an ASO that is about 12-30 nucleosides in length and comprises a nucleoside sequence that is complementary to about 12-30 consecutive nucleosides of a full-length human target mRNA sequence encoding the target protein, (i) the oligonucleotide comprises modifications that include modified nucleosides and / or modified internucleoside linkages, and / or (ii) the composition comprises a pharma- ceutically acceptable carrier.

[0136] 1. Oligonucleotide modification patterns In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, the oligonucleotide comprises a modification comprising a modified nucleoside and / or a modified internucleoside linkage, and / or (ii) the composition comprises a pharma- ceutically acceptable carrier. In some embodiments, the oligonucleotide comprises a modification comprising a modified nucleoside and / or a modified internucleoside linkage. In some embodiments, the oligonucleotide comprises a modified internucleoside linkage. In some embodiments, the modified internucleoside linkage comprises an alkyl phosphonate, phosphorothioate, methyl phosphonate, phosphorodithioate, alkyl phosphonothioate, phosphoramidate, carbamate, carbonate, phosphate triester, acetamidate, or carboxymethyl ester, or a combination thereof. In some embodiments, the modified internucleoside linkage comprises one or more phosphorothioate linkages. Benefits of modified internucleoside linkages may include reduced toxicity or improved pharmacokinetics.

[0137] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, the oligonucleotide comprises modified internucleoside linkages, the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 modified internucleoside linkages, or a range of modified internucleoside linkages defined by any two of the foregoing numbers. In some embodiments, the oligonucleotide comprises 18 or fewer modified internucleoside linkages. In some embodiments, the oligonucleotide comprises 20 or fewer modified internucleoside linkages. In some embodiments, the oligonucleotide has 2 or more modified internucleoside linkages, 3 or more modified internucleoside linkages, 4 or more modified internucleoside linkages, 5 or more modified internucleoside linkages, 6 or more modified internucleoside linkages, 7 or more modified internucleoside linkages, 8 or more modified internucleoside linkages, 9 or more modified internucleoside linkages, 10 or more modified internucleoside linkages, 11 or more modified internucleoside linkages, 12 or more modified internucleoside linkages, 13 or more modified internucleoside linkages, 14 or more modified internucleoside linkages, 15 or more modified internucleoside linkages, 16 or more modified internucleoside linkages, 17 or more modified internucleoside linkages, 18 or more modified internucleoside linkages, 19 or more modified internucleoside linkages, 20 or more modified internucleoside linkages, 21 or more modified internucleoside linkages, 22 or more modified internucleoside linkages, 23 or more modified internucleoside linkages, 24 or more modified internucleoside linkages, 25 or more modified internucleoside linkages, 26 or more modified internucleoside linkages, 27 or more modified internucleoside linkages, 28 or more modified internucleoside linkages, 29 or more modified internucleoside linkages, 30 or more modified internucleoside linkages, 31 or more modified internucleoside linkages, 32 or more modified internucleoside linkages, 33 or more modified internucleoside linkages, 34 or more modified internucleoside linkages, 35 or more modified internucleoside linkages, or more modified internucleoside linkages, 12 or more modified internucleoside linkages, 13 or more modified internucleoside linkages, 14 or more modified internucleoside linkages, 15 or more modified internucleoside linkages, 16 or more modified internucleoside linkages, 17 or more modified internucleoside linkages, 18 or more modified internucleoside linkages, 19 or more modified internucleoside linkages, or 20 or more modified internucleoside linkages.

[0138] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, the oligonucleotide comprising a modified nucleoside. In some embodiments, the modified nucleoside comprises a locked nucleic acid (LNA), a hexitol nucleic acid (HLA), a cyclohexene nucleic acid (CeNA), a 2'-methoxyethyl, a 2'-O-alkyl, a 2'-O-allyl, a 2'-fluoro, or a 2'-deoxy, or a combination thereof. In some embodiments, the modified nucleoside comprises an LNA. In some embodiments, the modified nucleoside comprises a 2',4' constrained ethyl nucleic acid. In some embodiments, the modified nucleoside comprises an HLA. In some embodiments, the modified nucleoside comprises a CeNA. In some embodiments, the modified nucleoside comprises a 2'-methoxyethyl group. In some embodiments, the modified nucleoside comprises a 2'-O-alkyl group. In some embodiments, the modified nucleoside comprises a 2'-O-allyl group. In some embodiments, the modified nucleoside comprises a 2'-fluoro group. In some embodiments, the modified nucleoside comprises a 2'-deoxy group. In some embodiments, the modified nucleoside comprises a 2'-O-methyl nucleoside, a 2'-deoxyfluoro nucleoside, a 2'-ON-methylacetamide (2'-O-NMA) nucleoside, a 2'-O-dimethylaminoethoxyethyl (2'-O-DMAEOE) nucleoside, a 2'-O-aminopropyl (2'-O-AP) nucleoside, or a 2'-ara-F, or a combination thereof. In some embodiments, the modified nucleoside comprises a 2'-O-methyl nucleoside. In some embodiments, the modified nucleoside comprises a 2'-deoxyfluoro nucleoside. In some embodiments, the modified nucleoside comprises a 2'-O-NMA nucleoside. In some embodiments, the modified nucleoside comprises a 2'-O-DMAEOE nucleoside. In some embodiments, the modified nucleoside comprises a 2'-O-aminopropyl (2'-O-AP) nucleoside. In some embodiments, the modified nucleoside comprises a 2'-ara-F.In some embodiments, the modified nucleoside comprises one or more 2'fluoro modified nucleosides. In some embodiments, the modified nucleoside comprises a 2'O-alkyl modified nucleoside. Advantages of the modified nucleoside may include reduced toxicity or improved pharmacokinetics.

[0139] In some embodiments, the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21 modified nucleosides, or a range of nucleosides defined by any two of the foregoing numbers. In some embodiments, the oligonucleotide comprises 19 or fewer modified nucleosides. In some embodiments, the oligonucleotide comprises 21 or fewer modified nucleosides. In some embodiments, an oligonucleotide comprises 2 or more modified nucleosides, 3 or more modified nucleosides, 4 or more modified nucleosides, 5 or more modified nucleosides, 6 or more modified nucleosides, 7 or more modified nucleosides, 8 or more modified nucleosides, 9 or more modified nucleosides, 10 or more modified nucleosides, 11 or more modified nucleosides, 12 or more modified nucleosides, 13 or more modified nucleosides, 14 or more modified nucleosides, 15 or more modified nucleosides, 16 or more modified nucleosides, 17 or more modified nucleosides, 18 or more modified nucleosides, 19 or more modified nucleosides, 20 or more modified nucleosides, or 21 or more modified nucleosides.

[0140] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, the oligonucleotide comprising a lipid attached to the 3' or 5' end of the oligonucleotide. In some embodiments, the lipid comprises cholesterol, myristoyl, palmitoyl, stearoyl, lithocholoyl, docosanoyl, docosahexaenoyl, myristyl, palmitylstearyl, or α-tocopherol, or a combination thereof.

[0141] In some embodiments, a composition comprises an oligonucleotide that inhibits expression of a target mRNA, the oligonucleotide comprising a sugar moiety. The sugar moiety may comprise an N-acetylgalactose moiety (e.g., an N-acetylgalactosamine (GalNAc) moiety), an N-acetylglucose moiety (e.g., an N-acetylglucosamine (GlcNAc) moiety), a fucose moiety, or a mannose moiety. The sugar moiety may comprise one, two, three, or more sugar molecules. The sugar moiety may be attached to the 3' or 5' end of the oligonucleotide. The sugar moiety may comprise an N-acetylgalactose moiety. The sugar moiety may comprise an N-acetylgalactosamine (GalNAc) moiety. The sugar moiety may comprise an N-acetylglucose moiety. The sugar moiety may comprise an N-acetylglucosamine (GlcNAc) moiety. The sugar moiety may comprise a fucose moiety. The sugar moiety may comprise a mannose moiety. N-acetylglucose, GlcNAc, fucose, or mannose can target or bind to mannose receptors such as CD206 and thus may be useful for targeting macrophages.

[0142] In some embodiments, the composition comprises an oligonucleotide that inhibits expression of a target mRNA, and the oligonucleotide comprises an N-acetylgalactosamine (GalNAc) moiety. GalNAc can be useful for hepatocyte targeting. In some embodiments, the composition comprises GalNAc. In some embodiments, the composition comprises a GalNAc derivative. The GalNAc moiety can comprise one, two, three, or more GalNAc molecules. The GalNAc moiety can comprise a bivalent or trivalent branched linker. The oligo can be linked to one, two, or three GalNAcs via a bivalent or trivalent branched linker. The GalNAc moiety can be linked to the 3' or 5' end of the oligonucleotide.

[0143] The oligonucleotide may contain purines. Examples of purines include adenine (A) or guanine (G), or modified versions thereof. The oligonucleotide may contain pyrimidines. Examples of pyrimidines include cytosine (C), thymine (T), or uracil (U), or modified versions thereof.

[0144] In some embodiments, the purines of the oligonucleotide comprise 2'fluoro modified purines. In some embodiments, the purines of the oligonucleotide comprise 2'-O-methyl modified purines. In some embodiments, the purines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, all of the purines of the oligonucleotide comprise 2'fluoro modified purines. In some embodiments, all of the purines of the oligonucleotide comprise 2'-O-methyl modified purines. In some embodiments, all of the purines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, 2'-O-methyl comprises 2'O-methyl.

[0145] In some embodiments, the pyrimidines of the oligonucleotide comprise 2'fluoro modified pyrimidines. In some embodiments, the pyrimidines of the oligonucleotide comprise 2'-O-methyl modified pyrimidines. In some embodiments, the pyrimidines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, all pyrimidines of the oligonucleotide comprise 2'fluoro modified pyrimidines. In some embodiments, all pyrimidines of the oligonucleotide comprise 2'-O-methyl modified pyrimidines. In some embodiments, all pyrimidines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines.

[0146] In some embodiments, the purines of the oligonucleotide comprise 2'fluoro modified purines and the pyrimidines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the oligonucleotide comprise 2'-O-methyl modified purines and the pyrimidines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the oligonucleotide comprise 2'fluoro modified purines and the pyrimidines of the oligonucleotide comprise 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the oligonucleotide comprise 2'-O-methyl modified purines and the pyrimidines of the oligonucleotide comprise 2'fluoro modified pyrimidines. In some embodiments, the pyrimidines of the oligonucleotide comprise 2'fluoro modified pyrimidines and the purines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the oligonucleotide comprise 2'-O-methyl modified pyrimidines and the purines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the oligonucleotide comprise 2'fluoro modified pyrimidines and the purines of the oligonucleotide comprise 2'-O-methyl modified purines.In some embodiments, the pyrimidines of the oligonucleotide comprise 2'-O-methyl modified pyrimidines and the purines of the oligonucleotide comprise 2'fluoro modified purines.

[0147] In some embodiments, all purines of the oligonucleotide comprise 2'fluoro modified purines and all pyrimidines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, all purines of the oligonucleotide comprise 2'-O-methyl modified purines and all pyrimidines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, all purines of the oligonucleotide comprise 2'fluoro modified purines and all pyrimidines of the oligonucleotide comprise 2'-O-methyl modified pyrimidines. In some embodiments, all purines of the oligonucleotide comprise 2'-O-methyl modified purines and all pyrimidines of the oligonucleotide comprise 2'fluoro modified pyrimidines. In some embodiments, all pyrimidines of the oligonucleotide comprise 2'fluoro modified pyrimidines and all purines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, all pyrimidines of the oligonucleotide comprise 2'-O-methyl modified pyrimidines and all purines of the oligonucleotide comprise a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, all pyrimidines of the oligonucleotide comprise 2'-fluoro modified pyrimidines and all purines of the oligonucleotide comprise 2'-O-methyl modified purines. In some embodiments, all pyrimidines of the oligonucleotide comprise 2'-O-methyl modified pyrimidines and all purines of the oligonucleotide comprise 2'-fluoro modified purines.

[0148] 2. siRNA modification pattern In some embodiments, the sense strand comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, or 29 modified internucleoside linkages, or a range of modified internucleoside linkages defined by any two of the foregoing integers. In some embodiments, the sense strand comprises 1-11 modified internucleoside linkages. In some embodiments, the sense strand comprises 2-6 modified internucleoside linkages. In some embodiments, the sense strand comprises 5 modified internucleoside linkages. In some embodiments, the sense strand comprises 4 modified internucleoside linkages.

[0149] In some embodiments, the antisense strand comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, or 29 modified internucleoside linkages, or a range of modified internucleoside linkages defined by any two of the foregoing integers. In some embodiments, the antisense strand comprises 1-11 modified internucleoside linkages. In some embodiments, the antisense strand comprises 2-6 modified internucleoside linkages. In some embodiments, the antisense strand comprises 5 modified internucleoside linkages. In some embodiments, the antisense strand comprises 4 modified internucleoside linkages.

[0150] In some embodiments, the sense strand comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 modified nucleosides, or a range of modified nucleosides defined by any two of the foregoing integers. In some embodiments, the sense strand comprises 12-19 modified nucleosides. In some embodiments, the sense strand comprises 12-21 modified nucleosides. In some embodiments, the sense strand comprises 19 modified nucleosides. In some embodiments, the sense strand comprises 21 modified nucleosides.

[0151] In some embodiments, the antisense strand comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 modified nucleosides, or a range of modified nucleosides defined by any two of the foregoing integers. In some embodiments, the antisense strand comprises 12-19 modified nucleosides. In some embodiments, the antisense strand comprises 12-21 modified nucleosides. In some embodiments, the antisense strand comprises 19 modified nucleosides. In some embodiments, the antisense strand comprises 21 modified nucleosides.

[0152] In some embodiments, the sense strand or antisense strand further comprises at least two additional nucleosides attached to the 3' end of the sense strand or antisense strand. In some embodiments, the sense strand or antisense strand comprises two additional nucleosides attached to the 3' end of the sense strand or antisense strand. As part of the sense strand, the additional nucleosides may or may not be complementary to the target mRNA. The additional nucleoside of the antisense strand may comprise uracil. The two additional nucleosides of the antisense strand may both comprise uracil.

[0153] In some embodiments, the sense strand or the sense strand further comprises at least two additional nucleosides attached to the 3' end of the sense strand or the sense strand. In some embodiments, the sense strand or the sense strand comprises two additional nucleosides attached to the 3' end of the sense strand or the sense strand. The additional nucleoside of the sense strand may comprise uracil. The two additional nucleosides of the sense strand may both comprise uracil.

[0154] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, the sense strand comprises a modification pattern. In some embodiments, the sense strand comprises a modification pattern 1S:5'-NfsnsNfnNfnNfNfNfNfnNfnNfnNfnNfnNfsnsn-3', where "Nf" is a 2'fluoro modified nucleoside, "n" is a 2'O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the sense strand comprises a modification pattern 1S#2:5'-NfnNfnNfnNfNfNfNfNfnNfnNfnNfnNfnNfsnsn-3', where "Nf" is a 2'fluoro modified nucleoside, "n" is a 2'O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the sense strand comprises the modification pattern 2S:5'-nsnsnnNfnNfNfNfnnnnnnnnnnsnsn-3', where "Nf" is a 2'fluoro modified nucleoside, "n" is a 2'O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the sense strand comprises the modification pattern 2S#2:5'-nnnnNfnNfNfNfnnnnnnnnnnsnsn-3', where "Nf" is a 2'fluoro modified nucleoside, "n" is a 2'O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the sense strand comprises the modification pattern 3S:5'-nsnsnnNfnNfnNfnnnnnnnnnnsnsn-3', where "Nf" is a 2'fluoro modified nucleoside, "n" is a 2'O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the sense strand comprises the modification pattern 3S#2:5'-nnnnNfnNfnNfnnnnnnnnnnsnsn-3', where "Nf" is a 2'fluoro modified nucleoside, "n" is a 2'O-methyl modified nucleoside, and "s" is a phosphorothioate linkage.In some embodiments, the sense strand comprises the modification pattern 4S:5'-NfsnsNfnNfnNfNfNfNfnNfnNfnNfnNfnNfsnsnN-3', where "Nf" is a 2' fluoro modified nucleoside, "n" is a 2' O-methyl modified nucleoside, "s" is a phosphorothioate linkage, and N comprises one or more nucleosides. In some embodiments, the sense strand comprises the modification pattern 4S#2:5'-NfnNfnNfnNfNfNfNfNfnNfnNfnNfnNfnNfsnsnN-3', where "Nf" is a 2' fluoro modified nucleoside, "n" is a 2' O-methyl modified nucleoside, "s" is a phosphorothioate linkage, and N comprises one or more nucleosides. In some embodiments, the sense strand comprises the modification pattern 5S:5'-nsnsnnNfnNfNfNfnnnnnnnnnnsnsnN-3', where "Nf" is a 2'fluoro modified nucleoside, "n" is a 2'O-methyl modified nucleoside, "s" is a phosphorothioate linkage, and N comprises one or more nucleosides. In some embodiments, the sense strand comprises the modification pattern 5S#2:5'-nnnnNfnNfNfNfnnnnnnnnnnsnsnN-3', where "Nf" is a 2'fluoro modified nucleoside, "n" is a 2'O-methyl modified nucleoside, "s" is a phosphorothioate linkage, and N comprises one or more nucleosides. In some embodiments, the sense strand comprises the modification pattern 6S:5'-NfsnsNfnNfnNfnNfnNfnNfnNfnNfnNfnNfsnsn-3', where "Nf" is a 2' fluoro modified nucleoside, "n" is a 2' O-methyl modified nucleoside, "s" is a phosphorothioate linkage, and N comprises one or more nucleosides. In some embodiments, the sense strand comprises the modification pattern 6S#2:5'-NfnNfnNfnNfnNfnNfnNfnNfnNfnNfnNfnNfsnsn-3', where "Nf" is a 2' fluoro modified nucleoside, "n" is a 2' O-methyl modified nucleoside, "s" is a phosphorothioate linkage, and N comprises one or more nucleosides.In some embodiments, the sense strand comprises any one of modification patterns 1S, 2S, 3S, 4S, 5S, or 6S. In some embodiments, the sense strand comprises any one of modification patterns 1S#2, 2S#2, 3S#2, 4S#2, 5S#2, or 6S#2. In some embodiments, the sense strand comprises any one of modification patterns 1S, 3S, 4S, or 6S. In some embodiments, the sense strand comprises any one of modification patterns 1S#2, 3S#2, 4S#2, or 6S#2. Any one of the modification patterns 1S-6S may include a GalNAc ligand attached to the 3' terminus. Any one of the modification patterns 1S-6S may include a GalNAc ligand attached to the 5' terminus. Any one of the modification patterns 1S-6S#2 may include a GalNAc ligand attached to the 3' terminus. Any one of the modification patterns 1S-6S#2 may include a GalNAc ligand attached to the 5' terminus.

[0155] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, and the antisense strand comprises a modification pattern. In some embodiments, the antisense strand comprises a modification pattern 1AS:5'-nsNfsnNfnNfnNfnNfnnnNfnNfnNfnsnsn-3', where "Nf" is a 2'fluoro-modified nucleoside, "n" is a 2'O-methyl-modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the antisense strand comprises a modification pattern 2AS:5'-nsNfsnnnNfnNfNfNfnnnnNfnNfnnnsnsn-3', where "Nf" is a 2'fluoro-modified nucleoside, "n" is a 2'O-methyl-modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the antisense strand comprises the modification pattern 3AS:5'-nsNfsnnnNfnnnnnnnNfnNfnnnsnsn-3', where "Nf" is a 2' fluoro modified nucleoside, "n" is a 2' O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the antisense strand comprises the modification pattern 4AS:5'-nsNfsnNfnNfnnnnnnnNfnNfnnnsnsn-3', where "Nf" is a 2' fluoro modified nucleoside, "n" is a 2' O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the antisense strand comprises the modification pattern 5AS:5'-nsNfsnnnNfnNfnnnnnNfnNfnnnsnsn-3', where "Nf" is a 2'fluoro modified nucleoside, "n" is a 2'O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the antisense strand comprises the modification pattern 6AS:5'-nsNfsnNfnNfnNfnNfnNfnNfnNfnNfnNfnsnsn-3', where "Nf" is a 2'fluoro modified nucleoside, "n" is a 2'O-methyl modified nucleoside, and "s" is a phosphorothioate linkage.In some embodiments, the antisense strand comprises the modification pattern 7AS:5'-nsNfsnNfnNfnNfNfnnnnNfnNfnnnsnsn-3', where "Nf" is a 2' fluoro modified nucleoside, "n" is a 2' O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the antisense strand comprises the modification pattern 8AS:5'-nsNfsnnnnnnnnnnNfnnnnnsnsn-3', where "Nf" is a 2' fluoro modified nucleoside, "n" is a 2' O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the antisense strand comprises the modification pattern 9AS: 5'-nsNfsnnnNfnnnnnnnNfnNfnNfnsnsn-3', where "Nf" is a 2'fluoro modified nucleoside, "n" is a 2'O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. Any one of the modification patterns 1AS-9AS may comprise a GalNAc ligand attached to the 3' terminus. Any one of the modification patterns 1AS-9AS may comprise a GalNAc ligand attached to the 5' terminus.

[0156] The modification in any of the modification patterns may be optional. For example, one, two, three or more phosphorothioate bonds in any of the modification patterns 1S-6S, 1S#2-6S#2 or 1AS-9AS may be replaced with unmodified bonds or different modified bonds. In some cases, one, two, three or more modified nucleosides in any of the modification patterns 1S-6S or 1AS-9AS may be replaced with unmodified nucleosides or different modified nucleosides.

[0157] In some embodiments, the composition comprises an oligonucleotide that inhibits expression of a target mRNA, the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, the sense strand comprises a modification pattern 1S and the antisense strand comprises a modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises a modification pattern 2S and the antisense strand comprises a modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises a modification pattern 3S and the antisense strand comprises a modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises a modification pattern 4S and the antisense strand comprises a modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises modification pattern 5S and the antisense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises modification pattern 6S and the antisense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises modification pattern 1S#2 and the antisense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises modification pattern 2S#2 and the antisense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises modification pattern 3S#2 and the antisense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS.In some embodiments, the sense strand comprises modification pattern 4S#2 and the antisense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises modification pattern 5S#2 and the antisense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises modification pattern 6S#2 and the antisense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the antisense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand or the antisense strand comprises modification pattern ASO1.

[0158] Any combination of sense and antisense modification patterns may be used. In some embodiments, the sense strand comprises modification pattern 1S and the antisense strand comprises modification pattern 1AS. In some embodiments, the sense strand comprises modification pattern 2S and the antisense strand comprises modification pattern 2AS. In some embodiments, the sense strand comprises modification pattern 2S and the antisense strand comprises modification pattern 3AS. In some embodiments, the sense strand comprises modification pattern 3S and the antisense strand comprises modification pattern 1AS. In some embodiments, the sense strand comprises modification pattern 3S and the antisense strand comprises modification pattern 4AS. In some embodiments, the sense strand comprises modification pattern 3S and the antisense strand comprises modification pattern 5AS. In some embodiments, the sense strand comprises modification pattern 3S and the antisense strand comprises modification pattern 6AS. In some embodiments, the sense strand comprises modification pattern 3S and the antisense strand comprises modification pattern 7AS. In some embodiments, the sense strand comprises modification pattern 3S and the antisense strand comprises modification pattern 8AS. In some embodiments, the sense strand comprises modification pattern 6S and the antisense strand comprises modification pattern 1AS. In some embodiments, the sense strand comprises modification pattern 6S and the antisense strand comprises modification pattern 4AS. In some embodiments, the sense strand comprises modification pattern 6S and the antisense strand comprises modification pattern 5AS. In some embodiments, the sense strand comprises modification pattern 6S and the antisense strand comprises modification pattern 6AS. In some embodiments, the sense strand comprises modification pattern 6S and the antisense strand comprises modification pattern 7AS. In some embodiments, the sense strand comprises modification pattern 6S and the antisense strand comprises modification pattern 8AS. In some embodiments, the sense strand comprises modification pattern 1S#2 and the antisense strand comprises modification pattern 1AS. In some embodiments, the sense strand comprises modification pattern 2S#2 and the antisense strand comprises modification pattern 2AS. In some embodiments, the sense strand comprises modification pattern 2S#2 and the antisense strand comprises modification pattern 3AS.In some embodiments, the sense strand comprises modification pattern 3S#2 and the antisense strand comprises modification pattern 1AS. In some embodiments, the sense strand comprises modification pattern 3S#2 and the antisense strand comprises modification pattern 4AS. In some embodiments, the sense strand comprises modification pattern 3S#2 and the antisense strand comprises modification pattern 5AS. In some embodiments, the sense strand comprises modification pattern 3S#2 and the antisense strand comprises modification pattern 6AS. In some embodiments, the sense strand comprises modification pattern 3S#2 and the antisense strand comprises modification pattern 7AS. In some embodiments, the sense strand comprises modification pattern 3S#2 and the antisense strand comprises modification pattern 8AS. In some embodiments, the sense strand comprises modification pattern 6S#2 and the antisense strand comprises modification pattern 1AS. In some embodiments, the sense strand comprises modification pattern 6S#2 and the antisense strand comprises modification pattern 4AS. In some embodiments, the sense strand comprises modification pattern 6S#2 and the antisense strand comprises modification pattern 5AS. In some embodiments, the sense strand comprises modification pattern 6S#2 and the antisense strand comprises modification pattern 6AS. In some embodiments, the sense strand comprises modification pattern 6S#2 and the antisense strand comprises modification pattern 7AS, hi some embodiments, the sense strand comprises modification pattern 6S#2 and the antisense strand comprises modification pattern 8AS.

