GALNAC composition for improving siRNA bioavailability

Abstract

Provided herein are compositions comprising a GalNAc moiety that can be conjugated to an oligonucleotide. The oligonucleotide may be a small interfering RNA or an antisense oligonucleotide. Also provided herein are methods of treatment comprising administering the compositions to a subject.

Description

[0001] Cross-references

[0002] This application claims the benefit of U.S. Provisional Application No. 63 / 320,431 filed on March 16, 2022, U.S. Provisional Application No. 63 / 354,359 filed on June 22, 2022, and U.S. Provisional Application No. 63 / 430,542 filed on December 6, 2022, which are incorporated herein by reference.

[0003] Incorporation by Reference of Sequence Listing

[0004] This application is accompanied by a sequence listing submitted in electronic format. The sequence listing is provided as a file named "54462-735601_PCT.xml", which was created on March 13, 2023 and is 922,453 bytes in size. The information in the electronic format of the sequence listing is incorporated herein by reference in its entirety. Background Art

[0005] Cardiovascular, metabolic, and liver-related conditions are numerous and affect a wide range of populations. Improved therapies are needed to treat these conditions. Summary of the invention

[0006] Disclosed herein is a compound represented by formula (I) or (II):

[0007]

[0008] or a salt thereof, wherein

[0009] J is an oligonucleotide;

[0010] Each w is independently selected from any value from 1 to 20;

[0011] Each v is independently selected from any value from 1 to 20;

[0012] n is selected from any value from 1 to 20;

[0013] m is selected from any value from 1 to 20;

[0014] z is selected from any value from 1 to 3, wherein

[0015] If z is 3, then Y is C;

[0016] If z is 2, then Y is CR 6 ,or

[0017] If z is 1, then Y is C(R 6 )2;

[0018] Q is selected from:

[0019] C optionally substituted by one or more substituents 3-10 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 Alkyl, where C 1-6 The alkyl group is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2;

[0020] R 1 is a linker selected from the group consisting of:

[0021] -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)(OR7 )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 )2NR 7 -;

[0022] Each R 2 Independently selected from:

[0023] C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from 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 ;

[0024] R 3 and R 4 Each independently selected from:

[0025] -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)R7 ;

[0026] Each R 5 Independently selected from:

[0027] -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;

[0028] Each R 6 Independently selected from:

[0029] hydrogen;

[0030] Halogen, -CN, -NO2, -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 ;as well as

[0031] C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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(R7 )2、-N(R 7 )C(O)OR 7 、-C(O)OR 7 、-OC(O)R 7 and-S(O)R 7 ;

[0032] Each R 7 Independently selected from:

[0033] hydrogen;

[0034] C 1-6 Alkyl, C 2-6 Alkenyl and C 2-6 Alkynyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 Carbocyclic rings and 3- to 10-membered heterocyclic rings; and

[0035] C 3-10 carbocyclic and 3- to 10-membered heterocyclic rings, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -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 ring, 3- to 10-membered heterocyclic ring and C 1-6 Halogenated alkyl.

[0036] 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 C optionally substituted with one or more substituents. 5-6 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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, Q is selected from C optionally substituted with one or more substituents. 5-6 In some embodiments, Q is selected from phenyl and cyclohexyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2. In some embodiments, Q is selected from phenyl. In some embodiments, Q is selected from cyclohexyl. In some embodiments, R 1 Selected from -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 )2NR 7 In some embodiments, R 1 Selected from -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 )O-. In some embodiments, R 1 Selected from -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-. In some embodiments, R 1 Selected from -OP(O)(OR 7 )O- and -OP(OR 7 )O-. In some embodiments, R 2 is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from halogen, -OR 7 、-OC(O)R 7 、-SR 7、-N(R 7 )2. -C(O)R 7 and-S(O)R 7 In some embodiments, R 2 is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from -OR 7 、-OC(O)R 7 、-SR 7 and -N(R 7 )2. In some embodiments, R 2 is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from -OR 7 and -OC(O)R 7 In some embodiments, R 3 Selected from 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 Select from -OR 7 、-SR 7 、-OC(O)R 7 and -N(R 7 )2. In some embodiments, R 3 Select from -OR 7 -and-OC(O)R 7 In some embodiments, R 4 Selected from 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 Select from -OR 7 、-SR 7 、-OC(O)R 7 and -N(R 7 )2. In some embodiments, R 4 Select from -OR 7 -and-OC(O)R 7 In some embodiments, R 5 Selected from -OC(O)R 7 、-OC(O)N(R 7 )2、-N(R7 )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 Selected from -OC(O)R 7 and -N(R 7 )C(O)R 7 In some embodiments, each R 7 are independently selected from: hydrogen; and C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 In some embodiments, each R 7 are independently selected from C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2 and -NH(C 1-6 In some embodiments, each R 7 are independently selected from C optionally substituted by one or more substituents 1-6 alkyl, the one or more substituents being independently selected from halogen, -CN, -OH, and -SH. In some embodiments, 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 optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, and C 1-3 Alkyl; R 1 Selected from -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 2is a C1 alkyl substituted by -OH or -OC(O)CH3;

[0037] R 3 is -OH or -OC(O)CH3; R 4 is -OH or -OC(O)CH3; and R 5 is -NH(O)CH3. In some embodiments, the compound comprises:

[0038]

[0039]

[0040]

[0041]

[0042]

[0043] In some embodiments, the oligonucleotide (J) is attached at the 5' end or 3' end of the oligonucleotide. In some embodiments, the oligonucleotide includes DNA. In some embodiments, the oligonucleotide includes RNA. In some embodiments, the oligonucleotide includes one or more modified internucleoside linkages. In some embodiments, one or more modified internucleoside linkages include alkylphosphonates, phosphorothioates, methylphosphonates, phosphorodithioates, alkylthiophosphonates, phosphoramidates, carbamates, carbonates, phosphotriesters, acetimidates or carboxymethyl esters, or a combination thereof. In some embodiments, the oligonucleotide includes 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 includes one or more modified nucleosides. In some embodiments, one or more modified nucleosides include locked nucleic acids (LNA), hexitol nucleic acids (HLA), cyclohexene nucleic acids (CeNA), 2'-methoxyethyl, 2'-O-alkyl, 2'-O-allyl, 2'-O-allyl, 2'-fluoro or 2'-deoxy, or a combination thereof. In some embodiments, one or more modified nucleosides include 2',4'-constrained ethyl nucleosides, 2'-O-methyl nucleosides, 2'-deoxyfluoro nucleosides, 2'-ON-methylacetylamino (2'-O-NMA) nucleosides, 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' end or 5' end of the oligonucleotide. In some embodiments, the lipid comprises cholesterol, myristoyl, palmitoyl, stearoyl, lithocholyl, behenoyl, docosahexenoyl, myristyl, palmityl, stearyl or alpha-tocopherol, or a combination thereof. In some embodiments, the oligonucleotide comprises an arginine-glycine-aspartic acid (RGD) peptide attached at the 3' end or 5' end of the oligonucleotide. In some embodiments, the RGD peptide comprises cyclic (-Arg-Gly-Asp-D-Phe-Cys), cyclic (-Arg-Gly-Asp-D-Phe-Lys), cyclic (-Arg-Gly-Asp-D-Phe-azido), aminobenzoic acid-derivatized RGD, 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 double helix. In some embodiments, the first base pair of the double-stranded RNA double helix 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 1, 2 or more nucleosides. In some embodiments, the sense strand comprises any one of the modification patterns 1S to 6S or 1S#2 to 6S#2. In some embodiments, the antisense strand comprises any one of the modification patterns 1AS to 9AS. In some embodiments, the oligonucleotide comprises an antisense oligonucleotide (ASO). In some embodiments, the ASO has a length of 12-30 nucleosides. In some embodiments, the ASO comprises the modification pattern ASO1. In some embodiments, the compound binds to the asialoglycoprotein receptor. In some embodiments, the compound targets hepatocytes. Disclosed herein is a pharmaceutical composition comprising any one of the compounds described herein and a pharmaceutically acceptable carrier, excipient or diluent. In some embodiments, the pharmaceutical composition is sterile. In some embodiments, the pharmaceutical composition comprises a pharmaceutically acceptable carrier. In some embodiments, the pharmaceutically acceptable carrier comprises water, a buffer, or saline. In some embodiments, the oligonucleotide targets a target mRNA and, when administered to a subject in an effective amount, reduces the target mRNA or target protein by at least 10%.

[0044] Disclosed herein is a method for reducing a target mRNA or target protein of an object in need, the method comprising administering an effective amount of a pharmaceutical composition of any compound described herein. In some embodiments, the effective amount reduces the measured value of the target mRNA or target protein of the object relative to the baseline target mRNA or target protein measured value. In some embodiments, the effective amount treats the disease of the object. In some embodiments, the effective amount reduces the measured value of the symptom or parameter associated with the disease of the object relative to the baseline symptom or parameter measured value. In some embodiments, the measured value of the symptom or parameter associated with the disease of the object is reduced by at least 10 days. In some embodiments, the measured value of the symptom or parameter associated with the disease of the object is reduced by at least 100 days. In some embodiments, the disease is a metabolic disease. In some embodiments, the disease is a liver disease.

[0045] Disclosed herein is a compound represented by formula (A) or (B):

[0046]

[0047] or a salt thereof, wherein

[0048] Each w is independently selected from any value from 1 to 20;

[0049] Each v is independently selected from any value from 1 to 20;

[0050] n is selected from any value from 1 to 20;

[0051] m is selected from any value from 1 to 20;

[0052] z is selected from any value from 1 to 3, wherein

[0053] If z is 3, then Y is C

[0054] If z is 2, then Y is CR 6 ,or

[0055] If z is 1, then Y is C(R 6 )2;

[0056] Q is selected from:

[0057] C optionally substituted by one or more substituents 3-10 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 Alkyl, where C 1-6 The alkyl group is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2;

[0058] R 1 Selected from:

[0059] -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)2R 7 、-OS(O)2R 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;

[0060] Each R 2 Independently selected from:

[0061] C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from halogen, -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 ;

[0062] R 3 and R 4 Each independently selected from:

[0063] -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 ;

[0064] Each R 5 Independently selected from:

[0065] -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;

[0066] Each R 6 Independently selected from:

[0067] hydrogen;

[0068] Halogen, -CN, -NO2, -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 ;as well as

[0069] C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 ;

[0070] Each R 7 Independently selected from:

[0071] hydrogen;

[0072] C 1-6 Alkyl, C 2-6 Alkenyl and C 2-6 Alkynyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 Carbocyclic rings and 3- to 10-membered heterocyclic rings; and

[0073] C 3-10 carbocyclic and 3- to 10-membered heterocyclic rings, each of which is optionally substituted by one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -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 ring, 3- to 10-membered heterocyclic ring and C 1-6 Halogenated alkyl.

[0074] 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 C optionally substituted with one or more substituents. 5-6 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 and-S(O)R 7 In some embodiments, Q is selected from C optionally substituted with one or more substituents. 5-6 In some embodiments, Q is selected from phenyl and cyclohexyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2. In some embodiments, Q is selected from phenyl. In some embodiments, Q is selected from cyclohexyl. In some embodiments, R 1 Selected from -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 Selected from -OP(O)(OR 7 )2 and -OP(OR 7 )N(R 7 )2. In some embodiments, R 1 is selected from -OP(O)(OCH2CH3)OH and -OP(OCH2CH2CN)N(CH(CH3)2)2. In some embodiments, R 1 is -OP(OCH2CH2CN)N(CH(CH3)2)2. In some embodiments, R 2 is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from halogen, -OR 7 、-OC(O)R 7 、-SR 7 、-N(R 7 )2. -C(O)R 7 and-S(O)R 7 In some embodiments, R 2 is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from -OR 7 、-OC(O)R 7 、-SR 7 and -N(R 7 )2. In some embodiments, R 2is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from -OR 7 and -OC(O)R 7 In some embodiments, R 3 Selected from 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 Select from -OR 7 、-SR 7 、-OC(O)R 7 and -N(R 7 )2. In some embodiments, R 3 Select from -OR 7 and -OC(O)R 7 In some embodiments, R 4 Selected from 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 Select from -OR 7 、-SR 7 、-OC(O)R 7 and -N(R 7 )2. In some embodiments, R 4 Select from -OR 7 and -OC(O)R 7 In some embodiments, R 5 Selected from -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 Selected from -OC(O)R 7 and -N(R 7 )C(O)R 7 In some embodiments, each R 7are independently selected from: hydrogen; and C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 In some embodiments, each R 7 are independently selected from C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2 and -NH(C 1-6 In some embodiments, each R 7 are independently selected from C optionally substituted by one or more substituents 1-6 alkyl, the one or more substituents being independently selected from halogen, -CN, -OH and -SH. In some embodiments, 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 optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2 and C 1-3 Alkyl; R 1 Selected from -OP(O)(OR 7 )2 and -OP(OR 7 )N(R 7 )2; R 2 is a C1 alkyl group substituted by -OH or -OC(O)CH3; R 3 is -OH or -OC(O)CH3; R 4 is -OH or -OC(O)CH3; and R 5 is -NH(O)CH3. In some embodiments, the compound comprises:

[0075]

[0076]

[0077]

[0078]

[0079] DETAILED DESCRIPTION

[0080] 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 present on hepatocytes. Therefore, GalNAc can target oligonucleotides to hepatocytes or the liver.

[0081] Provided herein are GalNAc moieties. These GalNAc moieties can be conjugated to oligonucleotides such as small interfering RNA (siRNA) or antisense oligonucleotides (ASO). Oligonucleotides conjugated to the GalNAc moiety can be administered to a subject, targeting the liver or hepatocytes, or used to treat a subject's liver-related disorders.

[0082] I. Composition

[0083] In some embodiments, provided herein are compositions comprising oligonucleotides and N-acetylgalactosamine (GalNAc) moieties. In some embodiments, compositions comprise oligonucleotides. The oligonucleotides can inhibit target genes or oligonucleotides. The oligonucleotides can bind to target oligonucleotides. In some embodiments, compositions are used in methods described herein.

[0084] In some embodiments, provided herein is a compound comprising an oligonucleotide and a GalNAc moiety. In some embodiments, the compound comprises an oligonucleotide. The oligonucleotide can be combined with a target oligonucleotide. In some embodiments, the compound is used in the methods described herein. In some embodiments, the compound is included in the compositions described herein.

[0085] The oligonucleotides of the compounds or compositions described herein may comprise small interfering RNA (siRNA) or antisense oligonucleotides (ASO).

[0086] Some embodiments include a composition comprising a GalNAc moiety and an oligonucleotide, which reduces the target mRNA or protein level in the cell, liquid or tissue of the object when applied to the object in an effective amount. Some embodiments include a composition comprising a GalNAc moiety and an oligonucleotide, which reduces the target mRNA or protein level in liver tissue or hepatocytes when applied to the object in an effective amount. In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide, which reduces the level of the target (e.g., mRNA) in the cell or tissue when applied to the object in an effective amount. In some embodiments, the cell is a hepatocyte. In some embodiments, the tissue is liver tissue. Some embodiments include a composition comprising a GalNAc moiety and an oligonucleotide, which reduces the target mRNA level in liver tissue when applied to the object in an effective amount. Some embodiments include a composition comprising a GalNAc moiety and an oligonucleotide, which reduces the target mRNA level in hepatocytes when applied to the object in an effective amount.

[0087] In some embodiments, the reduction in target oligonucleotide levels is specific to hepatocytes relative to other cell types. In some embodiments, the reduction in target RNA levels is specific to hepatocytes relative to other cell types. In some embodiments, the reduction in target mRNA levels is specific to hepatocytes relative to other cell types. In some embodiments, the reduction in target protein levels is specific to hepatocytes relative to other cell types. In some embodiments, the reduction in target oligonucleotide levels is specific to liver tissue relative to other cell types. In some embodiments, the reduction in target RNA levels is specific to the liver relative to other cell types. In some embodiments, the reduction in target mRNA levels is specific to the liver relative to other cell types. In some embodiments, the reduction in target protein levels is specific to the liver relative to other cell types.

[0088] In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide bound to a target oligonucleotide, which reduces the level of the target oligonucleotide in a cell or tissue when applied to a subject in an effective amount. 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 no more than about 2.5%, no more than about 5% or no more than about 7.5% compared to before administration. In some embodiments, the target oligonucleotide level is reduced by no more than about 10% compared to before administration. In some embodiments, the target oligonucleotide level is reduced by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100% 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 the range of reduction is defined by any two of the above percentages.

[0089] In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide bound to a target mRNA, which reduces the level of target mRNA in a cell or tissue when applied to a subject in an effective amount. 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 no more than about 2.5%, no more than about 5% or no more than about 7.5% compared to before administration. In some embodiments, the target mRNA level is reduced by no more than about 10% compared to before administration. In some embodiments, the target mRNA level is no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90% or no more than about 100% 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 the range of reduction is defined by any two of the above percentages.

[0090] In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide bound to an oligonucleotide encoding a target protein, and the composition reduces the level of the target protein in a cell or tissue when 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 no more than about 2.5%, no more than about 5% or no more than about 7.5% compared to before administration. In some embodiments, the target protein level is reduced by no more than about 10% compared to before administration. In some embodiments, the target protein level is reduced by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100% 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 the range of reduction is defined by any two of the above percentages.

[0091] In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide bound to a target oligonucleotide (e.g., mRNA) that reduces an adverse phenotype (e.g., a disease symptom associated with a target oligonucleotide) when administered to a subject in an effective amount. In some embodiments, the adverse 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 adverse phenotype is reduced by about 10% or more compared to before administration. In some embodiments, the adverse 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 adverse phenotype is reduced by no more than about 2.5%, no more than about 5%, or no more than about 7.5% compared to before administration. In some embodiments, the adverse phenotype is reduced by no more than about 10% compared to before administration. In some embodiments, the adverse phenotype is reduced by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100% compared to before administration. In some embodiments, the adverse 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 the range of reduction is defined by any two of the above percentages.

[0092] In some embodiments, the composition comprises a GalNAc moiety and an oligonucleotide that binds to a target oligonucleotide (e.g., mRNA) that increases a protective phenotype (e.g., protection against a condition) 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 no more than about 2.5%, no more than about 5%, or no more than about 7.5% compared to before administration. In some embodiments, the protective phenotype is increased by no more than about 10% compared to before administration. In some embodiments, the protective phenotype is increased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100% compared to before administration. In some embodiments, the protective phenotype is increased by no more than about 200%, no more than about 300%, no more than about 400%, no more than about 500%, no more than about 600%, no more than about 700%, no more than about 800%, no more than about 900%, or no more than about 1000% compared to before administration. In some embodiments, the protective phenotype increases 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 the range of increase is defined by any two of the above percentages.

[0093] A. GalNAc Moieties and Compounds

[0094] In some embodiments, compositions comprising a GalNAc moiety are provided herein. In some embodiments, the present scheme also provides compositions comprising a GalNAc moiety and an oligonucleotide. In some embodiments, the compositions comprising a GalNAc moiety and an oligonucleotide are described by compounds of formula (I) or formula (II). In some embodiments, the oligonucleotide in formula (I) or (II) is J. In some embodiments, the GalNAc moiety in formula (I) or formula (II) is a molecular moiety that is bound to J. The oligonucleotide may comprise a small interfering RNA (siRNA) or an antisense oligonucleotide (ASO).

[0095] In some embodiments, provided herein are compounds represented by formula (I) or (II):

[0096] or a salt thereof, wherein

[0097] J is an oligonucleotide;

[0098] Each w is independently selected from any value from 1 to 20;

[0099] Each v is independently selected from any value from 1 to 20;

[0100] n is selected from any value from 1 to 20;

[0101] m is selected from any value from 1 to 20;

[0102] z is selected from any value from 1 to 3, wherein

[0103] If z is 3, then Y is C;

[0104] If z is 2, then Y is CR 6 ,or

[0105] If z is 1, then Y is C(R 6 )2;

[0106] Q is selected from:

[0107] C optionally substituted by one or more substituents 3-10 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 Alkyl, where C 1-6 The alkyl group is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2;

[0108] R 1 is a linker selected from the group consisting of:

[0109] -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 )2NR 7 -;

[0110] Each R2 Independently selected from:

[0111] C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from 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 ;

[0112] R 3 and R 4 Each independently selected from:

[0113] -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 ;

[0114] Each R 5 Independently selected from:

[0115] -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;

[0116] Each R 6 Independently selected from:

[0117] hydrogen;

[0118] Halogen, -CN, -NO2, -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 ;as well as

[0119] C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 ;

[0120] Each R 7 Independently selected from:

[0121] hydrogen;

[0122] C1-6 Alkyl, C 2-6 Alkenyl and C 2-6 Alkynyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 Carbocyclic rings and 3- to 10-membered heterocyclic rings; and

[0123] C 3-10 carbocyclic and 3- to 10-membered heterocyclic rings, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -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 ring, 3- to 10-membered heterocyclic ring and C 1-6 Halogenated alkyl.

[0124] 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.

[0125] 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.

[0126] In some embodiments, n is selected from any value from 1 to 20. In some embodiments, n is selected from any value from 1 to 15. 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 9. In some embodiments, n is selected from any value from 1 to 8. In some embodiments, n is selected from any value from 1 to 7. In some embodiments, n is selected from any value from 1 to 6. In some embodiments, n is selected from any value from 1 to 5. In some embodiments, n is selected from any value from 1 to 4. In some embodiments, n is selected from any value from 2 to 4. In some embodiments, n is selected from any value from 1 to 3. In some embodiments, n is 2 or 3. In some embodiments, n is 3. In some embodiments, n is selected from any value from 1 to 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.

[0127] In some embodiments, m is selected from any value from 1 to 20. In some embodiments, m is selected from any value from 1 to 15. 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 9. In some embodiments, m is selected from any value from 1 to 8. In some embodiments, m is selected from any value from 1 to 7. In some embodiments, m is selected from any value from 3 to 7. In some embodiments, m is selected from any value from 1 to 6. In some embodiments, m is selected from any value from 2 to 6. In some embodiments, m is selected from any value from 3 to 6. In some embodiments, m is selected from any value from 4 to 6. In some embodiments, m is 6. In some embodiments, m is selected from any value from 1 to 5. In some embodiments, m is selected from any value from 3 to 5. In some embodiments, m is 5. In some embodiments, m is 4 or 5. In some embodiments, m is selected from any value from 1 to 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.

[0128] 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(R 6 )2.

[0129] In some embodiments, Q is selected from C optionally substituted with one or more substituents. 3-10 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 Alkyl, where C 1-6 The alkyl group is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2. In some embodiments, Q is selected from C 3-6 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 Alkyl, where C 1-6 The alkyl group is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2. In some embodiments, Q is selected from C 5-6 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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, Q is selected from a C6 carbocycle optionally substituted with one or more substituents independently selected from halogen, -CN, -NO2, -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, Q is selected from a C5 carbocycle optionally substituted with one or more substituents independently selected from halogen, -CN, -NO2, -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, Q is selected from C optionally substituted with one or more substituents. 5-6In some embodiments, Q is selected from phenyl, cyclohexyl, cyclopentadiene and cyclopentyl, each of which is optionally substituted by one or more substituents, and the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2. In some embodiments, Q is selected from phenyl and cyclohexyl, each of which is optionally substituted by one or more substituents, and the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2. In some embodiments, Q is selected from phenyl and cyclohexyl. In some embodiments, Q is phenyl. In some embodiments, Q is cyclohexyl.

[0130] In some embodiments, R 1 is a linker selected from the group consisting of: -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 )2NR 7 --. In some embodiments, R 1 is a linker selected from the group consisting of: -O-, -S-, -N(R 7 )-、-C(O)-、-C(O)N(R 7 )-、-N(R 7 )C(O)-、-N(R7 )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 )2NR 7 -. In some embodiments, R 1 is selected from -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 )2NR 7 In some embodiments, R 1 Selected from -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 )O-. In some embodiments, R 1 Selected from -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-. In some embodiments, R 1 Selected from -OP(O)(OR 7 )O- and -OP(OR 7 )O-. In some embodiments, R 1 is a linker selected from the group consisting of: -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, R 1 Selected from -OP(O)(OR 7 )O-、-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 )2NR 7 -. In some embodiments, R 1 Selected from -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 - )S-. In some embodiments, R 1 Selected from -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 )S-. In some embodiments, R 1 Select from -OP(S)(OR 7 )O-、-OP(O)(SR 7 )O- and -OP(O)(OR 7 )S-. In some embodiments, R 1 Select from -OP(S)(OR 7 )O-、-OP(O)(SR 7 )O-、-OP(S)(O - )O- and -OP(O)(S - )O-. In some embodiments, R 1 Select from -OP(S)(OR 7 )O-and-OP(O)(SR 7 )O-. In some embodiments, R 1 Selected from -OP(O)(OR 7 )O-、-OP(OR 7 )N(R 7 )- and -OPN(R 7 )2O-. In some embodiments, R 1is -OP(O)(OH)O-, -OP(O)(OCH2CH3)O-, -OP(OCH2CH2CN)N(CH(CH3)2)-, or -OPN(CH(CH3)2)2O-. In some embodiments, R 1 Selected from -OP(O)(OH)O- and OP(O)(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 Yes -S-.

[0131] In some embodiments, each R 2 are independently selected from C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from 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 In some embodiments, each R 2 are independently selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from halogen, -OR 7 、-OC(O)R 7 、-SR 7 、-N(R 7 )2. -C(O)R 7 and-S(O)R 7 In some embodiments, each R 2 are independently selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from -OR7 、-OC(O)R 7 、-SR 7 and -N(R 7 )2. In some embodiments, each R 2 independently selected from C1 alkyl substituted by one or more substituents, the one or more substituents being independently selected from -OR 7 and -OC(O)R 7 In some embodiments, each R 2 Independently selected from -CH2OH and -CH2OC(O)CH3.

[0132] In some embodiments, each R 3 Independently selected from -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, each R 3 are independently selected from 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, each R 3 Independently selected from -OR 7 、-OC(O)R 7 、-SR 7 and -N(R 7 )2. In some embodiments, each R 3 Independently selected from -OR 7 and -OC(O)R 7 In some embodiments, R 3 Independently selected from -OH and -OC(O)CH3.

[0133] In some embodiments, each R 4 Independently selected from -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 In some embodiments, each R 4 are independently selected from 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, each R 4 Independently selected from -OR 7 、-OC(O)R 7 、-SR 7 and -N(R 7 )2. In some embodiments, each R 4 Independently selected from -OR 7 and -OC(O)R 7 In some embodiments, R 4 Independently selected from -OH and -OC(O)CH3.

[0134] In some embodiments, each R 5 Independently selected from -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, each R 5 Selected from -OC(O)R 7 、-OC(O)N(R7 )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, each R 5 Independently selected from -OC(O)R 7 and -N(R 7 )C(O)R 7 In some embodiments, each R 5 Independently selected from -N(H)C(O)CH3.

[0135] In some embodiments, each R 6 are independently selected from 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 C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from 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 In some embodiments, each R6 are independently selected from hydrogen, halogen, -CN, -OR 7 、-SR 7 、-N(R 7 )2 and C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from halogen, -OR 7 、-SR 7 and -N(R 7 )2. In some embodiments, each R 6 are independently selected from hydrogen, halogen, -CN, -OH, -SH and -NH2.

[0136] In some embodiments, each R 7 Independently selected from: hydrogen, C 1-6 Alkyl, C 2-6 Alkenyl and C 2-6 Alkynyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 Carbocyclic and 3- to 10-membered heterocyclic rings; and C 3-10 carbocyclic and 3- to 10-membered heterocyclic rings, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -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, 3- to 10-membered heterocyclic and C 1-6 In some embodiments, each R 7 are independently selected from: hydrogen; and C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C3-10 In some embodiments, each R 7 are independently selected from C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2 and -NH(C 1-6 In some embodiments, R 7 are independently selected from C optionally substituted by one or more substituents 1-6 alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH and -SH. 7 In some embodiments, each R 7 are independently selected from C optionally substituted by one or more substituents 1-3 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2 and -NH(C 1-6 alkyl).

[0137] In some embodiments, 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 optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2 and C 1-3 Alkyl; R 1 Selected from -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 a C1 alkyl group substituted by -OH or -OC(O)CH3; R 3 is -OH or -OC(O)CH3; R 4 is -OH or -OC(O)CH3; and R 5 It is -NH(O)CH3.

[0138] In some embodiments, 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 optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2 and C 1-3 Alkyl; R 1 Selected from -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 a C1 alkyl group substituted by -OH or -OC(O)CH3; R 3 is -OH or -OC(O)CH3; R 4 is -OH or -OC(O)CH3; and R 5 is -NH(O)CH3. In some embodiments, 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 optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2 and C 1-3 Alkyl; R 1 Selected from -OP(O)(OH)O-, -OP(S)(OH)O- and -OP(OH)O-; R 2 is a C1 alkyl group substituted by -OH or -OC(O)CH3; R 3 is -OH or -OC(O)CH3; R 4 is -OH or -OC(O)CH3; and R 5 It is -NH(O)CH3.

[0139] In some embodiments, the compound of formula (I) is selected from:

[0140]

[0141]

[0142]

[0143]

[0144]

[0145]

[0146] In some embodiments, the compound of formula (II) is selected from:

[0147]

[0148]

[0149]

[0150]

[0151]

[0152] 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 hepatocyte receptor. In some embodiments, the compound binds to a hepatocyte receptor. In some embodiments, the compound targets a hepatocyte.

[0153] In embodiments provided herein, the compositions described herein comprise a GalNAc compound. In some embodiments, the GalNAc compound describes a compound of formula (A) or formula (B):

[0154] or a salt thereof, wherein:

[0155] Each w is independently selected from any value from 1 to 20;

[0156] Each v is independently selected from any value from 1 to 20;

[0157] n is selected from any value from 1 to 20;

[0158] m is selected from any value from 1 to 20;

[0159] z is selected from any value from 1 to 3, wherein

[0160] If z is 3, then Y is C;

[0161] If z is 2, then Y is CR 6 ,or

[0162] If z is 1, then Y is C(R 6 )2;

[0163] Q is selected from:

[0164] C optionally substituted by one or more substituents 3-10 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -OR 7 、-SR 7 、-N(R7 )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 Alkyl, where C 1-6 The alkyl group is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2;

[0165] R 1 Selected from:

[0166] -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)2R 7 、-OS(O)2R 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;

[0167] Each R 2 Independently selected from:

[0168] C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from 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 ;

[0169] R 3 and R 4 Each independently selected from:

[0170] -OR 7 、-SR 7 、-N(R 7 )2. -C(O)R 7 、-C(O)N(R 7 )2、-N(R7 )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 ;

[0171] Each R 5 Independently selected from:

[0172] -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;

[0173] Each R 6 Independently selected from:

[0174] hydrogen;

[0175] Halogen, -CN, -NO2, -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 ;as well as

[0176] C optionally substituted by one or more substituents 1-6Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 ;

[0177] Each R 7 Independently selected from:

[0178] hydrogen;

[0179] C 1-6 Alkyl, C 2-6 Alkenyl and C 2-6 alkynyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN,

[0180] -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 alkyl,

[0181] -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 Carbocyclic rings and 3- to 10-membered heterocyclic rings; and

[0182] C 3-10 carbocyclic ring and 3- to 10-membered heterocyclic ring, each of which is optionally substituted by one or more substituents independently selected from halogen, -CN,

[0183] -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 alkyl,

[0184] -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 1-6 Alkyl, C 2-6Alkenyl, C 2-6 Alkynyl, C 3-10 Carbocyclic ring, 3- to 10-membered heterocyclic ring and C 1-6 Halogenated alkyl.

[0185] 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.

[0186] 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 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.

[0187] In some embodiments, n is selected from any value from 1 to 20. In some embodiments, n is selected from any value from 1 to 15. 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 9. In some embodiments, n is selected from any value from 1 to 8. In some embodiments, n is selected from any value from 1 to 7. In some embodiments, n is selected from any value from 1 to 6. In some embodiments, n is selected from any value from 1 to 5. In some embodiments, n is selected from any value from 1 to 4. In some embodiments, n is selected from any value from 2 to 4. In some embodiments, n is selected from any value from 1 to 3. In some embodiments, n is 2 or 3. In some embodiments, n is 3. In some embodiments, n is selected from any value from 1 to 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.

[0188] In some embodiments, m is selected from any value from 1 to 20. In some embodiments, m is selected from any value from 1 to 15. 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 9. In some embodiments, m is selected from any value from 1 to 8. In some embodiments, m is selected from any value from 1 to 7. In some embodiments, m is selected from any value from 3 to 7. In some embodiments, m is selected from any value from 1 to 6. In some embodiments, m is selected from any value from 2 to 6. In some embodiments, m is selected from any value from 3 to 6. In some embodiments, m is selected from any value from 4 to 6. In some embodiments, m is 6. In some embodiments, m is selected from any value from 1 to 5. In some embodiments, m is selected from any value from 3 to 5. In some embodiments, m is 5. In some embodiments, m is 4 or 5. In some embodiments, m is selected from any value from 1 to 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.

[0189] 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 6In some embodiments, z is 1 and Y is C(R 6 )2.

[0190] In some embodiments, Q is selected from C optionally substituted with one or more substituents. 3-10 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 Alkyl, where C 1-6 The alkyl group is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2. In some embodiments, Q is selected from C 3-6 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 Alkyl, where C 1-6The alkyl group is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2. In some embodiments, Q is selected from C 5-6 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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, Q is selected from a C6 carbocycle optionally substituted with one or more substituents independently selected from halogen, -CN, -NO2, -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, Q is selected from a C5 carbocycle optionally substituted with one or more substituents independently selected from halogen, -CN, -NO2, -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, Q is selected from C optionally substituted with one or more substituents. 5-6 In some embodiments, Q is selected from phenyl, cyclohexyl, cyclopentadiene and cyclopentyl, each of which is optionally substituted by one or more substituents, and the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2. In some embodiments, Q is selected from phenyl and cyclohexyl, each of which is optionally substituted by one or more substituents, and the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2. In some embodiments, Q is selected from phenyl and cyclohexyl. In some embodiments, Q is phenyl. In some embodiments, Q is cyclohexyl.

[0191] In some embodiments, R 1 Select from -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)2R 7 、-OS(O)2R 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 is selected from -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 is selected from -SR 7 , -S(O)R 7 , -S(O)2R7 、 -OS(O)2R 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 is selected from -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 Selected from -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 Selected from -OP(O)(OR 7 )2 and -OP(OR 7 )N(R 7 )2. In some embodiments, R 1 is selected from -OP(O)(OCH2CH3)OH and -OP(OCH2CH2CN)N(CH(CH3)2)2. In some embodiments, R 1 It is -OP(OCH2CH2CN)N(CH(CH3)2)2.

[0192] In some embodiments, each R 2 are independently selected from C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from 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 In some embodiments, each R 2 are independently selected from C substituted by one or more substituents 1-3Alkyl, the one or more substituents are independently selected from halogen, -OR 7 、-OC(O)R 7 、-SR 7 、-N(R 7 )2. -C(O)R 7 and-S(O)R 7 In some embodiments, each R 2 are independently selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from -OR 7 、-OC(O)R 7 、-SR 7 and -N(R 7 )2. In some embodiments, each R 2 independently selected from C1 alkyl substituted by one or more substituents, the one or more substituents being independently selected from -OR 7 and -OC(O)R 7 In some embodiments, each R 2 Independently selected from -CH2OH and -CH2OC(O)CH3.

[0193] In some embodiments, each R 3 Independently selected from -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, each R 3 are independently selected from 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, each R 3 Independently selected from -OR 7 、-OC(O)R7 、-SR 7 and -N(R 7 )2. In some embodiments, each R 3 Independently selected from -OR 7 and -OC(O)R 7 In some embodiments, R 3 Independently selected from -OH and -OC(O)CH3.

[0194] In some embodiments, each R 4 Independently selected from -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, each R 4 are independently selected from 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, each R 4 Independently selected from -OR 7 、-OC(O)R 7 、-SR 7 and -N(R 7 )2. In some embodiments, each R 4 Independently selected from -OR 7 and -OC(O)R 7 In some embodiments, R 4 Independently selected from -OH and -OC(O)CH3.

[0195] In some embodiments, each R 5 Independently selected from -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, each R 5 Selected from -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, each R 5 Independently selected from -OC(O)R 7 and -N(R 7 )C(O)R 7 In some embodiments, each R 5 Independently selected from -N(H)C(O)CH3.

[0196] In some embodiments, each R 6 are independently selected from 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 C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from 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 In some embodiments, each R 6 are independently selected from hydrogen, halogen, -CN-OR 7 、-SR 7 、-N(R 7 )2 and C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from halogen, -OR 7 、-SR 7 and -N(R 7 )2. In some embodiments, each R 6 are independently selected from hydrogen, halogen, -CN, -OH, -SH and -NH2.

[0197] In some embodiments, each R 7 Independently selected from: hydrogen; C 1-6 Alkyl, C 2-6 Alkenyl and C 2-6 Alkynyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 Carbocyclic and 3- to 10-membered heterocyclic rings; and C 3-10 carbocyclic and 3 to 10 membered heterocyclic rings, each of which is optionally substituted by one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -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 ring, 3- to 10-membered heterocyclic ring and C1-6 In some embodiments, each R 7 are independently selected from: hydrogen; and C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 In some embodiments, each R 7 are independently selected from C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2 and -NH(C 1-6 In some embodiments, each R 7 In some embodiments, each R 7 are independently selected from C optionally substituted by one or more substituents 1-3 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2 and -NH(C 1-6 In some embodiments, each R 7 are independently selected from C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from halogen, -CN, -OH and -SH.

[0198] In some embodiments, the compound of formula (A) is selected from:

[0199]

[0200]

[0201]

[0202] In some embodiments, provided herein is a compound represented by formula (B):

[0203]

[0204]

[0205]

[0206] In some embodiments, the phosphate is deprotonated to form a salt of Formula (I), (II), (A), or (B). 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 includes Na + In some embodiments, the metal cation comprises K + In some embodiments, the metal cation includes Mg 2+ In some embodiments, the metal cation includes Ca 2+ In some embodiments, the cation is an organic or inorganic small molecule. In some embodiments where the compound is a deprotonated form of Formula (I) or (II), the cation is a positively charged nucleic acid present in an oligonucleotide.

[0207] Some embodiments include the following, wherein J is an oligonucleotide: J may include one or more additional phosphates or one or more phosphorothioates connected to the oligonucleotide. J may include one or more additional phosphates connected to the oligonucleotide. J may include one or more phosphorothioates connected to the oligonucleotide.

[0208] Some embodiments include the following, wherein J is an oligonucleotide:

[0209] J may include one or more additional phosphates or one or more phosphorothioates connected to the oligonucleotide. J may include one or more additional phosphates connected to the oligonucleotide. J may include one or more phosphorothioates connected to the oligonucleotide.

[0210] Some embodiments include the following, wherein J is an oligonucleotide: J may include one or more phosphates or thiophosphates that are connected to an oligonucleotide. J may include one or more phosphates that are connected to an oligonucleotide. J may include a phosphate that is connected to an oligonucleotide. J may include one or more thiophosphates that are connected to an oligonucleotide. J may include a thiophosphate that is connected to an oligonucleotide.

[0211] Some embodiments include the following, wherein J is an oligonucleotide: The structure attached to the oligonucleotide (J) in this compound is an example of a GalNAc portion. J may include one or more phosphates or thiophosphates connected to the oligonucleotide. J may include one or more phosphates connected to the oligonucleotide. J may include a phosphate connected to the oligonucleotide. J may include one or more thiophosphates connected to the oligonucleotide. J may include a thiophosphate connected to the oligonucleotide.

[0212] Some embodiments include the following, wherein J is an oligonucleotide: J may include one or more additional phosphates or one or more phosphorothioates connected to the oligonucleotide. J may include one or more additional phosphates connected to the oligonucleotide. J may include one or more phosphorothioates connected to the oligonucleotide.

[0213] Some embodiments include the following, wherein J is an oligonucleotide:

[0214]

[0215] J may include one or more additional phosphates or one or more phosphorothioates connected to the oligonucleotide. J may include one or more additional phosphates connected to the oligonucleotide. J may include one or more phosphorothioates connected to the oligonucleotide.

[0216] Some embodiments include the following, wherein J is an oligonucleotide: J may include one or more phosphates or thiophosphates that are connected to an oligonucleotide. J may include one or more phosphates that are connected to an oligonucleotide. J may include a phosphate that is connected to an oligonucleotide. J may include one or more thiophosphates that are connected to an oligonucleotide. J may include a thiophosphate that is connected to an oligonucleotide.

[0217] Some embodiments include the following, wherein J is an oligonucleotide:

[0218]

[0219] The structure of the compound attached to the oligonucleotide (J) can be referred to as "ETL17", which is an example of a GalNAc portion. J can include one or more phosphates or phosphorothioates attached to the oligonucleotide. J can include one or more phosphates attached to the oligonucleotide. J can include a phosphate attached to the oligonucleotide. J can include one or more phosphorothioates attached to the oligonucleotide. J can include a phosphorothioate attached to the oligonucleotide.

[0220] Some embodiments include the following, wherein J is an oligonucleotide:

[0221]

[0222] The structure attached to the oligonucleotide (J) in this compound is an example of a GalNAc part. J can include one or more phosphates or thiophosphates connected to the oligonucleotide. J can include one or more phosphates connected to the oligonucleotide. J can include a phosphate connected to the oligonucleotide. J can include one or more thiophosphates connected to the oligonucleotide. J can include a thiophosphate connected to the oligonucleotide. In any embodiment, J can include an additional joint.

[0223] 1. Analogs

[0224] Chemical entities with carbon-carbon double bonds or carbon-nitrogen double bonds can exist in Z- or E-form (or cis- or trans-form). In addition, some chemical entities can exist in different tautomeric forms. Unless otherwise stated, the compounds described herein also include all Z-, E- and tautomeric forms.

[0225] "Tautomer" refers to a molecule or moiety in which a proton may be transferred from one atom of a molecule to another atom in the same molecule. The compounds presented herein exist in some embodiments as tautomers. Where tautomerism is possible, a chemical equilibrium of the tautomers will exist. The exact ratio of the tautomers depends on a number of factors, including physical state, temperature, solvent, and pH. Some examples of tautomeric equilibrium include:

[0226]

[0227] The compounds and moieties disclosed herein, in some embodiments, are in different isotopically enriched forms, e.g. 2 H. 3 H. 11 C. 13 C and / or 14 C. In a specific embodiment, the compound is deuterated at at least one position. These deuterated forms can be prepared by the procedures described in U.S. Pat. Nos. 5,846,514 and 6,334,997. As described in U.S. Pat. Nos. 5,846,514 and 6,334,997, deuteration can increase metabolic stability and / or potency, thereby extending the duration of action of the drug.

[0228] Unless otherwise indicated, 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, a given structure may have a hydrogen replaced by a deuterium or tritium, or a carbon replaced by a 13 C or 14 C-enriched carbon-substituted compounds all fall within the scope of the present disclosure.

[0229] The compounds and moieties of the present disclosure optionally contain unnatural ratios of atomic isotopes at one or more atoms that constitute the compound. For example, the compound may be deuterated with isotopes such as deuterium ( 2 H), tritium ( 3 H), iodine-125 ( 125 I) or carbon-14 ( 14 C) Marking. 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 Isotopic substitutions of 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.

[0230] In some embodiments, some or all of the disclosed compounds and moieties 1 H atoms are 2 H atom substitution. Methods for synthesizing deuterium-containing compounds are known in the art and include, by way of non-limiting example only, the following synthetic methods.

[0231] 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, 110 pp; 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.

[0232] Deuterated starting materials are readily available and processed by the synthetic methods described herein to provide the synthesis of deuterated compounds. A large number of deuterated reagents and building blocks are commercially available through chemical suppliers such as Aldrich Chemical Co.

[0233] The disclosure herein includes salts of the compounds described herein, particularly pharmaceutically acceptable salts. Compounds disclosed herein having functional groups that are sufficiently acidic, sufficiently basic, or both can react with a variety of inorganic bases, inorganic acids, and organic acids to form salts. Alternatively, compounds having an intrinsic charge, such as compounds containing quaternary ammonium nitrogen, can form salts with appropriate counterions, such as bromides, chlorides, or fluorides, particularly bromides.

[0234] The compounds described herein may exist in some cases as diastereomers, enantiomers or other stereoisomeric forms. The compounds provided herein include all diastereomers, enantiomers and epimers, and their appropriate mixtures. The separation of stereoisomers can be carried out by chromatography, or by forming diastereomers and separating by recrystallization or chromatography, or any combination thereof. (Jean Jacques, Andre Collet, Samuel H. Wilen, "Enantiomers, Racemates and Resolutions", John Wiley And Sons, Inc., 1981, incorporated herein by reference). Stereoisomers can also be obtained by stereoselective synthesis.

[0235] The methods and compositions described herein encompass the use of amorphous forms and crystalline forms (also referred to as polymorphs). The compounds described herein may exist in the form of pharmaceutically acceptable salts. In addition, in some embodiments, active metabolites of these compounds having the same activity are also included within the scope of the present disclosure. In addition, the compounds described herein may exist in a solvent-free or solvated form, wherein a pharmaceutically acceptable solvent is, for example, water, ethanol, etc. The solvated forms of the compounds described herein are also considered to be disclosed.

[0236] B. Oligonucleotides

[0237] In some embodiments, provided herein are compositions or compounds comprising oligonucleotides. The oligonucleotides can be conjugated to the GalNAc moiety. The oligonucleotides can be directly connected to the joint connected to the GalNAc moiety. The oligonucleotides can be used in the methods described herein.

[0238] In some embodiments, the oligonucleotide is combined with a target oligonucleotide. Examples of target oligonucleotides include target RNA or target DNA. In some embodiments, the oligonucleotide is combined with a target DNA. In some embodiments, the oligonucleotide is combined with a target DNA and suppresses RNA (e.g., mRNA) expression from the target DNA. In some embodiments, the oligonucleotide is combined with a target RNA. The target RNA may include a target mRNA. In some embodiments, the oligonucleotide is combined with a target mRNA. In some embodiments, the oligonucleotide is, for example, by reducing the amount of the target mRNA, causing degradation of the target mRNA, or reducing or preventing the translation of the target mRNA to suppress the target mRNA. In some embodiments, the oligonucleotide is, for example, by suppressing the target mRNA to reduce the amount of the target protein produced from the target mRNA. The oligonucleotide may include a small interfering RNA (siRNA). The oligonucleotide may include an antisense oligonucleotide (ASO).

[0239] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide and inhibits the 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 the 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 the expression of a target protein encoded by the target mRNA.

[0240] 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.

[0241] The target oligonucleotide can be identified in a variety of ways. In some cases, the target oligonucleotide includes mRNA, the expression level of which is related to the onset of a disease (e.g., liver disease). In some cases, the target oligonucleotide includes mRNA encoded by a gene with a specific genotype associated with the disease. Large-scale human genetics data can improve the success rate of drug discovery and development. Genome-wide association studies (GWAS) can detect the association between genetic variants and traits in population samples. GWAS can enable a better understanding of the biology of the disease and provide applicable treatment methods. GWAS can utilize genotyping and / or sequencing data, and generally involves the evaluation of millions of relatively evenly distributed genetic variants on the genome. The most common GWAS design is a case-control study, which involves comparing the variant frequencies in cases relative to controls. If the variant has a significantly different frequency in the case relative to the control, the variant is said to be associated with the disease. The association statistics that can be used in GWAS are p-values ​​as a measure of statistical significance; odds ratios (OR) as a measure of effect size; or beta coefficients (β) as a measure of effect size. Researchers typically assume an additive genetic model and calculate the allelic advantage ratio, which is the increased (or decreased) disease risk conferred by each additional copy of the allele (compared to not carrying this allele copy). Another concept in GWAS design and interpretation is linkage disequilibrium, which is the non-random association of alleles. The presence of linkage disequilibrium may confuse which variant is "causal".

[0242] Functional annotation of variants and / or wet lab experiments can identify causal genetic variants identified via GWAS, and in many cases may lead to the identification of disease-causing genes. In particular, understanding the functional effects of causal genetic variants (e.g., loss of protein function, gain of protein function, increase in gene expression, or reduction in gene expression) can allow this variant to be used as a representative for therapeutic regulation of a target gene, or to gain insight into the potential therapeutic efficacy and safety of a therapeutic agent that regulates this target.

[0243] The identification of such gene-disease associations has provided insights into disease biology and can be used to identify novel therapeutic targets for the pharmaceutical industry. In order to transform therapeutic insights derived from human genetics, disease biology in patients can be exogenously "programmed" to replicate observations from human genetics. There are several potential therapeutic options that can be utilized in converting therapeutic targets identified via human genetics into novel drugs. These can include well-established therapeutic modalities, such as small molecules and monoclonal antibodies; mature methods, such as oligonucleotides; and emerging methods, such as gene therapy and gene editing. The choice of therapeutic modality can depend on several factors, including the location of the target (e.g., intracellular, extracellular or secreted), related tissues (e.g., liver) and related indications. Such studies can be used to identify specific targets associated with liver disorders for inhibition of siRNA or ASO by compositions or compounds described herein.