[0159] In some embodiments, the purines in the sense strand comprise 2'fluoro modified purines. In some embodiments, the purines in the sense strand comprise 2'-O-methyl modified purines. In some embodiments, the purines in the sense strand comprise a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, all purines in the sense strand comprise 2'fluoro modified purines. In some embodiments, all purines in the sense strand comprise 2'-O-methyl modified purines. In some embodiments, all purines in the sense strand comprise a mixture of 2'fluoro and 2'-O-methyl modified purines.

[0160] In some embodiments, the pyrimidines of the sense strand comprise 2'fluoro modified pyrimidines. In some embodiments, the pyrimidines of the sense strand comprise 2'-O-methyl modified pyrimidines. In some embodiments, the pyrimidines of the sense strand comprise a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, all pyrimidines of the sense strand comprise 2'fluoro modified pyrimidines. In some embodiments, all pyrimidines of the sense strand comprise 2'-O-methyl modified pyrimidines. In some embodiments, all pyrimidines of the sense strand comprise a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines.

[0161] In some embodiments, the purines of the sense strand include 2'fluoro modified purines and the pyrimidines of the sense strand include a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the sense strand include 2'-O-methyl modified purines and the pyrimidines of the sense strand include a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the sense strand include 2'fluoro modified purines and the pyrimidines of the sense strand include 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the sense strand include 2'-O-methyl modified purines and the pyrimidines of the sense strand include 2'fluoro modified pyrimidines. In some embodiments, the pyrimidines of the sense strand include 2'fluoro modified pyrimidines and the purines of the sense strand include a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the sense strand include 2'-O-methyl modified pyrimidines and the purines of the sense strand include a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the sense strand comprise 2'-fluoro modified pyrimidines and the purines of the sense strand comprise 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the sense strand comprise 2'-O-methyl modified pyrimidines and the purines of the sense strand comprise 2'-fluoro modified purines.

[0162] In some embodiments, all purines in the sense strand comprise 2'-fluoro modified purines and all pyrimidines in the sense strand comprise a mixture of 2'-fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, all purines in the sense strand comprise 2'-O-methyl modified purines and all pyrimidines in the sense strand comprise a mixture of 2'-fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, all purines in the sense strand comprise 2'-fluoro modified purines and all pyrimidines in the sense strand comprise 2'-O-methyl modified pyrimidines. In some embodiments, all purines in the sense strand comprise 2'-O-methyl modified purines and all pyrimidines in the sense strand comprise 2'-fluoro modified pyrimidines. In some embodiments, all pyrimidines in the sense strand comprise 2'-fluoro modified pyrimidines and all purines in the sense strand comprise a mixture of 2'-fluoro and 2'-O-methyl modified purines. In some embodiments, all pyrimidines in the sense strand comprise 2'-O-methyl modified pyrimidines and all purines in the sense strand comprise a mixture of 2'-fluoro and 2'-O-methyl modified purines. In some embodiments, all pyrimidines in the sense strand comprise 2'-fluoro modified pyrimidines and all purines in the sense strand comprise 2'-O-methyl modified purines. In some embodiments, all pyrimidines in the sense strand comprise 2'-O-methyl modified pyrimidines and all purines in the sense strand comprise 2'-fluoro modified purines.

[0163] In some embodiments, the purines in the antisense strand comprise 2'fluoro modified purines. In some embodiments, the purines in the antisense strand comprise 2'-O-methyl modified purines. In some embodiments, the purines in the antisense strand comprise a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, all purines in the antisense strand comprise 2'fluoro modified purines. In some embodiments, all purines in the antisense strand comprise 2'-O-methyl modified purines. In some embodiments, all purines in the antisense strand comprise a mixture of 2'fluoro and 2'-O-methyl modified purines.

[0164] In some embodiments, the pyrimidines of the antisense strand comprise 2'fluoro modified pyrimidines. In some embodiments, the pyrimidines of the antisense strand comprise 2'-O-methyl modified pyrimidines. In some embodiments, the pyrimidines of the antisense strand comprise a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, all pyrimidines of the antisense strand comprise 2'fluoro modified pyrimidines. In some embodiments, all pyrimidines of the antisense strand comprise 2'-O-methyl modified pyrimidines. In some embodiments, all pyrimidines of the antisense strand comprise a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines.

[0165] In some embodiments, the purines of the antisense strand include 2'fluoro modified purines and the pyrimidines of the antisense strand include a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the antisense strand include 2'-O-methyl modified purines and the pyrimidines of the antisense strand include a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the antisense strand include 2'fluoro modified purines and the pyrimidines of the antisense strand include 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the antisense strand include 2'-O-methyl modified purines and the pyrimidines of the antisense strand include 2'fluoro modified pyrimidines. In some embodiments, the pyrimidines of the antisense strand include 2'fluoro modified pyrimidines and the purines of the antisense strand include a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the antisense strand include 2'-O-methyl modified pyrimidines and the purines of the antisense strand include a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the antisense strand include 2'fluoro modified pyrimidines and the purines of the antisense strand include 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the antisense strand include 2'-O-methyl modified pyrimidines and the purines of the antisense strand include 2'fluoro modified purines.

[0166] In some embodiments, all purines in the antisense strand comprise 2'-fluoro modified purines and all pyrimidines in the antisense strand comprise a mixture of 2'-fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, all purines in the antisense strand comprise 2'-O-methyl modified purines and all pyrimidines in the antisense strand comprise a mixture of 2'-fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, all purines in the antisense strand comprise 2'-fluoro modified purines and all pyrimidines in the antisense strand comprise 2'-O-methyl modified pyrimidines. In some embodiments, all purines in the antisense strand comprise 2'-O-methyl modified purines and all pyrimidines in the antisense strand comprise 2'-fluoro modified pyrimidines. In some embodiments, all pyrimidines in the antisense strand comprise 2'-fluoro modified pyrimidines and all purines in the antisense strand comprise a mixture of 2'-fluoro and 2'-O-methyl modified purines. In some embodiments, all pyrimidines in the antisense strand comprise 2'-O-methyl modified pyrimidines and all purines in the antisense strand comprise a mixture of 2'fluoro and 2'-O-methyl modified purines. In some embodiments, all pyrimidines in the antisense strand comprise 2'fluoro modified pyrimidines and all purines in the antisense strand comprise 2'-O-methyl modified purines. In some embodiments, all pyrimidines in the antisense strand comprise 2'-O-methyl modified pyrimidines and all purines in the antisense strand comprise 2'fluoro modified purines.

[0167] In some cases, the oligonucleotide comprises a specific modification pattern. In some embodiments, the 9th position counting from the 5' end of the oligonucleotide strand can have a 2'F modification. In some embodiments, if the 9th position of the oligonucleotide strand is a pyrimidine, all purines in the oligonucleotide strand have a 2'OMe modification. In some embodiments, if the 9th position is the only pyrimidine between the 5th and 11th positions of the sense strand, the 9th position is the only position in the oligonucleotide strand that has a 2'F modification. In some embodiments, if the 9th position of the oligonucleotide strand and one other base between the 5th and 11th positions are only pyrimidines, both of these pyrimidines are the only two positions in the oligonucleotide strand that have a 2'F modification. In some embodiments, if the 9th position of the oligonucleotide strand and two other bases between the 5th and 11th positions are only pyrimidines, and these two other pyrimidines are adjacent positions, so that there are not three 2'F modifications in a row, any combination of 2'F modifications that results in a total of three 2'F modifications can be made. In some embodiments, if there are more than two pyrimidines between positions 5 and 11 of an oligonucleotide strand, all combinations of pyrimidines with 2'F modifications are possible, with a total of 3-5 2'F modifications, provided that no oligonucleotide strand has three 2'F modifications in a row. In some cases, either oligonucleotide strand of the siRNA contains a modification pattern that fits any or all of these strands of oligonucleotide rules.

[0168] In some embodiments, if position 9 of the oligonucleotide strand is a purine, all purines in the oligonucleotide strand have a 2'OMe modification. In some embodiments, if position 9 is the only purine between positions 5 and 11 of the sense strand, then position 9 is the only position in the oligonucleotide strand with a 2'F modification. In some embodiments, if position 9 of the oligonucleotide strand and one other base between positions 5 and 11 are only purines, both of these purines are the only two positions in the oligonucleotide strand with a 2'F modification. In some embodiments, if position 9 of the oligonucleotide strand and two other bases between positions 5 and 11 are only purines, and these two other purines are in adjacent positions, such that there are not three 2'F modifications in a row, any combination of 2'F modifications resulting in a total of three 2'F modifications can be made. In some embodiments, if there are more than two purines between positions 5 and 11 of the oligonucleotide strand, then all combinations of purines with 2'F modifications are possible, with a total of three to five 2'F modifications, provided that the oligonucleotide strand does not have three 2'F modifications in a row. In some cases, either oligonucleotide strand of the siRNA contains a modification pattern that conforms to any or all of these strands of oligonucleotide rules.

[0169] In some cases, the 9th position of the oligonucleotide strand can be 2'deoxy.In these cases, the 2'F and 2'OMe modifications can occur at other positions of the oligonucleotide strand.In some cases, either oligonucleotide strand of siRNA contains a modification pattern that conforms to these oligonucleotide strand rules.

[0170] In some embodiments, position 9 of the sense strand comprises a 2'fluoro modified pyrimidine. In some embodiments, all purines of the sense strand comprise 2'-O-methyl modified purines. In some embodiments, 1, 2, 3, 4, or 5 pyrimidines between positions 5 and 11 comprise 2'fluoro modified pyrimidines, provided that there are not three 2'fluoro modified pyrimidines in a row. In some embodiments, odd-numbered positions of the antisense strand comprise 2'-O-methyl modified nucleotides. In some embodiments, even-numbered positions of the antisense strand comprise 2'fluoro modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, even-numbered positions of the antisense strand comprise 2'fluoro modified nucleotides, 2'-O-methyl modified nucleotides, and unmodified deoxyribonucleotides. In some embodiments, position 9 of the sense strand comprises a 2' fluoro modified pyrimidine, all purines of the sense strand comprise 2'-O-methyl modified purines, 1, 2, 3, 4, or 5 pyrimidines between positions 5 and 11 comprise 2' fluoro modified pyrimidines, provided that there are not three 2' fluoro modified pyrimidines in a row, odd-numbered positions of the antisense strand comprise 2'-O-methyl modified nucleotides, and even-numbered positions of the antisense strand comprise 2' fluoro modified nucleotides and unmodified deoxyribonucleotides.

[0171] In some embodiments, position 9 of the sense strand comprises a 2'fluoro modified purine. In some embodiments, all pyrimidines of the sense strand comprise a 2'-O-methyl modified purine. In some embodiments, 1, 2, 3, 4, or 5 purines between positions 5 and 11 comprise a 2'fluoro modified purine, provided that there are not three 2'fluoro modified purines in a row. In some embodiments, odd-numbered positions of the antisense strand comprise a 2'-O-methyl modified nucleotide. In some embodiments, even-numbered positions of the antisense strand comprise a 2'fluoro modified nucleotide and an unmodified deoxyribonucleotide. In some embodiments, even-numbered positions of the antisense strand comprise a 2'fluoro modified nucleotide, a 2'-O-methyl modified nucleotide, and an unmodified deoxyribonucleotide. In some embodiments, position 9 of the sense strand comprises a 2'fluoro modified purine, all pyrimidines of the sense strand comprise 2'-O-methyl modified pyrimidines, 1, 2, 3, 4, or 5 purines between positions 5 and 11 comprise 2'fluoro modified purines, provided that there are not three 2'fluoro modified purines in a row, odd positions of the antisense strand comprise 2'-O-methyl modified nucleotides, and even positions of the antisense strand comprise 2'fluoro modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, there are not three 2'fluoro modified purines in a row. In some embodiments, there are not three 2'fluoro modified pyrimidines in a row.

[0172] In some embodiments, position 9 of the sense strand comprises unmodified deoxyribonucleotides. In some embodiments, positions 5, 7, and 8 of the sense strand comprise 2'fluoro modified nucleotides. In some embodiments, all pyrimidines at positions 10-21 of the sense strand comprise 2'-O-methyl modified pyrimidines and all purines at positions 10-21 of the sense strand comprise 2'-O-methyl modified purines or 2'fluoro modified purines. In some embodiments, odd-numbered positions of the antisense strand comprise 2'-O-methyl modified nucleotides. In some embodiments, even-numbered positions of the antisense strand comprise 2'fluoro modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, even-numbered positions of the antisense strand comprise 2'fluoro modified nucleotides, 2'-O-methyl modified nucleotides, and unmodified deoxyribonucleotides. In some embodiments, position 9 of the sense strand comprises unmodified deoxyribonucleotides, positions 5, 7, and 8 of the sense strand comprise 2' fluoro modified nucleotides, all pyrimidines at positions 10-21 of the sense strand comprise 2'-O-methyl modified pyrimidines and all purines at positions 10-21 comprise 2'-O-methyl modified purines or 2' fluoro modified purines, odd positions of the antisense strand comprise 2'-O-methyl modified nucleotides, and even positions of the antisense strand comprise 2' fluoro modified nucleotides and unmodified deoxyribonucleotides.

[0173] In some embodiments, position 9 of the sense strand comprises unmodified deoxyribonucleotides. In some embodiments, positions 5, 7, and 8 of the sense strand comprise 2'fluoro modified nucleotides. In some embodiments, all purines at positions 10-21 of the sense strand comprise 2'-O-methyl modified purines and all pyrimidines at positions 10-21 comprise 2'-O-methyl modified pyrimidines or 2'fluoro modified pyrimidines. In some embodiments, odd-numbered positions of the antisense strand comprise 2'-O-methyl modified nucleotides. In some embodiments, even-numbered positions of the antisense strand comprise 2'fluoro modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, even-numbered positions of the antisense strand comprise 2'fluoro modified nucleotides, 2'-O-methyl modified nucleotides, and unmodified deoxyribonucleotides. In some embodiments, position 9 of the sense strand comprises unmodified deoxyribonucleotides, positions 5, 7, and 8 of the sense strand comprise 2' fluoro modified nucleotides, all purines at positions 10-21 of the sense strand comprise 2'-O-methyl modified purines and all pyrimidines at positions 10-21 comprise 2'-O-methyl modified pyrimidines or 2' fluoro modified pyrimidines, odd positions of the antisense strand comprise 2'-O-methyl modified nucleotides, and even positions of the antisense strand comprise 2' fluoro modified nucleotides and unmodified deoxyribonucleotides.

[0174] 3. ASO modification pattern In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, and the oligonucleotide comprises an antisense oligonucleotide (ASO). In some embodiments, the ASO comprises the modification pattern ASO1:5'nsnsnsnsnsdNsdNsdNsdNsdNsdNsdNsdNsdNsdNsdNsnsnsnsn-3', where "dN" is any deoxynucleotide, "n" is a 2'O-methyl or 2'O-methoxyethyl modified nucleoside, and "s" is a phosphorothioate linkage. In some embodiments, the ASO comprises the modification pattern 1S, 2S, 3S, 4S, 5S, 6S, 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS.

[0175] In some embodiments, the ASO is conjugated to a GalNAc moiety. The GalNAc moiety can be conjugated to the ASO at the 5' or 3' end. In some embodiments, the GalNAc moiety is conjugated to the 5' end of the ASO. In some embodiments, the GalNAc moiety is conjugated to the 3' end of the ASO.

[0176] C. Preparation In some embodiments, the composition is a pharmaceutical composition.In some embodiments, the composition is sterile.In some embodiments, the composition further comprises a pharma- ceutically acceptable carrier.The formulation can comprise a compound such as GalNAc moiety and the oligonucleotide conjugated to GalNAc moiety as described herein.

[0177] In some embodiments, the pharma- ceutically acceptable carrier comprises water. In some embodiments, the pharma- ceutically acceptable carrier comprises a buffer. In some embodiments, the pharma- ceutically acceptable diluent is saline. In some embodiments, the pharma- ceutically acceptable carrier comprises water, a buffer, or saline. In some embodiments, the composition comprises a liposome. In some embodiments, the pharma- ceutically acceptable carrier comprises a liposome, a lipid, a nanoparticle, a protein, a protein-antibody complex, a peptide, cellulose, a nanogel, or a combination thereof.

[0178] II. Methods and Uses Disclosed herein, in some embodiments, is a method of administering the compositions described herein to a subject. Some embodiments relate to the use of the compositions described herein, such as administering the compositions to a subject.

[0179] Some embodiments relate to a method of treating a disorder in a subject in need of treatment. Some embodiments relate to the use of a composition described herein in a treatment method. Some embodiments include administering a composition described herein to a subject suffering from the disorder. In some embodiments, the administration treats the disorder in the subject. In some embodiments, the composition treats the disorder in the subject.

[0180] In some embodiments, treatment includes preventing, inhibiting, or reversing a disorder in a subject. Some embodiments relate to the use of a composition described herein in a method of preventing, inhibiting, or reversing a disorder. Some embodiments relate to a method of preventing, inhibiting, or reversing a disorder in a subject. Some embodiments include administering a composition described herein to a subject suffering from a disorder. In some embodiments, the administration prevents, inhibits, or reverses a disorder in the subject. In some embodiments, the composition prevents, inhibits, or reverses a disorder in the subject.

[0181] Some embodiments relate to a method of preventing a disorder in a subject. Some embodiments relate to the use of a composition described herein in a method of preventing a disorder. Some embodiments include administering a composition described herein to a subject suffering from the disorder. In some embodiments, the administration prevents the disorder in the subject. In some embodiments, the composition prevents the disorder in the subject.

[0182] Some embodiments relate to a method of inhibiting a disorder in a subject. Some embodiments relate to the use of a composition described herein in a method of inhibiting a disorder. Some embodiments include administering a composition described herein to a subject suffering from the disorder. In some embodiments, the administration inhibits the disorder in the subject. In some embodiments, the composition inhibits the disorder in the subject.

[0183] Some embodiments relate to a method of reversing a disorder in a subject. Some embodiments relate to the use of a composition described herein in a method of reversing a disorder. Some embodiments include administering a composition described herein to a subject suffering from a disorder. In some embodiments, the administration reverses the disorder in the subject. In some embodiments, the composition reverses the disorder in the subject.

[0184] A. Disability Some embodiments of the methods described herein include treating a disorder in a subject in need of treatment. In some embodiments, the disorder is a liver disorder. Non-limiting examples of liver disorders include hepatitis, liver cancer, liver fibrosis, cholestasis, gallbladder disease, biliary tree disease, alcoholic liver disease, non-alcoholic steatohepatitis, liver infection, or genetic liver disorder. In some embodiments, the liver disorder includes hepatitis. In some embodiments, the liver disorder includes liver cancer. In some embodiments, the liver disorder includes liver fibrosis. In some embodiments, the liver disorder includes cholestasis. In some embodiments, the liver disorder includes gallbladder disease. In some embodiments, the liver disorder includes biliary tree disease. In some embodiments, the liver disorder is alcoholic liver disease. In some embodiments, the liver disorder includes non-alcoholic steatohepatitis.

[0185] In some embodiments, the liver disorder comprises a liver infection. In some embodiments, the liver infection comprises Hepatitis A. In some embodiments, the liver infection comprises Hepatitis B. In some embodiments, the liver infection comprises Hepatitis C.

[0186] In some embodiments, the liver disorder comprises a genetic liver disorder. In some embodiments, the genetic liver disorder comprises hemochromatosis. In some embodiments, the genetic liver disorder comprises Wilson's disease.

[0187] B. Target Some embodiments of the methods described herein include treatment of a subject. Non-limiting examples of subjects include vertebrates, animals, mammals, dogs, cats, cows, rodents, mice, rats, primates, monkeys, and humans. In some embodiments, the subject is a vertebrate. In some embodiments, the subject is an animal. In some embodiments, the subject is a mammal. In some embodiments, the subject is a dog. In some embodiments, the subject is a cat. In some embodiments, the subject is a cow. In some embodiments, the subject is a mouse. In some embodiments, the subject is a rat. In some embodiments, the subject is a primate. In some embodiments, the subject is a monkey. In some embodiments, the subject is an animal, mammal, dog, cat, cow, rodent, mouse, rat, primate, or monkey. In some embodiments, the subject is a human.

[0188] In some embodiments, the subject is male. In some embodiments, the subject is female.

[0189] In some embodiments, the subject has a Body Mass Index (BMI) of 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, or greater, or in a range defined by any two of the foregoing integers. In some embodiments, the subject is overweight. In some embodiments, the subject has a BMI of 25 or greater. In some embodiments, the subject has a BMI between 25 and 29. In some embodiments, the subject is obese. In some embodiments, the subject has a BMI of 30 or greater. In some embodiments, the subject has a BMI between 30 and 39. In some embodiments, the subject has a BMI between 40 and 50. In some embodiments, the subject has a BMI between 25 and 50.

[0190] In some embodiments, the subject is 90 years of age or older. In some embodiments, the subject is 85 years of age or older. In some embodiments, the subject is 80 years of age or older. In some embodiments, the subject is 70 years of age or older. In some embodiments, the subject is 60 years of age or older. In some embodiments, the subject is 50 years of age or older. In some embodiments, the subject is 40 years of age or older. In some embodiments, the subject is 30 years of age or older. In some embodiments, the subject is 20 years of age or older. In some embodiments, the subject is 10 years of age or older. In some embodiments, the subject is 1 year of age or older. In some embodiments, the subject is zero years of age or older.

[0191] In some embodiments, the subject is 100 years old or younger. In some embodiments, the subject is 90 years old or younger. In some embodiments, the subject is 85 years old or younger. In some embodiments, the subject is 80 years old or younger. In some embodiments, the subject is 70 years old or younger. In some embodiments, the subject is 60 years old or younger. In some embodiments, the subject is 50 years old or younger. In some embodiments, the subject is 40 years old or younger. In some embodiments, the subject is 30 years old or younger. In some embodiments, the subject is 20 years old or younger. In some embodiments, the subject is 10 years old or younger. In some embodiments, the subject is 1 year old or younger.

[0192] In some embodiments, the subject is 0-100 years old. In some embodiments, the subject is 20-90 years old. In some embodiments, the subject is 30-80 years old. In some embodiments, the subject is 40-75 years old. In some embodiments, the subject is 50-70 years old. In some embodiments, the subject is 40-85 years old.

[0193] C. Baseline Measurements Some embodiments of the method described herein include obtaining baseline measurements from a subject.For example, in some embodiments, baseline measurements are obtained from a subject before treating the subject.Non-limiting examples of baseline measurements include baseline symptom (e.g., liver damage symptom) measurements, baseline protective phenotype measurements, baseline target oligonucleotide (e.g., mRNA) measurements, or baseline target protein measurements.