[0244] Some embodiments include methods for preparing oligonucleotides or siRNA using the methods disclosed herein. For example, any aspect of the examples herein including a synthesis step can be used. Some embodiments include preparing a GalNAc moiety, or preparing an oligonucleotide having a GalNAc moiety.

[0245] 1.siRNA

[0246] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide (e.g., mRNA), wherein 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), wherein the oligonucleotide comprises an 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.

[0247] In some embodiments, the sense strand is 12 to 30 nucleosides in length. In some embodiments, the sense strand is 14 to 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 no more than 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 no more than 12 nucleotides in length. In some embodiments, the sense strand is no more than 14 nucleotides in length. In some embodiments, the sense strand is no more than 16 nucleotides in length. In some embodiments, the sense strand is no more than 18 nucleotides in length. In some embodiments, the sense strand is no more than 20 nucleotides in length. In some embodiments, the sense strand is no more than 22 nucleotides in length. In some embodiments, the sense strand is no more than 24 nucleotides in length. In some embodiments, the sense strand is no more than 26 nucleotides in length. In some embodiments, the sense strand is no more than 28 nucleotides in length. In some embodiments, the sense strand is no longer than 30 nucleotides.

[0248] In some embodiments, the antisense strand is 12 to 30 nucleosides in length. In some embodiments, the antisense strand is 14 to 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 length of the antisense strand is at least 28 nucleotides. In some embodiments, the length of the antisense strand is at least 30 nucleotides. In some embodiments, the length of the antisense strand is no more than 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 some embodiments, the length of the antisense strand is no more than 12 nucleotides. In some embodiments, the length of the antisense strand is no more than 14 nucleotides. In some embodiments, the length of the antisense strand is no more than 16 nucleotides. In some embodiments, the length of the antisense strand is no more than 18 nucleotides. In some embodiments, the length of the antisense strand is no more than 20 nucleotides. In some embodiments, the length of the antisense strand is no more than 22 nucleotides. In some embodiments, the length of the antisense strand is no more than 24 nucleotides. In some embodiments, the length of the antisense strand is no more than 26 nucleotides. In some embodiments, the length of the antisense strand is no more than 28 nucleotides. In some embodiments, the length of the antisense strand is no more than 30 nucleotides. In some embodiments, the antisense strand is the same length as the sense strand.

[0249] In some embodiments, the composition comprises an oligonucleotide that inhibits expression of a target oligonucleotide (e.g., mRNA), wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the sense strand is 12 to 30 nucleosides in length. In some embodiments, the composition comprises a sense strand of 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 any two of the above numbers. In some embodiments, the composition comprises an antisense strand of 12 to 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 a range defined by any two of the foregoing numbers.

[0250] In some embodiments, the composition comprises an oligonucleotide that inhibits expression of a target oligonucleotide (e.g., mRNA), wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, each strand independently being about 14 to 30 nucleosides in length, and at least one of the sense strand and the antisense strand comprises a nucleoside sequence comprising about 12 to 30 consecutive nucleosides of a full-length human target mRNA sequence. In some embodiments, at least one of the sense strand and the antisense strand comprises a nucleoside sequence comprising 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 or more consecutive nucleosides of one of the full-length human target mRNA sequences.

[0251] In some embodiments, the composition comprises an oligonucleotide that inhibits expression of a target protein, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein 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.

[0252] 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 above 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 above numbers. In some embodiments, the 5' overhang comprises 1, 2 or more nucleosides. In some embodiments, the 5' overhang comprises 2 nucleosides.

[0253] 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 above 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 above numbers. In some embodiments, the 5' overhang comprises 1, 2 or more nucleosides. In some embodiments, the 5' overhang comprises 2 nucleosides.

[0254] In some embodiments, the composition comprises an oligonucleotide that inhibits the expression of a target protein, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the siRNA binds to a 19-mer in a human target mRNA encoding the target protein. In some embodiments, the siRNA binds to a 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 a human target mRNA.

[0255] In some embodiments, the composition comprises an oligonucleotide that inhibits the expression of a target protein, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the siRNA binds to a 17-mer in a non-human primate target mRNA encoding the target protein. In some embodiments, the siRNA binds to a 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 a non-human primate target mRNA.

[0256] In some embodiments, the composition comprises an oligonucleotide that inhibits the expression of a target protein, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the siRNA binds to a 19-mer in a human target mRNA encoding the target protein, or a combination thereof. In some embodiments, the siRNA binds to a 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 a human target mRNA.

[0257] 2. ASO

[0258] In some embodiments, compositions include oligonucleotides that inhibit the expression of target oligonucleotides (e.g., mRNA), wherein the oligonucleotides include antisense oligonucleotides (ASOs). In some embodiments, the length of ASO is 12-30 nucleosides. In some embodiments, the length of ASO is 14-30 nucleosides. In some embodiments, the length of 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, or the scope defined by any two of the above numbers. In some embodiments, the length of ASO is 15-25 nucleosides. In some embodiments, the length of ASO is 20 nucleosides. In some embodiments, ASO includes DNA.

[0259] 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 no more than 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 no more than 12 nucleotides in length. In some embodiments, the ASO is no more than 14 nucleotides in length. In some embodiments, the ASO is no more than 16 nucleotides in length. In some embodiments, the ASO is no more than 18 nucleotides in length. In some embodiments, the ASO is no more than 20 nucleotides in length. In some embodiments, the ASO is no more than 22 nucleotides in length. In some embodiments, the ASO is no more than 24 nucleotides in length. In some embodiments, the ASO is no more than 26 nucleotides in length. In some embodiments, the ASO is no more than 28 nucleotides in length. In some embodiments, the ASO is no more than 30 nucleotides in length.

[0260] In some embodiments, the composition comprises an oligonucleotide that inhibits the expression of a target protein, wherein the oligonucleotide comprises an ASO of about 12-30 nucleosides in length and comprises a nucleoside sequence that is complementary to about 12-30 consecutive nucleosides of a human full-length pre-mRNA target sequence encoding the target protein; wherein (i) the oligonucleotide comprises a modification comprising a modified nucleoside and / or a modified internucleoside linkage, and / or (ii) the composition comprises a pharmaceutically acceptable carrier.

[0261] In some embodiments, the composition comprises an oligonucleotide that inhibits the expression of a target protein, wherein the oligonucleotide comprises an ASO of about 12-30 nucleosides in length and comprises a nucleoside sequence that is complementary to about 12-30 consecutive nucleosides in a human full-length target mRNA sequence encoding the target protein; wherein (i) the oligonucleotide comprises a modification comprising a modified nucleoside and / or a modified internucleoside linkage, and / or (ii) the composition comprises a pharmaceutically acceptable carrier.

[0262] 1. Oligonucleotide modification mode

[0263] In some embodiments, the composition comprises an oligonucleotide bound to a target oligonucleotide, wherein the oligonucleotide comprises a modification, the modification comprising a modified nucleoside and / or a modified internucleoside linkage, and / or (ii) the composition comprises a pharmaceutically acceptable carrier. In some embodiments, the oligonucleotide comprises a modification, the 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 alkylphosphonate, a thiophosphate, a methylphosphonate, a dithiophosphate, an alkylthiophosphonate, an aminophosphorate, a carbamate, a carbonate, a phosphotriester, an acetimidate or a carboxymethyl ester, or a combination thereof. In some embodiments, the modified internucleoside linkage comprises one or more thiophosphate linkages. The benefits of the modified internucleoside linkage can include reduced toxicity or improved pharmacokinetics.

[0264] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, wherein the oligonucleotide comprises a modified internucleoside linkage, 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, or a range of modified internucleoside linkages defined by any two of the foregoing numbers. In some embodiments, the oligonucleotide comprises no more than 18 modified internucleoside linkages. In some embodiments, the oligonucleotide comprises no more than 20 modified internucleoside linkages. In some embodiments, the oligonucleotide comprises 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, a plurality of 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.

[0265] In some embodiments, the composition comprises an oligonucleotide bound to a target oligonucleotide, wherein the oligonucleotide comprises 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. In some embodiments, the modified nucleoside comprises a 2'-O-alkyl. In some embodiments, the modified nucleoside comprises a 2'-O-allyl. 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 includes 2'-O-methyl nucleosides, 2'-deoxyfluoro nucleosides, 2'-ON-methylacetamido (2'-O-NMA) nucleosides, 2'-O-dimethylaminoethoxyethyl (2'-O-DMAEOE) nucleosides, 2'-O-aminopropyl (2'-O-AP) nucleosides or 2'-ara-F, or a combination thereof. In some embodiments, the modified nucleoside includes 2'-O-methyl nucleosides. In some embodiments, the modified nucleoside includes 2'-deoxyfluoro nucleosides. In some embodiments, the modified nucleoside includes 2'-O-NMA nucleosides. In some embodiments, the modified nucleoside includes 2'-O-DMAEOE nucleosides. In some embodiments, the modified nucleoside includes 2'-O-aminopropyl (2'-O-AP) nucleosides. In some embodiments, the modified nucleoside includes 2'-ara-F. In some embodiments, the modified nucleoside includes one or more 2' fluorine modified nucleosides. In some embodiments, modified nucleosides include 2'O-alkyl modified nucleosides.Benefits of modified nucleosides may include reduced toxicity or improved pharmacokinetics.

[0266] 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 aforementioned numbers. In some embodiments, the oligonucleotide comprises no more than 19 modified nucleosides. In some embodiments, the oligonucleotide comprises no more than 21 modified nucleosides. In some embodiments, the 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.

[0267] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, wherein the oligonucleotide comprises a lipid attached at the 3' end or the 5' end of the oligonucleotide. In some embodiments, the lipid comprises cholesterol, myristoyl, palmitoyl, stearoyl, lithocholyl, behenoyl, docosahexaenoyl, myristyl, palmitoyl, stearyl, or alpha-tocopherol, or a combination thereof.

[0268] In some embodiments, the composition comprises an oligonucleotide that inhibits the expression of a target mRNA, wherein the oligonucleotide comprises a sugar moiety. The sugar moiety may include 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 include 1, 2, 3, or more sugar molecules. The sugar moiety may be attached to the 3' end or the 5' end of the oligonucleotide. The sugar moiety may include an N-acetylgalactose moiety. The sugar moiety may include an N-acetylgalactosamine (GalNAc) moiety. The sugar moiety may include an N-acetylglucose moiety. The sugar moiety may include an N-acetylglucosamine (GlcNAc) moiety. The sugar moiety may include a fucose moiety. The sugar moiety may include a mannose moiety. N-acetylglucose, GlcNAc, fucose, or mannose may be used to target macrophages because they may target or bind to mannose receptors, such as CD206.

[0269] In some embodiments, the composition comprises an oligonucleotide that inhibits the expression of a target mRNA, wherein the oligonucleotide comprises an N-acetylgalactosamine (GalNAc) moiety. GalNAc can be used to target hepatocytes. In some embodiments, the composition comprises GalNAc. In some embodiments, the composition comprises a GalNAc derivative. The GalNAc moiety may include 1, 2, 3 or more GalNAc molecules. The GalNAc moiety may include a divalent or trivalent branched joint. The oligonucleotide may be attached to 1, 2 or 3 GalNAcs by a divalent or trivalent branched joint. The GalNAc moiety may be attached to the 3' or 5' end of the oligonucleotide.

[0270] Oligonucleotides may include purines. Examples of purines include adenine (A) or guanine (G) or modified forms thereof. Oligonucleotides may include pyrimidines. Examples of pyrimidines include cytosine (C), thymine (T) or uracil (U) or modified forms thereof.

[0271] In some embodiments, the purines of the oligonucleotide include 2' fluorine-modified purines. In some embodiments, the purines of the oligonucleotide include 2'-O-methyl-modified purines. In some embodiments, the purines of the oligonucleotide include a mixture of 2' fluorine and 2'-O-methyl-modified purines. In some embodiments, all purines of the oligonucleotide include 2' fluorine-modified purines. In some embodiments, all purines of the oligonucleotide include 2'-O-methyl-modified purines. In some embodiments, all purines of the oligonucleotide include a mixture of 2' fluorine and 2'-O-methyl-modified purines. In some embodiments, 2'-O-methyl includes 2'O-methyl.

[0272] In some embodiments, the pyrimidines of the oligonucleotides include 2' fluoro-modified pyrimidines. In some embodiments, the pyrimidines of the oligonucleotides include 2'-O-methyl-modified pyrimidines. In some embodiments, the pyrimidines of the oligonucleotides include a mixture of 2' fluoro- and 2'-O-methyl-modified pyrimidines. In some embodiments, all pyrimidines of the oligonucleotides include 2' fluoro-modified pyrimidines. In some embodiments, all pyrimidines of the oligonucleotides include 2'-O-methyl-modified pyrimidines. In some embodiments, all pyrimidines of the oligonucleotides include a mixture of 2' fluoro- and 2'-O-methyl-modified pyrimidines.

[0273] In some embodiments, the purines of the oligonucleotides include 2'-fluoro modified purines, and the pyrimidines of the oligonucleotides include a mixture of 2'-fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the oligonucleotides include 2'-O-methyl modified purines, and the pyrimidines of the oligonucleotides include a mixture of 2'-fluoro and 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the oligonucleotides include 2'-fluoro modified purines, and the pyrimidines of the oligonucleotides include 2'-O-methyl modified pyrimidines. In some embodiments, the purines of the oligonucleotides include 2'-O-methyl modified purines, and the pyrimidines of the oligonucleotides include 2'-fluoro modified pyrimidines. In some embodiments, the pyrimidines of the oligonucleotides include 2'-fluoro modified pyrimidines, and the purines of the oligonucleotides include a mixture of 2'-fluoro and 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the oligonucleotides include 2'-O-methyl modified pyrimidines, and the purines of the oligonucleotides include a mixture of 2' fluoro and 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the oligonucleotides include 2' fluoro modified pyrimidines, and the purines of the oligonucleotides include 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the oligonucleotides include 2'-O-methyl modified pyrimidines, and the purines of the oligonucleotides include 2' fluoro modified purines.

[0274] In some embodiments, all purines of the oligonucleotide include 2' fluorine-modified purines, and all pyrimidines of the oligonucleotide include a mixture of 2' fluorine and 2'-O-methyl modified pyrimidines. In some embodiments, all purines of the oligonucleotide include 2'-O-methyl modified purines, and all pyrimidines of the oligonucleotide include a mixture of 2' fluorine and 2'-O-methyl modified pyrimidines. In some embodiments, all purines of the oligonucleotide include 2' fluorine-modified purines, and all pyrimidines of the oligonucleotide include 2'-O-methyl modified pyrimidines. In some embodiments, all purines of the oligonucleotide include 2' fluorine-modified purines, and all pyrimidines of the oligonucleotide include 2' fluorine-modified pyrimidines. In some embodiments, all pyrimidines of the oligonucleotide include 2' fluorine-modified pyrimidines, and all purines of the oligonucleotide include a mixture of 2' fluorine and 2'-O-methyl modified purines. In some embodiments, all pyrimidines of the oligonucleotide include 2'-O-methyl modified pyrimidines, and all purines of the oligonucleotide include a mixture of 2' fluorine 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.

[0275] 2. siRNA modification patterns

[0276] 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 modified internucleoside linkage within a range defined by any two of the above 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.

[0277] 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 modified internucleoside linkages within the range defined by any two integers above. 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.

[0278] 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 modified nucleoside within the range defined by any two integers above. 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.

[0279] 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 modified nucleosides within the range defined by any two integers above. 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.

[0280] In some embodiments, the sense strand or antisense strand further comprises at least 2 additional nucleosides attached to the 3' end of the sense strand or antisense strand. In some embodiments, the sense strand or antisense strand comprises 2 additional nucleosides attached to the 3' end of the sense strand or antisense strand. As part of the sense strand, these additional nucleosides may or may not be complementary to the target mRNA. The additional nucleosides of the antisense strand may include uracil. The 2 additional nucleosides of the antisense strand may both include uracil.

[0281] In some embodiments, the sense strand or sense strand further comprises at least 2 additional nucleosides attached to the sense strand or the 3' end of the sense strand. In some embodiments, the sense strand or sense strand comprises 2 additional nucleosides attached to the sense strand or the 3' end of the sense strand. The additional nucleosides of the sense strand may include uracil. The 2 additional nucleosides of the sense strand may both include uracil.

[0282] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, wherein the oligonucleotide comprises a siRNA comprising a sense strand and an antisense strand, wherein the sense strand comprises a modification pattern. In some embodiments, the sense strand comprises a modification pattern 1S: 5'-NfsnsNfnNfnNfnNfnNfnNfnNfnNfnNfsnsn-3', wherein "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'-NfnNfnNfnNfnNfnNfnNfnNfnNfsnsn-3', wherein "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 modification pattern 2S: 5'-nsnsnnNfnNfNfNfnnnnnnnnnnsnsn-3', wherein "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 modification pattern 2S#2: 5'-nnnnNfnNfNfnnnnnnnnnnsnsn-3', wherein "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 modification pattern 3S: 5'-nsnsnnNfnNfnNfnnnnnnnnnnsnsn-3', wherein "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 modification pattern 3S#2: 5'-nnnnNfnNfnNfnnnnnnnnsnsn-3', wherein "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 modification pattern 4S: 5'-NfsnsNfnNfnNfNfnNfnNfnNfnNfnNfnNfsnsnN-3', wherein "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 modification pattern 4S#2: 5'-NfnNfnNfnNfnNfnNfnNfnNfnNfnNfsnsnN-3', wherein "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 a modification pattern 5S: 5'-nsnsnnNfnNfNfNfnnnnnnnnnnsnsnN-3', wherein "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 a modification pattern 5S#2: 5'-nnnnNfnNfNfNfnnnnnnnnnnsnsnN-3', wherein "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 a modification pattern 6S: 5'-NfsnsNfnNfnNfnNfnNfnNfnNfnNfnNfsnsn-3', wherein "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 a modification pattern 6S#2: 5'-NfnNfnNfnNfnNfnNfnNfnNfnNfsnsn-3', wherein "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 the 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 the modification patterns 1S, 3S, 4S, or 6S. In some embodiments, the sense strand comprises any one of the modification patterns 1S#2, 3S#2, 4S#2, or 6S#2. Any one of the modification patterns 1S-6S may comprise a GalNAc ligand attached to the 3' end. Any one of the modification patterns 1S-6S may comprise a GalNAc ligand attached to the 5' end. Any one of the modification patterns 1S-6S#2 may comprise a GalNAc ligand attached to the 3' end. Any one of the modification patterns 1S-6S#2 may comprise a GalNAc ligand attached to the 5' end.

[0283] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the antisense strand comprises a modification pattern. In some embodiments, the antisense strand comprises modification pattern 1AS: 5'-nsNfsnNfnNfnNfnNfnNfnNfnNfnsnsn-3', wherein "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 modification pattern 2AS: 5'-nsNfsnnnNfnNfnnnnNfnNfnnnsnsn-3', wherein "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 modification pattern 3AS: 5'-nsNfsnnnNfnnnnnnnfnNfnnnsnsn-3', wherein "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 modification pattern 4AS: 5'-nsNfsnNfnNfnnnnnnnfnNfnnnsnsn-3', wherein "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'-nsNfsnnnNfnNfnnnnnfnNfnnnsnsn-3', wherein "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'-nsNfsnNfnNfnNfnNfnNfnNfnNfnNfnsnsn-3', wherein "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 modification pattern 7AS: 5'-nsNfsnNfnNfnNfnNfnnnnNfnNfnnnsnsn-3', wherein "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 modification pattern 8AS: 5'-nsNfsnnnnnnnnnnnNfnnnnnsnsn-3', wherein "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 9AS: 5'-nsNfsnnnNfnnnnnnnfnNfnNfnNfnsnsn-3', wherein "Nf" is a 2'-fluoro modified nucleoside, "n" is a 2'-O-methyl modified nucleoside, and "s" is a phosphorothioate linkage. Any of the modification patterns 1AS-9AS may include a GalNAc ligand attached to the 3' end. Any of the modification patterns 1AS-9AS may include a GalNAc ligand attached to the 5' end.

[0284] The modification in any modification pattern can be optional. For example, 1, 2, 3 or more phosphorothioate linkages in any of the modification patterns 1S-6S, 1S#2-6S#2 or 1AS-9AS can be replaced by unmodified linkages or different modified linkages. In some cases, 1, 2, 3 or more modified nucleosides in any of the modification patterns 1S-6S or 1AS-9AS can be replaced by unmodified nucleosides or different modified nucleosides.

[0285] In some embodiments, the composition comprises an oligonucleotide that inhibits the expression of a target mRNA, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the sense strand comprises a modification pattern of 1S, and the antisense strand comprises a modification pattern of 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises a modification pattern of 2S, and the antisense strand comprises a modification pattern of 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises a modification pattern of 3S, and the antisense strand comprises a modification pattern of 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises a modification pattern of 4S, and the antisense strand comprises a modification pattern of 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises a modification pattern of 5S and the antisense strand comprises a modification pattern of 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises a modification pattern of 6S and the antisense strand comprises a modification pattern of 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises a modification pattern of 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises a modification pattern of 1S#2 and the antisense strand comprises a modification pattern of 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, or 9AS. In some embodiments, the sense strand comprises a modification pattern of 2S#2 and the antisense strand comprises a modification pattern of 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 the modification pattern ASO1.

[0286] Any combination of sense and antisense modification patterns can 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. In some embodiments, the sense strand comprises modification pattern 6S#2 and the antisense strand comprises modification pattern 8AS.

[0287] In some embodiments, the purines of the sense strand comprise 2'-fluoro modified purines. In some embodiments, the purines of the sense strand comprise 2'-O-methyl modified purines. In some embodiments, the purines of the sense strand comprise a mixture of 2'-fluoro modified purines and 2'-O-methyl modified purines. In some embodiments, all purines of the sense strand comprise 2'-fluoro modified purines. In some embodiments, all purines of the sense strand comprise 2'-O-methyl modified purines. In some embodiments, all purines of the sense strand comprise a mixture of 2'-fluoro modified purines and 2'-O-methyl modified purines.

[0288] 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-modified pyrimidines 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-modified pyrimidines and 2'-O-methyl-modified pyrimidines.

[0289] In some embodiments, the purines of the sense strand comprise 2'-fluoro-modified purines, and the pyrimidines of the sense strand comprise a mixture of 2'-fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines. In some embodiments, the purines of the sense strand comprise 2'-O-methyl-modified purines, and the pyrimidines of the sense strand comprise a mixture of 2'-fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines. In some embodiments, the purines of the sense strand comprise 2'-fluoro-modified purines, and the pyrimidines of the sense strand comprise 2'-O-methyl-modified pyrimidines. In some embodiments, the purines of the sense strand comprise 2'-O-methyl-modified purines, and the pyrimidines of the sense strand comprise 2'-fluoro-modified pyrimidines. In some embodiments, the pyrimidines of the sense strand comprise 2'-fluoro-modified pyrimidines, and the purines of the sense strand comprise a mixture of 2'-fluoro-modified purines and 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 a mixture of 2'-fluoro modified purines 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.

[0290] In some embodiments, all purines of the sense strand comprise 2'-fluoro-modified purines, and all pyrimidines of the sense strand comprise a mixture of 2'-fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines. In some embodiments, all purines of the sense strand comprise 2'-O-methyl-modified purines, and all pyrimidines of the sense strand comprise a mixture of 2'-fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines. In some embodiments, all purines of the sense strand comprise 2'-fluoro-modified purines, and all pyrimidines of the sense strand comprise 2'-O-methyl-modified pyrimidines. In some embodiments, all purines of the sense strand comprise 2'-O-methyl-modified purines, and all pyrimidines of the sense strand comprise 2'-fluoro-modified pyrimidines. In some embodiments, all pyrimidines of the sense strand comprise 2'-fluoro-modified pyrimidines, and all purines of the sense strand comprise a mixture of 2'-fluoro-modified purines and 2'-O-methyl-modified purines. In some embodiments, all pyrimidines of the sense strand comprise 2'-O-methyl modified pyrimidines, and all purines of the sense strand comprise a mixture of 2'-fluoro modified purines and 2'-O-methyl modified purines. In some embodiments, all pyrimidines of the sense strand comprise 2'-fluoro modified pyrimidines, and all purines of the sense strand comprise 2'-O-methyl modified purines. In some embodiments, all pyrimidines of the sense strand comprise 2'-O-methyl modified pyrimidines, and all purines of the sense strand comprise 2'-fluoro modified purines.