[0194] In some embodiments, the baseline measurements are obtained directly from the subject. In some embodiments, the baseline measurements are obtained by observation, for example, by observation of the subject or tissues of the subject. In some embodiments, the baseline measurements are obtained non-invasively using an imaging device. In some embodiments, the baseline measurements are obtained from a sample of the subject. In some embodiments, the baseline measurements are obtained in one or more histological tissue sections. In some embodiments, the baseline measurements are obtained by performing an assay, such as an immunoassay, a colorimetric assay, or a fluorescent assay, on a sample obtained from the subject. In some embodiments, the baseline measurements are obtained by an immunoassay, a colorimetric assay, or a fluorescent assay. In some embodiments, the baseline measurements are obtained by PCR.

[0195] In some embodiments, the baseline measure is a baseline symptom measure. The symptom can be a symptom of a disorder associated with the target oligonucleotide. The disorder can be a liver disorder.

[0196] In some embodiments, the baseline measurement is a baseline protective phenotype measurement. The protective phenotype may protect the subject from a disorder associated with the target oligonucleotide. The protective phenotype may be inversely correlated with the incidence of the disorder.

[0197] In some embodiments, the baseline measurement is a baseline target protein measurement. In some embodiments, the baseline target protein measurement comprises a baseline target protein level. In some embodiments, the baseline target protein level is expressed as mass or percentage of target protein per sample weight. In some embodiments, the baseline target protein level is expressed as mass or percentage of target protein per sample volume. In some embodiments, the baseline target protein level is expressed as mass or percentage of target protein per total protein in the sample. In some embodiments, the baseline target protein measurement is a baseline liver or hepatocyte target protein measurement. In some embodiments, the baseline target protein measurement is a baseline circulating target protein measurement. In some embodiments, the baseline target protein measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescent assay.

[0198] In some embodiments, the baseline measurement is a baseline target mRNA measurement. In some embodiments, the baseline target mRNA measurement is a baseline target mRNA level. In some embodiments, the baseline target mRNA level is expressed as mass or percentage of target mRNA per sample weight. In some embodiments, the baseline target mRNA level is expressed as mass or percentage of target mRNA per sample volume. In some embodiments, the baseline target mRNA level is expressed as mass or percentage of target mRNA per total mRNA in the sample. In some embodiments, the baseline target mRNA level is expressed as mass or percentage of target mRNA per total nucleic acid in the sample. In some embodiments, the baseline target mRNA level is expressed relative to another mRNA level, such as the mRNA level of a housekeeping gene in the sample. In some embodiments, the baseline target mRNA measurement is a baseline liver or hepatocyte target mRNA measurement. In some embodiments, the baseline target mRNA measurement is obtained by an assay, such as a polymerase chain reaction (PCR) assay. In some embodiments, the PCR comprises quantitative PCR (qPCR). In some embodiments, the PCR comprises reverse transcription of the target mRNA.

[0199] Some embodiments of the methods described herein include obtaining a sample from a subject. In some embodiments, a baseline measurement is obtained in a sample obtained from the subject. In some embodiments, a sample is obtained from a subject prior to administration or treatment of the subject with a composition described herein. In some embodiments, a baseline measurement is obtained in a sample obtained from a subject prior to administration of a composition to the subject. In some embodiments, a sample is obtained from a subject in a fasted state. In some embodiments, a sample is obtained from a subject after an overnight fasting period. In some embodiments, a sample is administered to a subject in a fed state.

[0200] In some embodiments, the sample comprises a fluid. In some embodiments, the sample is a fluid sample. In some embodiments, the sample is a blood, plasma, or serum sample. In some embodiments, the sample comprises blood. In some embodiments, the sample is a blood sample. In some embodiments, the sample is a whole blood sample. In some embodiments, the blood is fractionated or centrifuged. In some embodiments, the sample comprises plasma. In some embodiments, the sample is a plasma sample. In some embodiments, the sample comprises serum. In some embodiments, the sample is a serum sample.

[0201] In some embodiments, the sample comprises tissue. In some embodiments, the sample is a tissue sample. In some embodiments, the sample comprises liver tissue. In some embodiments, the sample is a liver sample. In some embodiments, the sample comprises hepatocytes. In some embodiments, the sample consists of hepatocytes. For example, the baseline target mRNA measurement or baseline target protein measurement may be obtained in a patient's liver sample or hepatocyte sample. In some embodiments, the sample comprises adipose tissue. In some embodiments, the sample is an adipose sample. The adipose sample may comprise or consist of white adipose tissue. The adipose sample may comprise or consist of brown adipose tissue. In some embodiments, the sample comprises renal tissue. In some embodiments, the sample is a renal sample. In some embodiments, the sample comprises cardiac tissue, such as ventricular tissue or atrial tissue. In some embodiments, the sample is a cardiac sample. In some embodiments, the sample comprises intestinal tissue, such as small intestinal tissue. In some embodiments, the sample is a small intestinal sample. In some embodiments, the sample comprises lymph node tissue, such as mesenteric lymph node. In some embodiments, the sample is a mesenteric lymph node sample. In some embodiments, the sample comprises muscle tissue. In some embodiments, the sample is a muscle sample. In some embodiments, the tissue sample comprises liver, adipose, kidney, or heart tissue. In some embodiments, the tissue sample comprises brown adipose tissue, white adipose tissue, kidney tissue, muscle tissue, intestinal tissue, mesenteric lymph node, or muscle tissue.

[0202] D. Effects In some embodiments, the composition or administration of the composition affects a measurement, such as a symptom (e.g., a liver damage symptom) measurement, a protective phenotype measurement, a target oligonucleotide (e.g., mRNA) measurement, or a target protein measurement (e.g., liver tissue target protein level) compared to a baseline measurement.

[0203] Some embodiments of the method described herein include obtaining a measurement from a subject. For example, the measurement may be obtained from the subject after the subject is treated. In some embodiments, the measurement is obtained in a second sample (such as a fluid or tissue sample described herein) obtained from the subject after the composition is administered to the subject. In some embodiments, the measurement is indicative of the disorder being treated.

[0204] In some embodiments, the measurements are obtained directly from the subject. In some embodiments, the measurements are obtained non-invasively using an imaging device. In some embodiments, the measurements are obtained in a second sample from the subject. In some embodiments, the measurements are obtained in one or more histological tissue sections. In some embodiments, the measurements are obtained by performing an assay on the second sample obtained from the subject. In some embodiments, the measurements are obtained by an assay, such as the assays described herein. In some embodiments, the assay is an immunoassay, a colorimetric assay, a fluorometric assay, or a PCR assay. In some embodiments, the measurements are obtained by an assay, such as an immunoassay, a colorimetric assay, or a fluorometric assay. In some embodiments, the measurements are obtained by PCR. In some embodiments, the measurements are obtained by histological examination. In some embodiments, the measurements are obtained by observation. In some embodiments, further measurements are provided in a third sample, a fourth sample, a fifth sample, etc.

[0205] In some embodiments, the measurements are obtained within 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 12 hours, 18 hours, or 24 hours after administration of the composition. In some embodiments, the measurements are obtained within 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days after administration of the composition. In some embodiments, the measurements are obtained within 1 week, 2 weeks, 3 weeks, 1 month, 2 months, 3 months, 6 months, 1 year, 2 years, 3 years, 4 years, or 5 years after administration of the composition. In some embodiments, the measurements are obtained 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 12 hours, 18 hours, or 24 hours after administration of the composition. In some embodiments, the measurements are obtained 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days after administration of the composition. In some embodiments, measurements are obtained 1 week, 2 weeks, 3 weeks, 1 month, 2 months, 3 months, 6 months, 1 year, 2 years, 3 years, 4 years, or 5 years after administration of the composition.

[0206] In some embodiments, the measurement of a symptom or parameter associated with the disorder in a subject is reduced or affected over an extended period of time compared to a baseline measurement. In some embodiments, the measurement is maintained for at least about 1 day, about 2 days, about 3 days, about 4 days, about 5 days, about 6 days, about 7 days, about 8 days, about 9 days, about 10 days, about 11 days, about 12 days, about 13 days, about 14 days, about 15 days, about 16 days, about 17 days, about 18 days, about 19 days, about 20 days, about 21 days, about 22 days, about 23 days, about 24 days, about 25 days, about 26 days, about 27 days, about 28 days, about 29 days, about 30 days, about 31 days, about 32 days, about 33 days, about 34 days, about 35 days, about 36 days, about 37 days, about 38 days, about 39 days, about 40 days, about 41 days, about 42 days, about 43 days, about 44 days, about 45 days, about 46 days, about 47 days, about 48 days, about 49 days, about 50 days, about 51 days, about 52 days, about 53 days, about 54 days, about 55 days, about 56 days, about 57 days, about 58 days, about 59 days, about 60 days, about 61 days, about 62 days, about 63 days, about 64 days, about 65 days, about 66 days, about 67 days, about 68 days, about 69 days, about 70 days, about 71 days, about 72 days, about 73 days, about 74 days, about 75 days, about 76 days, about 77 days, about 78 days, about 79 days, about The decreased or affected range of time after administration includes about 29 days, about 30 days, about 35 days, about 40 days, about 45 days, about 50 days, about 55 days, about 60 days, about 65 days, about 70 days, about 75 days, about 80 days, about 85 days, about 90 days, about 95 days, about 100 days, about 105 days, about 110 days, about 115 days, or about 120 days, or a range defined by any two of the foregoing numbers of days. In some embodiments, measurements are taken for at least 1 day, at least 2 days, at least 3 days, at least 4 days, at least 5 days, at least 6 days, at least 7 days, at least 8 days, at least 9 days, at least 10 days, at least 11 days, at least 12 days, at least 13 days, at least 14 days, at least 15 days, at least 16 days, at least 17 days, at least 18 days, at least 19 days, at least 20 days, at least 21 days, at least 22 days, at least 23 days, at least 24 days, at least 25 days, at least 26 days, at least 27 days, at least 28 days, at least 29 days, at least 30 days, at least 31 days, at least 32 days, at least 33 days, at least 34 days, at least 35 days, at least 36 days, at least 37 days, at least 38 days, at least 39 days, at least 40 days, at least 41 days, at least 42 days, at least 43 days, at least 44 days, at least 45 days, at least 46 days, at least 47 days, at least 48 days, at least 49 days, at least 50 days, at least 51 days, at least 52 days, at least 53 days, at least 54 days, at least 55 days, at least 56 days, at least 57 days, at least 58 days, at least 59 days, at least 60 days, at least 61 days, at least 62 days, at least 63 days, at least 64 days, at least 65 days, at least 66 days, at least 67 days, at least 68 days, at least 69 days, at least 70 days, at least 71 days, at least 72 days, at least 73 days, at reduced or affected for 25 days, at least 26 days, at least 27 days, at least 28 days, at least 29 days, at least 30 days, at least 35 days, at least 40 days, at least 45 days, at least 50 days, at least 55 days, at least 60 days, at least 65 days, at least 70 days, at least 75 days, at least 80 days, at least 85 days, at least 90 days, at least 95 days, at least 100 days, at least 105 days, at least 110 days, at least 115 days, or at least 120 days.In some embodiments, the measurements are taken 1 day or less, 2 days or less, 3 days or less, 4 days or less, 5 days or less, 6 days or less, 7 days or less, 8 days or less, 9 days or less, 10 days or less, 11 days or less, 12 days or less, 13 days or less, 14 days or less, 15 days or less, 16 days or less, 17 days or less, 18 days or less, 19 days or less, 20 days or less, 21 days or less, 22 days or less, 23 days or less, 24 days or less, 25 days or less, 2 ... The measurement is decreased or affected for 6 days or less, 27 days or less, 28 days or less, 29 days or less, 30 days or less, 35 days or less, 40 days or less, 45 days or less, 50 days or less, 55 days or less, 60 days or less, 65 days or less, 70 days or less, 75 days or less, 80 days or less, 85 days or less, 90 days or less, 95 days or less, 100 days or less, 105 days or less, 110 days or less, 115 days or less, or 120 days or less. In some embodiments, the measurement is decreased or affected for at least about 5 days. In some embodiments, the measurement is decreased or affected for at least about 10 days. In some embodiments, the measurement is decreased or affected for at least about 15 days. In some embodiments, the measurement is decreased or affected for at least about 20 days. In some embodiments, the measurement is decreased or affected for at least about 25 days. In some embodiments, the measurement is decreased or affected for at least about 30 days. In some embodiments, the measurement is decreased or affected for at least about 40 days. In some embodiments, the measurements are decreased or affected for at least about 50 days. In some embodiments, the measurements are decreased or affected for at least about 60 days. In some embodiments, the measurements are decreased or affected for at least about 70 days. In some embodiments, the measurements are decreased or affected for at least about 80 days. In some embodiments, the measurements are decreased or affected for at least about 90 days. In some embodiments, the measurements are decreased or affected for at least about 100 days. In some embodiments, the measurements are decreased or affected for at least about 110 days. In some embodiments, the measurements are decreased or affected for at least about 120 days.Examples of measurements of symptoms or parameters associated with a disorder may include target mRNA measurements, target protein measurements, biomarker measurements, or physiological measurements. Measurements may be in tissues (e.g., liver) or biological fluids (e.g., blood, serum, or plasma).

[0207] In some embodiments, the composition reduces the symptom measure compared to a baseline symptom measure. In some embodiments, the reduction is measured in a second tissue sample obtained from the subject after administering the composition to the subject. In some embodiments, the reduction is measured directly in the subject after administering the composition to the subject. In some embodiments, the symptom measure is reduced by about 2.5% or more, about 5% or more, or about 7.5% or more compared to the baseline symptom measure. In some embodiments, the symptom measure is reduced by about 10% or more compared to the baseline symptom measure. In some embodiments, the symptom measure is reduced by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more compared to the baseline symptom measure. In some embodiments, the symptom measure is reduced by about 2.5% or less, about 5% or less, or about 7.5% or less compared to the baseline symptom measure. In some embodiments, the symptom measure is reduced by about 10% or less compared to the baseline symptom measure. In some embodiments, the symptom measure is reduced by about 20% or less, about 30% or less, about 40% or less, about 50% or less, about 60% or less, about 70% or less, about 80% or less, about 90% or less, or about 100% or less compared to the baseline symptom measure. In some embodiments, the symptom measure is reduced by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or a range defined by any of the two aforementioned percentages.

[0208] In some embodiments, the composition increases a protective phenotype measure compared to a baseline protective phenotype measure. In some embodiments, the increase is measured in a second tissue sample obtained from the subject after administering the composition to the subject. In some embodiments, the increase is measured directly in the subject after administering the composition to the subject. In some embodiments, the protective phenotype measure increases by about 2.5% or more, about 5% or more, or about 7.5% or more compared to the baseline protective phenotype measure. In some embodiments, the protective phenotype measure increases by about 10% or more compared to the baseline protective phenotype measure. In some embodiments, the protective phenotype measure increases by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more compared to the baseline protective phenotype measure. In some embodiments, the protective phenotype measure is increased by about 100% or more, about 250% or more, about 500% or more, about 750% or more, or about 1000% or more compared to the baseline protective phenotype measure. In some embodiments, the protective phenotype measure is increased by about 2.5% or less, about 5% or less, or about 7.5% or less compared to the baseline protective phenotype measure. In some embodiments, the protective phenotype measure is increased by about 10% or less compared to the baseline protective phenotype measure. In some embodiments, the protective phenotype measure is increased by about 20% or less, about 30% or less, about 40% or less, about 50% or less, about 60% or less, about 70% or less, about 80% or less, about 90% or less, or about 100% or less compared to the baseline protective phenotype measure. In some embodiments, the protective phenotype measure is increased by about 100% or less, about 250% or less, about 500% or less, about 750% or less, or about 1000% or less compared to the baseline protective phenotype measure. In some embodiments, the protective phenotype measure is increased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 250%, 500%, 750%, or 1000%, or a range defined by any of the two aforementioned percentages.

[0209] In some embodiments, the measurements are target protein measurements. In some embodiments, the target protein measurements include target protein levels. In some embodiments, the target protein levels are presented as mass or percentage of target protein per sample weight. In some embodiments, the target protein levels are presented as mass or percentage of target protein per sample volume. In some embodiments, the target protein levels are presented as mass or percentage of target protein per total protein in the sample. In some embodiments, the target protein measurements are cellular (e.g., hepatocyte) target protein measurements. In some embodiments, the target protein measurements are tissue (e.g., liver tissue) target protein measurements. In some embodiments, the target protein measurements are circulating target protein measurements. In some embodiments, the baseline target protein measurements are obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescent assay.

[0210] In some embodiments, the composition reduces the target protein measurement compared to a baseline target protein measurement. In some embodiments, the composition reduces tissue target protein levels (such as but not limited to liver tissue target protein levels) compared to a baseline target protein measurement. In some embodiments, the composition reduces cellular target protein levels (such as but not limited to hepatocyte target protein levels) compared to a baseline target protein measurement. In some embodiments, the composition reduces circulating target protein levels compared to a baseline target protein measurement. In some embodiments, the reduction in target protein levels is measured in a second sample obtained from the subject after administering the composition to the subject.

[0211] In some embodiments, the target protein measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more as compared to the baseline target protein measurement. In some embodiments, the target protein measurement is decreased by about 10% or more as compared to the baseline target protein measurement. In some embodiments, the target protein measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more as compared to the baseline target protein measurement. In some embodiments, the target protein measurement is decreased by about 2.5% or less, about 5% or less, or about 7.5% or less as compared to the baseline target protein measurement. In some embodiments, the target protein measurement is decreased by about 10% or less as compared to the baseline target protein measurement. In some embodiments, the target protein measurement is decreased by about 20%, about 30% or less, about 40% or less, about 50% or less, about 60% or less, about 70% or less, about 80% or less, about 90% or less, or about 100% or less compared to the baseline target protein measurement. In some embodiments, the target protein measurement is decreased by 2.5%, 5%, 7.5%, 19%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or a range defined by any of the two aforementioned percentages. The target protein measurement can be decreased over an extended period of time. In some embodiments, the target protein measurement is decreased over 7 days, 14 days, 28 days, 42 days, 56 days, 70 days, 77 days, 84 days, 91 days, 98 days, 105 days, or a range therebetween. In some embodiments, the target protein measurements are decreased over about 7 days, about 14 days, about 28 days, about 42 days, about 56 days, about 70 days, about 77 days, about 84 days, about 91 days, about 98 days, about 105 days, or a range therebetween.

[0212] In some embodiments, the measurements are target protein measurements. In some embodiments, the target mRNA measurements include target mRNA levels. In some embodiments, the target mRNA levels are presented as mass or percentage of target mRNA per sample weight. In some embodiments, the target mRNA levels are presented as mass or percentage of target mRNA per sample volume. In some embodiments, the target mRNA levels are presented as mass or percentage of target mRNA per total mRNA in the sample. In some embodiments, the target mRNA levels are presented as mass or percentage of target mRNA per total nucleic acid in the sample. In some embodiments, the target mRNA levels are presented relative to another mRNA level, such as the mRNA level of a housekeeping gene in the sample. In some embodiments, the target mRNA measurements are obtained by an assay, such as a PCR assay. In some embodiments, the PCR comprises qPCR. In some embodiments, the PCR comprises reverse transcription of the target mRNA.

[0213] In some embodiments, the composition reduces the target mRNA measurement compared to a baseline target mRNA measurement. In some embodiments, the target mRNA measurement is obtained in a second sample obtained from the subject after administering the composition to the subject. In some embodiments, the composition reduces the target mRNA level relative to the baseline target mRNA level. In some embodiments, the reduction in the target mRNA level is measured in a second sample obtained from the subject after administering the composition to the subject. In some embodiments, the second sample is a second liver sample. In some embodiments, the second sample is a second hepatocyte sample.

[0214] In some embodiments, the target mRNA measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more as compared to the baseline target mRNA measurement. In some embodiments, the target mRNA measurement is decreased by about 10% or more as compared to the baseline target mRNA measurement. In some embodiments, the target mRNA measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more as compared to the baseline target mRNA measurement. In some embodiments, the target mRNA measurement is decreased by about 2.5% or less, about 5% or less, or about 7.5% or less as compared to the baseline target mRNA measurement. In some embodiments, the target mRNA measurement is decreased by about 10% or less as compared to the baseline target mRNA measurement. In some embodiments, the target mRNA measurement is decreased by about 20% or less, about 30% or less, about 40% or less, about 50% or less, about 60% or less, about 70% or less, about 80% or less, about 90% or less, or about 100% compared to the baseline target mRNA measurement. In some embodiments, the target mRNA measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or a range defined by any of the two aforementioned percentages. The target mRNA measurement can be decreased over an extended period of time. In some embodiments, the target mRNA measurement is decreased for 7 days, 14 days, 28 days, 42 days, 56 days, 70 days, 77 days, 84 days, 91 days, 98 days, 105 days, or a range therebetween. In some embodiments, target protein measurements are decreased for about 7 days, about 14 days, about 28 days, about 42 days, about 56 days, about 70 days, about 77 days, about 84 days, about 91 days, about 98 days, about 105 days, or a range therebetween. Some embodiments include decreasing RNA measurements in addition to mRNA measurements.

[0215] III. Definition Unless otherwise defined, all terminology, notations, and other technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the claimed subject matter belongs. In some cases, terms having a commonly understood meaning are defined herein for clarity and / or ready reference, and the inclusion of such definitions herein should not necessarily be construed as indicating that they are substantially different from those commonly understood in the art.

[0216] Throughout this application, various embodiments may be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the disclosure. Thus, the description of a range should be considered to have specifically disclosed all possible subranges and individual numerical values ​​within that range. For example, the description of a range such as 1-6 should be considered to have specifically disclosed subranges such as 1-3, 1-4, 1-5, 2-4, 2-6, 3-6, and individual numerical values ​​within that range, such as 1, 2, 3, 4, 5, and 6. This applies regardless of the breadth of the range.

[0217] As used in this specification and claims, "a," "an," and "the" include plural references unless the content clearly dictates otherwise. For example, the term "a sample" includes multiple samples, including mixtures thereof.

[0218] Some examples refer to a sequence listing. To the extent that the sequence listing conflicts with the disclosure herein, the present specification controls.

[0219] "Determining," "measuring," "evaluating," "assessing," "assaying," and "analyzing" are often used interchangeably herein to refer to forms of measurement. The terms include determining whether an element is present (e.g., detecting). Such terms can include quantitative, qualitative, or quantitative and qualitative determinations. Assessing can be relative or absolute. "Detecting the presence of" can include determining the amount of something present in addition to determining whether something is present or absent depending on the context.

[0220] The terms "subject" and "patient" may be used interchangeably. A "subject" may be a biological entity that contains expressed genetic material. The biological entity may be, for example, a plant, an animal, or a microorganism, including bacteria, viruses, fungi, and protozoa. The subject may be a mammal. The mammal may be a human. The subject may be diagnosed or suspected to be at high risk for a disease. In some cases, the subject is not necessarily diagnosed or suspected to be at high risk for a disease.

[0221] As used herein, a number followed by the term "about" refers to a number that is plus or minus 10% of that number. A range followed by the term "about" refers to a range of minus 10% of the minimum value and plus 10% of the maximum value.

[0222] As used herein, the term "treatment" or "treating" is used in reference to a pharmaceutical or other interventional regimen to obtain a beneficial or desired outcome in a recipient. A beneficial or desired outcome includes, but is not limited to, a therapeutic benefit and / or a prophylactic benefit. A therapeutic benefit may also refer to the eradication or amelioration of the condition or underlying disease being treated. Similarly, a therapeutic benefit may be achieved by the eradication or amelioration of one or more of the physiological symptoms associated with the underlying disease, such that an improvement in the subject is observed, even though the subject may still be affected by the underlying disease. A prophylactic effect includes delaying, preventing, or eliminating the appearance of a disease or illness, delaying or eliminating the onset of symptoms of a disease or illness, slowing, stopping, or reversing the progression of a disease or illness, or any combination thereof. With regard to a prophylactic benefit, a subject at risk of developing a particular disease or reporting one or more physiological symptoms of a disease may receive treatment, even if a diagnosis of the disease has not been made.

[0223] "Treatment" or "treating" may include an approach to obtain a beneficial or desired result with respect to a disease, disorder, or medical condition, including, but not limited to, a therapeutic benefit and / or a prophylactic benefit. A therapeutic benefit may include, for example, eradication or amelioration of the underlying disease being treated. Similarly, a therapeutic benefit may include, for example, eradication or amelioration of one or more physiological symptoms associated with an underlying disease, such that an improvement in the subject is observed, even though the subject may still be affected by the underlying disease. In certain embodiments, with respect to a prophylactic benefit, the composition is administered to a subject at risk of developing a particular disease, or to a subject reporting one or more physiological symptoms of a disease, even if a diagnosis of the disease has not been made. Treatment by administration of the compounds described herein does not necessarily require the involvement of a medical professional.

[0224] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

[0225] "C x-y " or "C x -C y The term "alkyl," when used with a chemical moiety such as alkyl, alkynyl, or alkenyl, means that the group contains from x to y carbons in the chain. For example, "C 1-6 The term "alkyl" refers to substituted or unsubstituted saturated hydrocarbon groups, including straight-chain and branched-chain alkyl groups, containing 1 to 6 carbons. x-y " or "C x -C y The term "C" is not meant to limit the number of carbon atoms that may be attached to a chemical moiety when the chemical moiety is substituted with a second chemical moiety. For example, "C 1-6 Alkyl" or "C 1 -C 6 The term "alkyl" refers to saturated, substituted, or unsubstituted hydrocarbon groups, including straight chain alkyl groups (e.g., linear alkyl groups) and branched alkyl groups, containing 1, 2, 3, 4, 5, or 6 carbon atoms, although many of the carbon atoms may be C 1-6 It may be present in any of the substituents of the alkyl. For example, C 1-6 When the alkyl is optionally substituted with a second chemical moiety containing two carbon atoms, C 1-6 It is understood that an alkyl can contain from 1 to 8 carbon atoms.

[0226] "C x-y alkenyl" and "C x-y The term "alkynyl" refers to substituted or unsubstituted unsaturated aliphatic groups analogous in length and possible substitution to the alkyls described above, but which contain at least one double or triple bond respectively.

[0227] "Amino" is -NH 2 Refers to a part.

[0228] "Cyano" refers to a -CN moiety.

[0229] "Nitro" is -NO 2 Refers to a part.

[0230] "Oxa" refers to an -O- moiety.

[0231] "Oxo" refers to the =O moiety.

[0232] "Thioxo" refers to the =S moiety.

[0233] "Imino" refers to the =NH moiety.

[0234] "Oximo" refers to the =N-OH moiety.

[0235] "Hydrazino" is =N-NH 2 Refers to a part.

[0236] "Alkyl" refers to a straight or branched chain hydrocarbon moiety that is fully saturated and consists solely of carbon and hydrogen atoms. In certain embodiments, an "alkyl" contains 1 to 15 carbon atoms (e.g., C 1 -C 15 In certain embodiments, alkyl comprises 1 to 13 carbon atoms (e.g., C 1 -C 13 In certain embodiments, alkyl comprises 1 to 8 carbon atoms (e.g., C 1 -C 8 In certain embodiments, alkyl contains 1 to 6 carbon atoms (e.g., C 1-6 In other embodiments, alkyl contains 1 to 5 carbon atoms (e.g., C 1 -C 5 In other embodiments, alkyl contains 1 to 4 carbon atoms (e.g., C 1 -C 4 In other embodiments, alkyl contains 1 to 3 carbon atoms (e.g., C 1 -C 3In other embodiments, alkyl contains 1 to 2 carbon atoms (e.g., C 1 -C 2 In other embodiments, alkyl is an alkyl group having one carbon atom (e.g., C 1 In other embodiments, the alkyl group contains 5 to 15 carbon atoms (e.g., C 5 -C 15 In other embodiments, alkyl contains 5 to 8 carbon atoms (e.g., C 5 -C 8 In other embodiments, alkyl contains 2 to 5 carbon atoms (e.g., C 2 -C 5 In other embodiments, alkyl contains 3 to 5 carbon atoms (e.g., C 3 -C 5 In other embodiments, the alkyl group is selected from methyl, ethyl, 1-propyl (n-propyl), 1-methylethyl (2-propyl, iso-propyl), 1-butyl (n-butyl), 1-methylpropyl (sec-butyl), 2-methylpropyl (iso-butyl), 1,1-dimethylethyl (tert-butyl), and 1-pentyl (n-pentyl). The alkyl is attached to the remainder of the molecule by a single bond.

[0237] "Aminoalkyl" refers to a moiety attached through a nitrogen atom of the form -N(H)(alkyl) or N(alkyl)(alkyl), where the moiety is N(alkyl)(alkyl), the two alkyl groups attached to the nitrogen can be the same alkyl group or different alkyl groups.

[0238] "Alkoxy" refers to a moiety attached through an oxygen atom of the formula -O-alkyl, where alkyl is an alkyl chain as defined above.

[0239] "Alkenyl" refers to a straight or branched chain hydrocarbon moiety consisting solely of carbon and hydrogen atoms containing at least one carbon-carbon double bond. In certain embodiments, an alkenyl contains 2-12 carbon atoms. In certain embodiments, an alkenyl contains 2-8 carbon atoms. In other embodiments, an alkenyl contains 2-4 carbon atoms. An alkenyl is attached to the remainder of the molecule by a single bond, such as, for example, ethenyl (i.e., vinyl), prop-1-enyl (i.e., allyl), but-1-enyl, pent-1-enyl, penta-1,4-dienyl, and the like.

[0240] "Alkynyl" refers to a straight or branched chain hydrocarbon moiety consisting solely of carbon and hydrogen atoms, containing at least one carbon-carbon triple bond, having from 2 to 12 carbon atoms, and optionally further containing at least one carbon-carbon double bond. In certain embodiments, an alkynyl contains from 2 to 8 carbon atoms. In other embodiments, an alkynyl contains from 2 to 6 carbon atoms. In other embodiments, an alkynyl contains from 2 to 4 carbon atoms. An alkynyl is attached to the remainder of the molecule by a single bond, for example, ethynyl, propynyl, butynyl, pentynyl, hexynyl, and the like.

[0241] An "alkylene" or "alkylene chain" refers to a divalent hydrocarbon moiety that is linear (e.g., straight) or branched. An "alkylene" or "alkylene chain" can link one part of a molecule to a second part. An "alkylene" or "alkylene chain" consists of only carbon and hydrogen atoms (substitution of the alkylene with one or more substituents containing atoms other than hydrogen, such as N, O, and S, can be specified). An "alkylene" or "alkylene chain" can contain no unsaturation (despite the point of attachment of the alkylene to the rest of the molecule). In certain embodiments, an "alkylene" or "alkylene chain" contains 1 to 12 carbon atoms, e.g., methylene, ethylene, propylene, n-butylene, etc. An alkylene chain can be attached to one part of a molecule through a single bond and to a second part through a single bond. The points of attachment of the alkylene chain to the rest of the molecule and to the second part can be through one carbon in the alkylene chain or through any two carbons in the alkylene. In certain embodiments, alkylene contains 1 to 8 carbon atoms (e.g., C 1 -C 8 In other embodiments, the alkylene contains 1 to 5 carbon atoms (e.g., C 1 -C 5 In other embodiments, the alkylene contains 1 to 4 carbon atoms (e.g., C 1 -C 4 In other embodiments, the alkylene contains 1 to 3 carbon atoms (e.g., C 1 -C 3 In other embodiments, the alkylene contains 1 to 2 carbon atoms (e.g., C 1 -C 2 In other embodiments, the alkylene contains one carbon atom (e.g., C 1 In other embodiments, the alkylene contains 5 to 8 carbon atoms (e.g., C 5 -C 8 In other embodiments, the alkylene contains 2 to 5 carbon atoms (e.g., C 2 -C 5In other embodiments, the alkylene contains 3 to 5 carbon atoms (e.g., C 3 -C 5 alkylene).

[0242] An "alkenylene" or "alkenylene chain" refers to a divalent hydrocarbon moiety that is linear (e.g., straight) or branched. An "alkenylene" or "alkenylene chain" can link one part of a molecule to a second part. An "alkenylene" or "alkenylene chain" consists only of carbon and hydrogen atoms (substitution of the alkenylene with one or more substituents, including atoms other than hydrogen, such as N, O, and S, can be specified). An "alkenylene" or "alkenylene chain" contains at least one carbon-carbon double bond. In certain embodiments, an "alkenylene" or "alkenylene chain" contains 2 to 12 carbon atoms. An alkenylene chain can be attached to one part of a molecule through a single bond and to a second part through a single bond. The point of attachment of the alkenylene chain to the rest of the molecule and to the second part can be through one carbon in the alkenylene chain or through any two carbons in the alkenylene chain. In certain embodiments, alkenylene contains 2 to 8 carbon atoms (e.g., C 2 -C 8 In other embodiments, the alkenylene contains 2 to 5 carbon atoms (e.g., C 2 -C 5 In other embodiments, the alkenylene contains 2 to 4 carbon atoms (e.g., C 2 -C 4 In other embodiments, the alkenylene contains 2 to 3 carbon atoms (e.g., C 2 -C 3 In other embodiments, the alkenylene contains 5 to 8 carbon atoms (e.g., C 5 -C 8 In other embodiments, the alkenylene contains 2 to 5 carbon atoms (e.g., C 2 -C 5 In other embodiments, the alkenylene contains 3 to 5 carbon atoms (e.g., C3 -C 5 alkenylene).

[0243] "Alkynylene" or "alkynylene chain" refers to a divalent hydrocarbon moiety that is linear (e.g., straight) or branched. An "alkynylene" or "alkynylene chain" can link one part of a molecule to a second part. An "alkynylene" or "alkynylene chain" consists only of carbon and hydrogen (substitution of an alkynylene with one or more substituents containing atoms other than hydrogen, such as N, O, and S, can be specified). An "alkynylene" or "alkynylene chain" contains at least one carbon-carbon triple bond. In certain embodiments, an "alkynylene" or "alkynylene chain" contains 2 to 12 carbon atoms. An alkynylene chain can be attached to one part of a molecule through a single bond and to a second part through a single bond. The points of attachment of the alkynylene chain to the rest of the molecule and to the second part can be through one carbon in the alkynylene chain or through any two carbons in the alkynylene chain. In certain embodiments, alkynylene contains 2 to 8 carbon atoms (e.g., C 2 -C 8 In other embodiments, the alkynylene contains 2 to 5 carbon atoms (e.g., C 2 -C 5 In other embodiments, the alkynylene contains 2 to 4 carbon atoms (e.g., C 2 -C 4 In other embodiments, the alkynylene contains 2 to 3 carbon atoms (e.g., C 2 -C 3 In other embodiments, the alkynylene contains two carbon atoms (e.g., C 2 In other embodiments, the alkynylene contains 5 to 8 carbon atoms (e.g., C 5 -C 8 In other embodiments, the alkynylene contains 3 to 5 carbon atoms (e.g., C 3 -C 5 alkynylene).

[0244] The term "carbocycle" as used herein refers to a saturated, unsaturated, or aromatic ring in which each atom of the ring is carbon. Carbocycles include 3-10 membered monocyclic rings, 5-12 membered bicyclic rings, 5-12 membered spiro bicyclic rings, and 5-12 membered bridged rings. Each ring of a bicyclic carbocycle may be selected from a saturated ring, an unsaturated ring, and an aromatic ring. In an exemplary embodiment, an aromatic ring, e.g., phenyl, may be fused to a saturated or unsaturated ring, e.g., cyclohexane, cyclopentane, or cyclohexene. Bicyclic carbocycles include any combination of saturated, unsaturated, and aromatic bicyclic rings, as valences permit. Bicyclic carbocycles further include spiro bicyclic rings, e.g., spiropentane. Bicyclic carbocycles include any combination of ring sizes, such as 3-3 spiro ring systems, 4-4 spiro ring systems, 4-5 fused ring systems, 5-5 fused ring systems, 5-6 fused ring systems, 6-6 fused ring systems, 5-7 fused ring systems, 6-7 fused ring systems, 5-8 fused ring systems, and 6-8 fused ring systems. Exemplary carbocycles include cyclopentyl, cyclohexyl, cyclohexenyl, adamantyl, phenyl, indanyl, naphthyl, and bicyclo[1.1.1]pentanyl. "Carbocycle" can include "aryl" and "cycloalkyl."

[0245] The term "aryl" refers to an aromatic monocyclic or aromatic polycyclic hydrocarbon ring system. An aromatic monocyclic or aromatic polycyclic hydrocarbon ring system contains only hydrogen and carbon from 5 to 18 carbon atoms, and at least one of the rings in the ring system is aromatic, i.e., it contains a cyclic delocalized (4n+2) π-electron system according to Hückel theory. Ring systems from which aryl groups are derived include, but are not limited to, groups such as benzene, fluorene, indane, indene, tetralin, and naphthalene. In some embodiments, the aryl substituent is positively or negatively charged. In some embodiments, the aryl substituent is neutral. In some embodiments, the aryl substituent is zwitterionic, alternatively or in addition, in some embodiments, the aryl substituent is uncharged. In some embodiments, the aryl substituent has no charge. In some embodiments, the aryl substituent has no net charge. In some embodiments, the aryl substituent has no net charge and is not zwitterionic.

[0246] The term "cycloalkyl" refers to a saturated ring in which each atom of the ring is carbon. Cycloalkyls can include monocyclic and polycyclic rings, such as 3-10 membered monocyclic rings, 5-12 membered bicyclic rings, 5-12 membered spiro bicyclic rings, and 5-12 membered bridged rings. In certain embodiments, cycloalkyls contain 3-10 carbon atoms. In other embodiments, cycloalkyls contain 5-7 carbon atoms. Cycloalkyls can be attached to the remainder of the molecule by a single bond. Examples of monocyclic cycloalkyls include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. Polycyclic cycloalkyl radicals include, for example, adamantyl, spiropentane, norbornyl (i.e., bicyclo[2.2.1]heptanyl), decalinyl, 7,7-dimethylbicyclo[2.2.1]heptanyl, bicyclo[1.1.1]pentanyl, and the like.

[0247] The term "cycloalkenyl" refers to a saturated ring in which each atom of the ring is carbon and there is at least one double bond between two ring carbons. Cycloalkenyls can include monocyclic and polycyclic rings, such as 3-10 membered monocyclic rings, 6-12 membered bicyclic rings, and 5-12 membered bridged rings. In other embodiments, cycloalkenyls contain 5-7 carbon atoms. Cycloalkenyls can be attached to the remainder of the molecule by a single bond. Examples of monocyclic cycloalkenyls include, for example, cyclopentenyl, cyclohexenyl, cycloheptenyl, and cyclooctenyl.

[0248] The term "halo," or alternatively, "halogen" or "halide," means fluoro, chloro, bromo, or iodo. In some embodiments, halo is fluoro, chloro, or bromo.

[0249] The term "haloalkyl" refers to an alkyl radical, as defined above, that is substituted by one or more halo radicals, e.g., trifluoromethyl, dichloromethyl, bromomethyl, 2,2,2-trifluoroethyl, 1-chloromethyl-2-fluoroethyl, etc. In some embodiments, the alkyl portion of the haloalkyl radical is optionally further substituted as described herein.

[0250] The term "heterocycle" refers to a saturated, unsaturated, or aromatic ring containing one or more heteroatoms. Exemplary heteroatoms include N, O, Si, P, B, and S atoms. Heterocycles include 3-10 membered monocyclic rings, 6-12 membered bicyclic rings, 5-12 membered spiro bicyclic rings, and 5-12 membered bridged rings. Bicyclic heterocycles include any combination of saturated, unsaturated, and aromatic bicyclic rings, as valences permit. In an exemplary embodiment, an aromatic ring, e.g., pyridyl, can be fused to a saturated or unsaturated ring, e.g., cyclohexane, cyclopentane, morpholine, piperidine, or cyclohexene. Bicyclic heterocycles include any combination of ring sizes, such as 4-5 fused ring systems, 5-5 fused ring systems, 5-6 fused ring systems, 6-6 fused ring systems, 5-7 fused ring systems, 6-7 fused ring systems, 5-8 fused ring systems, and 6-8 fused ring systems. Bicyclic heterocycles further include spiro bicyclic rings, such as 5-12 membered spiro bicycles, such as 2-oxa-6-azaspiro[3.3]heptane. "Heterocycle" can include "heteroaryl" and "heterocycloalkyl".

[0251] "Heteroaryl" refers to a radical derived from a 5-18 membered aromatic ring radical containing 2-17 carbon atoms and 1-6 heteroatoms selected from nitrogen, oxygen, and sulfur. As used herein, a heteroaryl radical is a monocyclic, bicyclic, tricyclic, or tetracyclic ring system in which at least one of the rings in the ring system is aromatic, i.e., contains a cyclic delocalized (4n+2) π-electron system according to Huckel theory. Heteroaryl includes fused or bridged ring systems. The heteroatoms in a heteroaryl radical are optionally oxidized. One or more nitrogen atoms, if present, are optionally quaternized. The heteroaryl is attached to the remainder of the molecule through any atom of the ring. Examples of heteroaryl include, but are not limited to, azepinyl, acridinyl, benzimidazolyl, benzindolyl, 1,3-benzodioxolyl, benzofuranyl, benzoxazolyl, benzo[d]thiazolyl, benzothiadiazolyl, benzo[b][1,4]dioxepinyl, benzo[b][1,4]oxazinyl, 1,4-benzodioxanyl, benzonaphthofuranyl, benzoxazolyl, benzodioxolyl, benzodioxinyl, benzopyranyl, benzopyranonyl, benzofuranyl, benzofuranonyl, benzothienyl (benzothiophenyl), benzothieno[3,2-d]pyrimidinyl, benzotriazolyl, benzo[4,6]iphenyl, midazo[1,2-a]pyridinyl, carbazolyl, cinnolinyl, cyclopenta[d]pyrimidinyl, 6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3-d]pyrimidinyl, 5,6-dihydrobenzo[h]quinazolinyl, 5,6-dihydrobenzo[h]cinnolinyl, 6,7-dihydro-5H-benzo[6,7]cyclohepta[1,2-c]pyridazinyl, dibenzofuranyl, dibenzothiophenyl, furanyl, furanonyl, furo[3,2-c]pyridinyl, 5,6,7,8,9,10-hexahydrocycloocta[d]pyrimidinyl, 5,6,7,8,9,10-hexahydrocycloocta[d]pyridazinyl, 5,6,7,8,9,10-Hexahydrocycloocta[d]pyridinyl, isothiazolyl, imidazolyl, indazolyl, indolyl, indazolyl, isoindolyl, indolinyl, isoindolinyl, isoquinolyl, indolizinyl, isoxazolyl, 5,8-methano-5,6,7,8-tetrahydroquinazolinyl, naphthyridinyl, 1,6-naphthyridinonyl, oxadiazolyl, 2-oxazolyl, Xoazepinyl, oxazolyl, oxiranyl, 5,6,6a,7,8,9,10,10a-octahydrobenzo[h]quinazolinyl, 1-phenyl-1H-pyrrolyl, phenazinyl, phenothiazinyl, phenoxazinyl, phthalazinyl, pteridinyl, purinyl, pyrrolyl, pyrazolyl, pyrazolo[3,4-d]pyrimidinyl, pyridinyl, pyrido[3,2-d]pyrimidinyl quinolinyl, 5,6,7,8-tetrahydroquinazolinyl, 5,6,7,8-tetrahydrobenzo[4,5]thieno[2,3-d]pyrimidinyl, 6,7,8,9-tetrahydro-5H-cycloquinazolinyl, 5,6,7,8-tetrahydrobenzo[4,5]thieno[2,3-d]pyrimidinyl, 6,7,8,9-tetrahydro-5H-cycloquinazin ... Hepta[4,5]thieno[2,3-d]pyrimidinyl, 5,6,7,8-tetrahydropyrido[4,5-c]pyridazinyl, thiazolyl, thiadiazolyl, triazolyl, tetrazolyl, triazinyl, thieno[2,3-d]pyrimidinyl, thieno[3,2-d]pyrimidinyl, thieno[2,3-c]pyridinyl, and thiophenyl (i.e., thienyl).

[0252] One or more nitrogen atoms, if present, may be optionally quaternized. In some embodiments, the heterocyclic substituent is positively or negatively charged. In some embodiments, the heterocyclic substituent is neutral. In some embodiments, the heterocyclic substituent is zwitterionic, alternatively or in addition, in some embodiments, the heterocyclic substituent is uncharged. In some embodiments, the heterocyclic substituent does not have a charge. In some embodiments, the heterocyclic substituent does not have a net charge. In some embodiments, the heterocyclic substituent does not have a net charge and is not zwitterionic.

[0253] The term "heterocycloalkyl" refers to a saturated ring having carbon atoms and at least one heteroatom. Typical heteroatoms include atoms of N, O, Si, P, B, and S. Heterocycloalkyls can include monocyclic and polycyclic rings, such as 3-10 membered monocyclic rings, 6-12 membered bicyclic rings, 5-12 membered spiro bicyclic rings, and 5-12 membered bridged rings. The heteroatoms in the heterocycloalkyl radical are optionally oxidized. One or more nitrogen atoms, if present, are optionally quaternized. The heterocycloalkyl is attached to the remainder of the molecule through any atom of the heterocycloalkyl, such as any carbon or nitrogen atom of the heterocycloalkyl, where valence allows. Examples of heterocycloalkyl radicals include, but are not limited to, dioxolanyl, thienyl[1,3]dithianyl, decahydroisoquinolyl, imidazolinyl, imidazolidinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl, piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl, thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl, thiomorpholinyl, thiamorpholinyl, 1-oxo-thiomorpholinyl, 2-oxa-6-azaspiro[3.3]heptane, and 1,1-dioxo-thiomorpholinyl. In some embodiments, the heterocycloalkyl contains one heteroatom. In some embodiments, the heterocycloalkyl contains one heteroatom selected from N, O, and S. In some embodiments, the heterocycloalkyl contains multiple heteroatoms. In some embodiments, the heterocycloalkyl contains multiple heteroatoms selected from N, O, and S.

[0254] The term "heterocycloalkenyl" refers to an unsaturated ring having carbon atoms and at least one heteroatom, and at least one double bond exists between two ring carbons. Heterocycloalkenyl does not include heteroaryl rings. Typical heteroatoms include N, O, Si, P, B, and S atoms. Heterocycloalkenyl can include monocyclic and polycyclic rings, such as 3-10 membered monocyclic rings, 6-12 membered bicyclic rings, and 5-12 membered bridged rings. In other embodiments, heterocycloalkenyl contains 5-7 ring atoms. Heterocycloalkenyl can be attached to the remainder of the molecule by a single bond. Examples of monocyclic cycloalkenyls include, for example, pyrroline (dihydropyrrole), pyrazoline (dihydropyrazole), imidazoline (dihydroimidazole), triazoline (dihydrotriazole), dihydrofuran, dihydrothiophene, oxazoline (dihydrooxazole), isoxazoline (dihydroisoxazole), thiazoline (dihydrothiazole), isothiazolin (dihydroisothiazole), oxadiazoline (dihydrooxadiazole), thiadiazoline (dihydrothiadiazole), dihydropyridine, tetrahydropyridine, dihydropyridazine, tetrahydropyridazine, dihydropyrimidine, tetrahydropyrimidine, dihydropyrazine, tetrahydropyrazine, pyran, dihydropyran, thiopyran, dihydrothiopyran, dioxin, dihydrodioxin, oxazine, dihydrooxazine, thiazine, and dihydrothiazine.

[0255] The term "substituted" refers to the replacement of one or more carbon or hydrogen on a substitutable heteroatom, e.g., NH or NH of a compound. 2"Substituted" or "substituted with" includes the implicit proviso that such substitution is in accordance with the allowed valences of the substituted atom and substituent, and results in a stable compound, i.e., a compound that does not spontaneously undergo transformation by rearrangement, cyclization, elimination, and the like. In certain embodiments, substituted refers to a moiety having a substituent replacing two hydrogen atoms on the same carbon atom (e.g., replacing two hydrogen atoms on one carbon with an oxo, imino, or thioxo group). As used herein, the term "substituted" is intended to include all permissible substituents of organic compounds. In a broad aspect, the permissible substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and nonaromatic substituents of organic compounds. The permissible substituents can be one or more and the same or different for appropriate organic compounds.