[0291] In some embodiments, the purines of the antisense strand comprise 2'-fluoro modified purines. In some embodiments, the purines of the antisense strand comprise 2'-O-methyl modified purines. In some embodiments, the purines of the antisense strand comprise a mixture of 2'-fluoro modified purines and 2'-O-methyl modified purines. In some embodiments, all purines of the antisense strand comprise 2'-fluoro modified purines. In some embodiments, all purines of the antisense strand comprise 2'-O-methyl modified purines. In some embodiments, all purines of the antisense strand comprise 2'-fluoro modified purines and a mixture of 2'-O-methyl modified purines.

[0292] 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-modified pyrimidines 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-modified pyrimidines and 2'-O-methyl-modified pyrimidines.

[0293] In some embodiments, the purines of the antisense strand comprise 2'-fluoro-modified purines, and the pyrimidines of the antisense strand comprise a mixture of 2'-fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines. In some embodiments, the purines of the antisense strand comprise 2'-O-methyl-modified purines, and the pyrimidines of the antisense strand comprise a mixture of 2'-fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines. In some embodiments, the purines of the antisense strand comprise 2'-fluoro-modified purines, and the pyrimidines of the antisense strand comprise 2'-O-methyl-modified purines. In some embodiments, the purines of the antisense strand comprise 2'-O-methyl-modified purines, and the pyrimidines of the antisense strand comprise 2'-fluoro-modified pyrimidines. In some embodiments, the pyrimidines of the antisense strand comprise 2'-fluoro-modified pyrimidines, and the purines of the antisense strand comprise a mixture of 2'-fluoro-modified purines and 2'-O-methyl-modified purines. In some embodiments, the pyrimidines of the antisense strand comprise 2'-O-methyl modified pyrimidines, and the purines of the antisense strand comprise a mixture of 2'-fluoro modified purines and 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the antisense strand comprise 2'-fluoro modified pyrimidines, and the purines of the antisense strand comprise 2'-O-methyl modified purines. In some embodiments, the pyrimidines of the antisense strand comprise 2'-O-methyl modified pyrimidines, and the purines of the antisense strand comprise 2'-fluoro modified purines.

[0294] In some embodiments, all purines of the antisense strand comprise 2' fluoro-modified purines, and all pyrimidines of the antisense strand comprise a mixture of 2' fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines. In some embodiments, all purines of the antisense strand comprise 2'-O-methyl-modified purines, and all pyrimidines of the antisense strand comprise a mixture of 2' fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines. In some embodiments, all purines of the antisense strand comprise 2' fluoro-modified purines, and all pyrimidines of the antisense strand comprise 2'-O-methyl-modified purines. In some embodiments, all purines of the antisense strand comprise 2'-O-methyl-modified purines, and all pyrimidines of the antisense strand comprise 2' fluoro-modified pyrimidines. In some embodiments, all pyrimidines of the antisense strand comprise 2' fluoro-modified pyrimidines, and all purines of the antisense strand comprise a mixture of 2' fluoro-modified pyrimidines and 2'-O-methyl-modified purines. In some embodiments, all pyrimidines of the antisense strand comprise 2'-O-methyl modified pyrimidines, and all purines of the antisense strand comprise a mixture of 2' fluoro-modified pyrimidines and 2'-O-methyl modified purines. In some embodiments, all pyrimidines of the antisense strand comprise 2' fluoro-modified pyrimidines, and all purines of the antisense strand comprise 2'-O-methyl modified purines. In some embodiments, all pyrimidines of the antisense strand comprise 2'-O-methyl modified pyrimidines, and all purines of the antisense strand comprise 2' fluoro-modified purines.

[0295] In some cases, the oligonucleotide comprises a specific modification pattern. In some embodiments, position 9, counted from the 5' end of the oligonucleotide chain, can have a 2'F modification. In some embodiments, when position 9 of the oligonucleotide chain is a pyrimidine, all purines in the oligonucleotide chain have a 2'OMe modification. In some embodiments, when position 9 is the only pyrimidine between positions 5 and 11 of the sense chain, position 9 is the only position with a 2'F modification in the oligonucleotide chain. In some embodiments, when only another base between position 9 and position 5 and 11 of the oligonucleotide chain is a pyrimidine, these two pyrimidines are the only two positions with a 2'F modification in the oligonucleotide chain. In some embodiments, when only two other bases between position 9 and position 5 and 11 of the oligonucleotide chain are pyrimidines, and these other two pyrimidines are in adjacent positions so that there are no continuous three 2'F modifications, then any combination of 2'F modifications that produce a total of three 2'F modifications can be performed. In some embodiments, when there are more than 2 pyrimidines between positions 5 and 11 of an oligonucleotide strand, then all combinations of pyrimidines with 2'F modifications are allowed to have a total of 3 to 5 2'F modifications, provided that the oligonucleotide strand does not have 3 consecutive 2'F modifications. In some cases, the oligonucleotide strand of any siRNA comprises a modification pattern that complies with any or all of these oligonucleotide strand rules.

[0296] In some embodiments, when position 9 of the oligonucleotide chain is a purine, all purines in the oligonucleotide chain have 2'OMe modifications. In some embodiments, when position 9 is the only purine between positions 5 and 11 of the sense chain, position 9 is the only position in the oligonucleotide chain with a 2'F modification. In some embodiments, when only another base between position 9 and position 5 and 11 of the oligonucleotide chain is a purine, these two purines are the only two positions in the oligonucleotide chain with a 2'F modification. In some embodiments, when only two other bases between position 9 and position 5 and 11 of the oligonucleotide chain are purines, and these other two purines are in adjacent positions so that there are no three consecutive 2'F modifications, any combination of 2'F modifications that produce three 2'F modifications can be performed. In some embodiments, when there are more than 2 purines between positions 5 and 11 of the oligonucleotide chain, all combinations of purines with 2'F modifications are allowed to have a total of 3 to 5 2'F modifications, provided that the oligonucleotide chain does not have 3 consecutive 2'F modifications. In some cases, the oligonucleotide strand of any siRNA comprises a modification pattern that conforms to any or all of these oligonucleotide strand rules.

[0297] In some cases, position 9 of the oligonucleotide chain can be 2'deoxy. In these cases, 2'F and 2'OMe modifications can occur at other positions of the oligonucleotide chain. In some cases, the oligonucleotide chain of any siRNA contains a modification pattern that conforms to these oligonucleotide chain rules.

[0298] In some embodiments, position 9 of the sense strand comprises a 2' fluorine-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' fluorine-modified pyrimidines, provided that there are no three consecutive 2' fluorine-modified pyrimidines. In some embodiments, the odd positions of the antisense strand comprise 2'-O-methyl-modified nucleotides. In some embodiments, the even positions of the antisense strand comprise 2' fluorine-modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, the even positions of the antisense strand comprise 2' fluorine-modified nucleotides, 2'-O-methyl-modified nucleotides, and unmodified deoxyribonucleotides. In some embodiments, position 9 of the sense strand comprises a 2' fluorine-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' fluorine-modified pyrimidines, provided that there are not three consecutive 2' fluorine-modified pyrimidines; the odd-numbered positions of the antisense strand comprise 2'-O-methyl-modified nucleotides; and the even-numbered positions of the antisense strand comprise 2' fluorine-modified nucleotides and unmodified deoxyribonucleotides.

[0299] In some embodiments, position 9 of the sense strand includes a 2' fluorine-modified purine. In some embodiments, all pyrimidines of the sense strand include 2'-O-methyl-modified purines. In some embodiments, 1, 2, 3, 4 or 5 purines between positions 5 and 11 include 2' fluorine-modified purines, provided that there are no three consecutive 2' fluorine-modified purines. In some embodiments, the odd positions of the antisense strand include 2'-O-methyl-modified nucleotides. In some embodiments, the even positions of the antisense strand include 2' fluorine-modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, the even positions of the antisense strand include 2' fluorine-modified nucleotides, 2'-O-methyl-modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, position 9 of the sense strand comprises a 2' fluorine-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' fluorine-modified purines, provided that there are not three consecutive 2' fluorine-modified purines; the odd positions of the antisense strand comprise 2'-O-methyl-modified nucleotides; and the even positions of the antisense strand comprise 2' fluorine-modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, there are not three consecutive 2' fluorine-modified purines. In some embodiments, there are not three consecutive 2' fluorine-modified pyrimidines.

[0300] In some embodiments, position 9 of the sense strand comprises an unmodified deoxyribonucleotide. In some embodiments, positions 5, 7, and 8 of the sense strand comprise 2' fluorine-modified nucleotides. In some embodiments, all pyrimidines at positions 10 to 21 of the sense strand comprise 2'-O-methyl-modified pyrimidines, and all purines at positions 10 to 21 of the sense strand comprise 2'-O-methyl-modified purines or 2' fluorine-modified purines. In some embodiments, the odd positions of the antisense strand comprise 2'-O-methyl-modified nucleotides. In some embodiments, the even positions of the antisense strand comprise 2' fluorine-modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, the even positions of the antisense strand comprise 2' fluorine-modified nucleotides, 2'-O-methyl-modified nucleotides, and unmodified deoxyribonucleotides. In some embodiments, position 9 of the sense strand comprises an unmodified deoxyribonucleotide; positions 5, 7, and 8 of the sense strand comprise 2' fluorine-modified nucleotides; all pyrimidines at positions 10 to 21 of the sense strand comprise 2'-O-methyl-modified pyrimidines, and all purines at positions 10 to 21 comprise 2'-O-methyl-modified purines or 2' fluorine-modified purines; the odd-numbered positions of the antisense strand comprise 2'-O-methyl-modified nucleotides; and the even-numbered positions of the antisense strand comprise 2' fluorine-modified nucleotides and unmodified deoxyribonucleotides.

[0301] In some embodiments, position 9 of the sense strand comprises an unmodified deoxyribonucleotide. In some embodiments, positions 5, 7, and 8 of the sense strand comprise 2' fluorine-modified nucleotides. In some embodiments, all purines at positions 10 to 21 of the sense strand comprise 2'-O-methyl-modified purines, and all pyrimidines at positions 10 to 21 of the sense strand comprise 2'-O-methyl-modified pyrimidines or 2' fluorine-modified pyrimidines. In some embodiments, the odd positions of the antisense strand comprise 2'-O-methyl-modified nucleotides. In some embodiments, the even positions of the antisense strand comprise 2' fluorine-modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, the even positions of the antisense strand comprise 2' fluorine-modified nucleotides, 2'-O-methyl-modified nucleotides, and unmodified deoxyribonucleotides. In some embodiments, position 9 of the sense strand comprises an unmodified deoxyribonucleotide; positions 5, 7, and 8 of the sense strand comprise 2' fluorine-modified nucleotides; all purines at positions 10 to 21 of the sense strand comprise 2'-O-methyl-modified purines, and all pyrimidines at positions 10 to 21 comprise 2'-O-methyl-modified pyrimidines or 2' fluorine-modified pyrimidines; the odd-numbered positions of the antisense strand comprise 2'-O-methyl-modified nucleotides; and the even-numbered positions of the antisense strand comprise 2' fluorine-modified nucleotides and unmodified deoxyribonucleotides.

[0302] 3. ASO modification mode

[0303] In some embodiments, the composition comprises an oligonucleotide that binds to a target oligonucleotide, wherein the oligonucleotide comprises an antisense oligonucleotide (ASO). In some embodiments, the ASO comprises a modification pattern ASO1:

[0304] 5'-nsnsnsnsnsdNsdNsdNsdNsdNsdNsdNsdNsdNsdNsdNsnsnsnsn-3', wherein "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.

[0305] In some embodiments, the ASO is conjugated to a GalNAc moiety. The GalNAc moiety can be conjugated to the ASO at the 5' end or the 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.

[0306] C. Preparation

[0307] In some embodiments, the composition is a pharmaceutical composition. In some embodiments, the composition is sterile. In some embodiments, the composition further comprises a pharmaceutically acceptable carrier. The preparation may include compounds such as GalNAc moieties, and oligonucleotides conjugated to the GalNAc moieties described herein.

[0308] In some embodiments, the pharmaceutically acceptable carrier comprises water. In some embodiments, the pharmaceutically acceptable carrier comprises a buffer. In some embodiments, the pharmaceutically acceptable carrier comprises a saline solution. In some embodiments, the pharmaceutically acceptable carrier comprises water, a buffer, or a saline solution. In some embodiments, the composition comprises liposomes. In some embodiments, the pharmaceutically acceptable carrier comprises liposomes, lipids, nanoparticles, proteins, protein-antibody complexes, peptides, cellulose, nanogels, or combinations thereof.

[0309] II. Methods and Uses

[0310] Disclosed herein, in some embodiments, are methods of administering a composition described herein to a subject.Some embodiments relate to using a composition described herein, such as administering the composition to a subject.

[0311] Some embodiments relate to a method of treating a condition in a subject in need thereof. Some embodiments relate to the use of a composition as described herein in a method of treatment. Some embodiments include administering a composition as described herein to a subject suffering from a condition. In some embodiments, administering treats a condition in a subject. In some embodiments, the composition treats a condition in a subject.

[0312] In some embodiments, treatment includes preventing, inhibiting, or reversing a condition in a subject. Some embodiments relate to the use of a composition as described herein in a method for preventing, inhibiting, or reversing a condition. Some embodiments relate to a method for preventing, inhibiting, or reversing a condition in a subject in need thereof. Some embodiments include administering a composition as described herein to a subject having a condition. In some embodiments, administration prevents, inhibits, or reverses a condition in a subject. In some embodiments, the composition prevents, inhibits, or reverses a condition in a subject.

[0313] Some embodiments relate to a method of preventing a condition in a subject in need thereof. Some embodiments relate to the use of a composition described herein in a method of preventing a condition. Some embodiments include administering a composition described herein to a subject having a condition. In some embodiments, administration prevents the condition in a subject. In some embodiments, the composition prevents the condition in a subject.

[0314] Some embodiments relate to a method of inhibiting a condition in a subject in need thereof. Some embodiments relate to the use of a composition described herein in a method of inhibiting a condition. Some embodiments include administering a composition described herein to a subject having a condition. In some embodiments, administration inhibits the condition of the subject. In some embodiments, the composition inhibits the condition of the subject.

[0315] Some embodiments relate to a method of reversing a condition in a subject in need thereof. Some embodiments relate to the use of a composition as described herein in a method of reversing a condition. Some embodiments include administering a composition as described herein to a subject having a condition. In some embodiments, administration reverses the condition of the subject. In some embodiments, the composition reverses the condition of the subject.

[0316] A. Symptoms

[0317] Some embodiments of the methods described herein include treating a condition in a subject in need thereof. In some embodiments, the condition is a liver condition. Non-limiting examples of liver conditions include hepatitis, liver cancer, liver fibrosis, cholestasis, gallbladder disease, biliary disease, alcoholic liver disease, nonalcoholic fatty hepatitis, liver infection, or a hereditary liver condition. In some embodiments, liver conditions include hepatitis. In some embodiments, liver conditions include liver cancer. In some embodiments, liver conditions include liver fibrosis. In some embodiments, liver conditions include cholestasis. In some embodiments, liver conditions include gallbladder disease. In some embodiments, liver conditions include biliary disease. In some embodiments, liver conditions include alcoholic liver disease. In some embodiments, liver conditions include nonalcoholic fatty hepatitis.

[0318] 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.

[0319] In some embodiments, the liver disorder comprises an inherited liver disorder. In some embodiments, the inherited liver disorder comprises hemochromatosis. In some embodiments, the inherited liver disorder comprises Wilson's disease.

[0320] B. Object

[0321] 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, a mammal, a dog, a cat, a cow, a rodent, a mouse, a rat, a primate, or a monkey. In some embodiments, the subject is a human.

[0322] In some embodiments, the subject is male. In some embodiments, the subject is female.

[0323] 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 more, or a range defined by any two of the above integers. In some embodiments, the subject is overweight. In some embodiments, the subject has a BMI of 25 or more. In some embodiments, the subject has a BMI of 25-29. In some embodiments, the subject is obese. In some embodiments, the subject has a BMI of 30 or more. In some embodiments, the subject has a BMI of 30-39. In some embodiments, the subject has a BMI of 40-50. In some embodiments, the subject has a BMI of 25-50.

[0324] In some embodiments, the subject is ≥90 years old. In some embodiments, the subject is ≥85 years old. In some embodiments, the subject is ≥80 years old. In some embodiments, the subject is ≥70 years old. In some embodiments, the subject is ≥60 years old. In some embodiments, the subject is ≥50 years old. In some embodiments, the subject is ≥40 years old. In some embodiments, the subject is ≥30 years old. In some embodiments, the subject is ≥20 years old. In some embodiments, the subject is ≥10 years old. In some embodiments, the subject is ≥1 year old. In some embodiments, the subject is ≥0 years old.

[0325] In some embodiments, the subject is ≤100 years old. In some embodiments, the subject is ≤90 years old. In some embodiments, the subject is ≤85 years old. In some embodiments, the subject is ≤80 years old. In some embodiments, the subject is ≤70 years old. In some embodiments, the subject is ≤60 years old. In some embodiments, the subject is ≤50 years old. In some embodiments, the subject is ≤40 years old. In some embodiments, the subject is ≤30 years old. In some embodiments, the subject is ≤20 years old. In some embodiments, the subject is ≤10 years old. In some embodiments, the subject is ≤1 year old.

[0326] In some embodiments, the subject is between 0 and 100 years old. In some embodiments, the subject is between 20 and 90 years old. In some embodiments, the subject is between 30 and 80 years old. In some embodiments, the subject is between 40 and 75 years old. In some embodiments, the subject is between 50 and 70 years old. In some embodiments, the subject is between 40 and 85 years old.

[0327] C. Baseline Measurements

[0328] Some embodiments of the methods described herein include obtaining a baseline measurement from a subject. For example, in some embodiments, a baseline measurement is obtained from a subject prior to treating the subject. Non-limiting examples of baseline measurements include baseline symptom (e.g., liver disorder symptoms) measurements, baseline protective phenotype measurements, baseline target oligonucleotide (e.g., mRNA) measurements, or baseline target protein measurements.

[0329] In some embodiments, the baseline measurement value is obtained directly from the object. In some embodiments, the baseline measurement value is obtained by observation, such as by observing the object or the tissue of the object. In some embodiments, the baseline measurement value is obtained in a non-invasive manner using an imaging device. In some embodiments, the baseline measurement value is obtained in a sample from the object. In some embodiments, the baseline measurement value is obtained in one or more histological tissue sections. In some embodiments, the baseline measurement value is obtained by performing an assay such as an immunoassay, a colorimetric assay, or a fluorometric assay on a sample obtained from the object. In some embodiments, the baseline measurement value is obtained by an immunoassay, a colorimetric assay, or a fluorometric assay. In some embodiments, the baseline measurement value is obtained by PCR.

[0330] In some embodiments, the baseline measurement is a baseline symptom measurement. The symptom can be a symptom of a disorder associated with the target oligonucleotide. The disorder can be a liver disorder.

[0331] In some embodiments, the baseline measurement is a baseline protective phenotype measurement. The protective phenotype can protect the subject from developing a disease associated with the target oligonucleotide. The protective phenotype can be negatively correlated with the incidence of the disease.

[0332] In some embodiments, the baseline measurement is a baseline target protein measurement. In some embodiments, the baseline target protein measurement includes a baseline target protein level. In some embodiments, the baseline target protein level is expressed as the mass or percentage of the target protein per sample weight. In some embodiments, the baseline target protein level is expressed as the mass or percentage of the target protein per sample volume. In some embodiments, the baseline target protein level is expressed as the mass or percentage of the 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 fluorometric assay.

[0333] In some embodiments, the baseline measurement is a baseline target mRNA measurement. In some embodiments, the baseline target mRNA measurement includes a baseline target mRNA level. In some embodiments, the baseline target mRNA level is expressed as a target mRNA mass or percentage per sample weight. In some embodiments, the baseline target mRNA level is expressed as a target mRNA mass or percentage per sample volume. In some embodiments, the baseline target mRNA level is expressed as a target mRNA mass or percentage per total mRNA in the sample. In some embodiments, the baseline target mRNA level is expressed as a target mRNA mass or percentage of 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 measuring such as polymerase chain reaction (PCR). In some embodiments, PCR includes quantitative PCR (qPCR). In some embodiments, PCR includes reverse transcription of the target mRNA.

[0334] Some embodiments of the methods described herein include obtaining a sample from a subject. In some embodiments, the baseline measurement is obtained in a sample obtained from a subject. In some embodiments, a sample is obtained from a subject before administering a composition as described herein or treating a subject with a composition as described herein. In some embodiments, a baseline measurement is obtained in a sample obtained from a subject before administering a composition to a subject. In some embodiments, the sample is obtained from a subject in a fasting state. In some embodiments, the sample is obtained from a subject after an overnight fast. In some embodiments, the sample is obtained from a subject in a fed state.

[0335] In some embodiments, the sample comprises a liquid. In some embodiments, the sample is a liquid 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.

[0336] In some embodiments, the sample includes tissue. In some embodiments, the sample is a tissue sample. In some embodiments, the sample includes liver tissue. In some embodiments, the sample is a liver sample. In some embodiments, the sample includes hepatocytes. In some embodiments, the sample consists of hepatocytes. For example, baseline target mRNA measurements or baseline target protein measurements can be obtained in liver or hepatocyte samples from patients. In some embodiments, the sample includes adipose tissue. In some embodiments, the sample is a fat sample. The fat sample may include white adipose tissue or consist of white adipose tissue. The fat sample may include brown adipose tissue or consist of brown adipose tissue. In some embodiments, the sample includes kidney tissue. In some embodiments, the sample is a kidney sample. In some embodiments, the sample includes heart tissue, such as ventricular or atrial tissue. In some embodiments, the sample is a heart sample. In some embodiments, the sample includes intestinal tissue, such as small intestinal tissue. In some embodiments, the sample is a small intestinal sample. In some embodiments, the sample includes lymph node tissue, such as mesenteric lymph node tissue. In some embodiments, the sample is a mesenteric lymph node sample. In some embodiments, the sample includes muscle tissue. In some embodiments, the sample is a muscle sample. In some embodiments, the tissue sample comprises liver, fat, kidney or heart tissue. In some embodiments, the tissue sample comprises brown adipose tissue, white adipose tissue, kidney tissue, intestinal tissue, mesenteric lymph node or muscle tissue.

[0337] D. Efficacy

[0338] In some embodiments, the composition or administration of the composition affects a measurement such as a symptom measurement (e.g., a liver disorder symptom), a protective phenotype measurement, a target oligonucleotide (e.g., an mRNA) measurement, or a target protein measurement (e.g., liver tissue target protein levels) relative to a baseline measurement.

[0339] Some embodiments of the methods described herein include obtaining a measurement from a subject. For example, the measurement can be obtained from the subject after the subject is treated. In some embodiments, after the composition is applied to the subject, the measurement is obtained in a second sample (such as a fluid or tissue sample described herein) obtained from the subject. In some embodiments, the measurement is an indication that the condition has been treated.

[0340] In some embodiments, the measured value is obtained directly from the object. In some embodiments, the measured value is obtained using an imaging device in a non-invasive manner. In some embodiments, the measured value is obtained in a second sample from the object. In some embodiments, the measured value is obtained in one or more histological tissue sections. In some embodiments, the measured value is obtained by measuring the second sample obtained from the object. In some embodiments, the measured value is obtained by measuring (such as the measurement described herein). In some embodiments, it is determined as an immunoassay, a colorimetric assay, a fluorometric assay or a PCR assay. In some embodiments, the measured value is obtained by measuring such as an immunoassay, a colorimetric assay or a fluorometric assay. In some embodiments, the measured value is obtained by PCR. In some embodiments, the measured value is obtained by histology. In some embodiments, the measured value is obtained by observation. In some embodiments, additional measurements are performed, such as in the 3rd sample, the 4th sample or the 5th sample.

[0341] In some embodiments, the measured value is obtained within 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 12 hours, 18 hours, or 24 hours after applying the composition. In some embodiments, the measured value is obtained within 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days after applying the composition. In some embodiments, the measured value is 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 applying the composition. In some embodiments, the measured value is obtained after 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 12 hours, 18 hours, or 24 hours after applying the composition. In some embodiments, the measured value is obtained after 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days after applying the composition. In some embodiments, the measurements are obtained after 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.

[0342] In some embodiments, the measured value of a symptom or a parameter associated with a condition of a subject is reduced or affected over a longer period of time compared to a baseline measured value. In some embodiments, the measured value is reduced or affected 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 6 days, about 27 days, about 28 days, 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 time range after administration that continues to include the range defined by any two of the above days. In some embodiments, the measured value is reduced or affected 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 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 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 measured value is reduced or affected for no more than 1 day, no more than 2 days, no more than 3 days, no more than 4 days, no more than 5 days, no more than 6 days, no more than 7 days, no more than 8 days, no more than 9 days, no more than 10 days, no more than 11 days, no more than 12 days, no more than 13 days, no more than 14 days, no more than 15 days, no more than 16 days, no more than 17 days, no more than 18 days, no more than 19 days, no more than 20 days, no more than 21 days, no more than 22 days, no more than 23 days, no more than 24 days, no more than 25 days, no more than 26 days, no more than 27 days, no more than 28 days, no more than 29 days, no more than 30 days, no more than 31 days, no more than 32 days, no more than 33 days, no more than 34 days, no more than 35 days, no more than 36 days, no more than 37 days, no more than 38 days, no more than 39 days, no more than 40 days, no more than 41 days, no more than 42 days, no more than 43 days, no more than 44 days, no more than 45 days, no more than 46 days, no more than 47 days, no more than 48 days, no more than 49 days, no more than 50 days, no more than 51 days, no more than 52 days, no more than 53 days, no more than 54 days, no more than 55 days, no more than 56 days, no more than 57 days, no more than 58 days, no more than 59 days, no more than 60 days, no more than 61 days, no more than 62 days, no more than 63 days, no more than In some embodiments, the measured value is reduced or affected for at least about 5 days. In some embodiments, the measured value is reduced or affected for at least about 10 days. In some embodiments, the measured value is reduced or affected for at least about 15 days. In some embodiments, the measured value is reduced or affected for at least about 20 days. In some embodiments, the measured value is reduced or affected for at least about 25 days. In some embodiments, the measured value is reduced or affected for at least about 30 days. In some embodiments, the measured value is reduced or affected for at least about 40 days. In some embodiments, the measured value is reduced or affected for at least about 50 days. In some embodiments, the measured value is reduced or affected for at least about 60 days. In some embodiments, the measured value is reduced or affected for at least about 70 days. In some embodiments, the measured value is reduced or affected for at least about 80 days. In some embodiments, the measured value is reduced or affected for at least about 90 days. In some embodiments, the measured value is reduced or affected for at least about 100 days. In some embodiments, the measured value is reduced or affected for at least about 110 days. In some embodiments, the measured value is reduced or affected for at least about 120 days. Examples of the measured values ​​of the symptoms or parameters associated with the disease may include target mRNA measured values, target protein measured values, biomarker measured values, or physiological measured values. The measured value may be in a tissue (e.g., liver) or in a biological fluid (e.g., blood, serum, or plasma).

[0343] In some embodiments, the composition reduces the symptom measurement relative to the baseline symptom measurement. In some embodiments, after applying the composition to the object, the reduction is measured in the second tissue sample obtained from the object. In some embodiments, the reduction is measured directly in the object after applying the composition to the object. In some embodiments, the symptom measurement is reduced by about 2.5% or more, about 5% or more, or about 7.5% or more relative to the baseline symptom measurement. In some embodiments, the symptom measurement is reduced by about 10% or more relative to the baseline symptom measurement. In some embodiments, the symptom measurement 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 relative to the baseline symptom measurement. In some embodiments, the symptom measurement is reduced by no more than about 2.5%, no more than about 5%, or no more than about 7.5% relative to the baseline symptom measurement. In some embodiments, the symptom measurement is reduced by no more than about 10% relative to the baseline symptom measurement. In some embodiments, the symptom measurement value is reduced by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100% relative to the baseline symptom measurement value. In some embodiments, the symptom measurement value 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 two of the above percentages.

[0344] In some embodiments, the composition increases the protective phenotype measurement relative to the baseline protective phenotype measurement. In some embodiments, the increase is measured in a second tissue sample obtained from the object after the composition is applied to the object. In some embodiments, the increase is measured directly from the object after the composition is applied to the object. In some embodiments, the protective phenotype measurement is increased by about 2.5% or more, about 5% or more, or about 7.5% or more relative to the baseline protective phenotype measurement. In some embodiments, the protective phenotype measurement is increased by about 10% or more relative to the baseline protective phenotype measurement. In some embodiments, the protective phenotype measurement 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 relative to the baseline protective phenotype measurement. In some embodiments, the protective phenotype measurement is increased by about 100% or more, about 250% or more, about 500% or more, about 750% or more, or about 1000% or more relative to the baseline protective phenotype measurement. In some embodiments, the protective phenotype measurement value increases by no more than about 2.5%, no more than about 5%, or no more than about 7.5% relative to the baseline protective phenotype measurement value. In some embodiments, the protective phenotype measurement value increases by no more than about 10% relative to the baseline protective phenotype measurement value. In some embodiments, the protective phenotype measurement value increases by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100% relative to the baseline protective phenotype measurement value. In some embodiments, the protective phenotype measurement value increases by no more than about 100%, no more than about 250%, no more than about 500%, no more than about 750%, or no more than about 1000% relative to the baseline protective phenotype measurement value. In some embodiments, the protective phenotypic measurement value increases 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 two of the above percentages.

[0345] In some embodiments, the measured value is a target protein measured value. In some embodiments, the target protein measured value includes the target protein level. In some embodiments, the target protein level is expressed as the mass or percentage of the target protein per sample weight. In some embodiments, the target protein level is expressed as the mass or percentage of the target protein per sample volume. In some embodiments, the target protein level is expressed as the mass or percentage of the target protein per total protein in the sample. In some embodiments, the target protein measured value is a cell (e.g., hepatocyte) target protein measured value. In some embodiments, the target protein measured value is a tissue (e.g., liver tissue) target protein measured value. In some embodiments, the target protein measured value is a circulating target protein measured value. In some embodiments, the baseline target protein measured value is obtained by measuring such as immunoassay, colorimetric assay or fluorometric assay.

[0346] In some embodiments, the composition reduces the target protein measurement relative to the baseline target protein measurement. In some embodiments, the composition reduces the tissue target protein level (such as but not limited to the liver tissue target protein level) relative to the baseline target protein measurement. In some embodiments, the composition reduces the cell target protein level (such as but not limited to the hepatocyte target protein level) relative to the baseline target protein measurement. In some embodiments, the composition reduces the circulating target protein level relative to the baseline target protein measurement. In some embodiments, the reduced target protein level is measured in a second sample obtained from the subject after the composition is administered to the subject.

[0347] In some embodiments, the target protein measurement value is reduced by about 2.5% or more, about 5% or more, or about 7.5% or more relative to the baseline target protein measurement value. In some embodiments, the target protein measurement value is reduced by about 10% or more relative to the baseline target protein measurement value. In some embodiments, the target protein measurement value 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% relative to the baseline target protein measurement value. In some embodiments, the target protein measurement value is reduced by no more than about 2.5%, no more than about 5%, or no more than about 7.5% relative to the baseline target protein measurement value. In some embodiments, the target protein measurement value is reduced by no more than about 10% relative to the baseline target protein measurement value. In some embodiments, the target protein measurement value is reduced by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline target protein measurement value. In some embodiments, the target protein measurement value decreases by 2.5%, 5%, 7.5%, 19%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 100%, or a range defined by any two of the above percentages. The target protein measurement value can be reduced over a longer period of time. In some embodiments, the target protein measurement value decreases 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, the target protein measurement value decreases 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.

[0348] In some embodiments, the measured value is a target mRNA measured value. In some embodiments, the target mRNA measured value includes the target mRNA level. In some embodiments, the target mRNA level is expressed as the target mRNA mass or percentage per sample weight. In some embodiments, the target mRNA level is expressed as the target mRNA mass or percentage per sample volume. In some embodiments, the target mRNA level is expressed as the target mRNA mass or percentage per total mRNA in the sample. In some embodiments, the target mRNA level is expressed as the target mRNA mass or percentage per total nucleic acid in the sample. In some embodiments, the target mRNA level is expressed relative to another mRNA level in the sample (such as the mRNA level of a housekeeping gene). In some embodiments, the target mRNA measured value is obtained by measuring such as PCR measurement. In some embodiments, PCR includes qPCR. In some embodiments, PCR includes reverse transcription of the target mRNA.

[0349] In some embodiments, the composition reduces the target mRNA measurement relative to the baseline target mRNA measurement. In some embodiments, the target mRNA measurement is obtained from a second sample obtained from the object after the composition is administered to the object. In some embodiments, the composition reduces the target mRNA level relative to the baseline target mRNA level. In some embodiments, the reduced target mRNA level is measured in a second sample obtained from the object after the composition is administered to the object. In some embodiments, the second sample is a second liver sample. In some embodiments, the second sample is a second hepatocyte sample.

[0350] In some embodiments, the target mRNA measured value is reduced by about 2.5% or more, about 5% or more, or about 7.5% or more relative to the baseline target mRNA measured value. In some embodiments, the target mRNA measured value is reduced by about 10% or more relative to the baseline target mRNA measured value. In some embodiments, the target mRNA measured value 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% relative to the baseline target mRNA measured value. In some embodiments, the target mRNA measured value is reduced by no more than about 2.5%, no more than about 5%, or no more than about 7.5% relative to the baseline target mRNA measured value. In some embodiments, the target mRNA measured value is reduced by no more than about 10% relative to the baseline target mRNA measured value. In some embodiments, the target mRNA measured value is no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100% relative to the baseline target mRNA measured value. In some embodiments, the target mRNA measured value is reduced by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or the scope defined by any two of the above percentages. The target mRNA measured value can be reduced in a long period of time. In some embodiments, the target mRNA measured value is reduced by 7 days, 14 days, 28 days, 42 days, 56 days, 70 days, 77 days, 84 days, 91 days, 98 days, 105 days or the scope therebetween. In some embodiments, the target protein measurement is reduced 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 reducing RNA measurements other than mRNA measurements.

[0351] III. Definitions

[0352] Unless otherwise defined, all technical terms, symbols, and other technical and scientific terms or terminology used herein are intended to have the same meaning as commonly understood by those of ordinary skill in the art to which the claimed subject matter belongs. In some cases, terms with commonly understood meanings are defined herein for clarity and / or for ease of reference, and such definitions contained herein should not necessarily be construed as indicating a substantial difference from what is generally understood in the art.

[0353] Throughout this application, various embodiments can be presented in the form of ranges. It should be understood that the description of the range form is only for convenience and brevity, and should not be interpreted as a hard limit to the scope of the present disclosure. Therefore, the description of the range should be regarded as having all possible sub-ranges disclosed and the individual numerical values ​​within this range. For example, the description of a range such as 1 to 6 should be regarded as having the sub-ranges disclosed, such as 1 to 3, 1 to 4, 1 to 5, 2 to 4, 2 to 6, 3 to 6, etc., and the individual numbers within this range, such as 1, 2, 3, 4, 5 and 6. Regardless of the width of the scope, this applies.

[0354] As used in the specification and claims, the singular forms "a", "an", and "the" include plural referents unless the context clearly dictates otherwise. For example, the term "a sample" includes a plurality of samples, including mixtures thereof.

[0355] Certain examples involve sequences. If there is a conflict between the sequence listing and the disclosure in the specification, the specification shall prevail.

[0356] The terms "determine," "measure," "evaluate," "assess," "measure," and "analyze" are generally used interchangeably herein to refer to forms of measurement. The terms include determining whether an element is present (e.g., detecting). These terms may include quantitative, qualitative, or quantitative and qualitative determinations. Assessments may be relative or absolute. In addition to determining whether something is present or absent based on context, "detecting the presence of" may also include determining the amount of something that is present.

[0357] The terms "subject" and "patient" are used interchangeably herein. A "subject" can be a biological entity containing expressed genetic material. A biological entity can be a plant, an animal, or a microorganism, including, for example, bacteria, viruses, fungi, and protozoa. A subject can be a mammal. A mammal can be a human. A subject may be diagnosed or suspected of being at high risk for a disease. In some cases, a subject is not necessarily diagnosed or suspected of being at high risk for a disease.

[0358] As used herein, the term "about" a number refers to the number plus or minus 10% of the number. The term "about" a range refers to the range minus 10% of its lower value and plus 10% of its upper value.

[0359] As used herein, the term "treatment" is used to refer to a drug or other intervention regimen for obtaining a beneficial or desired result in a recipient. Beneficial or desired results include, but are not limited to, therapeutic benefits and / or preventive benefits. Therapeutic benefits may refer to eradicating or improving the underlying condition or its symptoms being treated. In addition, therapeutic benefits may be achieved by eradicating or improving one or more physiological symptoms associated with the underlying condition so that improvements are observed in the subject, although the subject may still be troubled by the underlying condition. Preventive effects include delaying, preventing or eliminating the appearance of a disease or condition, delaying or eliminating the onset of symptoms of a disease or condition, slowing down, stopping or reversing the progress of a disease or condition, or any combination thereof. For preventive benefits, treatment may be performed on an object at risk of a specific disease or on an object reporting one or more physiological symptoms of a disease (even if the diagnosis of the disease may not yet have been made).

[0360] "Treatment" or "treatment" may include measures to obtain beneficial or desired results (including but not limited to therapeutic benefits and / or preventive benefits) for a disease, disorder, or medical condition. Therapeutic benefits can include, for example, eradication or improvement of the underlying disorder being treated. In addition, therapeutic benefits can also include, for example, eradication or improvement of one or more physiological symptoms associated with the underlying disorder, such that improvement is observed in the subject, even though the subject may still suffer from the underlying disorder. In certain embodiments, for preventive 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 though the disease has not yet been diagnosed. Treatment by administering the compounds described herein does not necessarily require the involvement of a medical professional.

[0361] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

[0362] When the term "C x-y " or "C x -C y " when used with a chemical moiety such as alkyl, alkenyl or alkynyl, means a chain group containing x to y carbons. For example, the term "C 1-6 "Alkyl" refers to substituted or unsubstituted saturated hydrocarbon groups, including straight chain alkyl and branched chain alkyl groups containing 1 to 6 carbon atoms. The term "C x-y " or "C x -C y " does not limit the number of carbon atoms that can be attached to a chemical moiety when the chemical moiety is substituted with a second chemical group. For example, the term "C 1-6 "C1 to C6 alkyl" refers to a saturated, substituted or unsubstituted hydrocarbon group, including straight-chain alkyl groups (e.g., linear alkyl groups) and branched alkyl groups containing 1, 2, 3, 4, 5 or 6 carbon atoms, as well as C1 to C6 alkyl groups.1-6 How many carbon atoms can be present in any substituent of the alkyl group? 1-6 The alkyl group is optionally substituted with a second chemical group comprising two carbon atoms, then it is understood that the C 1-6 The alkyl group may contain 1 to 8 carbon atoms.

[0363] The term "C x-y "Alkenyl" and "C x-y "Alkynyl" refers to a substituted or unsubstituted unsaturated aliphatic group similar in length and possible substitution to the alkyl described above, but containing at least one double or triple bond, respectively.

[0364] "Amino" refers to a -NH2 moiety.

[0365] "Cyano" refers to a -CN moiety.

[0366] "Nitro" refers to the -NO2 moiety.

[0367] "Oxa" refers to the moiety -O-.

[0368] "Oxo" refers to the moiety =0.

[0369] "Thio" refers to the =S moiety.

[0370] "Imino" refers to a =NH moiety.

[0371] "Oximo" refers to a =N-OH moiety.

[0372] "Hydrazino" refers to the moiety =N-NH2.

[0373] "Alkyl" refers to a fully saturated straight or branched chain hydrocarbon moiety consisting only of carbon and hydrogen atoms. In some embodiments, "alkyl" contains from one to fifteen carbon atoms (e.g., C1-C 15 In some embodiments, the alkyl group contains from one to thirteen carbon atoms (e.g., C1-C 13 In some embodiments, the alkyl group comprises one to eight carbon atoms (e.g., C1-C8 alkyl). In some embodiments, the alkyl group comprises one to six carbon atoms (e.g., C1-C6 alkyl). In other embodiments, the alkyl group comprises one to five carbon atoms (e.g., C1-C5 alkyl). In other embodiments, the alkyl group comprises one to four carbon atoms (e.g., C1-C4 alkyl). In other embodiments, the alkyl group comprises one to three carbon atoms (e.g., C1-C3 alkyl). In other embodiments, the alkyl group comprises one to two carbon atoms (e.g., C1-C2 alkyl). In other embodiments, the alkyl group comprises one carbon atom (e.g., C1 alkyl, e.g., methyl). In other embodiments, the alkyl group comprises five to fifteen carbon atoms (e.g., C5-C15 In other embodiments, the alkyl group comprises five to eight carbon atoms (e.g., C5-C8 alkyl). In other embodiments, the alkyl group comprises two to five carbon atoms (e.g., C2-C5 alkyl). In other embodiments, the alkyl group comprises three to five carbon atoms (e.g., C3-C5 alkyl). In other embodiments, the alkyl group comprises methyl, ethyl, 1-propyl (n-propyl), 1-methylethyl (2-propyl, isopropyl), 1-butyl (n-butyl), 1-methylpropyl (sec-butyl), 2-methylpropyl (isobutyl), 1,1-dimethylethyl (tert-butyl), and 1-pentyl (n-pentyl). The alkyl group is attached to the rest of the molecule by a single bond.

[0374] "Aminoalkyl" refers to a moiety bonded through a nitrogen atom and is of the form -N(H)(alkyl) or N(alkyl)(alkyl), wherein when the moiety is N(alkyl)(alkyl), the two alkyl groups bonded to the nitrogen can be the same alkyl group or different alkyl groups.

[0375] "Alkoxy" refers to a moiety of the formula -O-alkyl bonded through an oxygen atom, wherein alkyl is an alkyl chain as defined above.

[0376] "Alkenyl" refers to a straight or branched hydrocarbon group consisting only of carbon and hydrogen atoms, containing at least one carbon-carbon double bond. In some embodiments, the alkenyl group contains two to twelve carbon atoms. In some embodiments, the alkenyl group contains two to eight carbon atoms. In other embodiments, the alkenyl group contains two to four carbon atoms. The alkenyl group is attached to the rest of the molecule by a single bond, such as vinyl (i.e., ethylene), prop-1-enyl (i.e., allyl), but-1-enyl, pent-1-enyl, pent-1,4-dienyl, etc.

[0377] "Alkynyl" refers to a straight or branched hydrocarbon group consisting only of carbon and hydrogen atoms, containing at least one carbon-carbon triple bond, having two to twelve carbon atoms, and optionally further containing at least one carbon-carbon double bond. In some embodiments, the alkynyl group contains two to eight carbon atoms. In other embodiments, the alkynyl group contains two to six carbon atoms. In other embodiments, the alkynyl group contains two to four carbon atoms. The alkynyl group is attached to the rest of the molecule by a single bond, such as ethynyl, propynyl, butynyl, pentynyl, hexynyl, etc.

[0378] "Alkylene" or "alkylene chain" refers to a linear (e.g., straight chain) or branched divalent hydrocarbon group. "Alkylene" or "alkylene chain" can connect a certain part of a molecule to a second part. "Alkylene" or "alkylene chain" consists only of carbon and hydrogen atoms (it can be specified that alkylene is replaced by one or more substituents containing atoms other than hydrogen such as nitrogen, oxygen and sulfur). "Alkylene" or "alkylene chain" may not contain unsaturation (although alkylene has an attachment point to the rest of the molecule). In some embodiments, "alkylene" or "alkylene chain" comprises one to twelve carbon atoms, such as methylene, ethylene, propylene, n-butane, etc. Alkylene chain can be attached to a certain part of a molecule by a single bond, and attached to the second part by a single bond. The attachment point of an alkylene chain to the rest of the molecule and the second part can be through a carbon in the alkylene chain or through any two carbons in the alkylene. In some embodiments, an alkylene comprises one to eight carbon atoms (e.g., C1-C8 alkylene). In other embodiments, the alkylene group comprises one to five carbon atoms (e.g., C1-C5 alkylene). In other embodiments, the alkylene group comprises one to four carbon atoms (e.g., C1-C4 alkylene). In other embodiments, the alkylene group comprises one to three carbon atoms (e.g., C1-C3 alkylene). In other embodiments, the alkylene group comprises one to two carbon atoms (e.g., C1-C2 alkylene). In other embodiments, the alkylene group comprises one carbon atom (e.g., C1 alkylene). In other embodiments, the alkylene group comprises five to eight carbon atoms (e.g., C5-C8 alkylene). In other embodiments, the alkylene group comprises two to five carbon atoms (e.g., C2-C5 alkylene). In other embodiments, the alkylene group comprises three to five carbon atoms (e.g., C3-C5 alkylene).

[0379] "Alkenylene" or "alkenylene chain" refers to a linear (e.g., straight chain) or branched divalent hydrocarbon moiety."Alkenylene" or "alkenylene chain" can connect a portion of a molecule to a second portion."Alkenylene" or "alkenylene chain" consists only of carbon and hydrogen atoms (alkenylene can be specified to be substituted by one or more substituents comprising atoms other than hydrogen such as nitrogen, oxygen and sulfur). "Alkenylene" or "alkenylene chain" includes at least one carbon-carbon double bond. In certain embodiments, "alkenylene" or "alkenylene chain" includes two to twelve carbon atoms. The alkenylene chain can be attached to the portion of the molecule by a single bond and is attached to the second portion by a single bond. The attachment point of the alkenylene chain to the other portions of the molecule and the second portion can be through one carbon in the alkenylene chain or any two carbons in the alkenylene chain. In certain embodiments, the alkenylene includes two to eight carbon atoms (e.g., C2-C8 alkenylene). In other embodiments, the alkenylene includes two to five carbon atoms (e.g., C2-C5 alkenylene). In other embodiments, the alkenylene group comprises two to four carbon atoms (e.g., C2-C4 alkenylene). In other embodiments, the alkenylene group comprises two to three carbon atoms (e.g., C2-C3 alkenylene). In other embodiments, the alkenylene group comprises five to eight carbon atoms (e.g., C5-C8 alkenylene). In other embodiments, the alkenylene group comprises two to five carbon atoms (e.g., C2-C5 alkenylene). In other embodiments, the alkenylene group comprises three to five carbon atoms (e.g., C3-C5 alkenylene).

[0380] "Alkynylidene" or "alkynylene chain" refers to a linear (e.g., straight chain) or branched divalent hydrocarbon moiety. "Alkynylidene" or "alkynylene chain" can connect a portion of a molecule to a second portion. "Alkynylidene" or "alkynylene chain" consists only of carbon and hydrogen (alkynylidene can be specified to be substituted by one or more substituents comprising atoms other than hydrogen such as nitrogen, oxygen and sulfur). "Alkynylidene" or "alkynylene chain" includes at least one carbon-carbon triple bond. In certain embodiments, "alkynyllidene" or "alkynylene chain" includes two to twelve carbon atoms. Alkynylidene chain can be attached to the portion of the molecule by a single bond and attached to the second portion by a single bond. The attachment point of the alkynylene chain to the rest of the molecule and the second portion can be through a carbon atom in the alkynylene chain or through any two carbon atoms in the alkynylene chain. In certain embodiments, the alkynylene includes two to eight carbon atoms (e.g., C2-C8 alkynylene). In other embodiments, the alkynylene includes two to five carbon atoms (e.g., C2-C5 alkynylene). In other embodiments, the alkynylene group comprises two to four carbon atoms (e.g., C2-C4 alkynylene). In other embodiments, the alkynylene group comprises two to three carbon atoms (e.g., C2-C3 alkynylene). In other embodiments, the alkynylene group comprises two carbon atoms (e.g., C2 alkenylene). In other embodiments, the alkynylene group comprises five to eight carbon atoms (e.g., C5-C8 alkynylene). In other embodiments, the alkynylene group comprises three to five carbon atoms (e.g., C3-C5 alkynylene).

[0381] As used herein, the term "carbocycle" refers to a saturated ring, an unsaturated ring or an aromatic ring, wherein each atom of the ring is carbon.Carbocycles include 3-10-membered monocycles, 5-12-membered bicyclic rings, 5-12-membered spiral bicyclic rings and 5-12-membered bridged rings.Each ring of the bicyclic carbocycle can be selected from saturated rings, unsaturated rings and aromatic rings.In an exemplary embodiment, an aromatic ring, such as a phenyl, can be fused with a saturated or unsaturated ring, such as cyclohexane, cyclopentane or cyclohexene.Where valence permits, the bicyclic carbocycle includes any combination of saturated bicyclic rings, unsaturated bicyclic rings and aromatic bicyclic rings.Bicyclic carbocycles also include spiral bicyclic rings, such as spiropentane. Bicyclic carbocycles include any combination of ring sizes, such as 3-3 spiro ring system, 4-4 spiro ring system, 4-5 fused ring system, 5-5 fused ring system, 5-6 fused ring system, 6-6 fused ring system, 5-7 fused ring system, 6-7 fused ring system, 5-8 fused ring system and 6-8 fused ring system. Exemplary carbocycles include cyclopentyl, cyclohexyl, cyclohexenyl, adamantyl, phenyl, indenyl, naphthyl and bicyclo[1.1.1]pentanyl. "Carbocycle" can include "aryl" and "cycloalkyl".