[0256] In some embodiments, a substituent may be any of the substituents described herein, for example, halogen, hydroxy, oxo (=O), thioxo (=S), cyano (-CN), nitro (-NO 2 ), imino (=NH), oximo (=N-OH), hydrazino (=N-NH 2 ), -R b -OR a , -R b -OC(O)-R a , -R b -OC(O)-OR a , -R b -OC(O)-N(R a ) 2 , -R b -N(R a ) 2 , -R b -C(O)R a , -R b -C(O)OR a , -R b -C(O)N(R a ) 2 , -R b -OR c -C(O)N(Ra ) 2 , -R b -N(R a )C(O)OR a , -R b -N(R a )C(O)R a , -R b -N(R a )S(O) t R a (t is 1 or 2), -R b -S(O) t R a (t is 1 or 2), -R b -S(O) t OR a (t is 1 or 2), and -R b -S(O) t N(R a ) 2 (t is 1 or 2), as well as alkyl, alkenyl, alkynyl, aryl, aralkyl, aralkenyl, aralkynyl, cycloalkyl, cycloalkylalkyl, heterocycloalkyl, heterocycloalkylalkyl, heteroaryl, and heteroarylalkyl, any of which may be alkyl, alkenyl, alkynyl, halogen, haloalkyl, haloalkenyl, haloalkynyl, oxo (=O), thioxo (=S), cyano (-CN), nitro (-NO 2 ), imino (=NH), oximo (=N-OH), hydrazine (=N-NH 2 ), -R b -OR a , -R b -OC(O)-R a , -R b -OC(O)-OR a , -R b -OC(O)-N(R a ) 2 , -R b -N(R a ) 2 , -R b -C(O)R a , -R b -C(O)OR a , -R b -C(O)N(R a )2 , -R b -OR c -C(O)N(R a ) 2 , -R b -N(R a )C(O)OR a , -R b -N(R a )C(O)R a , -R b -N(R a )S(O) t R a (t is 1 or 2), -R b -S(O) t R a (t is 1 or 2), -R b -S(O) t OR a (t is 1 or 2), and -R b -S(O) t N(R a ) 2 (t is 1 or 2), where R a are each independently selected from hydrogen, alkyl, cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl, heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl, where R a are each, as far as valence allows, alkyl, alkenyl, alkynyl, halogen, haloalkyl, haloalkenyl, haloalkynyl, oxo (=O), thioxo (=S), cyano (-CN), nitro (-NO 2 ), imino (=NH), oximo (=N-OH), hydrazine (=N-NH 2 ), -R b -OR a , -R b -OC(O)-R a , -R b -OC(O)-OR a , -R b -OC(O)-N(R a ) 2 , -R b -N(R a ) 2 , -R b -C(O)Ra , -R b -C(O)OR a , -R b -C(O)N(R a ) 2 , -R b -OR c -C(O)N(R a ) 2 , -R b -N(R a )C(O)OR a , -R b -N(R a )C(O)R a , -R b -N(R a )S(O) t R a (t is 1 or 2), -R b -S(O) t R a (t is 1 or 2), -R b -S(O) t OR a (t is 1 or 2), and -R b -S(O) t N(R a ) 2 (t is 1 or 2), and wherein R b are each independently selected from a direct bond or a linear or branched alkylene, alkenylene, or alkynylene chain; R c are each a straight or branched alkylene chain, an alkenylene chain, or an alkynylene chain.

[0257] A double bond to an oxygen atom, such as an oxo group, is represented herein as both "=O" and "(O)". A double bond to a nitrogen atom is represented herein as both "=NR" and "(NR)". A double bond to a sulfur atom is represented herein as both "=S" and "(S)".

[0258] The phrases "parenteral administration" and "administered parenterally" as used herein mean modes of administration other than enteral and topical administration, usually by injection, and include, but are not limited to, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, and substernal injection and infusion.

[0259] The phrase "pharmacologically acceptable" is used herein to refer to those compounds, materials, compositions, and / or dosage forms that are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without undue toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit-risk ratio.

[0260] The phrases "pharmacologically acceptable excipient" or "pharmacologically acceptable carrier" as used herein refer to a pharma- ceutically acceptable material, composition, or vehicle, such as a liquid or solid filler, diluent, excipient, solvent, or encapsulating material. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not harmful to the patient. Some examples of substances that can function as pharma- ceutically acceptable carriers include: (1) sugars, such as lactose, glucose, and sucrose; (2) starches, such as corn starch and potato starch; (3) cellulose and its derivatives, such as sodium carboxymethylcellulose, ethylcellulose, and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, such as cocoa butter and suppository wax; (9) peanut oil, cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil, and the like. (10) glycols, such as propylene glycol, (11) polyols, such as glycerin, sorbitol, mannitol, and polyethylene glycol, (12) esters, such as ethyl oleate and ethyl laurate, (13) agar, (14) buffers, such as magnesium hydroxide and aluminum hydroxide, (15) alginic acid, (16) pyrogen-free water, (17) isotonic saline, (18) Ringer's solution, (19) ethyl alcohol, (20) phosphate buffer, and (21) other non-toxic compatible substances utilized in pharmaceutical formulations.

[0261] The term "salt" or "pharmaceutically acceptable salt" refers to salts derived from various organic and inorganic counterions well known in the art. Pharmaceutically acceptable acid addition salts can be formed with inorganic and organic acids. Inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like. Organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like. Pharmaceutically acceptable acid addition salts can be formed with inorganic and organic bases. Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like. Organic bases from which salts may be derived include, for example, primary, secondary, and tertiary amines, naturally occurring substituted amines, cyclic amines, substituted amines including basic ion exchange resins, and the like, specifically, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. In some embodiments, the pharma-ceutically acceptable base addition salts are selected from ammonium, potassium, sodium, calcium, and magnesium salts.

[0262] VI. Working Examples Example 1: Identification of variants in target oligonucleotides associated with increased or decreased risk of a disorder

[0263] Approximately 30 million imputed variants are analyzed in approximately 375,000 individuals from the Biobank cohort for association with liver disorders such as non-alcoholic steatohepatitis.

[0264] Protective or maladaptive associations are observed between certain allelic variants of various target genes and liver disease.In some cases, these associations suggest that therapeutic inhibition or modification of the target protein encoded by any of the target genes can be an effective genetically based treatment method for any of liver disorders.

[0265] Example 2: Bioinformatics Selection of Sequences to Identify Therapeutic siRNAs to Downregulate Expression of Target mRNAs

[0266] Based on bioinformatics analysis, screening sets are defined. Therapeutic siRNAs are designed to bind to human target mRNAs and target mRNA sequences of at least one toxicologically relevant species, such as non-human primate (NHP) species (e.g., rhesus or cynomolgus macaques). The drivers for the design of screening sets are the predicted specificity of the siRNAs to the transcriptome of the relevant species and the cross-reactivity between species. The predicted specificity in human, rhesus, cynomolgus, mouse, and rat is determined for the sense (S) and antisense (AS) strands. These are assigned a "specificity score", which takes into account the number and position of mismatches as well as the possibility of unintended downregulation of any other transcripts due to full or partial complementarity of the siRNA strands (up to four mismatches within positions 2-18). In this way, off-targets of the antisense and sense strands of each siRNA are identified. In addition, the number of possible off-targets is used as an additional specificity factor in the specificity score. If identified, siRNAs with high specificity and a low number of predicted off-targets confer the advantage of increasing the specificity of targeting.

[0267] In addition to selecting siRNA sequences with high sequence specificity for the target mRNA, the siRNA sequences in the seed region are analyzed for similarity to the seed regions of known miRNAs. siRNAs can function like miRNAs through base pairing with complementary sequences in the 3'-UTR of the mRNA molecule. The complementarity usually encompasses the 5'-base at positions 2-7 (seed region) of the miRNA. To avoid siRNAs that act through functional miRNA binding sites, siRNA strands that contain natural miRNA seed regions are avoided. Seed regions identified in miRNAs from human, mouse, rat, rhesus monkey, dog, rabbit, and pig are called "conserved". Combining the "specificity score" with the miRNA seed analysis results in a "specificity category", which is divided into categories 1-4, with 1 being the most specific and 4 being the least specific. Each strand of the siRNA is assigned to a specificity category.

[0268] Species cross-reactivity will be evaluated for human, cynomolgus monkey, rhesus monkey, mouse, and rat. Analysis will be based on canonical siRNA designs using 19 and 17 bases (without considering positions 1 and 19) and will be evaluated for cross-reactivity. Analysis of perfect matches and single mismatches will also be included.

[0269] Analysis of the human single nucleotide polymorphism (SNP) database (NCBI-DB-SNP) to identify siRNA targeting regions with known SNPs is also performed to identify siRNAs that may be non-functional in individuals containing the SNP. This analysis provides information on the location of the SNP within the target sequence, as well as the minor allele frequency (MAF) in case data.

[0270] The above method can be used to identify therapeutic siRNA for downregulating the expression of target mRNA. Bioinformatics method can also be used to identify ASO that binds to and downregulates the expression of target mRNA.

[0271] Example 3: Chemically modified siRNA

[0272] siRNAs that bind to target mRNAs can be synthesized by chemical modification with a sense strand having a modification such as modification pattern 1S, and an antisense strand having a modification such as modification pattern 1AS. In addition, an adenosine can be placed at position 19 of the sense strand and a uridine at position 1 of the antisense strand.

[0273] The siRNA that binds to the target mRNA can also be synthesized by chemical modification with a sense strand having modification pattern 2S and an antisense strand having modification pattern 3AS. In addition, adenosine can be placed at position 19 of the sense strand and uridine at position 1 of the antisense strand.

[0274] The siRNA that binds to the target mRNA can also be synthesized by chemical modification with a sense strand having a modification pattern 2S and an antisense strand having a modification pattern 9AS. In addition, adenosine can be placed at position 19 of the sense strand and uridine at position 1 of the antisense strand.

[0275] The siRNA targeted to bind to the target mRNA can also be synthesized by chemical modification with the sense strand having modification pattern 3S and the antisense strand having modification pattern 3AS. In addition, adenosine can be placed at position 19 of the sense strand and uridine can be placed at position 1 of the antisense strand.

[0276] The siRNA targeted to bind to the target mRNA may further be chemically modified with a sense strand having any of the following: all purines, including 2'fluoro modified purines, and all pyrimidines, including a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines, all purines, including 2'-O-methyl modified purines, and all pyrimidines, including a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines, all purines, including 2'fluoro modified purines, and all pyrimidines, including 2'-O-methyl modified pyrimidines, all pyrimidines, including 2'fluoro modified pyrimidines, and all purines, including a mixture of 2'fluoro and 2'-O-methyl modified purines, all pyrimidines, including 2'fluoro modified pyrimidines, and all purines, including 2'O-methyl modified pyrimidines, and all purines, including a mixture of 2'fluoro and 2'-O-methyl modified purines, or all pyrimidines, including 2'fluoro modified pyrimidines, and all purines, including 2'fluoro modified pyrimidines, and all purines, including 2'-O-methyl modified pyrimidines and further can be synthesized by chemical modification with an antisense strand having any of the following: all purines, including 2'fluoro modified purines, and all pyrimidines, including a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines, all purines, including 2'-O-methyl modified purines, and all pyrimidines, including a mixture of 2'fluoro and 2'-O-methyl modified pyrimidines, all purines, including 2'-O-methyl modified purines, and all pyrimidines, including 2'fluoro modified pyrimidines, all pyrimidines, including 2'fluoro modified pyrimidines, and all purines, including a mixture of 2'fluoro and 2'-O-methyl modified purines, all pyrimidines, including 2'fluoro modified pyrimidines, and all purines, including a mixture of 2'fluoro and 2'-O-methyl modified purines, all pyrimidines, including 2'O-methyl modified pyrimidines, and all purines, including a mixture of 2'fluoro and 2'-O-methyl modified purines, or all pyrimidines, including 2'O-methyl modified pyrimidines, and all purines, including 2'fluoro modified purines.

[0277] Example 4: Screening siRNAs for activity in cells in culture

[0278] Chemically modified siRNA derived from the sequence in the previous example is assayed for target mRNA knockdown activity in cells in culture. Cell lines expressing target mRNA are seeded in 96-well tissue culture plates at a density of 10,000 cells per well in DMEM supplemented with 10% fetal bovine serum and incubated overnight at 37°C in a water-jacketed humidified incubator under an air + 5% carbon dioxide atmosphere. siRNA is transfected individually into cells in duplicate wells at a final concentration of 10 nM using 0.3 μL per well of Lipofectamine RNAiMax (Fisher). Silencer Select Negative Control #1 (ThermoFisher, Cat. No. 4390843) and positive control siRNA are transfected at a final concentration of 10 nM as controls. After 48 hours of incubation at 37°C, total RNA was harvested from each well and cDNA was prepared using the TaqMan® Fast Advanced Cells-to-CT™ Kit (ThermoFisher, Cat. No. A35374) according to the manufacturer's instructions. The levels of target mRNA from each well are measured in triplicate by real-time qPCR on an Applied Biosystems 7500 Fast Real-Time PCR machine using the TaqMan Gene Expression Assay for human target mRNA. The levels of PPIA mRNA are measured using the TaqMan Gene Expression Assay (ThermoFisher) and used to determine the relative target mRNA levels in each well using the delta-delta Ct method. Data are normalized to the relative target mRNA levels in untreated cells.

[0279] The siRNA that shows the greatest degree of knockdown of target mRNA at 10 nM is tested in a second screening for activity at a concentration of 1 nM using the transfection procedure described above.Similar experiments can be carried out with ASO.Thus, the siRNA and ASO that most effectively downregulate the expression of target mRNA can be identified.

[0280] Example 5: GalNAc Ligands for Hepatocyte Targeting of Oligonucleotides

[0281] Without limiting the disclosure to these individual methods, there are at least two general methods for the attachment of polyvalent N-acetylgalactosamine (GalNAc) ligands to oligonucleotides: solid-phase or solution-phase conjugation. GalNAc ligands can be attached to solid-phase resins for 3' conjugation or at the 5' end using GalNAc phosphoramidite reagents. GalNAc phosphoramidites can be attached to solid phase at any position in the sequence, as well as other nucleosides in oligonucleotide sequences. Non-limiting examples of phosphoramidite reagents for GalNAc conjugation to oligonucleotides at the 5' end are shown in Table 1.

[0282] [Table 1]

[0283] Example 6: Synthesis of GalNAc Ligands Scheme for the preparation of NAcegal-linker-TMSOTf

[0284] [ka]

[0285] General procedure for the preparation of compound 2A

[0286] [ka]

[0287] To a solution of compound 1A (500 g, 4.76 mol, 476 mL) in 2-methyl-THF (2.00 L) was added CbzCl (406 g, 2.38 mol, 338 mL) in 2-methyl-THF (750 mL) dropwise at 0° C. The mixture was subjected to N 2 The mixture was stirred under atmospheric pressure for 2 hours at 25° C. TLC (DCM:MeOH=20:1, PMA) showed that CbzCl was completely consumed and one new spot (R f =0.43) was formed. HCl / EtOAc (1N, 180 mL) was added to the reaction mixture, stirred for 30 min, the white solid was removed by filtration through Celite, and the filtrate was concentrated in vacuo to give compound 2A (540 g, 2.26 mol, 47.5% yield) as a pale yellow oil, which was used in the next step without further purification. 1 H NMR:δ 7.28-7.41(m,5H),5.55(br s,1H),5.01-5.22(m,2H),3.63-3.80(m,2H),3.46-3.59(m,4H),3.29-3.44(m,2H),2.83-3.02(m,1H).

[0288] General procedure for the preparation of compound 4A

[0289] [ka]

[0290] To a solution of compound 3A (1.00 kg, 4.64 mol, HCl) in pyridine (5.00 L) was added acetyl acetate (4.73 kg, 46.4 mol, 4.34 L) under N 2 The mixture was poured into a 25 mL flask under atmospheric pressure at 0°C. 2 The mixture was stirred at 25° C. for 16 hours under atmospheric pressure. TLC (DCM:MeOH=20:1, PMA) showed that compound 3A was completely consumed and two new spots (R f=0.35). The reaction mixture was added to cold water (30.0 L) and stirred for 0.5 h at 0° C., a white solid was formed, which was filtered and dried to give compound 4A (1.55 kg, 3.98 mol, 85.8% yield) as a white solid, which was used in the next step without further purification. 1 H NMR:δ 7.90(d,J=9.29Hz,1H),5.64(d,J=8.78Hz,1H),5.26(d,J=3.01Hz,1H),5.06(dd,J=11.29,3.26Hz,1H),4. 22(t,J=6.15Hz,1H),3.95-4.16(m,3H),2.12(s,3H),2.03(s,3H),1.99(s,3H),1.90(s,3H),1.78(s,3H).

[0291] General procedure for the preparation of compound 5A

[0292] [ka]

[0293] To a solution of compound 4A (300 g, 771 mmol) in DCE (1.50 L), TMSOTf (257 g, 1.16 mol, 209 mL) was added and stirred at 60° C. for 2 h, then at 25° C. for 1 h. Compound 2A (203 g, 848 mmol) was dissolved in DCE (1.50 L) and N 2 Powdered 4 Å molecular sieves (150 g) were added with stirring under atmospheric pressure for 30 min. Then, a solution of compound 4A in DCE was added dropwise to the mixture at 0° C. The mixture was then purged with N 2 The mixture was stirred at 25° C. for 16 hours under atmospheric pressure. TLC (DCM:MeOH=25:1, PMA) showed that compound 4A was completely consumed and new spots (R f =0.24) was formed. The reaction mixture was filtered and saturated NaHCO 3 (2.00 L), water (2.00 L), and saturated brine (2.00 L). The organic layer was washed with anhydrous Na 2 SO 4The mixture was dried at 40° C., filtered, and concentrated under reduced pressure to give a residue which was triturated with 2-Me-THE / heptane (5 / 3, v / v, 1.80 L) for 2 h, filtered, and dried to give compound 5A (225 g, 389 mmol, 50.3% yield, 98.4% purity) as a white solid. 1 H NMR:δ 7.81(d,J=9.29Hz,1H),7.20-7.42(m,6H),5.21(d,J=3.26Hz,1H),4.92-5. 05(m,3H),4.55(d,J=8.28Hz,1H),3.98-4.07(m,3H),3.82-3.93(m,1H),3. 71-3.81(m,1H),3.55-3.62(m,1H),3.43-3.53(m,2H),3.37-3.43(m,2H),3 .14(q,J=5.77Hz,2H),2.10(s,3H),1.99(s,3H),1.89(s,3H),1.77(s,3H).

[0294] General procedure for the preparation of NAcegal-Linker-Tosylate Salts

[0295] [ka]

[0296] To a solution of compound 5A (200 g, 352 mmol) in THF (1.0 L), dried Pd / C (15.0 g, 10% purity) and TsOH (60.6 g, 352 mmol) were added under N 2 The suspension was degassed under vacuum and diluted with H 2 The mixture was purged multiple times with H 2 The mixture was stirred at 25° C. for 3 hours under an atmosphere of (45 psi). TLC (DCM:MeOH=10:1, PMA) showed that compound 5A was completely consumed and one new spot (R f=0.04) was formed. The reaction mixture was filtered and concentrated under reduced pressure (below 40 °C) to give a residue. The residue was diluted with anhydrous DCM (500 mL, dried over 4 Å molecular sieves overnight (dried at 300 °C for 12 h) and concentrated to give a Karl Fischer titration (KF) to check water content. This was repeated three times with dilutions of anhydrous DCM (500 mL) and concentrated to give NAcegal-Linker-TMSOTf (205 g, 95.8% yield, TsOH salt) as a foamy white solid. 1 H NMR:δ 7.91(d,J=9.03Hz,1H),7.53-7.86(m,2H),7.49(d,J=8.03Hz,2H),7.13(d,J=8.03Hz,2H),5.22(d,J=3.26Hz,1H),4.98(dd,J=1 1.29,3.26Hz,1H),4.57(d,J=8.53Hz,1H),3.99-4.05(m,3H),3.87-3.94(m,1H),3.79-3.85(m,1H),3.51-3.62(m,5H),2.96(br t,J=5.14Hz,2H),2.29(s,3H),2.10(s,3H),2.00(s,3H),1.89(s,3H),1.78(s,3H).

[0297] Scheme for the preparation of TRIS-PEG2-CBZ

[0298] [ka]

[0299] General procedure for the preparation of compound 5B

[0300] [ka]

[0301] To a solution of compound 4B (400 g, 1.67 mol, 1.00 equiv.) and NaOH (10 M, 16.7 mL, 0.10 equiv.) in THF (2.00 L), compound 4B_2 (1.07 kg, 8.36 mol, 1.20 L, 5.00 equiv.) was added and the mixture was stirred at 30° C. for 2 h. LCMS showed that the desired MS was given. Five batches of the solution were combined into one batch, and then the mixture was diluted with water (6.00 L) and ethyl acetate (3.00 L). * 3), and the combined organic layers were washed with brine (3.00 L) and 2 SO 4 The crude was purified by column chromatography (SiO 2 , petroleum ether: ethyl acetate = 100: 1 to 10: 1, R f =0.5) to give compound 5B (2.36 kg, 6.43 mol, 76.9% yield) as a pale yellow oil. HNMR: δ 7.31-7.36 (m, 5H), 5.38 (s, 1H), 5.11-5.16 (m, 2H), 3.75 (t, J=6.4 Hz), 3.54-3.62 (m, 6H), 3.39 (d, J=5.2 Hz), 2.61 (t, J=6.0 Hz).

[0302] General Procedure for the Preparation of 3-Oxo-1-phenyl-2,7,10-trioxa-4-azatridecan-13-oic acid (Compound 2B below)

[0303] [ka]

[0304] To a solution of compound 5B (741 g, 2.02 mol, 1.00 equiv) in DCM (2.80 L) was added TFA (1.43 kg, 12.5 mol, 928 mL, 6.22 equiv) and the mixture was stirred for 3 h at 25° C. LCMS showed the desired MS was given. The mixture was diluted with DCM (5.00 L) and water (3.00 L) was added. * 3), and then washed with brine (2.00 L). The combined organic layer was extracted with Na 2 SO 4The mixture was dried at 40° C., filtered, and concentrated in vacuo to give compound 2B (1800 g, crude) as a pale yellow oil. HNMR: δ 9.46(s,5H), 7.27-7.34(m,5H), 6.50-6.65(m,1H), 5.71(s,1H), 5.10-5.15(m,2H), 3.68-3.70(m,14H), 3.58-3.61(m,6H), 3.39(s,2H), 2.55(s,6H), 2.44(s,2H).

[0305] General procedure for the preparation of compound 3B

[0306] [ka]

[0307] To a solution of compound 2B (375 g, 999 mmol, 83.0% purity, 1.00 equiv.) in DCM (1.80 L), HATU (570 g, 1.50 mol, 1.50 equiv.) and DIEA (258 g, 2.00 mol, 348 mL, 2.00 equiv.) were added at 0° C. and the mixture was stirred at 0° C. for 30 min, after which compound 1B (606 g, 1.20 mol, 1.20 equiv.) was added and the mixture was stirred at 25° C. for 1 h. LCMS showed that the desired MS was given. The mixture was combined in one batch, then the mixture was diluted with DCM (5.00 L) and 1N aqueous HCl (2.00 L) was added. * 2), and then the organic layer was washed with saturated Na 2 CO 3 Aqueous solution (2.00L * 2) and brine (2.00 L), and the organic layer was washed with Na 2 SO 4 The mixture was dried at 40° C., filtered and concentrated in vacuo to give compound 3B (3.88 kg, crude) as a yellow oil.