[0382] The term "aryl" refers to an aromatic monocyclic or polycyclic hydrocarbon ring system. An aromatic monocyclic or polycyclic hydrocarbon ring system contains only hydrogen and carbon and contains from 5 to 18 carbon atoms, wherein at least one ring in the ring system is aromatic, i.e., it contains a cyclic, delocalized (4n+2)π-electron system according to the 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 carries a positive or negative charge. In some embodiments, the aryl substituent is neutral. In some embodiments, the aryl substituent is zwitterionic; alternatively or additionally, in some embodiments, the aryl substituent is uncharged. In some embodiments, the aryl substituent is uncharged. In some embodiments, the aryl substituent has no net charge. In some embodiments, the aryl substituent has no net charge and is not zwitterionic.

[0383] The term "cycloalkyl" refers to a saturated ring in which each atom of the ring is carbon. Cycloalkyl can include monocycles and polycycles, such as 3-10 monocycles, 5-12 bicycles, 5-12 spiral bicycles, and 5-12 bridged rings. In certain embodiments, cycloalkyl comprises 3 to 10 carbon atoms. In other embodiments, cycloalkyl comprises 5 to 7 carbon atoms. Cycloalkyl 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 groups include, for example, adamantyl, spiropentane, bornyl (i.e., bicyclo [2.2.1] heptyl), decahydronaphthyl, 7,7-dimethylbicyclo [2.2.1] heptyl, bicyclo [1.1.1] pentyl, etc.

[0384] The term "cycloalkenyl" refers to a saturated ring, wherein each atom of the ring is carbon, and there is at least one double bond between two ring carbons. Cycloalkenyl can include monocycles and polycycles, such as 3 to 10 monocycles, 6 to 12 bicycles, and 5 to 12 bridged rings. In other embodiments, the cycloalkenyl comprises 5 to 7 carbon atoms. The cycloalkenyl 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.

[0385] The term "halo" or alternatively "halogen" or "halide" means fluoro, chloro, bromo or iodo. In some embodiments, halo is fluoro, chloro or bromo.

[0386] The term "haloalkyl" refers to an alkyl group as defined above, substituted with one or more halo groups, for example, trifluoromethyl, dichloromethyl, bromomethyl, 2,2,2-trifluoroethyl, 1-chloromethyl, 2-fluoroethyl, etc. In some embodiments, the alkyl portion of the haloalkyl is optionally further substituted as described herein.

[0387] As used herein, the term "heterocycle" refers to a saturated ring, an unsaturated ring or an aromatic ring containing one or more heteroatoms. Exemplary heteroatoms include N, O, Si, P, B and S atoms. Heterocycles include 3-10-membered monocycles, 6-12-membered bicyclic rings, 5-12-membered spiral bicyclic rings and 5-12-membered bridged rings. Where valence permits, bicyclic heterocycles include any combination of saturated bicyclic rings, unsaturated bicyclic rings and aromatic bicyclic rings. In an exemplary embodiment, an aromatic ring, such as a pyridyl group, can be fused with a saturated or unsaturated ring, such as 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 also include spiro bicycles, for example 5- to 12-membered spiro bicycles, such as 2-oxa-6-azaspiro[3.3]heptane. "Heterocycle" may include "heteroaryl" and "heterocycloalkyl".

[0388] The term "heteroaryl" refers to a group derived from a 5- to 18-membered aromatic ring group containing 2 to 17 carbon atoms and 1 to 6 heteroatoms selected from nitrogen, oxygen and sulfur. As used herein, a heteroaryl group is a monocyclic, bicyclic, tricyclic or tetracyclic ring system, wherein at least one ring in the ring system is aromatic, i.e., according to the Hückel theory, it contains a cyclic, delocalized (4n+2) π-electron system. Heteroaryl includes fused or bridged ring systems. One or more heteroatoms in the heteroaryl 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 groups include, but are not limited to, azepine, 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, benzopyrone, benzofuranyl, benzofuranone, benzothiophenyl, benzothieno[3,2-d]pyrimidinyl, benzotriazolyl, benzo[4,6]imidazo[1,2-a ]pyridyl, carbazolyl, cinnolinyl, cyclopenta[d]pyrimidinyl, 6,7-dihydro5H-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]cyclopenta[1,2-c]pyridazinyl, dibenzofuranyl, dibenzothiophenyl, furanyl, furanonyl, furano[3,2-c]pyridinyl, 5,6,7,8,9,10-hexahydrocyclooctatetraeno[d]pyrimidinyl, 5,6,7,8,9,10-hexahydrocyclooctatetraeno[d]pyridazinyl, 5,6,7,8,9,10-hexahydrocyclooctatetraeno[d]pyridazinyl, tetraenol[d]pyridinyl, isothiazolyl, imidazolyl, indazolyl, indolyl, indazolyl, isoindolyl, indolyl, isoindolyl, isoquinolyl, indolizinyl, isoxazolyl, 5,8-methano-5,6,7,8-tetrahydroquinazolinyl, naphthyridinyl, 1,6-naphthyridinonyl, oxadiazolyl, 2-oxoazepine, 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, pyrido[3,4-d]pyrimidinyl, pyrazinyl, pyrimidinyl, pyridazinyl, pyrrolyl, quinazolinyl, quinoxalinyl, quinolinyl, isoquinolinyl, tetrahydroquinolinyl, 5,6,7,8-tetrahydroquinazolinyl, 5,6,7,8-tetrahydrobenzo[4,5]thieno[2,3-d]pyrimidinyl, 6,7,8,9-tetrahydro5Hcyclopenta[4,5]thieno[2,3-d]pyrimidinyl, 5,6,7,8-tetrahydropyrido[4,5-c]pyridazinyl, thiazolyl, thiadiazolyl, triazolyl, tetrazolyl, triazinyl, thieno[2,3d]pyrimidinyl, thieno[3,2-d]pyrimidinyl, thieno[2,3-c]pyridinyl, and thienyl.

[0389] If one or more nitrogen atoms are present, they may be selectively quaternized. In some embodiments, the heterocyclic substituent carries a positive or negative charge. In some embodiments, the heterocyclic substituent is neutral. In some embodiments, the heterocyclic substituent is zwitterionic; alternatively or additionally, in some embodiments, the heterocyclic substituent is uncharged. In some embodiments, the heterocyclic substituent is uncharged. In some embodiments, the heterocyclic substituent has no net charge. In some embodiments, the heterocyclic substituent has no net charge and is not zwitterionic.

[0390] The term "heterocycloalkyl" refers to a saturated ring with carbon atoms and at least one heteroatom. Exemplary heteroatoms include N, O, Si, P, B and S atoms. Heterocycloalkyl can include monocycles and polycycles, such as 3 to 10 monocycles, 6 to 12 bicycles, 5 to 12 spiral bicycles and 5 to 12 bridged rings. The heteroatoms in the heterocycloalkyl group are optionally oxidized. One or more nitrogen atoms (if present) are optionally quaternized. Where valence permits, heterocycloalkyl is attached to the remainder of the molecule by any atom of heterocycloalkyl (such as any carbon or nitrogen atom of heterocycloalkyl). Examples of heterocycloalkyl groups include, but are not limited to, dioxolane, thienyl [1,3] dithianyl, decahydroisoquinolinyl, imidazolinyl, imidazolidinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl, piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl, thiazolidinyl, tetrahydrofuranyl, trithianyl, tetrahydropyranyl, thiomorpholinyl, thiamorpholinyl, 1-oxothiomorpholinyl, 2-oxa-6-azaspiro [3.3] heptane, and 1,1-dioxothiomorpholinyl. In some embodiments, the heterocycloalkyl group contains one heteroatom. In some embodiments, the heterocycloalkyl group contains one heteroatom selected from N, O, and S. In some embodiments, the heterocycloalkyl group comprises multiple heteroatoms. In some embodiments, the heterocycloalkyl group comprises multiple heteroatoms selected from N, O, and S.

[0391] The term "heterocycloalkenyl" refers to an unsaturated ring having carbon atoms and at least one heteroatom, and at least one double bond between two ring carbons. Heterocycloalkenyl does not include heteroaryl rings. Exemplary 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, the heterocycloalkenyl includes 5 to 7 ring atoms. The heterocycloalkenyl can be attached to the remainder of the molecule by a single bond. Examples of monocyclic cycloalkenyl groups include, for example, pyrroline (dihydropyrrole), pyrazoline (dihydropyrazole), imidazoline (dihydroimidazole), triazoline (dihydrotriazole), dihydrofuran, dihydrothiophene, oxazoline (dihydrooxazole), isoxazoline (dihydroisoxazole), thiazoline (dihydrothiazole), isothiazolin (dihydroisothiazole), oxadiazolin (dihydrooxadiazole), thiadiazoline (dihydrothiadiazole), dihydropyridine, tetrahydropyridine, dihydropyridazine, tetrahydropyridazine, dihydropyrimidine, tetrahydropyrimidine, dihydropyrazine, tetrahydropyrazine, pyran, dihydropyran, thiopyran, dihydrothiopyran, dioxadiene, dihydrodioxine, oxazine, dihydrooxazine, thiazine, and dihydrothiazine.

[0392] The term "substituted" refers to a substituent having a hydrogen or substitutable heteroatom (e.g., NH or NH2 of a compound) that replaces one or more carbons. It should be understood that "substituted" or "substituted by..." includes implicit prerequisites, i.e., such substitutions meet the allowed valences of the substituted atom and the substituent, and the substitution produces a stable compound, i.e., a compound that does not spontaneously undergo transformations such as rearrangement, cyclization, elimination, etc. In certain embodiments, substituted refers to a substituent having a substituent that replaces two hydrogen atoms on the same carbon atom, such as replacing two hydrogen atoms on a single carbon with an oxo group, an imino group, or a thio group. As used herein, it is contemplated that the term "substituted" includes all allowed substituents of an organic compound. In a broad sense, allowed substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and non-aromatic substituents of an organic compound. For appropriate organic compounds, allowed substituents may be one or more and may be identical or different.

[0393] In some embodiments, the substituents may include any of the substituents described herein, for example, halogen, hydroxyl, oxo (=O), thio (=S), cyano (-CN), nitro (-NO2), imino (=NH), oxime (=N-OH), hydrazine (=N-NH2), -R b -OR a , -R b -OC(O)-R a , -R b -OC(O)-OR a , -R b -OC(O)-N(Ra 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 (where t is 1 or 2), -R b -S(O) t R a (where t is 1 or 2), -R b -S(O) t OR a (where t is 1 or 2) and -R b -S(O) t N(R a )2 (wherein t is 1 or 2); and alkyl, alkenyl, alkynyl, aryl, aralkyl, aralkenyl, aralkynyl, cycloalkyl, cycloalkylalkyl, heterocycloalkyl, heterocycloalkylalkyl, heteroaryl and heteroarylalkyl, each of which may be optionally substituted by alkyl, alkenyl, alkynyl, halogen, haloalkyl, haloalkenyl, haloalkynyl, oxo (=O), thio (=S), cyano (-CN), nitro (-NO2), imino (=NH), oxime (=N-OH), hydrazine (=N-NH2), -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 (where t is 1 or 2), -R b -S(O) t R a (where t is 1 or 2), -R b -S(O) t OR a (where t is 1 or 2) and -R b -S(O) t N(R a )2 (wherein t is 1 or 2); wherein each R a is independently selected from hydrogen, alkyl, cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl, heterocycloalkylalkyl, heteroaryl or heteroarylalkyl, wherein each R a The alkyl radical may be optionally substituted with alkyl, alkenyl, alkynyl, halogen, haloalkyl, haloalkenyl, haloalkynyl, oxo (=O), thio (=S), cyano (-CN), nitro (-NO2), imino (=NH), oxime (=N-OH), hydrazine (=N-NH2), -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(Ra )C(O)OR a , -R b -N(R a )C(O)R a , -R b -N(R a )S(O) t R a (where t is 1 or 2), -R b -S(O) t R a (where t is 1 or 2), -R b -S(O) t OR a (where t is 1 or 2) and -R b -S(O) t N(R a )2 (wherein t is 1 or 2); and wherein each R b are independently selected from a direct bond or a straight or branched alkylene, alkenylene or alkynylene chain, and each R c is a straight or branched alkylene, alkenylene or alkynylene chain.

[0394] Double bonds to oxygen atoms, such as oxo groups, are represented herein as both "=O" and "(O)". Double bonds to nitrogen atoms are represented as both "=NR" and "(NR)". Double bonds to sulfur atoms are represented as both "=S" and "(S)".

[0395] As used herein, the phrases "parenteral administration" and "administered parenterally" refer to 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, subcutaneous, intraarticular, subcapsular, subarachnoid, intraspinal, and intrasternal injection and infusion.

[0396] The phrase "pharmaceutically acceptable" as used herein refers to compounds, materials, compositions and / or dosage forms that are, within the scope of sound medical judgment, suitable for contact with the tissues of humans and animals without excessive toxicity, irritation, allergic response or other problems or complications, and are commensurate with a reasonable benefit / risk ratio.

[0397] As used herein, the phrase "pharmaceutically acceptable excipient" or "pharmaceutically acceptable carrier" means a pharmaceutically 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 materials that can serve as pharmaceutically 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, ethyl cellulose, and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients such as cocoa butter and suppository waxes; (9) oils such as peanut oil, cottonseed oil, , safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols such as propylene glycol; (11) polyols such as glycerol, 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) ethanol; (20) phosphate buffer solution; and (21) other nontoxic compatible substances used in pharmaceutical preparations.

[0398] The term "salt" or "pharmaceutically acceptable salt" refers to salts derived from a variety of 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, etc. 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, etc. Pharmaceutically acceptable base 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, etc. Organic bases from which salts can be derived include, for example, primary, secondary and tertiary amines, substituted amines (including naturally occurring substituted amines), cyclic amines, basic ion exchange resins, and the like, particularly such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine and ethanolamine. In some embodiments, pharmaceutically acceptable base addition salts are selected from ammonium, potassium, sodium, calcium and magnesium salts.

[0399] VI. Examples

[0400] Example 1: Identification of variants in target oligonucleotides associated with increased or decreased risk of a disorder

[0401] Approximately 30,000,000 putative variants from approximately 375,000 individuals in the Biobank cohort will be analyzed to investigate their association with liver conditions such as nonalcoholic steatohepatitis.

[0402] Protective or maladaptive associations were observed between specific allelic variants of many target genes and liver disorders. These associations would suggest that, in some cases, therapeutic inhibition or modulation of the target protein encoded by any target gene may be an effective genetically informed approach to treat any liver disorder.

[0403] Example 2: Bioinformatic selection of sequences to identify therapeutic siRNAs that downregulate expression of target mRNAs

[0404] Screening set will be defined based on bioinformatics analysis.Therapeutic siRNA is designed to bind to human target mRNA and target mRNA sequence of at least one toxicology-related species, such as non-human primate (NHP) species (e.g., rhesus monkey or cynomolgus monkey).The driving factor of screening set design is the predicted specificity of siRNA to the transcriptome of related species and the cross-reactivity between species.The predicted specificity of sense strand (S) and antisense strand (AS) in people, rhesus monkey, cynomolgus monkey, mouse and rat is determined.These are assigned "specificity score", which takes into account the possibility of accidentally lowering any other transcript by the complete or partial complementarity of siRNA strand (up to 4 mispairings in position 2-18) and the number and position of mispairing.Therefore, the miss-target of antisense strand and sense strand of every kind of siRNA is identified.In addition, the number of possible miss-targets is used as the additional specificity factor in the specificity score.As identified, siRNA with high specificity and a small amount of predicted miss-targets provides the benefit of increased targeting specificity.

[0405] In addition to selecting siRNA sequences with high sequence specificity to target mRNA, the similarity of the siRNA sequence in the seed region to the seed region of known miRNA is also analyzed. siRNA can play a role in a miRNA-like manner via base pairing with the complementary sequence in the 3'-UTR of the mRNA molecule. Complementarity generally covers the 5'-base at the position 2-7 (seed region) of miRNA. In order to avoid that siRNA works via functional miRNA binding sites, the siRNA chain containing the natural miRNA seed region is avoided. The seed region identified in the miRNA from people, mice, rats, rhesus monkeys, dogs, rabbits and pigs is referred to as "conserved". "Specificity score" is combined with miRNA seed analysis to produce a "specificity category". This is divided into categories 1-4, wherein 1 has the highest specificity, and 4 has the lowest specificity. Each chain of siRNA is assigned to a specificity category.

[0406] Species cross-reactivity was assessed for humans, cynomolgus monkeys, rhesus monkeys, mice, and rats. The analysis was based on classical siRNA design using 19 bases and 17 bases (positions 1 and 19 were not considered) for cross-reactivity. Both perfect match analysis and single mismatch analysis were included.

[0407] A human single nucleotide polymorphism (SNP) database (NCBI-DB-SNP) analysis for identifying siRNA targeting regions with known SNPs was also performed to identify siRNAs that may not be functional in individuals containing the SNPs. Information about the position of the SNP within the target sequence and the minor allele frequency (MAF) in the case data was obtained in this analysis.

[0408] The above methods can be used to identify therapeutic siRNAs that downregulate expression of a target mRNA. Bioinformatics methods can also be used to identify ASOs that bind to and downregulate expression of a target mRNA.

[0409] Example 3: Chemically modified siRNA

[0410] The siRNA binding to the target mRNA can be synthesized with chemical modifications, wherein the sense strand has a modification such as modification pattern 1S, and the antisense strand has a modification such as modification pattern 1AS. In addition, adenosine can be located at position 19 of the sense strand, and uridine can be located at position 1 of the antisense strand.

[0411] siRNAs that bind to target mRNAs can also be synthesized with chemical modifications, wherein the sense strand has a modification pattern of 2S and the antisense strand has a modification pattern of 3AS. In addition, adenosine can be located at position 19 of the sense strand and uridine at position 1 of the antisense strand.

[0412] siRNAs that bind to target mRNAs can also be synthesized with chemical modifications, wherein the sense strand has a modification pattern of 2S and the antisense strand has a modification pattern of 9AS. In addition, adenosine can be located at position 19 of the sense strand and uridine at position 1 of the antisense strand.

[0413] siRNA targeting binding to target mRNA can also be synthesized with chemical modifications, wherein the sense strand has a modification pattern of 3S and the antisense strand has a modification pattern of 3AS. In addition, adenosine can be located at position 19 of the sense strand and uridine at position 1 of the antisense strand.

[0414] siRNA targeting that binds to a target mRNA can also be synthesized with chemical modifications in which the sense strand has any of the following: all purines comprising 2' fluoro-modified purines, and all pyrimidines comprising a mixture of 2' fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines; all purines comprising 2'-O-methyl-modified purines, and all pyrimidines comprising a mixture of 2' fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines; all purines comprising 2' fluoro-modified purines, and all pyrimidines comprising 2'-O-methyl-modified pyrimidines; all pyrimidines comprising 2' fluoro-modified pyrimidines, and all purines comprising a mixture of 2' fluoro-modified purines and 2'-O-methyl-modified pyrimidines; all pyrimidines comprising 2'-O-methyl-modified pyrimidines, and all purines comprising a mixture of 2' fluoro-modified purines and 2'-O-methyl-modified pyrimidines; or all pyrimidines comprising 2' fluoro-modified pyrimidines, and all pyrimidines comprising 2'-O-methyl-modified pyrimidines. all purines containing 2'-fluoro-modified purines; and further wherein the antisense strand has any of the following: all purines containing 2'-fluoro-modified purines, and all pyrimidines containing a mixture of 2'-fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines; all purines containing 2'-O-methyl-modified purines, and all pyrimidines containing a mixture of 2'-fluoro-modified pyrimidines and 2'-O-methyl-modified pyrimidines; all purines containing 2'-O-methyl-modified purines, and all pyrimidines containing 2'-fluoro-modified purines; all pyrimidines containing 2'-fluoro-modified pyrimidines, and all purines containing a mixture of 2'-fluoro-modified purines and 2'-O-methyl-modified purines; all pyrimidines containing 2'-O-methyl-modified pyrimidines, and all purines containing a mixture of 2'-fluoro-modified purines and 2'-O-methyl-modified purines; or all pyrimidines containing 2'-O-methyl-modified pyrimidines, and all purines containing 2'-fluoro-modified purines.

[0415] Example 4: Screening for siRNA activity in cells in culture

[0416] Chemically modified siRNAs derived from the sequences in the previous examples will be assayed for target mRNA knockdown activity in cultured cells. Cell lines expressing target mRNAs were seeded in 96-well tissue culture plates at a cell density of 10,000 cells / well in DMEM supplemented with 10% fetal bovine serum and incubated overnight in a water-jacketed humidified incubator at 37°C in an atmosphere consisting of air plus 5% carbon dioxide. siRNAs were transfected individually into cells in duplicate wells at a final concentration of 10 nM using 0.3 μL Lipofectamine RNAiMax (Fisher) / well. Silencer Select negative control #1 (ThermoFisher, catalog number 4390843) and positive control siRNA were transfected at a final concentration of 10 nM as controls. After incubation for 48 hours at 37°C, total RNA was harvested from each well and used according to the manufacturer's instructions. Fast Advanced Cells-to-CT TM cDNA was prepared using a kit (ThermoFisher, catalog number A35374). The level of target mRNA in each well was measured in triplicate by real-time qPCR on an Applied Biosystems 7500 fast real-time PCR instrument using TaqMan gene expression assays for human target mRNA. The level of PPIA mRNA was measured using TaqMan gene expression assays (ThermoFisher) and used to determine the relative target mRNA level in each well using the Δ-ΔCt method. The data were normalized for the relative target mRNA levels in untreated cells.

[0417] The siRNAs that show the highest degree of target mRNA knockdown at 10 nM will be tested in a second activity screen at 1 nM concentration using the transfection procedure described above. Similar experiments can be performed using ASOs. Thus, the siRNAs and ASOs that most effectively downregulate target mRNA expression can be identified.

[0418] Example 5: GalNAc ligands for hepatocyte targeting of oligonucleotides

[0419] The present disclosure is not limited to these individual methods, and there are at least two general methods for attaching multivalent N-acetylgalactosamine (GalNAc) ligands to oligonucleotides: solid phase or solution phase conjugation. The GalNAc ligand can be attached to a solid phase resin for 3' conjugation, or it can be attached at the 5' end using a GalNAc phosphoramidite reagent. The GalNAc phosphoramidite can be coupled on the solid phase at any position in the sequence, just like other nucleosides in the oligonucleotide sequence. Non-limiting examples of phosphoramidite reagents are shown in Table 1 for conjugating GalNAc to 5' terminal oligonucleotides.

[0420] Table 1. GalNAc conjugation reagents

[0421]

[0422] Example 6: Synthesis of GalNAc ligand

[0423] Preparation scheme of NAcegal-linker-TMSOTf

[0424]

[0425] General procedure for the preparation of compound 2A

[0426]

[0427] 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 stirred at 25°C for 2 hours under N2 atmosphere. TLC (DCM: MeOH = 20: 1, PMA) showed that CbzCl was completely consumed and a new spot (R f =0.43). HCl / EtOAc (1N, 180 mL) was added to the reaction mixture and stirred for 30 minutes. The white solid was removed by filtration through celite and the filtrate was concentrated under vacuum to obtain compound 2A (540 g, 2.26 mol, 47.5% yield) as a light yellow oil, which was used directly in the next step without further purification. 1 H NMR: δ7.28-7.41(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).

[0428] General procedure for the preparation of compound 4A

[0429]

[0430] To a solution of compound 3A (1.00 kg, 4.64 mol, HCl) in pyridine (5.00 L) was added ethyl acetate (4.73 kg, 46.4 mol, 4.34 L) dropwise under N2 atmosphere at 0°C. The mixture was stirred at 25°C for 16 hours under N2 atmosphere. TLC (DCM:MeOH=20:1, PMA) indicated 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 at 0°C for 0.5 h to form a white solid, 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 directly in the next reaction without further purification. 1 H NMR: δ7.90(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).

[0431] General procedure for the preparation of compound 5A

[0432]

[0433] To a solution of compound 4A (300 g, 771 mmol) in DCE (1.50 L) was added TMSOTf (257 g, 1.16 mol, 209 mL) and stirred at 60 °C for 2 hours and then at 25 °C for 1 hour. Compound 2A (203 g, 848 mmol) was dissolved in DCE (1.50 L) and added under N2 atmosphere. Powdered molecular sieve (150g), stirred for 30 minutes. Then at 0°C, the DCE solution of compound 4A was added dropwise to the mixture. Under N2 atmosphere, the mixture was stirred at 25°C for 16 hours. TLC (DCM: MeOH = 25: 1, PMA) indicated that compound 4A was completely consumed and a new spot (Rf = 0.24) was formed. The reaction mixture was filtered and washed with saturated NaHCO3 (2.00L), water (2.00L) and saturated brine (2.00L). The organic layer was dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to obtain a residue. The residue was ground together with 2-Me-THE / heptane (5 / 3, v / v, 1.80L) for 2 hours, filtered and dried to obtain compound 5A (225g, 389mmol, 50.3% yield, 98.4% purity), which was a white solid. 1H NMR: δ7.81(d,J=9.29Hz,1H),7.20-7.42(m,6H),5.21(d,J=3.26Hz,1H),4.9 2-5.05(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).

[0434] General procedure for the preparation of NAcegal-linker-tosylate

[0435]

[0436] To a solution of compound 5A (200 g, 352 mmol) in THF (1.0 L) was added dry Pd / C (15.0 g, 10% purity) and TsOH (60.6 g, 352 mmol) under N2 atmosphere. The suspension was degassed under vacuum and purged with H2 several times. Under H2 (45 psi) atmosphere, the mixture was stirred at 25°C for 3 hours. TLC (DCM:MeOH=10:1, PMA) indicated that compound 5A was completely consumed and a new spot (R f =0.04). The reaction mixture was filtered and concentrated under reduced pressure (≤40°C) to give a residue. The residue was washed with anhydrous DCM (500 mL, with The residue was dried over 500 molecular sieves overnight (dried at 300° C. for 12 hours) and diluted and concentrated to give a residue, which was then subjected to Karl Fischer (KF) to check the water content. The dilution and concentration with anhydrous DCM (500 mL) was repeated 3 times to give NAcegal-linker-TMSOTf (205 g, 95.8% yield, TsOH salt) as a foamy white solid. 1H NMR: δ7.91(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=11.29,3.26Hz,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).

[0437] Preparation of TRIS-PEG2-CBZ

[0438]

[0439]

[0440] General procedure for the preparation of compound 5B

[0441]

[0442] Compound 4B_2 (1.07 kg, 8.36 mol, 1.20 L, 5.00 eq) was added to a solution of compound 4B (400 g, 1.67 mol, 1.00 eq) and NaOH (10 M, 16.7 mL, 0.10 eq) in THF (2.00 L), and the mixture was stirred at 30 ° C for 2 hours. LCMS showed that the desired MS was obtained. Five batches of solution were combined into one batch, and then the mixture was diluted with water (6.00 L), extracted with ethyl acetate (3.00 L * 3), and the combined organic layer was washed with brine (3.00 L), dried over Na2SO4, filtered and concentrated under vacuum. The crude product was purified by column chromatography (SiO2, petroleum ether: ethyl acetate = 100: 1-10: 1, R f =0.5), to obtain compound 5B (2.36 kg, 6.43 mol, 76.9% yield) as a light yellow oil. HNMR: δ7.31-7.36 (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).

[0443] General procedure for the preparation of 3-oxo-1-phenyl-2,7,10-trioxa-4-azatridecane-13-oic acid (hereinafter referred to as compound 2B)

[0444]

[0445] To a solution of compound 5B (741 g, 2.02 mol, 1.00 eq) in DCM (2.80 L) was added TFA (1.43 kg, 12.5 mol, 928 mL, 6.22 eq), and the mixture was stirred at 25 ° C for 3 hours. LCMS showed that the desired MS was obtained. The mixture was diluted with DCM (5.00 L), washed with water (3.00 L * 3), brine (2.00 L), and the combined organic layers were dried over Na2SO4, filtered and concentrated under vacuum to give compound 2B (1800 g, crude) as a light yellow oil. HNMR: δ9.46(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).

[0446] General procedure for the preparation of compound 3B

[0447]

[0448] To a solution of compound 2B (375 g, 999 mmol, 83.0% purity, 1.00 eq) in DCM (1.80 L) was added HATU (570 g, 1.50 mol, 1.50 eq) and DIEA (258 g, 2.00 mol, 348 mL, 2.00 eq) at 0° C., the mixture was stirred at 0° C. for 30 minutes, then compound 1B (606 g, 1.20 mol, 1.20 eq) was added, and the mixture was stirred at 25° C. for 1 hour. LCMS showed that the desired MS was obtained. The mixture was combined into one batch, and then the mixture was diluted with DCM (5.00 L), washed with 1N HCl aqueous solution (2.00 L*2), and then the organic layer was washed with saturated Na2CO3 aqueous solution (2.00 L*2) and brine (2.00 L), the organic layer was dried over Na2SO4, filtered and concentrated under vacuum to give compound 3B (3.88 kg, crude) as a yellow oil.

[0449] General procedure for preparing TRIS-PEG2-CBZ

[0450]

[0451] A solution of compound 3B (775 g, 487 mmol, purity 50.3%, 1.00 eq) in HCl / dioxane (4 M, 2.91 L, 23.8 eq) was stirred at 25 °C for 2 hours. LCMS showed that the desired MS was obtained. The mixture was concentrated under vacuum to obtain a residue. The combined residue was then diluted with DCM (5.00 L), adjusted to pH = 8 with 2.5 M NaOH aqueous solution, and separated. The aqueous phase was extracted again with DCM (3.00 L), and the aqueous solution was then adjusted to pH = 3 with 1N HCl aqueous solution, then extracted with DCM (5.00 L*2), and the combined organic layers were washed with brine (3.00 L), dried over Na2SO4, filtered and concentrated in vacuo. The crude product was purified by column chromatography (SiO2, DCM:MeOH=0:1-12:1, 0.1% HOAc, R f =0.4). The residue was diluted with DCM (5.00 L), adjusted to pH=8 with 2.5 M NaOH aqueous solution, separated, the aqueous solution was extracted again with DCM (3.00 L), then adjusted to pH=3 with 6 N HCl aqueous solution, extracted with DCM:MeOH=10:1 (5.00 L*2), the combined organic layers were washed with brine (2.00 L), dried over Na2SO4, filtered and concentrated in vacuo to obtain a residue. The residue was then diluted with MeCN (5.00 L), concentrated in vacuo, and this procedure was repeated twice to remove water to obtain TRIS-PEG2-CBZ (1.25 kg, 1.91 mol, 78.1% yield, 95.8% purity) as a light yellow oil. 1 HNMR:400MHz,MeOD,δ

[0452] 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).

[0453] Preparation of TriNGal-TRIS-Peg2-Phosph8c

[0454]

[0455]

[0456] General procedure for the preparation of compound 3C from TriGNa1-TRIS-Peg2-Phosph 8c

[0457]

[0458] To a solution of compound 1C (155 g, 245 mmol, 1.00 eq) in ACN (1500 mL) was added TBTU (260 g, 811 mmol, 3.30 eq), DIEA (209 g, 1.62 mol, 282 mL, 6.60 eq) and compound 2C (492 g, 811 mmol, 3.30 eq, TsOH) at 0 ° C, and the mixture was stirred at 15 ° C for 16 hours. LCMS showed that the desired MS was obtained. The mixture was concentrated under vacuum to obtain a residue, and then the mixture was diluted with DCM (2000 mL), washed with 1N HCl aqueous solution (700 mL * 2), then washed with saturated NaHCO3 aqueous solution (700 mL * 2) and concentrated under vacuum. The crude product was purified by column chromatography to provide compound 3C (304 g, 155 mmol, 63.1% yield, 96.0% purity) as a yellow solid.

[0459] General procedure for the preparation of compound 4C

[0460]

[0461] 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 to two batches of solution of compound 3C (55.0 g, 29.2 mmol, 1.00 eq) in MeOH (1600 mL), and the mixture was degassed in vacuo and purged with H2. The mixture was stirred at 15 °C for 2 hours under H2 (15 psi). LCMS showed that the desired MS was obtained. The mixture was filtered and the filtrate was concentrated under vacuum to obtain compound 4C (106 g, 54.8 mmol, 93.7% yield, 96.2% purity, TFA) as a white solid.

[0462] General procedure for the preparation of compound 5C

[0463]

[0464] Two batches were carried out in parallel. Compound 4a (25.0 g, 150 mmol, 1.00 eq) was added dropwise to a solution of EDCI (28.8 g, 150 mmol, 1.00 eq) in DCM (125 mL) at 0 ° C, and then the mixture was added to compound 4 (25.0 g, 150 mmol, 1.00 eq) in DCM (125 mL) at 0 ° C, and then the mixture was stirred at 25 ° C for 1 hour. TLC (petroleum ether: ethyl acetate = 3: 1, Rf = 0.45) showed that the reactant had been consumed and a new spot was formed. The reaction mixture was diluted with DCM (100 mL), then washed with NaHCO3 aqueous solution (250 mL * 1) and brine (250 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to obtain a residue. The residue was purified by column chromatography (SiO2, petroleum ether: ethyl acetate = 100: 1 to 3: 1), TLC (SiO2, petroleum ether: ethyl acetate = 3: 1), Rf = 0.45, and then concentrated under reduced pressure to obtain a residue. Compound 5C (57.0 g, 176 mmol, 58.4% yield, 96.9% purity) was obtained as a colorless oil and purified by 1 HNMR confirmation: EW33072-2-P1A, 400 MHz, DMSOδ9.21 (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).

[0465] General procedure for the preparation of compound 6

[0466]

[0467] To a mixture of compound 3 (79.0 g, 41.0 mmol, 96.4% purity, 1.00 eq, TFA) and compound 6C (14.2 g, 43.8 mmol, 96.9% purity, 1.07 eq) in DCM (800 mL) was added TEA (16.6 g, 164 mmol, 22.8 mL, 4.00 eq) dropwise at 0° C., and the mixture was stirred for 16 hours at 15° C. LCMS (EW33072-12-P1B, Rt=0.844 min) showed that the desired mass was detected. The reaction mixture was diluted with DCM (400 mL) and washed with NaHCO3 aqueous solution (400 mL*1) and brine (400 mL*1), then the mixture was diluted with DCM (2.00 L) and washed with 0.7 M Na2CO3 (1000 mL*3) and brine (800 mL*3), dried over Na2SO4, filtered and concentrated under reduced pressure to obtain a residue. The residue was used directly in the next step without purification. Compound 6 (80.0 g, crude product) was obtained as a white solid and was purified by1 HNMR confirmation: EW33072-12-P1A, 400MHz, MeODδ7.02-7.04(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,3 H),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).

[0468] General procedure for preparation of TriGNal-TRIS-Peg2-Phosph 8c

[0469]

[0470] Two batches were synthesized in parallel. To a solution of compound 6c (40.0 g, 21.1 mmol, 1.00 eq) in DCM (600 mL) were added diisopropylammonium tetrazolium salt (3.62 g, 21.1 mmol, 1.00 eq) and compound 7c (6.37 g, 21.1 mmol, 6.71 mL, 1.00 eq) in DCM (8.00 mL) dropwise, the mixture was stirred at 30 °C for 1 hour, and then added dropwise in DCM (8 00mL) was added to the mixture, the mixture was stirred at 30°C for 30 minutes, then compound 7c (3.18 g, 10.6 mmol, 3.35 mL, 0.50 eq) in DCM (8.00 mL) was added dropwise, and the mixture was stirred at 30°C for 1.5 hours. LCMS (EW33072-17-P1C1, Rt=0 .921 minutes) showed that the desired MS+1 was detected. LCMS (EW33072-17-P1C2, Rt=0.919 minutes) showed that the desired MS+1 was detected. The two batches were combined for post-processing. The mixture was diluted with DCM (1.20 L), washed with saturated NaHCO3 aqueous solution (1.60 L*2), 3% DMF aqueous solution (1.60 L*2), H2O (1.60 L*3), brine (1.60 L), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM:MeOH:TEA=100:3:2) TLC (SiO2, DCM:MeOH=10:1, Rf=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 and purified by 1 HNMR confirmation: EW33072-19-P1C, 400MHz, MeOD

[0471] δ7.13-7.15(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=11.26,3.38Hz,3H),4.64(d,J

[0472] =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).

[0473] Example 7: In vitro hepatocyte targeting using GalNAc-conjugated siRNA and ASO

[0474] In this experiment, the anti-GFP siRNA or ASO conjugated with the GalNAc part is treated with a hepatocyte cell line expressing asialoglycoprotein receptor and GFP, and in contrast, the hepatocyte cell line is treated with the anti-GFP siRNA or ASO of the unconjugated GalNAc part in the control experiment. The expression of GFP mRNA and protein is measured, and the GFP mRNA or protein expression in the cells treated with the siRNA or ASO conjugated with GalNAc or the GFP mRNA or protein expression in the cells treated with the siRNA or ASO conjugated with GalNAc is normalized and compared. This can help determine the hepatocyte targeting ability of the GalNAc part. Multiple GalNAc parts can be conjugated to siRNA or ASO and compared to determine which GalNAc part is the best in hepatocyte targeting. The GalNAc part tested in these experiments can be the GalNAc part described herein.

[0475] Similar experiments can be performed in primary hepatocytes treated with siRNA or ASO conjugated or not to a GalNAc moiety, and target mRNAs or target proteins other than GFP can be assessed in primary hepatocytes.

[0476] Example 8: In vivo hepatocyte targeting using GalNAc-conjugated siRNA and ASO

[0477] In this experiment, siRNA or ASO targeting target mRNA will be conjugated with GalNAc moieties and applied to mice (n=5 / group), while in control experiments, mice are administered siRNA or ASO not conjugated with GalNAc. After 2 days, mice are killed, and the liver is frozen, then homogenized, and the expression of target mRNA and protein is tested. The target mRNA or protein expression in the liver of mice treated with siRNA or ASO conjugated with GalNAc is normalized and compared with the GFP mRNA or protein expression in the liver of mice treated with siRNA or ASO not conjugated with GalNAc. This can help determine the liver targeting ability of the GalNAc moiety. Multiple GalNAc moieties can be conjugated to siRNA or ASO and compared to determine which GalNAc moiety is best in terms of liver targeting. The GalNAc moieties included in this experiment can be those showing the strongest hepatocyte targeting degree. The GalNAc moieties tested in these experiments can be GalNAc moieties described herein.

[0478] Example 9: Inhibition of target mRNA using GalNAc-conjugated siRNA and ASO in a mouse liver disease model

[0479] In this experiment, a mouse liver disease model (in this case, fatty liver disease) will be used to evaluate the effect of siRNA or ASO inhibition of target mRNA. The target mRNA can encode any target protein that is overexpressed or overactivated in liver disease and has a pathological role. In the mouse model, fatty liver disease is induced by feeding mice a Western diet (WD) for 12 weeks, which contains 21.1% fat, 41% sucrose and 1.25% cholesterol (Teklad diet, TD.120528) and a high sugar solution (23.1g / L d-fructose (Sigma-Aldrich, G8270) and 18.9g / L d-glucose (Sigma-Aldrich, F0127) by weight. 4-week-old C57BL / 6J mice are fed a Western diet instead of regular food for 12 weeks. The GalNAc moiety used in this experiment can be the GalNAc moiety described herein.

[0480] 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 group mice fed with normal chow. Each group contained 8 mice (4 males, 4 females). The siRNAs and ASOs of groups 1-4 all contained a GalNAc moiety attached to the siRNA or ASO.

[0481] At week 12 of Western diet feeding, blood samples were collected from each group before the first treatment.

[0482] Administration of siRNA or ASO was performed 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, group 1 mice will receive subcutaneous injections of non-targeting control siRNA, group 2 mice will receive subcutaneous injections of non-targeting control ASO, group 3 mice will receive subcutaneous injections of siRNA1 targeting mouse target mRNA, group 4 mice will receive subcutaneous injections of ASO1 targeting mouse target mRNA, and group 5 mice will receive subcutaneous injections of vehicle. Mice from each group will be dosed as on day 0 every other week starting from day 14, for a total of 5 injections.

[0483] Blood will be collected weekly and serum and plasma will be separated. Serum ALT, AST, total cholesterol and triglyceride levels were measured using VITROS 5,1FS (Ortho Clinical Diagnostics). Non-fasting plasma insulin was measured using an ultrasensitive mouse insulin ELISA kit (Crystal Chem, 90080) according to the manufacturer's instructions. Non-fasting blood glucose was measured using One Touch Ultra (Life Scan). HOMA-IR and QUICKI were calculated.

[0484] At the end of 12 weeks of Western diet and siRNA / ASO treatment, mice were killed by cervical dislocation and then injected intraperitoneally with 0.3 ml of pentobarbital (5 mg / ml). Terminal serum samples were collected by cardiac puncture, and final serum ALT, AST, total cholesterol and triglyceride levels, as well as non-fasting plasma insulin and blood glucose were measured. The liver was removed and divided into three parts: one part was placed in RNAlater for mRNA isolation, one part was quickly frozen for protein isolation, and the other part was fixed in formalin and then embedded in paraffin.

[0485] PureLink kit (ThermoFisher lot number 12183020) was used to separate mRNA from tissues placed in RNAlater solution according to the manufacturer's protocol. Reverse transcriptase reaction was performed according to the manufacturer's protocol. Samples were stored at -80 ° C until real-time qPCR was performed in triplicate using TaqMan gene expression assay (Applied Biosystems FAM probe, using BioRad iCycler). Compared with the target mRNA level in the liver tissue of mice administered with nonspecific control siRNA and ASO, the expression of target mRNA in the liver tissue of mice administered with siRNA and ASO decreased. It is expected that the amount of SDF-1 in the liver tissue of mice receiving siRNA and ASO will be lower than the amount of SDF-1 in the liver tissue of mice receiving nonspecific control siRNA or ASO. These results show that siRNA and ASO cause the knockdown of target mRNA and target protein in liver tissue, and the reduction of target mRNA and target protein expression is associated with the reduction of SDF-1 generation.

[0486] Formalin-fixed and paraffin-embedded liver tissue sections were stained with hematoxylin-eosin (H&E) to evaluate liver histology, stained with Sirius red (Sigma, 365548-5G) / Fast green (Sigma, F258) to evaluate fibrosis, and stained with periodic acid-Schiff (PAS) to evaluate glycogen accumulation. NAFLD activity score (NAS) and fibrosis stage were assessed by pathologists according to the NASHCRN scoring system 13. Histological scoring was performed in a blinded manner, and the pathologists were unaware of the treatment received. These results indicate that siRNA and ASO cause knockdown of target mRNA and target protein in liver tissue, and that the reduction in target mRNA and target protein expression is associated with a decrease in NAS and NASH CRN.

[0487] Example 10: Inhibition of mouse liver disease model

[0488] In this experiment, the effect of siRNA or ASO to inhibit the target protein expressed in the liver will be evaluated using a mouse model of liver disease (in this case, hypertriglyceridemia) and compared with an anti-mouse target protein antibody. The mouse strain C57Bl / 6Apoetm1Unc mice will be maintained on a high-fat Western diet (Research Diets, D12492; 60% of calories from fat). The target protein can be any protein that is overexpressed or overactivated in liver disease and plays a pathological role. The GalNAc moiety used in this experiment can be the GalNAc moiety described herein.

[0489] Four groups of mice (n=16 per group) will be used in this study. During the study, animals will maintain a high-fat diet. On the -4th day before the first injection, food will be removed and fasted overnight. On the -3rd day before the first injection, all animals will be anesthetized and 300 μL of blood will be collected through the mandibular vein into a serum separation tube to assess baseline triglycerides, serum glucose, insulin sensitivity, total cholesterol levels, HDL cholesterol levels, liver function, and serum levels of target proteins. On the 0th day of the experiment, the first group of mice will be intraperitoneally injected with 600 μL of saline, the second group of mice will be intraperitoneally injected with 600 μg of anti-mouse target protein antibodies (600 μL), the third group of mice will be subcutaneously injected with 150 μg of GalNAc-siRNA targeting the mRNA encoding the target protein of the mouse in 200 μL of saline, and the fourth group of mice will be subcutaneously injected with 150 μg of GalNAc-ASO targeting the mRNA encoding the target protein in 200 μL of saline. On the afternoon of the 3rd day, all groups will remove food and fast overnight. On the 4th day, all groups of animals will be anesthetized, and 150 μL of blood will be collected through the mandibular vein into a serum separation tube to assess the levels of serum triglycerides, glucose, total cholesterol, HDL cholesterol and target protein. Afterwards, all groups of animals will undergo oral glucose tolerance test and insulin tolerance test to assess insulin sensitivity. After blood sampling and insulin sensitivity test, food will be re-provided as usual. Thereafter, every week from the 7th day, the animals of Group 2 will be administered as on Day 0, for a total of 15 injections. Thereafter, every other week from the 14th day, the animals of Groups 3 and 4 will be administered as on Day 0, for a total of 8 injections. Every other week from the 10th day, all groups of mice will be fasted (overnight), and blood will be collected (150 μL, into a serum separation tube) to assess serum triglycerides, glucose, total cholesterol, HDL cholesterol and target protein levels, and insulin sensitivity test will be performed. On the third day after the last injection, food will be removed from all groups and fast overnight. On the fourth day after the last injection, animals from all groups will be anesthetized, euthanized, and 500 μL of blood will be collected by cardiac puncture into serum separator tubes to evaluate serum levels of triglycerides, serum glucose, insulin sensitivity, total cholesterol levels, HDL cholesterol levels, liver function, and target proteins. Liver, small intestine, and mesenteric lymph node tissues will be collected from all animals and immersed in 10% neutral buffered formalin for histopathological analysis. Liver samples will also be collected from all animals and stored in RNAlater. Target mRNA levels will be evaluated by RT-qPCR using TaqMan assays for mouse target proteins and mouse housekeeping gene PPIA.

[0490] Compared to mice in Group 1 (saline), animals treated with antibody (Group 2), mice treated with GalNAc-siRNA (Group 3), and mice treated with GalNAc-ASO (Group 4) are expected to have decreased triglycerides, serum total cholesterol, serum glucose levels, and serum target protein levels, while HDL cholesterol and insulin sensitivity are expected to increase. Animals in Groups 2 and 3 are also expected to have decreased target mRNA in liver samples.

[0491] Example 11: Inhibition of target mRNA in non-human primates using GalNAc-siRNA and GalNAc-ASO

[0492] In this experiment, the NHP model of hypertriglyceridemia is used to evaluate the effect of siRNA or ASO on inhibiting target mRNA expressed in the liver. The target protein can be any target protein that is overexpressed or overactivated in liver disease and plays a pathological role. Three groups of cynomolgus monkeys (n=5 / group) will be used, and they will be placed on a high-fat diet (Western primate diet, 5S2T) before the start of the study. Alternatively, three groups of rhesus monkeys (n=5 / group) will be used, and they will be placed on a high-fructose diet before the start of the study. Animals will be given 7 subcutaneous injections of saline (Group 1), GalNAc-siRNA (Group 2) or GalNAc-ASO (Group 3) every two weeks. The modified GalNAc-siRNA sequence can include any modification pattern described herein. The GalNAc moiety used in this experiment can be the GalNAc moiety described herein. Blood samples for lipid and blood glucose measurements were collected at the baseline and 4th, 8th and 14th weeks of the study, and lipid content, serum glucose, insulin sensitivity and target proteins were analyzed. All animals in each group were necropsied 2 weeks after the last blood draw. Tissues from the liver, small intestine, and mesenteric lymph nodes were collected from all animals and immersed in 10% neutral buffered formalin for histopathological analysis. Liver samples were also collected from all animals and placed in RNAlater. Target mRNA levels were assessed by RT-qPCR using TaqMan assays for cynomolgus or rhesus target proteins and cynomolgus or rhesus housekeeping genes PPIA.

[0493] It is expected that animals treated with GalNAc-siRNA (Group 2) and animals treated with GalNAc-ASO (Group 3) will show reductions in triglycerides, serum total cholesterol, and serum glucose, as well as reductions in serum target protein levels, and increases in HDL cholesterol and insulin sensitivity compared to animals in Group 1 (saline). It is also expected that animals in Groups 1 and 3 will show reductions in target mRNA in liver samples.

[0494] Example 12: Inhibition of target mRNA using GalNAc-siRNA and GalNAc-ASO in clinical trials

[0495] In this study, human subjects with hypertriglyceridemia were used to evaluate the effect of siRNA or ASO inhibiting target mRNA expressed in the liver. The target protein can be any target protein that is overexpressed or overactivated in liver disease and plays a pathological role. The selection criteria for inclusion in the study are age 40-90 years old, BMI ≥ 30 and serum triglycerides ≥ 250 mg / dL. Three groups of subjects (n = 15 / group) will be included in the study. The subjects will be given a total of 5 weekly subcutaneous injections of saline (Group 1), GalNAc-siRNA (Group 2) or GalNAc-ASO (Group 3). The GalNAc moieties to be used in these experiments can include the GalNAc moieties described herein.