[0308] General procedure for preparation of TRIS-PEG2-CBZ

[0309] [ka]

[0310] A solution of compound 3B (775 g, 487 mmol, 50.3% purity, 1.00 equiv) in HCl / dioxane (4M, 2.91 L, 23.8 equiv) was stirred for 2 h at 25° C. LCMS showed the desired MS was given. The mixture was concentrated under vacuum to give a residue. The combined residue was then diluted with DCM (5.00 L), adjusted to pH=8 with 2.5 M aqueous NaOH, and separated. The aqueous phase was extracted again with DCM (3.00 L), then the aqueous solution was adjusted to pH=3 with 1 N aqueous HCl, then diluted with DCM (5.00 L). * 2), and the combined organic layers were washed with brine (3.00 L) and 2 SO 4 The crude was purified by column chromatography (SiO 2 , DCM:MeOH=0:1 to 12:1, 0.1% HOAc, R f The residue was diluted with DCM (5.00 L), adjusted to pH=8 with 2.5 M aqueous NaOH, separated, and the aqueous solution was extracted again with DCM (3.00 L), and then the aqueous solution was adjusted to pH=3 with 6 N aqueous HCl and purified with DCM:MeOH=10:1 (5.00 L). * 2), and the combined organic layers were washed with brine (2.00 L) and 2 SO 4 The residue was dried over 1000 ml of ethyl acetate, filtered, and concentrated in vacuo to give a residue which was then diluted with MeCN (5.00 L) and concentrated in vacuo, this procedure was repeated twice to remove water to give TRIS-PEG2-CBZ (1.25 kg, 1.91 mol, 78.1% yield, 95.8% purity) as a pale yellow oil. 1 H NMR:400 MHz,MeOD,δ 7.30-7.35(5H),5.07(s,2H),3.65-3.70(m,16H),3.59(s,4H),3.45(t,J=5.6Hz),2.51(t,J=6.0Hz),2.43(t,6.4Hz).

[0311] Scheme for the preparation of TriNGal-TRIS-Peg2-Phosph 8c

[0312] [ka]

[0313] [ka]

[0314] General procedure for the preparation of compound 3C

[0315] [ka]

[0316] To a solution of compound 1C (155 g, 245 mmol, 1.00 equiv) in ACN (1500 mL), TBTU (260 g, 811 mmol, 3.30 equiv), DIEA (209 g, 1.62 mol, 282 mL, 6.60 equiv), and compound 2C (492 g, 811 mmol, 3.30 equiv, TsOH) were added at 0° C., and the mixture was stirred at 15° C. for 16 h. LCMS showed that the desired MS was given. The mixture was concentrated under vacuum to give a residue, after which the mixture was diluted with DCM (2000 mL) and washed with 1N aqueous HCl (700 mL×2), followed by saturated NaHCO. 3 It was washed with aqueous solution (700 mL×2) and concentrated under vacuum. The crude was purified by column chromatography to give compound 3C (304 g, 155 mmol, yield 63.1%, purity 96.0%) as a yellow solid.

[0317] General procedure for the preparation of compound 4C

[0318] [ka]

[0319] To two batches of a solution of compound 3C (55.0 g, 29.2 mmol, 1.00 equiv.) in MeOH (1600 mL), Pd / C (6.60 g, 19.1 mmol, 10.0% purity) and TFA (3.34 g, 29.2 mmol, 2.17 mL, 1.00 equiv.) were added, and the mixture was degassed under vacuum and diluted with H. 2 The mixture was purged with H for 2 h at 15 °C. 2 (15 psi). LCMS showed the desired MS was given. The mixture was filtered and the filtrate was concentrated in vacuo to give compound 4C (106 g, 54.8 mmol, 93.7% yield, 96.2% purity, TFA) as a white solid.

[0320] General procedure for the preparation of compound 5C

[0321] [ka]

[0322] Two batches in parallel. Compound 4a (25.0 g, 150 mmol, 1.00 equiv.) was added dropwise to a solution of EDCI (28.8 g, 150 mmol, 1.00 equiv.) in DCM (125 mL) at 0° C., then the mixture was added to compound 4 (25.0 g, 150 mmol, 1.00 equiv.) in DCM (125 mL) at 0° C., and the mixture was stirred for 1 h at 25° C. TLC (petroleum ether:ethyl acetate=3:1, R f = 0.45) indicated that the reactant was consumed and one new spot had formed. The reaction mixture was diluted with DCM (100 mL) and then washed with saturated NaHCO 3 Wash with aqueous solution (250 mL × 1) and brine (250 mL) and add Na 2 SO 4 The residue was purified by column chromatography (SiO 2 , petroleum ether:ethyl acetate=100:1-3:1), TLC (SiO 2 , petroleum ether:ethyl acetate=3:1), R f= 0.45, and then concentrated under reduced pressure to give a residue. Compound 5C (57.0 g, 176 mmol, 58.4% yield, 96.9% purity) was obtained as a colorless oil. 1 Characterized by H NMR: EW33072-2-P1A, 400 MHz, DMSO δ 9.21(s, 1H), 7.07-7.09(m, 2H), 6.67-6.70(m, 2H), 3.02-3.04(m, 2H), 2.86-2.90(m, 2H).

[0323] General procedure for the preparation of compound 6

[0324] [ka]

[0325] To a mixture of compound 3 (79.0 g, 41.0 mmol, 96.4% purity, 1.00 equiv, TFA) and compound 6C (14.2 g, 43.8 mmol, 96.9% purity, 1.07 equiv) in DCM (800 mL), TEA (16.6 g, 164 mmol, 22.8 mL, 4.00 equiv) was added dropwise at 0 °C and the mixture was stirred for 16 h at 15 °C. LCMS (EW33072-12-P1B, Rt = 0.844 min) showed the desired mass was detected. The reaction mixture was diluted with DCM (400 mL) and saturated NaHCO 3 Aqueous solution (400mL * 1) and brine (400 mL * 1), and the mixture was diluted with DCM (2.00 L) and 0.7 M Na 2 CO 3 (1000mL * 3) and brine (800 mL * 3) and washed with Na 2 SO 4 The mixture was dried at 40° C., filtered and concentrated under reduced pressure to give a residue which was used directly in the next step without purification. Compound 6 (80.0 g, crude) was obtained as a white solid. 1Characterized by H NMR: EW33072-12-P1A, 400 MHz, MeOD δ 7.02-7.04(m, 2H), 6.68-6.70(m, 2H), 5.34-5.35(s, 3H), 5.07-5.08(d, J=4.00Hz, 3H), 4.62-4.64(d, J=8.00Hz, 3H), 3.71-4.16(m, 16H), 3.31-3.70(m, 44H), 2.80-2.83(m, 2H), 2.68(m, 2H), 2.46-2.47(m, 10H), 2.14(s, 9H), 2.03(s, 9H), 1.94-1.95(d, J=4.00Hz, 18H).

[0326] General procedure for preparation of TriGNal-TRIS-Peg2-Phosph 8c

[0327] [ka]

[0328] Two batches were synthesized in parallel. To a solution of compound 6C (40.0 g, 21.1 mmol, 1.00 equiv) in DCM (600 mL) was added diisopropylammonium tetrazolide (3.62 g, 21.1 mmol, 1.00 equiv) and compound 7c (6.37 g, 21.1 mmol, 6.71 mL, 1.00 equiv) in DCM (8.00 mL) dropwise and the mixture was stirred at 30° C. for 1 h, then compound 7c (3.18 g, 10.6 mmol, 3.35 mL, 0.50 equiv) in DCM (8.00 mL) was added dropwise and the mixture was stirred at 30° C. for 30 min, then compound 7c (3.18 g, 10.6 mmol, 3.35 mL, 0.50 equiv) in DCM (8.00 mL) was added dropwise and the mixture was stirred at 30° C. for 1.5 h. LCMS (EW33072-17-P1C1, Rt = 0.921 min) showed that the desired MS+1 was detected. LCMS (EW33072-17-P1C2, Rt = 0.919 min) showed that the desired MS+1 was detected. The two batches were combined for workup. The mixture was diluted with DCM (1.20 L) and saturated NaHCO 3 Aqueous solution (1.60L * 2) H2 3% DMF in O (1.60L * 2) H 2 O(1.60L * 3), washed with brine (1.60 L), and Na 2 SO 4 The residue was purified by column chromatography (SiO 2 , DCM:MeOH:TEA=100:3:2)TLC(SiO 2 , DCM:MeOH=10:1, R f =0.45) and then concentrated under reduced pressure to give a residue. Compound 8C (76.0 g, 34.8 mmol, 82.5% yield, 96.0% purity) was obtained as a white solid. 1 Confirmed by H NMR: EW33072-19-P1C, 400 MHz, MeOD δ 7.13-7.15(d,J=8.50Hz,2H),6.95-6.97(dd,J=8.38,1.13Hz,2H),5.34(d,J=2.88Hz,3H),.09(dd,J=11.2 6,3.38Hz,3H),4.64(d,J=8.50Hz,3H),3.99-4.20(m,12H),3.88-3.98(m,5H),3.66-3.83(m,20H),3.51-3. 65(m,17H),3.33-3.50(m,9H),2.87(t,J=7.63Hz,2H),2.76(t,J=5.94Hz,2H),2.42-2.50(m,10H),2.14(s, 9H),2.03(s,9H),1.94-1.95(d,J=6.13Hz,18H),1.24-1.26(d,J=6.75Hz,6H),1.18-1.20(d,J=6.75Hz,6H)

[0329] Example 7: In vitro hepatocyte targeting using GalNAc-conjugated siRNA and ASO

[0330] In this experiment, a hepatocyte cell line expressing asialoglycoprotein receptor and GFP is treated with anti-GFP siRNA or ASO conjugated to GalNAc moiety, compared to a control experiment in which the hepatocyte cell line is treated with anti-GFP siRNA or ASO not conjugated to GalNAc moiety. GFP mRNA and protein expression are measured, and the amount of GFP mRNA or protein expression in cells treated with GalNAc conjugated siRNA or ASO is normalized and compared to the amount of GFP mRNA or protein expression in cells treated with siRNA or ASO not conjugated to GalNAc. This allows the hepatocyte targeting ability of GalNAc moiety to be determined. Multiple GalNAc moieties can be conjugated to siRNA or ASO and compared to see which GalNAc moiety provides optimal hepatocyte targeting. The GalNAc moiety tested in these experiments can be the GalNAc moiety described herein.

[0331] Similar experiments can be performed in primary hepatocytes treated with siRNA or ASO conjugated or not to a GalNAc moiety, and target mRNAs or proteins other than GFP can be evaluated in primary hepatocytes.

[0332] Example 8: In Vivo Hepatocyte Targeting Using GalNAc-Conjugated siRNA and ASO

[0333] In this experiment, siRNA or ASO targeting a target mRNA is conjugated to a GalNAc moiety and administered to mice (n=5 / group) and compared to a control experiment in which mice are administered siRNA or ASO without GalNAc conjugation. Mice are sacrificed after 2 days, and livers are frozen and later homogenized and tested for target mRNA and protein expression. The amount of target mRNA or protein expression in the liver of mice treated with GalNAc-conjugated siRNA or ASO is normalized and compared to the amount of GFP mRNA or protein expression in the liver of mice treated with non-GalNAc-conjugated siRNA or ASO. This allows the liver targeting ability of the GalNAc moiety to be determined. Multiple GalNAc moieties can be conjugated to siRNA or ASO and compared to see which GalNAc moiety provides optimal liver targeting. The GalNAc moiety included in this experiment can be the one that shows the greatest degree of hepatocyte targeting. The GalNAc moiety tested in these experiments can be the GalNAc moiety described herein.

[0334] Example 9: Inhibition of target mRNA in mouse models of liver disease using GalNAc-conjugated siRNA and ASO

[0335] In this experiment, a mouse model of liver disease (in this case, fatty liver disease) is used to evaluate the effect of siRNA or ASO inhibition of a target mRNA. The target mRNA can code for any target protein whose overexpression or overactivation plays a pathological role in liver disease. In the mouse model, fatty liver disease is induced by feeding mice for 12 weeks a Western diet (WD) containing 21.1% fat by weight, 41% sucrose by weight, and 1.25% cholesterol by weight (Teklad diets, TD.120528) and a high sugar solution (23.1 g / L d-fructose (Sigma-Aldrich, G8270) and 18.9 g / L d-glucose (Sigma-Aldrich, F0127)). Four-week-old C57BL / 6J mice are fed the Western diet instead of their normal chow for 12 weeks. The GalNAc moiety tested in this experiment can be a GalNAc moiety described herein.

[0336] Briefly, mice were divided into five groups: Group 1: fatty liver disease group treated with non-targeting control siRNA, Group 2: fatty liver disease group treated with non-targeting control ASO, Group 3: fatty liver disease group treated with siRNA targeting target mRNA, Group 4: fatty liver disease group treated with ASO targeting target mRNA, Group 5: control mice on normal diet. Each group contained eight mice (4 males, 4 females). The siRNAs and ASOs of groups 1 to 4 each contained a GalNAc moiety bound to the siRNA or ASO.

[0337] At week 12 on the Western diet, blood samples are taken from each group prior to the first treatment.

[0338] Administration of siRNA or ASO is achieved by subcutaneous injection of 200 μL of naked siRNA or ASO resuspended in PBS at a concentration of 10 μM. On day 0 of the study, mice in group 1 are subcutaneously injected with non-targeting control siRNA, mice in group 2 are subcutaneously injected with non-targeting control ASO, mice in group 3 are subcutaneously injected with siRNA1 that targets target mRNA in mice, mice in group 4 are subcutaneously injected with ASO1 that targets target mRNA in mice, and mice in group 5 are subcutaneously injected with vehicle. Then, starting from day 14, every other week, animals in each group are injected a total of 5 times as on day 0.

[0339] Blood is drawn weekly and serum and plasma are isolated. Serum ALT, AST, total cholesterol, and triglyceride levels are measured using a VITROS 5,1 FS (Ortho Clinical Diagnostics). Non-fasting plasma insulin is measured using an Ultrasensitive Mouse Insulin ELISA kit (Crystal Chem, 90080) according to the manufacturer's instructions. Non-fasting blood glucose is assayed using a One Touch Ultra (Life Scan). HOMA IR and QUICKI are calculated.

[0340] At the end of 12 weeks of Western diet and siRNA / ASO treatment, mice are sacrificed by cervical dislocation after intraperitoneal injection of 0.3 ml of Nembutal (5 mg / ml). Final serum collection is collected via cardiac puncture, and terminal serum ALT, AST, total cholesterol, and triglyceride levels are measured along with non-fasting plasma insulin and glucose. Livers are removed and divided into three sections, one section placed in RNAlater for mRNA isolation, one section snap frozen for protein isolation, and one section fixed in formalin and then embedded in paraffin.

[0341] mRNA is isolated from tissues placed in RNAlater solution using the PureLink kit according to the manufacturer's protocol (ThermoFisher catalog number 12183020). Reverse transcriptase reactions are performed according to the manufacturer's protocol. Samples are stored at -80°C until real-time qPCR is performed in triplicate using TaqMan Gene Expression Assays (Applied Biosystems FAM probes using a BioRad iCycler). Reduction in target mRNA expression in liver tissue from mice administered siRNA and ASO compared to target mRNA levels in liver tissue from mice administered nonspecific control siRNA and ASO. Reduction in the amount of SDF-1 in liver tissue from mice administered siRNA and ASO is expected compared to the amount of SDF-1 in liver tissue from mice administered nonspecific control siRNA or ASO. These results show that siRNA and ASO induce knockdown of target mRNA and target protein in liver tissue, and that reduction in target mRNA and target protein expression correlates with reduction in SDF-1 production.

[0342] Formalin-fixed paraffin-embedded liver sections are stained with hematoxylin and eosin (H&E) for evaluation of liver histology, Sirius Red (Sigma, 365548-5G) / Fast Green (Sigma, F258) for evaluation of fibrosis, and Periodic Acid Schiff (PAS) for evaluation of glycogen accumulation. NAFLD activity score (NAS) and fibrosis stage are evaluated by expert pathologists according to the NASH CRN scoring system13. Histological scoring is performed blinded, with the pathologists blinded to the treatments received. These results show that siRNA and ASO induce knockdown of target mRNA and target protein in liver tissue, and that the reduction in expression of target mRNA and target protein correlates with the reduction in NAS and NASH CRN.

[0343] Example 10: Inhibition of mouse models of liver disease

[0344] In this experiment, a mouse model of liver disease (in this case hypertriglyceridemia) is used to evaluate the effect of siRNA or ASO inhibition of a target protein expressed in the liver, compared to anti-mouse target protein antibodies. Mouse strain C57B1 / 6 Apoetm1Unc mice are maintained on a high-fat Western diet (Research Diets, D12492; 60% fat by calories). The target protein can be any target protein whose overexpression or overactivation plays a pathological role in liver disease. The GalNAc moiety tested in this experiment can be the GalNAc moiety described herein.

[0345] Four groups of mice (n=16 / group) will be utilized in this study. Animals will be maintained on a high fat diet for the duration of the study. On day -4 prior to the first injection, chow will be removed to allow for overnight fasting. On day -3 prior to the first injection, all animals will be anesthetized and 300uL of blood will be collected from the submandibular vein in serum separator tubes to assess baseline triglycerides, serum glucose, insulin sensitivity, total cholesterol levels, HDL cholesterol levels, liver function, and serum levels of target proteins. On study day 0, mice in group 1 are injected intraperitoneally with 600 μL saline, mice in group 2 are injected intraperitoneally with 600 μg of anti-mouse target protein antibody in 600 μL, mice in group 3 are injected subcutaneously with 150 μg of GalNAc-siRNA targeting the mRNA encoding the target protein in mice in 200 μL saline, and mice in group 4 are injected subcutaneously with 150 μg of GalNAc-ASO targeting the mRNA encoding the target protein in mice in 200 μL saline. In the afternoon of day 3, remove chow from all groups for overnight fasting. On day 4, animals in all groups are anesthetized and 150 μL of blood is collected from the submandibular vein into serum separator tubes to assess serum triglyceride, glucose, total cholesterol, HDL cholesterol, and target protein levels. Animals in all groups are then subjected to oral glucose tolerance tests and insulin tolerance tests to evaluate insulin sensitivity. After blood collection and insulin sensitivity testing, animals are again fed chow as normal. Starting on day 7, animals in group 2 are given a total of 15 injections every week, as on day 0. Then, starting on day 14, animals in groups 3 and 4 are given a total of 8 injections every other week, as on day 0. Starting on day 10, mice in all groups are fasted (overnight) and blood is drawn (150 μL into serum separator tubes) to assess serum triglyceride, glucose, total cholesterol, HDL cholesterol, and target protein levels, and insulin sensitivity testing is performed. On the 3rd day after the last injection, chow is removed from all groups in order to fast overnight.Four days after the last injection, animals from all groups are anesthetized, euthanized, and bled by cardiac puncture to collect 500 μL of blood into serum separator tubes to assess triglycerides, serum glucose, insulin sensitivity, total cholesterol levels, HDL cholesterol levels, liver function, and serum levels of target proteins. Liver, small intestine, and mesenteric lymph node tissues are harvested from all animals and placed in 10% neutral buffered formalin for histopathological analysis. Liver samples are also harvested from all animals and placed into RNAlater. Target mRNA levels are assessed by RT-qPCR using TaqMan assays for mouse target protein and mouse housekeeping gene PPIA.

[0346] Antibody-treated animals (Group 2), GalNAc-siRNA-treated mice (Group 3), and GalNAc-ASO-treated mice (Group 4) are expected to have reduced triglycerides, total serum cholesterol, serum glucose, and serum target protein levels, as well as increased HDL cholesterol and insulin sensitivity, compared to mice in Group 1 (saline). Group 2 and Group 3 animals are also expected to have reduced target mRNA in liver samples.

[0347] Example 11: Inhibition of target mRNA in non-human primates using GalNAc-siRNA and GalNAc-ASO

[0348] In this experiment, a hypertriglyceridemic NHP model is used to evaluate the effect of siRNA or ASO inhibition of a target mRNA expressed in the liver. The target protein can be any target protein whose overexpression or overactivation plays a pathological role in liver disease. Three groups of cynomolgus monkeys (n=5 / group) are used that are fed a high-fat diet (Western Primate Diet, 5S2T) before the start of the study. Alternatively, three groups of rhesus monkeys (n=5 / group) are used that are fed a high-fructose diet before the start of the study. The animals are injected subcutaneously seven times every other week with saline (group 1), GalNAc-siRNA (group 2), or GalNAc-ASO (group 3). The modified GalNAc-siRNA sequence can also include any of the modification patterns described herein. The GalNAc moiety used in this experiment can include the GalNAc moiety described herein. Blood samples for lipid and glycemic measurements are collected at baseline and at weeks 4, 8, and 14 of the study and analyzed for lipid content, serum glucose, insulin sensitivity, and target protein. All animals in each group are necropsied 2 weeks after the last blood draw. Liver, small intestine, and mesenteric lymph node tissues are collected from all animals and placed in 10% neutral buffered formalin for histopathological analysis. Liver samples are also collected from all animals and placed in RNAlater. Target mRNA levels are assessed by RT-qPCR using TaqMan assays for the target protein in cynomolgus or rhesus monkeys, and the housekeeping gene PPIA in cynomolgus or rhesus monkeys.

[0349] Animals treated with GalNAc-siRNA (Group 2) and GalNAc-ASO (Group 3) are expected to show decreased triglycerides, total serum cholesterol, serum glucose, and decreased serum target protein levels, as well as increased HDL cholesterol and insulin sensitivity, compared to animals in Group 1 (saline). Animals in Groups 1 and 3 are also expected to show decreased target mRNA in liver samples.

[0350] Example 12: Inhibition of target mRNA in clinical trials using GalNAc-siRNA and GalNAc-ASO

[0351] This study uses human patients with hypertriglyceridemia to evaluate the effect of siRNA or ASO inhibition of a target mRNA expressed in the liver. The target protein can be any target protein whose overexpression or overactivation plays a pathological role in liver disease. The inclusion criteria for inclusion in this study are age 40-90 years, BMI ≥ 30, and serum triglycerides ≥ 250 mg / dL. The study includes three groups of subjects (n = 15 / group). Subjects are to receive subcutaneous injections of saline (group 1), GalNAc-siRNA (group 2), or GalNAc-ASO (group 3) five times per week. The GalNAc moieties used in these experiments can include the GalNAc moieties described herein.

[0352] The siRNA or ASO sequences are from a selection set that shows high activity in cells in culture or in the experiments described in other examples. Blood samples for lipid and blood glucose measurements are taken at baseline and at weeks 3, 6, and 12 of the study and analyzed for lipid content, serum glucose, insulin sensitivity, target proteins, and liver and kidney function.

[0353] Subjects treated with GalNAc-siRNA (group 2) and subjects treated with GalNAc-ASO (group 3) are expected to show decreases in triglycerides, total serum cholesterol, serum glucose, and serum target protein levels, as well as increases in HDL cholesterol and insulin sensitivity compared to subjects in group 1 (saline).

[0354] Example 13: Oligonucleotide synthesis

[0355] RNAi agents (e.g., siRNA) were synthesized according to the phosphoramidite technique on the solid phase used in oligonucleotide synthesis. A K&A oligonucleotide synthesizer was used. Synthesis was performed on a solid support made of controlled pore glass (CPG, 500 Å or 600 Å, obtained from AM Chemicals, Oceanside, CA, USA). All 2'-OMe and 2'-F phosphoramidites were purchased from Hongene Biotech (Union City, CA, USA). All phosphoramidites were dissolved in anhydrous acetonitrile (100 mM) and molecular sieves (3 Å) were added. 5-benzylthio-1H-tetrazole (BTT, 250 mM in acetonitrile) or 5-ethylthio-1H-tetrazole (ETT, 250 mM in acetonitrile) were used as activator solutions. Coupling times were 9–18 min (EmpGalNAc), 6 min (2'OMe and 2'F). To introduce phosphorothioate linkages, a 100 mM solution of 3-phenyl-1,2,4-dithiazolin-5-one (POS, available from PolyOrg, Inc., Leominster, Massachusetts, USA) in anhydrous acetonitrile was used.

[0356] After solid-phase synthesis, the dried solid support was treated with a 1:1 volume solution of 40 wt% methylamine and 28% ammonium hydroxide solution (Aldrich) in water for 2 hours at 30°C. The solution was evaporated and the solid residue was reconstituted in water and purified by anion-exchange HPLC using a TKSgel SuperQ-5PW 13u column. Buffer A was 20 mM Tris, 5 mM EDTA, pH 9.0, containing 20% ​​acetonitrile, and buffer B was the same as buffer A with the addition of 1 M sodium chloride. The UV trace at 260 nm was recorded. Appropriate fractions were pooled and then desalted using Sephadex G-25 medium.

[0357] Duplexes were prepared by combining equimolar amounts of sense and antisense strands. Duplex solutions were prepared in 0.1x PBS (phosphate buffered saline, 1x, Gibco). Duplex solutions were annealed at 95°C for 5 min and slowly cooled to room temperature. Duplex concentrations were determined by measuring solution absorbance with a UV-Vis spectrometer at 260 nm in 0.1x PBS. In some experiments, conversion factors were calculated from experimentally determined extinction coefficients.

[0358] Example 14: In Vivo Hepatocyte Targeting Using GalNAc-Conjugated siRNA and ASO

[0359] In this experiment, siRNA or ASO targeting a target mRNA is conjugated to a GalNAc moiety and administered to mice (n=5 / group) and compared to a control experiment in which mice are administered siRNA or ASO without GalNAc conjugation. Mice are sacrificed after 2 days, and livers are frozen and later homogenized and tested for target mRNA and protein expression. The amount of target mRNA or protein expression in the liver of mice treated with GalNAc-conjugated siRNA or ASO is normalized and compared to the amount of GFP mRNA or protein expression in the liver of mice treated with non-GalNAc-conjugated siRNA or ASO. This allows the liver targeting ability of the GalNAc moiety to be determined. Multiple GalNAc moieties can be conjugated to siRNA or ASO and compared to see which GalNAc moiety provides optimal liver targeting. The GalNAc moiety can be the one that shows the greatest hepatocyte targeting in Example 6. The GalNAc moiety tested in these experiments can include the GalNAc moiety described herein, such as compound 1 or compound 2.

[0360] Example 15: Knockdown of PLIN1 in mice with GalNAc-conjugated siRNA

[0361] An example of a GalNAc moiety, ETL17, was conjugated to siRNA targeting an example of target mRNA. The sequence is shown in Table 2. In the table, Nf (e.g., Af, Cf, Gf, Tf, or Uf) is a 2'fluoro-modified nucleoside, n (e.g., a, c, g, t, or u) is a 2'O-methyl-modified nucleoside, and "s" is a phosphorothioate bond. siRNA was tested for activity in mice.

[0362] On day -14, 6- to 8-week-old female mice (C57Bl / 6) were transfected with 10 µL of recombinant adeno-associated virus 8 (AAV8) vectors (1.7 × 10E 13 AAV8 was injected at 100 ug of genomic copies / mL via retro-orbital route. Recombinant AAV8 contained the sequence of human PLIN1 (NM_002666.5) under the control of the human thyroxine-binding globulin promoter in an AAV2 backbone packaged in an AAV8 capsid (AAV8-TBGh-PLIN1). On day 0, infected mice (n=4) were subcutaneously injected with a single 100ug dose of GalNAc-conjugated siRNA or PBS as a vehicle control.

[0363] Mice were euthanized 10 days after subcutaneous injection and liver samples were collected from each and placed in RNAlater (ThermoFisher Cat. No. AM7020) until processing. Total liver RNA was prepared by homogenizing liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10-second cycles. Total RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) following the manufacturer's recommendations. cDNA preparation was performed using Quanta qScript cDNA SuperMix (VWR, Cat. No. 95048-500) following the manufacturer's instructions. Relative levels of hepatic PLIN1 mRNA were assessed by RT-qPCR in triplicate on a QuantStudio™ 6Pro Real-Time PCR System using TaqMan assays for human PLIN1 (ThermoFisher, assay number Hs01106925_m1) and mouse housekeeping gene PPIA (ThermoFisher, assay number Mm02342430_g1), as well as PerfeCTa® qPCR FastMix®, Low ROX™ (VWR, catalog number 101419-222). Data were normalized to the average PLIN1 mRNA levels of animals administered PBS. Results are shown in Table 3. All siRNAs tested caused a decrease in average hepatic PLIN1 mRNA at day 10 compared to mice administered PBS. These data indicate that siRNAs conjugated to GalNAc moieties, such as ETL17, are useful for knocking down target mRNA in the liver.

[0364] [Table 2]

[0365] [Table 3]

[0366] Example 16: Knockdown of MST1 in mice with GalNAc-conjugated siRNA

[0367] ETL17 is conjugated to siRNA that targets another target mRNA example. siRNA is bound to GalNAc ligand ETL17, and then phosphorothioate-linked at the 5' end of sense strand. siRNA is described in Table 6. In the table, Nf (e.g., Af, Cf, Gf, Tf, or Uf) is 2'fluoro-modified nucleoside, dN (e.g., dA, dC, dG, dT, or dU) is 2'deoxy-modified nucleoside, n (e.g., a, c, g, t, or u) is 2'O-methyl-modified nucleoside, and "s" is phosphorothioate linkage.

[0368] Six to eight week old female mice (C57Bl / 6) were injected with 5 μL of recombinant adeno-associated virus 8 (AAV8) vector (2.7 × 10E13 genome copies / mL) via retro-orbital route. The recombinant AAV8 contained the open reading frame and most of the 3'UTR of the human MST1 sequence (NM_020998.4) under the control of the human thyroxine-binding globulin promoter in an AAV2 backbone enveloped in an AAV8 capsid (AAV8-TBG-h-MST1). On day 13 post-infection, serum was collected and the levels of human MSP in each mouse were measured using a human MSP / MST1 DuoSet ELISA from R&D (catalog no. DY352). The manufacturer's instructions were followed for all reagent preparations of buffers and solutions. A serum sample dilution of 1:50 was utilized for all test samples. A standard curve ranging from 10,000 pg / mL to 0 pg / mL was generated using recombinant MSP provided in the kit. The optical density of the plate was read at 450 nm using a PerkinElmer Envision multimode plate reader. The concentration of MSP in each mouse serum sample was calculated from the standard curve by interpolation using a least-squares fit (Prism version 9, Software MacKiev).

[0369] Mice were assigned to groups (n=3) so that the groups had similar serum levels of MSP, and then subcutaneously injected with a single 60 μg dose of GalNAc-conjugated siRNA or PBS as vehicle control. On days 0, 4, and 12 after injection, serum was collected to evaluate serum MSP concentration by ELISA using the method described above. MSP serum concentration at each time point was made relative to the level of MSP in the day 0 sample for each individual mouse. The results are shown in Table 4. These data indicate that siRNA conjugated to a GalNAc moiety such as ETL17 is useful for knocking down proteins secreted by additional target mRNAs in the liver.

[0370] Mice were sacrificed on day 12 and liver samples were collected from each and placed in RNAlater (ThermoFisher Cat. No. AM7020) until processing. Total liver RNA was prepared by homogenizing liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10-second cycles. Total RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) following the manufacturer's recommendations. cDNA preparation was performed using Quanta qScript cDNA SuperMix (VWR, Cat. No. 95048-500) following the manufacturer's instructions. Relative levels of hepatic MST1 mRNA were assessed by RT-qPCR in triplicate on a QuantStudio™ 6Pro Real-Time PCR System using TaqMan assays for human MST1 (ThermoFisher, assay number Hs00360684_m1) and mouse housekeeping gene PPIA (ThermoFisher, assay number Mm02342430_g1), as well as PerfeCTa® qPCR FastMix®, Low ROX™ (VWR, catalog number 101419-222). Data were normalized to the levels of animals administered PBS. Results are shown in Table 5. These data indicated that siRNAs conjugated to GalNAc moieties such as ETL17 are useful for knocking down additional target mRNAs in the liver.

[0371] [Table 4]

[0372] [Table 5]

[0373] [Table 6]

[0374] Example 17: Knockdown of additional target mRNAs in mice with GalNAc-conjugated siRNA

[0375] Three groups (n=4 / group) of 8-week-old male ICR mice (Invigo) were utilized in the study. On study day 0, mice in group 1 were subcutaneously injected with 100 μL of sterile PBS, mice in group 2 were subcutaneously injected with 60 μg of siRNA 1811 in 100 μL of sterile PBS, and mice in group 3 were subcutaneously injected with 200 μg of siRNA 1818 in 100 μL of sterile PBS. On study day 14, animals from all groups were anesthetized and then euthanized. Liver samples were collected from all animals and placed in RNAlater™ Stabilization Solution (Thermo Fisher, Cat. No. AM7020). Liver samples were processed in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using Soft Tissue Homogenizing Kit CK14 (Bertin Instruments, Cat. No. P000933-LYSK0-A) in a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10-s cycles. Total RNA from liver lysates was purified on a Maxwell RSC 48 platform (Promega Corporation) according to the manufacturer's recommendations. The relative levels of gene A mRNA in each liver sample were assessed by RT-qPCR on a QuantStudio 6Pro instrument (Applied Biosystems) using TaqMan assays for mouse gene A and mouse housekeeping gene PPIA, then normalized to the mean value of control mice (group 1) using the delta-delta Ct method.

[0376] In this example, an additional GalNAc moiety (ETL1) was compared to ETL17. Results of liver mRNA analysis are shown in Table 7. Compared to PBS-injected mice (group 1), animals treated with ETL1-targeting siRNA (ETD01811, group 2) had a 78% relative knockdown, while ETL17-targeting siRNA (siRNA1818, group 3) had an 83% relative knockdown of liver gene A mRNA levels. Thus, compounds with GalNAc moieties described herein may be superior, or at least as effective, in knockdown of a target, such as a target mRNA, as compared to another GalNAc moiety.

[0377] [Table 7]

[0378] ETL1 is shown below, where J indicates attachment to the oligonucleotide.

[0379] [ka]

[0380] Example 18. Testing the activity of MST1 siRNAs ETD01821, ETD01822, ETD01823, and ETD01826 in non-human primates

[0381] Four groups (n=3 / group) of male cynomolgus monkeys (Zhaoqing Chuangyao Biotechnology Co., Ltd, and Guangzhou Xianngguan Biotechnology Co., Ltd) weighing 3–6 kg were utilized in this study.

[0382] On study day 0, cynomolgus monkeys in group 1 were injected subcutaneously with a single 5 mg / kg dose (0.2 mL dose volume / kg body weight) of ETD01821 at an siRNA concentration of 25 mg / mL formulated in PBS, cynomolgus monkeys in group 2 were injected with a single 5 mg / kg dose (0.2 mL dose volume / kg body weight) of ETD01822 at an siRNA concentration of 25 mg / mL formulated in PBS, cynomolgus monkeys in group 3 were injected with a single 5 mg / kg dose (0.2 mL dose volume / kg body weight) of ETD01823 at an siRNA concentration of 25 mg / mL formulated in PBS, and cynomolgus monkeys in group 4 were injected with a single 5 mg / kg dose (0.2 mL dose volume / kg body weight) of ETD01826 at an siRNA concentration of 25 mg / mL formulated in PBS. The siRNA sequences are shown in Table 8, where "Nf" is a 2'fluoro-modified nucleoside, "n" is a 2'O-methyl-modified nucleoside, and "s" is a phosphorothioate linkage. Injections were generally well tolerated, as measured by clinical symptoms.

[0383] All cynomolgus monkeys had no abnormal clinical signs during the study period, except for animal no. 101, which was found dead 65 days after dosing. Necropsy revealed severe gastric perforation, which may have been the cause of death, which can occur spontaneously in cynomolgus monkeys.

[0384] Body weights were recorded on study days -8, -1, 7, 14, 21, and 28. The results are shown in Table 9. The data show that animals in groups 1-4 did not experience significant changes in body weight.

[0385] On study days -8, -2, 7, 14, and 28, blood was collected into anticoagulant-free tubes and serum was collected. Clinical chemistry parameters including ALT, AST, ALP, TBIL, DBIL, GLU, GGT, TP, TG, CHOL, HDL, LDL, BUN, and CREA were analyzed. Animals treated with ETD01821, ETD01822, ETD01823, or ETD01926 showed no significant changes in ALT, AST, ALP, TBIL, DBIL, GLU, GGT, TP, TG, CHOL, HDL, LDL, BUN, and CREA starting from study day 7 to study day 28 when compared to study day -8 and study day -2 before treatment. Clinical chemistry results showed that all siRNAs were generally well tolerated. Results are shown in Tables 10-14.

[0386] On study days -8, -2, 7, 14, and 28, approximately 1 mL of whole blood was collected into tubes containing EDTA-K2 as an anticoagulant. Hematological parameters including WBC, NEUT, LYMP, MONO, EOS, BASO, RBC, HGB, HCT, MCV, MCH, MCHC, RDW, PLT, MPV, PCT, and PDW were analyzed. Animals treated with ETD01821, ETD01822, ETD01823, or ETD01926 showed no significant changes in these hematological parameters starting from study day 7 to study day 28 when compared to study day -8 and study day -2 before treatment. Results of hematological analysis showed that all siRNAs were generally well tolerated. Results are shown in Tables 15-19.

[0387] On study days -8, -2, 7, 14, 28, 42, 56, 70, 77, 84, 91, 98, and 105, blood was collected in anticoagulant-free tubes, serum was harvested, and serum macrophage stimulating protein (MSP) levels were determined. Additional serum samples were collected at later time points, i.e., days 42, 56, 70, 77, 84, 91, 98, and 105. Individual macrophage stimulating protein (MSP) concentrations were assessed in monkey serum samples using a custom AlphaLISA assay (PerkinElmer). Briefly, 5 μL of serum sample diluted 1:50 in 1× AlphaLISA HiBlock was placed into a well of a 96-well plate, followed by the addition of 5 μL of 4× anti-MSP acceptor bead solution. After 30 minutes of incubation at room temperature, 5 μL of 4× biotinylated anti-MSP antibody solution was added and the plate was incubated for 60 minutes at room temperature. Then, 5 μL of 4× streptavidin donor bead solution was added and the plate was incubated for an additional 30 minutes at room temperature. The plate was analyzed on an Envision 2105 Multimode Plate Reader (PerkinElmer). A standard curve was generated using recombinant human MSP (R&D Systems). The MSP serum concentration of each individual at each time point was generated relative to the mean of that individual's MSP serum concentration on days −2 and −8. Group mean results are shown in Table 20 and individual values ​​are shown in Table 21. Serum levels of MSP were reduced in all animals following treatment with the test article starting on day 7 and remained reduced until at least day 28. Monkeys treated with ETD01821 had the greatest reduction in serum MSP levels compared to pre-dose levels and showed a reduction in mean serum MSP levels compared to pre-dose levels through day 105.

[0388] On study days -8 and 28, animals were anesthetized with Zoletil (1.5-5.0 mg / kg, im) and Xylazine (0.5-2.0 mg / kg, im) and 3-4 mg liver biopsies were taken. Biopsies were then placed in 10 v / v RNAlater for 20 seconds and stored at 4°C for 24 hours, then the RNAlater™ Stabilization Solution (Thermo Fisher, Cat. No. AM7020) was removed and liver tissue was stored in a freezer until shipped to Empirico. There were no abnormal clinical observations for all animals following liver biopsy collection on days -2 or 28. Liver samples were processed in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using the Soft Tissue Homogenizing Kit CK14 (Bertin Instruments, Cat. No. P000933-LYSK0-A) in a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10-second cycles. Total RNA from liver lysates was purified on a Maxwell RSC 48 platform (Promega Corporation) following the manufacturer's recommendations. cDNA preparation was performed using Quanta qScript cDNA SuperMix (VWR, Cat. No. 95048-500) according to the manufacturer's instructions. Relative levels of liver MST1 mRNA were assessed in biplexed reactions by RT-qPCR in triplicate using TaqMan assays for cynomolgus MST1 (ThermoFisher, Assay No. Mf01117426_g1) and cynomolgus housekeeping gene GAPDH (ThermoFisher, Assay No. Mf04392546_g1) in PerfeCTa qPCR FastMix Reaction Mix (VWR). Samples were assessed on a QuantStudio™ 6Pro Real-Time PCR System. Relative amounts of MST1 mRNA were calculated using the delta-delta Ct method. Group mean relative MST1 mRNA levels for day -8 are shown in Table 22.Consistent with the reduction in serum MSP levels measured by AlphaLISA, treatment with 5 mg / kg test articles ETD01821, ETD01822, ETD01823, or ETD01826 resulted in a decrease in hepatic levels of MST1 mRNA on day 28 compared to pre-dose levels on day -8.

[0389] [Table 8]

[0390] [Table 9]

[0391] [Table 10-1]

[0392] [Table 10-2]

[0393] [Table 11-1]

[0394] [Table 11-2]

[0395] [Table 12-1]

[0396] [Table 12-2]

[0397] [Table 13-1]

[0398]

Table 13-2

[0399]

Table 14-1

[0400]

Table 14-2

[0401]

Table 15-1

[0402]

Table 15-2

[0403]

Table 15-3

[0404]

Table 16-1

[0405]

Table 16-2

[0406]

Table 16-3

[0407]

Table 17-1

[0408]

Table 17-2

[0409]

Table 17-3

[0410]

Table 18-1

[0411]

Table 18-2

[0412]

Table 18-3

[0413]

Table 19-1

[0414]

Table 19-2

[0415]

Table 19-3

[0416]

Table 20

[0417]

Table 21

[0418] [Table 22]

[0419] Example 19: Screening of siRNAs targeting human and mouse MTRES1 in mice

[0420] The following example shows the in vivo utility of the GalNAc moiety described herein when combined with an oligonucleotide targeting an additional target (MTRES1). Several siRNAs designed to cross-react with human, mouse, and cynomolgus monkey MTRES1 mRNA were tested for activity in mice. The siRNAs were conjugated to GalNAc ligands ETL1 or ETL17. The siRNA sequences are shown in Table 23, where Nf is a 2'fluoro-modified nucleoside, n is a 2'O-methyl-modified nucleoside, "d" is a deoxynucleoside, and "s" is a phosphorothioate linkage.

[0421] Female mice 6-8 weeks old (ICR strain, n=3) were injected subcutaneously on day 0 with a single 200ug dose of GalNAc-conjugated siRNA or PBS as vehicle control.

[0422] Mice were euthanized 10 days after injection and liver samples were collected from each and placed in RNAlater (ThermoFisher Cat. No. AM7020) until processing. Total liver RNA was prepared by homogenizing liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10-second cycles. Total RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) following the manufacturer's recommendations. cDNA preparation was performed using Quanta qScript cDNA SuperMix (VWR, Cat. No. 95048-500) following the manufacturer's instructions. Relative levels of hepatic MTRES1 mRNA were assessed by RT-qPCR in triplicate on a QuantStudio™ 6 Pro Real-Time PCR System using TaqMan assays for mouse MTRES1 (ThermoFisher, Assay No. Mm01229834_m1) and mouse housekeeping gene PPIA (ThermoFisher, Assay No. Mm02342430_g1), as well as PerfeCTa® qPCR FastMix®, Low ROX™ (VWR, Catalog No. 101419-222). Data were normalized to the mean MTRES1 mRNA levels of animals administered PBS. Results are shown in Table 24. Mice injected with ETD01597, ETD01955, ETD01958 had substantially lower mean hepatic MTRES1 mRNA levels on day 10 compared to mice administered PBS.

[0423] [Table 23]

[0424] [Table 24]

[0425] While preferred embodiments of the present invention have been shown and described herein, it will be apparent to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will occur to those skilled in the art without departing from the invention. It is understood that various alternatives to the embodiments of the invention described herein may be utilized in practicing the invention. The following claims define the scope of the invention, and it is intended that methods and structures within the scope of the claims, and their equivalents, be covered thereby.

Claims

1. Formula (I) or (II) 【Chemistry 1】 or a salt thereof, wherein J is an oligonucleotide; Each w is independently selected from any value from 1 to 20; Each v is independently selected from any value from 1 to 20; n is selected from any value between 1 and 20; m is selected from any value between 1 and 20; z is selected from any value from 1 to 3; When z is 3, Y is C; When z is 2, Y is CR ６ or When z is 1, Y is C(R ６ ) ２ and Q is halogen, -CN, -NO ２ , -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ , -S(O)R ７ , and C １－６ C optionally substituted with one or more substituents independently selected from alkyl ３－１０ carbocyclic ring, C １－６ Alkyl is a halogen, -CN, -OH, -SH, -NO ２ and -NH ２ and optionally substituted with one or more substituents independently selected from R １ is -O-, -S-, -N(R ７ )-, -C(O)-, -C(O)N(R ７ ) -, -N(R ７ )C(O)-,-N(R ７ ) C(O)N(R ７ )-,-OC(O)N(R ７ ) -, -N(R ７ )C(O)O-, -C(O)O-, -OC(O)-, -S(O)-, -S(O) ２ -, -OS (O) ２ -, -OP(O)(OR ７ )O-,-SP(O)(OR ７ )O-,-OP(S)(OR ７ )O-, -OP(O)(SR ７ )O-, -OP(O)(OR ７ )S-,-OP(O)(O － )O-,-SP(O)(O － )O-,-OP(S)(O － )O-, -OP(O)(S － )O-, -OP(O)(O － )S-,-OP(O)(OR ７ ) N.R. ７ -, -OP(O)(N(R ７ ) ２ ) N.R. ７ -, -OP(OR ７ )O-, -OP(N(R ７ ) ２ )O-, -OP(OR ７ ) N (R ７ )-, and -OPN(R ７ ) ２ N.R. ７ - is a linker selected from R ２ Each independently represents a halogen, -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ and -S(O)R ７ C optionally substituted with one or more substituents independently selected from １－６ alkyl, R ３ and R ４ Each independently represents -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ and -S(O)R ７ is selected from R ５ are each independently -OC(O)R ７ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)R ７ , -C(O)OR ７ and -C(O)N(R ７ ) ２ is selected from R ６ are each independently hydrogen, Halogen, -CN, -NO ２ , -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ and -S(O)R ７ , and Halogen, -CN, -NO ２ , -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ and -S(O)R ７ C optionally substituted with one or more substituents independently selected from １－６ alkyl, R ７ are each independently hydrogen, C １－６ Alkyl, C ２－６ Alkenyl, and C ２－６ Alkynyl (each of which is halogen, -CN, -OH, -SH, -NO ２ , -NH ２ , =O, =S, -O-C １－６ Alkyl, -S-C １－６ Alkyl, -N(C １－６ Alkyl) ２ , -NH(C １－６ Alkyl), C ３－１０ optionally substituted with one or more substituents independently selected from carbocycle, and 3- to 10-membered heterocycle; and C ３－１０ Carbocycles and 3- to 10-membered heterocycles, each of which may be selected from the group consisting of halogen, -CN, -OH, -SH, -NO ２ , -NH ２ , =O, =S, -O-C １－６ Alkyl, -S-C １－６ Alkyl, -N(C １－６ Alkyl) ２ , -NH(C １－６ Alkyl), C １－６ Alkyl, C ２－６ Alkenyl, C ２－６ Alkynyl, C ３－１０ Carbocycles, 3- to 10-membered heterocycles, and C １－６ haloalkyl), or a salt thereof.

2. The compound according to claim 1, or a salt thereof, wherein each w is independently selected from any value from 1 to 10.

3. The compound according to claim 1, or a salt thereof, wherein each w is independently selected from any value from 1 to 5.

4. The compound or salt thereof according to claim 1 , wherein each w is 1.

5. The compound according to claim 1, or a salt thereof, wherein each v is independently selected from any value from 1 to 10.

6. The compound according to claim 1, or a salt thereof, wherein each v is independently selected from any value from 1 to 5.

7. The compound or salt thereof according to claim 1 , wherein each v is 1.

8. The compound according to claim 1, or a salt thereof, wherein n is selected from any value from 1 to 10.

9. The compound according to claim 1, or a salt thereof, wherein n is selected from any value from 1 to 5.

10. 2. The compound according to claim 1, or a salt thereof, wherein n is 2.

11. The compound according to claim 1, or a salt thereof, wherein m is selected from any value from 1 to 10.

12. The compound according to claim 1, or a salt thereof, wherein m is selected from any value from 1 to 5.

13. 2. The compound according to claim 1, or a salt thereof, wherein m is selected from 1 and 2.

14. 2. The compound of claim 1, wherein z is 3 and Y is C, or a salt thereof.

15. Q is halogen, -CN, -NO ２ , -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ and -S(O)R ７ C optionally substituted with one or more substituents independently selected from ５－６ 2. The compound of claim 1 or a salt thereof, wherein the ring is selected from carbocycle.

16. Q is a halogen, -CN, -OH, -SH, or -NO ２ and -NH ２ C optionally substituted with one or more substituents independently selected from ５－６ 2. The compound of claim 1 or a salt thereof, wherein the ring is selected from carbocycle.

17. Q is selected from phenyl and cyclohexyl, each of which is halogen, —CN, —OH, —SH, —NO ２ and -NH ２ 2. The compound of claim 1, or a salt thereof, optionally substituted with one or more substituents independently selected from:

18. 2. The compound of claim 1 or a salt thereof, wherein Q is selected from phenyl.

19. 2. The compound of claim 1 or a salt thereof, wherein Q is selected from cyclohexyl.

20. R １ is -OP(O)(OR ７ )O-,-SP(O)(OR ７ )O-,-OP(S)(OR ７ )O-, -OP(O)(SR ７ )O-, -OP(O)(OR ７ )S-,-OP(O)(O － )O-,-SP(O)(O － )O-,-OP(S)(O － )O-, -OP(O)(S － )O-, -OP(O)(O － )S-,-OP(O)(OR ７ ) N.R. ７ -, -OP(O)(N(R ７ ) ２ ) N.R. ７ -, -OP(OR ７ )O-, -OP(N(R ７ ) ２ )O-, -OP(OR ７ ) N (R ７ )-, and -OPN(R ７ ) ２ N.R. ７ 2. The compound of claim 1, or a salt thereof, selected from:

21. R １ is -OP(O)(OR ７ )O-,-SP(O)(OR ７ )O-,-OP(S)(OR ７ )O-, -OP(O)(SR ７ )O-, -OP(O)(OR ７ )S-,-OP(O)(O － )O-,-SP(O)(O － )O-,-OP(S)(O － )O-, -OP(O)(S － )O-, -OP(O)(O － )S-, and -OP(OR ７ 2. The compound of claim 1, or a salt thereof, wherein:

22. R １ is -OP(O)(OR ７ )O-,-OP(S)(OR ７ )O-, -OP(O)(O － )O-,-OP(S)(O － )O-, -OP(O)(S － )O-, and -OP(OR ７ 2. The compound of claim 1, or a salt thereof, wherein:

23. R １ is -OP(O)(OR ７ )O- and -OP(OR ７ 2. The compound of claim 1, or a salt thereof, wherein:

24. R ２ is halogen, -OR ７ , -OC(O)R ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ and -S(O)R ７ C substituted with one or more substituents independently selected from １－３ 2. The compound of claim 1 or a salt thereof, wherein the aryl group is selected from the group consisting of aryl, ... and alkyl.

25. R ２ is -OR ７ , -OC(O)R ７ , -SR ７ , and -N(R ７ ) ２ C substituted with one or more substituents independently selected from １－３ 2. The compound of claim 1 or a salt thereof, wherein the aryl group is selected from the group consisting of aryl, ... and alkyl.

26. R ２ is -OR ７ and -OC(O)R ７ C substituted with one or more substituents independently selected from １－３ 2. The compound of claim 1 or a salt thereof, wherein the aryl group is selected from the group consisting of aryl, ... and alkyl.

27. R ３ is halogen, -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -OC(O)R ７ and -S(O)R ７ 2. The compound of claim 1, or a salt thereof, selected from:

28. R ３ is -OR ７ , -SR ７ , -OC(O)R ７ , and -N(R ７ ) ２ 2. The compound of claim 1, or a salt thereof, selected from:

29. R ３ is -OR ７ - and -OC(O)R ７ 2. The compound of claim 1, or a salt thereof, selected from:

30. R ４ is halogen, -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -OC(O)R ７ and -S(O)R ７ 2. The compound of claim 1, or a salt thereof, selected from:

31. R ４ is -OR ７ , -SR ７ , -OC(O)R ７ , and -N(R ７ ) ２ 2. The compound of claim 1, or a salt thereof, selected from:

32. R ４ is -OR ７ - and -OC(O)R ７ 2. The compound of claim 1, or a salt thereof, selected from:

33. R ５ is -OC(O)R ７ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , and -N(R ７ ) C(O)OR ７ 2. The compound of claim 1, or a salt thereof, selected from:

34. R ５ is -OC(O)R ７ and -N(R ７ ) C(O)R ７ 2. The compound of claim 1, or a salt thereof, selected from:

35. R ７ are each independently Hydrogen, and Halogen, -CN, -OH, -SH, -NO ２ , -NH ２ , =O, =S, -O-C １－６ Alkyl, -S-C １－６ Alkyl, -N(C １－６ Alkyl) ２ , -NH(C １－６ Alkyl), C ３－１０ C optionally substituted with one or more substituents independently selected from a carbocycle, or a 3- to 10-membered heterocycle １－６ 2. The compound of claim 1 or a salt thereof, wherein the aryl group is selected from the group consisting of aryl, ... and alkyl.

36. R ７ each independently represents a halogen, —CN, —OH, —SH, or —NO ２ , -NH ２ , =O, =S, -O-C １－６ Alkyl, -S-C １－６ Alkyl, -N(C １－６ Alkyl) ２ , and -NH(C １－６ C optionally substituted with one or more substituents independently selected from １－６ 31. The compound or salt thereof according to any one of claims 1 to 30, wherein:

37. R ７ each independently represents a C optionally substituted with one or more substituents independently selected from halogen, —CN, —OH, and —SH; １－６ 35. The compound according to any one of claims 1 to 34, or a salt thereof, wherein:

38. w is 1, v is 1, n is 2; m is 1 or 2; z is 3 and Y is C; Q is phenyl or cyclohexyl, each of which is halogen, —CN, —OH, —SH, —NO ２ , -NH ２ , and C １－３ optionally substituted with one or more substituents independently selected from alkyl; R １ is -OP(O)(OR ７ )O-,-OP(S)(OR ７ )O-, -OP(O)(O － )O-,-OP(S)(O － )O-, -OP(O)(S － )O-, and -OP(OR ７ ) O-; R ２ is -OH or -OC(O)CH ３ C substituted with １ is alkyl, R ３ is -OH or -OC(O)CH ３ and R ４ is -OH or -OC(O)CH ３ and R ５ -NH(O)CH ３ 2. The compound according to claim 1, wherein:

39. The compound is 【Chemistry 2-1】 【Chemistry 2-2】 【Chemistry 2-3】 【Chemistry 2-4】 【Chemistry 2-5】 【Chemistry 2-6】 【Chemistry 2-7】 【Chemistry 2-8】 【Chemistry 2-9】 2. The compound of claim 1, comprising:

40. The compound or salt thereof according to claim 1 , wherein the oligonucleotide (J) is linked at the 5′ end or the 3′ end of the oligonucleotide.

41. The compound or salt thereof according to claim 1 , wherein the oligonucleotide comprises DNA.

42. The compound or salt thereof of claim 1 , wherein the oligonucleotide comprises RNA.

43. 2. The compound or salt thereof of claim 1, wherein the oligonucleotide comprises one or more modified internucleoside linkages.

44. 44. The compound of claim 43, or a salt thereof, wherein the one or more modified internucleoside linkages comprise an alkyl phosphonate, phosphorothioate, methyl phosphonate, phosphorodithioate, alkyl phosphonothioate, phosphoramidate, carbamate, carbonate, phosphate triester, acetamidate, or carboxymethyl ester, or a combination thereof.

45. 2. The compound of claim 1, or a salt thereof, wherein the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 modified internucleoside linkages.

46. The compound or salt thereof according to claim 1 , wherein the oligonucleotide comprises one or more modified nucleosides.

47. 47. The compound of claim 46, or a salt thereof, wherein the one or more modified nucleosides comprise a locked nucleic acid (LNA), a hexitol nucleic acid (HLA), a cyclohexene nucleic acid (CeNA), a 2'-methoxyethyl, a 2'-O-alkyl, a 2'-O-allyl, a 2'-O-allyl, a 2'-fluoro, or a 2'-deoxy, or a combination thereof.

48. 47. The compound of claim 46, or a salt thereof, wherein the one or more modified nucleosides comprise a 2',4' constrained ethyl nucleoside, a 2'-O-methyl nucleoside, a 2'-deoxyfluoro nucleoside, a 2'-O-N-methylacetamide (2'-O-NMA), a 2'-O-dimethylaminoethoxyethyl (2'-O-DMAEOE) nucleoside, a 2'-O-aminopropyl (2'-O-AP) nucleoside, a 2'-ara-F, a 2'fluoro, or a 2'O-alkyl, or a combination thereof.

49. 2. The compound of claim 1, or a salt thereof, wherein the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, or more modified nucleosides.

50. 2. The compound or salt thereof of claim 1, wherein the oligonucleotide comprises a lipid attached at the 3' or 5' end of the oligonucleotide.

51. 51. The compound or salt thereof of claim 50, wherein the lipid comprises cholesterol, myristoyl, palmitoyl, stearoyl, lithochloyl, docosanoyl, docosahexaenoyl, myristyl, palmitylstearyl, or α-tocopherol, or a combination thereof.

52. 2. The compound or salt thereof of claim 1, wherein the oligonucleotide comprises an arginine-glycine-aspartic acid (RGD) peptide attached at the 3' or 5' end of the oligonucleotide.

53. 53. The compound or salt thereof according to claim 52, wherein the RGD peptide comprises cyclo(-Arg-Gly-Asp-D-Phe-Cys), cyclo(-Arg-Gly-Asp-D-Phe-Lys), cyclo(-Arg-Gly-Asp-D-Phe-azide), RGD derived from aminobenzoic acid, or a combination thereof.

54. 2. The compound or salt thereof of claim 1, wherein the oligonucleotide comprises a small interfering RNA (siRNA) comprising a sense strand and an antisense strand.

55. 55. The compound or salt thereof according to claim 54, wherein the sense strand is 12 to 30 nucleosides in length.

56. 55. The compound or salt thereof according to claim 54, wherein the antisense strand is 12 to 30 nucleosides in length.

57. 55. The compound or salt thereof of claim 54, wherein the sense strand and the antisense strand form a double-stranded RNA duplex.

58. 58. The compound or salt thereof of claim 57, wherein the first base pair of the double-stranded RNA duplex is an AU base pair.

59. 55. The compound or salt thereof of claim 54, wherein the sense strand or the antisense strand comprises a 3' overhang.

60. 60. The compound or salt thereof of claim 59, wherein the 3' overhang comprises one, two, or more nucleosides.

61. 2. The compound or salt thereof of claim 1, wherein the oligonucleotide comprises an antisense oligonucleotide (ASO).

62. 62. The compound or salt thereof of claim 61, wherein the ASO is 12 to 30 nucleosides in length.

63. The compound or salt thereof according to claim 1 , wherein the compound binds to an asialoglycoprotein receptor.

64. 2. The compound or salt thereof of claim 1, wherein the compound targets hepatocytes.

65. 13. A pharmaceutical composition comprising a compound of claim 1 and a pharma- ceutically acceptable carrier, excipient, or diluent.

66. 66. The pharmaceutical composition of claim 65, wherein the pharmaceutical composition is sterile.

67. 66. The pharmaceutical composition of claim 65, wherein the pharmaceutical composition comprises a pharma- ceutically acceptable carrier.

68. 68. The pharmaceutical composition of claim 67, wherein the pharma- ceutically acceptable carrier comprises water, a buffer, or saline.

69. 66. The pharmaceutical composition of claim 65, wherein the oligonucleotide targets a target mRNA and, when administered to a subject in an effective amount, reduces the target mRNA or protein by at least 10%.

70. 66. A method for reducing a target mRNA or a target protein in a subject, comprising administering to the subject an effective amount of the pharmaceutical composition of claim 65.

71. 71. The method of claim 70, wherein the effective amount reduces the measurement of a target mRNA or target protein in the subject as compared to a baseline target mRNA or target protein measurement.

72. 71. The method of claim 70, wherein the effective amount treats a disorder in the subject.

73. 73. The method of claim 72, wherein the effective amount reduces the measurement of a symptom or parameter associated with the disorder in the subject compared to a baseline measurement of the symptom or parameter.

74. 73. The method of claim 72, wherein the measurement of the symptom or parameter associated with the disorder in the subject is reduced for at least 10 days.

75. 73. The method of claim 72, wherein the disorder comprises a metabolic disorder.

76. 73. The method of claim 72, wherein the disorder comprises a liver disorder.

77. Formula (A) or (B) 【Chemistry 3】 or a salt thereof, wherein Each w is independently selected from any value from 1 to 20; Each v is independently selected from any value from 1 to 20; n is selected from any value between 1 and 20; m is selected from any value between 1 and 20; z is selected from any value from 1 to 3; When z is 3, Y is C; When z is 2, Y is CR ６ or When z is 1, Y is C(R ６ ) ２ and Q is halogen, -CN, -NO ２ , -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ , -S(O)R ７ , and C １－６ C optionally substituted with one or more substituents independently selected from alkyl ３－１０ carbocyclic ring, C １－６ Alkyl is a halogen, -CN, -OH, -SH, -NO ２ and -NH ２ and optionally substituted with one or more substituents independently selected from R １ は、-OR ７ 、-SR ７ 、-N(R ７ ) ２ 、-C(O)R ７ 、-C(O)N(R ７ ) ２ 、-N(R ７ )C(O)R ７ 、-N(R ７ )C(O)N(R ７ ) ２ 、-OC(O)N(R ７ ) ２ 、-N(R ７ )C(O)OR ７ 、-C(O)OR ７ 、-OC(O)R ７ 、-S(O)R ７ 、-S(O) ２ R ７ 、-OS(O) ２ R ７ 、-OP(O)(OR ７ ) ２ 、-OP(S)(OR ７ ) ２ 、-SP(O)(OR ７ ) ２ 、-OP(O)(SR ７ )(OR ７ )、-OP(O)(OR ７ )N(R ７ ) ２ 、-OP(S)(OR ７ )N(R ７ ) ２ 、-SP(O)(OR ７ )N(R ７ ) ２ 、-OP(O)(SR ７ )N(R ７ ) ２ 、-OP(O)(N(R ７ ) ２ ) ２ 、-OP(S)(N(R ７ ) ２ ) ２ 、-SP(O)(N(R ７ ) ２ ) ２ 、-OP(OR ７ ) ２ 、-SP(OR ７ ) ２ , -OP(OR ７ ) (SR ７ ), -OP(OR ７ ) N (R ７ ) ２ , -OP(SR ７ ) N (R ７ ) ２ , -SP(OR ７ ) N (R ７ ) ２ , -OP(N(R ７ ) ２ ) ２ , and -SP(N(R ７ ) ２ ) ２ is selected from R ２ Each independently represents a halogen, -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ and -S(O)R ７ C optionally substituted with one or more substituents independently selected from １－６ alkyl, R ３ and R ４ Each independently represents -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ and -S(O)R ７ is selected from R ５ are each independently -OC(O)R ７ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)R ７ , -C(O)OR ７ and -C(O)N(R ７ ) ２ is selected from R ６ are each independently hydrogen, Halogen, -CN, -NO ２ , -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ and -S(O)R ７ , and Halogen, -CN, -NO ２ , -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ and -S(O)R ７ C optionally substituted with one or more substituents independently selected from １－６ alkyl, R ７ are each independently hydrogen, C １－６ Alkyl, C ２－６ Alkenyl, and C ２－６ Alkynyl (each of which is halogen, -CN, -OH, -SH, -NO ２ , -NH ２ , =O, =S, -O-C １－６ Alkyl, -S-C １－６ Alkyl, -N(C １－６ Alkyl) ２ , -NH(C １－６ Alkyl), C ３－１０ optionally substituted with one or more substituents independently selected from carbocycle, and 3- to 10-membered heterocycle; and C ３－１０ Carbocycles and 3- to 10-membered heterocycles, each of which may be selected from the group consisting of halogen, -CN, -OH, -SH, -NO ２ , -NH ２ , =O, =S, -O-C １－６ Alkyl, -S-C １－６ Alkyl, -N(C １－６ Alkyl) ２ , -NH(C １－６ Alkyl), C １－６ Alkyl, C ２－６ Alkenyl, C ２－６ Alkynyl, C ３－１０ Carbocycles, 3- to 10-membered heterocycles, and C １－６ haloalkyl), or a salt thereof.

78. 78. The compound according to claim 77, or a salt thereof, wherein each w is independently selected from any value from 1 to 10.

79. 78. The compound according to claim 77, or a salt thereof, wherein each w is independently selected from any value from 1 to 5.

80. 78. The compound or salt thereof of claim 77, wherein each w is 1.

81. 78. The compound according to claim 77, or a salt thereof, wherein each v is independently selected from any value from 1 to 10.

82. 78. The compound according to claim 77, or a salt thereof, wherein each v is independently selected from any value from 1 to 5.

83. 78. The compound of claim 77, or a salt thereof, wherein each v is 1.

84. 78. The compound according to claim 77, or a salt thereof, wherein n is selected from any value from 1 to 10.

85. 78. The compound according to claim 77, or a salt thereof, wherein n is selected from any value from 1 to 5.

86. 78. The compound or salt thereof according to claim 77, wherein n is 2.

87. 78. The compound according to claim 77, or a salt thereof, wherein m is selected from any value from 1 to 10.

88. 78. The compound according to claim 77, or a salt thereof, wherein m is selected from any value from 1 to 5.

89. 78. The compound of claim 77, or a salt thereof, wherein m is selected from 1 and 2.

90. 78. The compound of claim 77, or a salt thereof, wherein z is 3 and Y is C.

91. Q is halogen, -CN, -NO ２ , -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -C(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)OR ７ , -C(O)OR ７ , -OC(O)R ７ and -S(O)R ７ C optionally substituted with one or more substituents independently selected from ５－６ 78. The compound of claim 77 or a salt thereof, wherein said compound is selected from carbocycles.

92. Q is a halogen, -CN, -OH, -SH, or -NO ２ and -NH ２ C optionally substituted with one or more substituents independently selected from ５－６ 78. The compound of claim 77 or a salt thereof, wherein said compound is selected from carbocycles.

93. Q is selected from phenyl and cyclohexyl, each of which is halogen, —CN, —OH, —SH, —NO ２ and -NH ２ 78. The compound of claim 77, or a salt thereof, optionally substituted with one or more substituents independently selected from:

94. 78. The compound of claim 77, or a salt thereof, wherein Q is selected from phenyl.

95. 78. The compound of claim 77, or a salt thereof, wherein Q is selected from cyclohexyl.

96. R １ is -OP(O)(OR ７ ) ２ , -OP(O)(OR ７ ) N (R ７ ) ２ , -OP(O)(N(R ７ ) ２ ) ２ , -OP(OR ７ ) ２ , -OP(OR ７ ) N (R ７ ) ２ , and -OP((NR ７ ) ２ ) ２ 78. The compound of claim 77, or a salt thereof, selected from:

97. R １ is -OP(O)(OR ７ ) ２ and -OP(OR ７ ) N (R ７ ) ２ 78. The compound of claim 77, or a salt thereof, selected from:

98. R １ is -OP(O)(OCH ２ CH ３ )OH and -OP(OCH ２ CH ２ CN)N(CH(CH ３ ) ２ ) ２ 78. The compound of claim 77, or a salt thereof, selected from:

99. R １ is -OP(OCH ２ CH ２ CN)N(CH(CH ３ ) ２ ) ２ 78. The compound of claim 77, or a salt thereof.

100. R ２ is halogen, -OR ７ , -OC(O)R ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ and -S(O)R ７ C substituted with one or more substituents independently selected from １－３ 78. The compound of claim 77, or a salt thereof, wherein said compound is selected from alkyl.

101. R ２ is -OR ７ , -OC(O)R ７ , -SR ７ , and -N(R ７ ) ２ C substituted with one or more substituents independently selected from １－３ 78. The compound of claim 77, or a salt thereof, wherein said compound is selected from alkyl.

102. R ２ is -OR ７ and -OC(O)R ７ C substituted with one or more substituents independently selected from １－３ 78. The compound of claim 77, or a salt thereof, wherein said compound is selected from alkyl.

103. R ３ is halogen, -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -OC(O)R ７ and -S(O)R ７ 78. The compound of claim 77, or a salt thereof, selected from:

104. R ３ is -OR ７ , -SR ７ , -OC(O)R ７ , and -N(R ７ ) ２ 78. The compound of claim 77, or a salt thereof, selected from:

105. R ３ is -OR ７ - and -OC(O)R ７ 78. The compound of claim 77, or a salt thereof, selected from:

106. R ４ is halogen, -OR ７ , -SR ７ , -N(R ７ ) ２ , -C(O)R ７ , -OC(O)R ７ and -S(O)R ７ 78. The compound of claim 77, or a salt thereof, selected from:

107. R ４ is -OR ７ , -SR ７ , -OC(O)R ７ , and -N(R ７ ) ２ 78. The compound of claim 77, or a salt thereof, selected from:

108. R ４ is -OR ７ - and -OC(O)R ７ 78. The compound of claim 77, or a salt thereof, selected from:

109. R ５ is -OC(O)R ７ , -OC(O)N(R ７ ) ２ , -N(R ７ ) C(O)R ７ , -N(R ７ ) C(O)N(R ７ ) ２ , and -N(R ７ ) C(O)OR ７ 78. The compound of claim 77, or a salt thereof, selected from:

110. R ５ is -OC(O)R ７ and -N(R ７ ) C(O)R ７ 78. The compound of claim 77, or a salt thereof, selected from:

111. R ７ are each independently Hydrogen, and Halogen, -CN, -OH, -SH, -NO ２ , -NH ２ , =O, =S, -O-C １－６ Alkyl, -S-C １－６ Alkyl, -N(C １－６ Alkyl) ２ , -NH(C １－６ Alkyl), C ３－１０ C optionally substituted with one or more substituents independently selected from a carbocycle, or a 3- to 10-membered heterocycle １－６ 78. The compound of claim 77, or a salt thereof, wherein said compound is selected from alkyl.

112. R ７ each independently represents a halogen, —CN, —OH, —SH, or —NO ２ , -NH ２ , =O, =S, -O-C １－６ Alkyl, -S-C １－６ Alkyl, -N(C １－６ Alkyl) ２ , and -NH(C １－６ C optionally substituted with one or more substituents independently selected from １－６ 78. The compound of claim 77, or a salt thereof, wherein said compound is selected from alkyl.

113. R ７ each independently represents a C optionally substituted with one or more substituents independently selected from halogen, —CN, —OH, and —SH; １－６ 78. The compound of claim 77, or a salt thereof, wherein said compound is selected from alkyl.

114. w is 1, v is 1, n is 2; m is 1 or 2; z is 3 and Y is C; Q is phenyl or cyclohexyl, each of which is halogen, —CN, —OH, —SH, —NO ２ , -NH ２ , and C １－３ optionally substituted with one or more substituents independently selected from alkyl; R １ is -OP(O)(OR ７ ) ２ and -OP(OR ７ ) N (R ７ ) ２ is selected from R ２ is -OH or -OC(O)CH ３ C substituted with １ is alkyl, R ３ is -OH or -OC(O)CH ３ and R ４ is -OH or -OC(O)CH ３ and R ５ -NH(O)CH ３ 114. The compound of claim 113, wherein:

115. The compound is 【Chemistry 4-1】 【Chemistry 4-2】 【Chemistry 4-3】 【Chemistry 4-4】 【Chemistry 4-5】 【Chemistry 4-6】 78. The compound of claim 77, or a salt thereof.

116. 【Chemical 5】 wherein J comprises an oligonucleotide or comprises a bond to said oligonucleotide.

117. 117. The compound of claim 116, wherein J further comprises one or more phosphate groups attaching the structure to the oligonucleotide.

118. 117. The compound of claim 116, wherein J further comprises one or more phosphorothioate groups linking the structure to the oligonucleotide.

119. 117. The compound of claim 116, wherein the oligonucleotide comprises a 5' end and J comprises a bond to the 5' end of the oligonucleotide.

120. The compound of claim 116, wherein the oligonucleotide comprises an siRNA.

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