[0496] The siRNA or ASO sequences should be from a selection set that exhibits high activity in cultured cells or in the experiments described in other Examples. Blood samples for lipid and glycemic measurements will be collected at baseline and 3, 6, and 12 weeks of the study and analyzed for lipid content, serum glucose, insulin sensitivity, target proteins, and liver and kidney function.

[0497] It is expected that subjects treated with GalNAc-siRNA (Group 2) and subjects treated with GalNAc-ASO (Group 3) will have decreased triglycerides, serum total cholesterol, and serum glucose, decreased serum target protein levels, and increased HDL cholesterol and insulin sensitivity compared to subjects in Group 1 (saline).

[0498] Example 13: Oligonucleotide Synthesis

[0499] RNAi agents (e.g., siRNA) were synthesized using the phosphoramidite technique on a solid phase used in oligonucleotide synthesis. The K&A oligonucleotide synthesizer was used. The synthesis was performed on a controlled pore glass (CPG) or 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 were added. . 5-Benzylthio-1H-tetrazole (BTT, 250 mM in acetonitrile) or 5-ethylthio-1H-tetrazole (ETT, 250 mM in acetonitrile) was used as the activator solution. The coupling time was 9-18 min (EmpGalNAc), 6 min (2'OMe and 2'F). To introduce the phosphorothioate linkage, a 100 mM solution of 3-phenyl 1,2,4-dithiazolin-5-one (POS, obtained from PolyOrg, Inc., Leominster, Mass., USA) in anhydrous acetonitrile was used.

[0500] After solid phase synthesis, the dried solid support was treated with a 1:1 volume ratio solution of 40wt.% methylamine aqueous solution and 28% ammonium hydroxide solution (Aldrich) at 30°C for 2 hours. 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 20mM Tris, 5mMEDTA, pH 9.0, and contained 20% acetonitrile, and buffer B was the same as buffer A, but 1M sodium chloride was added. The UV spectrum at 260nm was recorded. The appropriate fractions were combined and then desalted using Sephadex G-25 medium.

[0501] Equimolar amounts of the sense strand and the antisense strand were combined to prepare a duplex. The duplex solution was prepared in 0.1× PBS (phosphate buffered saline, 1×, Gibco). The duplex solution was annealed at 95°C for 5 minutes and then slowly cooled to room temperature. The duplex concentration was determined by measuring the absorbance of the solution on a UV-visible spectrometer at 260 nm in 0.1× PBS. For some experiments, a conversion factor was calculated from the experimentally determined extinction coefficient.

[0502] Example 14: In vivo hepatocyte targeting using GalNAc-conjugated siRNA and ASO

[0503] In this experiment, siRNA or ASO targeting target mRNA will be conjugated with GalNAc moieties and administered to mice (n=5 / group), whereas mice in control experiments were administered siRNA or ASO not conjugated with GalNAc. Mice were killed 2 days later, and the livers were frozen, then homogenized, and target mRNA and protein expression was tested. The target mRNA or protein expression in the liver of mice treated with siRNA or ASO conjugated with GalNAc was normalized and compared with the GFP mRNA or protein expression in the liver of mice treated with siRNA or ASO not conjugated with GalNAc. This can help determine the liver targeting ability of the GalNAc moiety. Multiple GalNAc moieties can be conjugated to siRNA or ASO and compared to determine which GalNAc moiety is best in terms of liver targeting. The GalNAc moieties can be those that show the greatest degree of hepatocyte targeting in Example 6. The GalNAc moieties to be tested in these experiments can be GalNAc moieties as described herein, such as compound 1 or compound 2.

[0504] Example 15: Knockdown of PLIN1 in mice by GalNAc-conjugated siRNA

[0505] The example GalNAc moiety ETL17 was conjugated to siRNAs targeting example target mRNAs. The sequences are shown in Table 2. In the table, Nf (e.g., Af, Cf, Gf, Tf, or Uf) is a 2' fluorine-modified nucleoside, n (e.g., a, c, g, t, or u) is a 2' O-methyl-modified nucleoside, and "s" is a phosphorothioate linkage. The activity of the siRNAs was tested in mice.

[0506] On day -14, 6- to 8-week-old female mice (C57Bl / 6) were injected with 10 uL of recombinant adeno-associated virus 8 (AAV8) vector (1.7 × 10 E 13 The recombinant AAV8 contained the human PLIN1 (NM_002666.5) sequence, controlled by the human thyroxine binding globulin promoter in the AAV2 backbone packaged in the AAV8 capsid (AAV8-TBG-h-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.

[0507] Mice were euthanized on day 10 after subcutaneous injection, and liver samples from each mouse were collected and placed in RNAlater (ThermoFisher catalog number 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) according to the manufacturer's recommendations. cDNA was prepared using Quanta qScript cDNA SuperMix (VWR, catalog number 95048-500) according to the manufacturer's instructions. TaqMan assays and qPCR Low ROX TM (VWR, catalog number 101419-222) in QuantStudio TM The relative levels of liver PLIN1 mRNA were assessed in triplicate by RT-qPCR on the 6 Pro real-time PCR system. The data were normalized to the average PLIN1 mRNA levels in animals receiving PBS. The results are shown in Table 3. All tested siRNAs resulted in a decrease in average liver PLIN1 mRNA on day 10 compared to mice receiving PBS. These data suggest that siRNAs conjugated to GalNAc moieties such as ETL17 can be used to knock down target mRNA in the liver.

[0508] Table 2. Example siRNA sequences

[0509]

[0510] Table 3. Relative human PLIN1 mRNA levels in mouse liver

[0511]

[0512] Example 16: Knockdown of MST1 in mice by GalNAc-conjugated siRNA

[0513] ETL17 is conjugated with siRNA targeting another exemplary target mRNA. The siRNA is attached to the GalNAc ligand ETL17, followed by a phosphorothioate linkage at the 5' end of the sense strand. The siRNA is described in Table 6. In the table, Nf (e.g., Af, Cf, Gf, Tf, or Uf) is a 2' fluorine-modified nucleoside, dN (e.g., dA, dC, dG, dT, or dU) is a 2' deoxy-modified nucleoside, n (e.g., a, c, g, t, or u) is a 2' O-methyl-modified nucleoside, and "s" is a phosphorothioate linkage.

[0514] Six to eight-week-old female mice (C57Bl / 6) were injected with 5uL of recombinant adeno-associated virus 8 (AAV8) vector (2.7×10E 13 Genome copies / mL). Recombinant AAV8 contains an open reading frame and most of the 3'UTR of the human MST1 sequence (NM_020998.4), controlled by the human thyroxine binding globulin promoter (AAV8-TBG-h-MST1) in the AAV2 backbone packaged in the AAV8 capsid. On day 13 post-infection, serum was collected and the level of human MSP in each mouse was measured using the Human MSP / MST1 DuoSet ELISA from R&D (Catalog No. DY352). The manufacturer's instructions for all reagent preparations for buffers and solutions were followed. A 1:50 dilution of serum sample was used for all test samples. The recombinant MSP included in the kit was used to create a standard curve from 10,000 pg / mL to 0 pg / mL. The optical density of the plate was read at 450 nm using a PerkinElmer Envision multi-mode plate reader. The MSP concentration in each mouse serum sample was calculated from the standard curve by interpolation using a least squares fit (Prism version 9, SoftwareMacKiev).

[0515] Mice were assigned to groups (n=3) so that each group had similar serum MSP levels, and then subcutaneously injected with a single dose of 60ug of GalNAc-conjugated siRNA or PBS as a vehicle control. On days 0, 4, and 12 after injection, serum was collected and serum MSP concentrations were assessed by ELISA using the above method. The MSP serum concentration at each time point was obtained relative to the MSP level in the day 0 sample of each mouse. The results are shown in Table 4. These data indicate that siRNA conjugated to a GalNAc moiety such as ETL17 can be used to knock down proteins secreted by other target mRNAs in the liver.

[0516] Mice were sacrificed on day 12, and liver samples from each mouse were collected and placed in RNAlater (ThermoFisher Cat#AM7020) until processing. Total liver RNA was prepared by homogenizing liver tissue in homogenization buffer (MaxwellRSC simplyRNA tissue kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10 second cycles. Total RNA in the lysate was purified on a MaxwellRSC 48 platform (Promega Corporation) according to the manufacturer's recommendations. cDNA was prepared using Quanta qScript cDNA SuperMix (VWR, catalog number 95048-500) according to the manufacturer's instructions. TaqMan assays and qPCR Low ROX TM (VWR, catalog number 101419-222) in QuantStudio TM Relative levels of liver MST1 mRNA were assessed in triplicate by RT-qPCR on the 6 Pro Real-Time PCR System. Data were normalized to levels in animals receiving PBS. Results are shown in Table 5. These data suggest that siRNA conjugated to a GalNAc moiety such as ETL17 can be used to knock down other target mRNAs in the liver.

[0517] Table 4. Relative mean serum human MSP levels in AAV8-TBG-h-MST1 mice

[0518]

[0519] ND, not determined

[0520] Table 5. Relative human MST1 mRNA levels in liver of AAV8-TBG-h-MST1 mice

[0521]

[0522] Table 6. siRNAs screened for activity in AAV8-TBG-h-MST1 mice

[0523]

[0524] Example 17: Knockdown of additional target mRNAs in mice by GalNAc-conjugated siRNA

[0525] Three groups (n=4 / group) of 8-week-old male ICR mice (Invigo) were used in the study. On study day 0, mice in group 1 were injected subcutaneously with 100uL sterile PBS, mice in group 2 were injected subcutaneously with 60ug siRNA 1811 in 100uL sterile PBS, and mice in group 3 were injected subcutaneously with 200ug siRNA 1818 in 100uL sterile PBS. On study day 14, animals in all groups were anesthetized and then euthanized. Liver samples were collected from all animals and placed in RNAlater TM Stabilizing solution (ThermoFisher, catalog number AM7020).Using soft tissue homogenization kit CK14 (Bertin Instruments, catalog number P000933-LYSK0-A), the liver samples were processed in a homogenization buffer (Maxwell RSC simplyRNA tissue kit) in a Percellys 24 tissue homogenizer (Bertin Instruments) with a setting speed of 5000rpm and two 10-second cycles.According to the manufacturer's advice, total RNA from liver lysate was purified on a Maxwell RSC 48 platform (Promega Corporation).The relative level of gene A mRNA in each liver sample was assessed by RT-qPCR on a QuantStudio6Pro instrument (Applied Biosystems) using TaqMan assays for mouse gene A and mouse housekeeping gene PPIA, and then normalized to the mean value of control mice (Group 1) using the Δ-ΔCt method.

[0526] In this example, an additional GalNAc moiety (ETL1) was compared to ETL17. The results of the liver mRNA analysis are shown in Table 7. Compared to mice injected with PBS (Group 1), animals treated with ETL1 targeting siRNA (ETD01811, Group 2) had a relative knockdown of 78% in liver gene A mRNA levels, while animals treated with ETL17 targeting siRNA (siRNA1818, Group 3) had a relative knockdown of 83% in liver gene A mRNA levels. Therefore, the compounds described herein having a GalNAc moiety may be superior to or at least as effective as another GalNAc moiety in knocking down a target, such as a target mRNA.

[0527] Table 7: Gene A mRNA levels in liver of mice treated with siRNA targeting Gene A at day 14

[0528] Group# deal with average value 1 PBS 1.00 2 siRNA 1811 (with control GalNAc ETL1) 0.22 3 siRNA 1818 (with ETL17) 0.17

[0529] ETL1 is shown below, where J indicates attachment to the oligonucleotide:

[0530]

[0531] Example 18. Testing of MST1 siRNA ETD01821, ETD01822, Activity of ETD01823 and ETD01826

[0532] Four groups (n=3 / group) of 3-6 kg male cynomolgus monkeys (Zhaoqing Chuangyao Biotechnology Co., Ltd. and Guangzhou Xiangguan Biotechnology Co., Ltd.) were used in this study.

[0533] On day 0 of the study, group 1 cynomolgus monkeys were subcutaneously injected with a single dose of 5 mg / kg (0.2 mL dose volume / kg body weight) of ETD01821, whose siRNA concentration was 25 mg / mL, prepared in PBS; group 2 cynomolgus monkeys were subcutaneously injected with a single dose of 5 mg / kg (0.2 mL dose volume / kg body weight) of ETD01822, whose siRNA concentration was 25 mg / mL, prepared in PBS; group 3 cynomolgus monkeys were subcutaneously injected with a single dose of 5 mg / kg (0.2 mL dose volume / kg body weight) of ETD01822, whose siRNA concentration was 25 mg / mL, prepared in PBS; The fourth group of cynomolgus monkeys were subcutaneously injected with a single dose of 5 mg / kg (0.2 mL dose volume / kg body weight) of ETD01823, whose siRNA concentration was 25 mg / mL, formulated in PBS; the fourth group of cynomolgus monkeys were subcutaneously injected with a single dose of 5 mg / kg (0.2 mL dose volume / kg body weight) of ETD01826, whose siRNA concentration was 25 mg / mL, formulated in PBS, and the siRNA sequences are shown in Table 8, where "Nf" is a 2' fluorine-modified nucleoside, "n" is a 2' O-methyl-modified nucleoside, and "s" is a phosphorothioate linkage. The injections were generally well tolerated as measured by clinical symptoms.

[0534] During the study, all cynomolgus monkeys showed no abnormal clinical symptoms except animal No. 101, which was found dead on day 65 after dosing. Autopsy showed severe gastric perforation as the likely cause of death. This may occur spontaneously in cynomolgus monkeys.

[0535] Body weights were recorded on study days -8, -1, 7, 14, 21 and 28. The results are shown in Table 9. The data showed that there were no meaningful changes in body weights of animals in Groups 1 to 4.

[0536] In research the 8th, 2nd, 7th, 14th and 28th day, blood was collected in the tube without anticoagulant, and serum was collected. The clinical chemistry parameters including ALT, AST, ALP, TBIL, DBIL, GLU, GGT, TP, TG, CHOL, HDL, LDL, BUN and CREA were analyzed. Compared with research the 8th day and research the 2nd day before treatment, the animals treated with ETD01821, ETD01822, ETD01823 or ETD01926 did not show significant changes in ALT, AST, ALP, TBIL, DBIL, GLU, GGT, TP, TG, CHOL, HDL, LDL, BUN and CREA from the 7th day to the 28th day of research. Clinical chemistry results show that all siRNAs are generally well tolerated. The results are shown in Tables 10-14.

[0537] In the research of -8 days, -2 days, -7 days, 14 days and 28 days, about 1mL whole blood was collected in the tube with EDTA-K2 as anticoagulant. Hematological parameters were analyzed, including WBC, NEUT, LYMP, MONO, EOS, BASO, RBC, HGB, HCT, MCV, MCH, MCHC, RDW, PLT, MPV, PCT and PDW. Compared with the research of -8 days before treatment and the research of -2 days, the animals treated with ETD01821, ETD01822, ETD01823 or ETD01926 did not have significant changes in these hematological parameters from the research of the 7th day to the research of the 28th day. The results of hematological analysis show that all siRNAs are generally well tolerated. The results are shown in Tables 15-19.

[0538] Blood was collected into tubes without anticoagulant and serum was collected to determine serum macrophage stimulating protein (MSP) levels on days -8, -2, 7, 14, 28, 42, 56, 70, 77, 84, 91, 98 and 105 of the study. 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 in monkey serum samples were assessed using a custom AlphaLISA assay (PerkinElmer). In brief, 5uL serum samples diluted 1:50 in 1×AlphaLISA HiBlock were placed in wells of a 96-well plate, followed by the addition of 5uL 4× anti-MSP receptor bead solution. After incubation at room temperature for 30 minutes, 5uL 4× biotinylated anti-MSP antibody solution was added, and the plate was incubated at room temperature for 60 minutes. Next, 5uL of 4× streptavidin donor bead solution was added and the plate was incubated for another 30 minutes at room temperature. The plates were analyzed using 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 derived relative to the average of the MSP serum concentrations of that individual on days -2 and -8. The results of the group averages are shown in Table 20, and the individual values ​​are shown in Table 21. After treatment with the test article starting on day 7, serum MSP levels in all animals decreased and remained decreased until at least day 28. Compared with pre-dose levels, the serum MSP levels of monkeys treated with ETD01821 decreased the most, and the average serum MSP levels decreased until day 105 compared with pre-dose levels.

[0539] 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 collected. The biopsies were then placed in 10 v / v RNAlater within 20 seconds and stored at 4°C for 24 hours before the RNAlater was removed. TMStabilizing solution (Thermo Fisher, catalog number AM7020), and liver tissues were stored in a refrigerator until they were shipped to Empirico. After collecting liver biopsy collections on the -2nd day or the 28th day, no abnormal clinical observations were observed in all animals. Liver samples were processed in a homogenization buffer (Maxwell RSCsimplyRNA tissue kit) using a soft tissue homogenization kit CK14 (Bertin Instruments, catalog number P000933-LYSK0-A) in a Percellys 24 tissue homogenizer (Bertin Instruments) with a set speed of 5000rpm and two 10 second cycles. Total RNA from liver lysates was purified on a Maxwell RSC 48 platform (Promega Corporation) according to the manufacturer's advice. According to the manufacturer's instructions, cDNA was prepared using QuantaqScript cDNA SuperMix (VWR, catalog number 95048-500). Relative levels of liver MST1 mRNA were assessed by RT-qPCR in triplicate in duplex reactions using TaqMan assays for Macaca fascicularis MST1 (ThermoFisher, assay number Mf01117426_g1) and the Macaca fascicularis housekeeping gene GAPDH (ThermoFisher, assay number Mf04392546_g1) in PerfeCTa qPCRFastMix Reaction Mix (VWR). TM Samples were evaluated on the 6Pro real-time PCR system. The relative amount of MST1 mRNA was calculated using the Δ-ΔCt method. Table 22 shows the average relative MST1 mRNA levels of the groups relative to day -8. Consistent with the decrease in serum MSP levels measured by AlphaLISA, treatment with 5 mg / kg test article ETD01821, ETD01822, ETD01823 or ETD01826 resulted in a decrease in liver MST1 mRNA levels on day 28 compared to the levels on day -8 before dosing.

[0540] Table 8. Example siRNA sequences

[0541]

[0542] Table 9. Body weight (kg)

[0543]

[0544] Table 10. Individual and mean clinical chemistry parameter results before dosing (Day -8)

[0545]

[0546]

[0547] Note: ↓ next to the value indicates the result is slightly lower than other animals.

[0548] Table 11. Individual and mean clinical chemistry parameter results before dosing (Day -2)

[0549]

[0550]

[0551] Note: ↓ next to the value indicates that the result is slightly lower than other animals. '----' indicates that DBIL in some samples could not be detected due to low concentration.

[0552] Table 12. Individual and mean clinical chemistry parameter results on day 7 post-dose

[0553]

[0554]

[0555] Note: ↓ next to the value indicates that the result is slightly lower than other animals. '----' indicates that DBIL in some samples could not be detected due to low concentration.

[0556] Table 13. Individual and mean clinical chemistry parameter results on day 14 after dosing

[0557]

[0558]

[0559]

[0560] Note: ↓ next to the value indicates that the result is slightly lower than other animals. '----' indicates that DBIL in some samples could not be detected due to low concentration.

[0561] Table 14. Individual and mean clinical chemistry parameter results on day 28 after dosing

[0562]

[0563]

[0564] Note: ↓ next to the value indicates that the result is slightly lower than other animals. '----' indicates that DBIL in some samples could not be detected due to low concentration.

[0565] Table 15. Individual and mean hematology results before dosing (Day -8)

[0566]

[0567]

[0568]

[0569] Note: ↓ next to the value indicates the result is slightly lower than other animals.

[0570] Table 16. Individual and mean hematology results before dosing (Day -2)

[0571]

[0572]

[0573]

[0574] Note: ↓ next to the value indicates the result is slightly lower than other animals.

[0575] Table 17. Individual and mean hematology results on day 7 after dosing

[0576]

[0577]

[0578]

[0579] Note: ↓ next to the value indicates the result is slightly lower than other animals.

[0580] Table 18. Individual and mean hematology results on day 14 after dosing

[0581]

[0582]

[0583]

[0584] Note: ↓ next to the value indicates the result is slightly lower than other animals.

[0585] Table 19. Individual and mean hematology results on day 28 after dosing

[0586]

[0587]

[0588]

[0589] Note: ↓ next to the value indicates the result is slightly lower than other animals.

[0590] Table 20. Relative mean serum MSP levels in cynomolgus monkeys

[0591]

[0592] Table 21. Relative individual serum MSP levels in cynomolgus monkeys

[0593]

[0594]

[0595] Table 22. Relative MST1 mRNA levels in cynomolgus monkey liver

[0596]

[0597] Example 19: Screening of siRNAs targeting human and mouse MTRES1 in mice

[0598] The following examples demonstrate the usefulness of the GalNAc moiety described herein in vivo in combination with oligonucleotides targeting other targets (MTRES1). The activity of several siRNAs designed to have cross-reactivity with human, mouse and cynomolgus monkey MTRES1 mRNA was tested in mice. siRNA was attached to GalNAc ligands ETL1 or ETL17. The siRNA sequence is shown in Table 23, wherein Nf is a 2' fluorine-modified nucleoside, n is a 2' O-methyl-modified nucleoside, "d" is a deoxynucleoside, and "s" is a phosphorothioate linkage.

[0599] On day 0, 6-8 week old female mice (ICR strain, n=3) were injected subcutaneously with a single 200 ug dose of GalNAc-conjugated siRNA or PBS as a vehicle control.

[0600] On the 10th day, mice were euthanized after injection, and liver samples from each mouse were collected and placed in RNAlater (ThermoFisher, catalog number AM7020) until processed.Total liver RNA was prepared by homogenizing liver tissue in homogenization buffer (Maxwell RSC simplyRNA tissue test kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set to 5000rpm, two 10 second cycles.Total RNA from lysate was purified on Maxwell RSC 48 platforms (Promega Corporation) according to the manufacturer's advice.CDNA was prepared using Quanta qScript cDNA SuperMix (VWR, catalog number 95048-500) according to the manufacturer's instructions.TaqMan assays and qPCR Low ROX TM (VWR, catalog number 101419-222) and RT-qPCR in QuantStudio TM The relative levels of liver MTRES1 mRNA were assessed in triplicate on the 6 Pro real-time PCR system. The data were normalized to the average MTRES1 mRNA levels in animals receiving PBS. The results are shown in Table 24. On day 10, the average liver MTRES1 mRNA levels of mice injected with ETD01597, ETD01955, and ETD01958 were significantly lower relative to mice receiving PBS.

[0601] Table 23. Example siRNA sequences

[0602]

[0603] Table 24. Relative MTRES1 mRNA levels in mouse liver

[0604]

[0605] Although 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. Without departing from the present invention, those skilled in the art will now appreciate that various modifications, changes and substitutions will be appreciated. It should be understood that the various alternatives of the embodiments of the present invention described herein may be used to implement the present invention. The appended claims are intended to define the scope of the present invention and are intended to cover methods and structures and their equivalents within the scope of these claims.

Claims

1. A compound represented by formula (I) or (II): 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 from 1 to 20; m is selected from any value from 1 to 20; z is selected from any value from 1 to 3, wherein If z is 3, then Y is C; If z is 2, then Y is CR 6 ,or If z is 1, then Y is C(R 6 )2; Q is selected from: C optionally substituted by one or more substituents 3-10 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 Alkyl, wherein the C 1-6 The alkyl group is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2; R 1 is a linker selected from the group consisting of: -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 )2NR 7 -; Each R 2 Independently selected from: C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from 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 ; R 3 and R 4 Each independently selected from: -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 ; Each R 5 Independently selected from: -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; Each R 6 Independently selected from: hydrogen; halogen, -CN, -NO2, -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 C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 ; Each R 7 Independently selected from: hydrogen; C 1-6 Alkyl, C 2-6 Alkenyl and C 2-6 Alkynyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 Carbocyclic rings and 3- to 10-membered heterocyclic rings; and C 3-10 carbocyclic ring and 3- to 10-membered heterocyclic ring, each of which is optionally substituted by one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -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 ring, 3- to 10-membered heterocyclic ring and C 1-6 Halogenated alkyl.

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

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

4. The compound or salt of claim 1, wherein each w is 1.

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

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

7. The compound or salt of claim 1, wherein each v is 1.

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

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

10. The compound or salt of claim 1, wherein n is 2.

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

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

13. The compound or salt of claim 1, wherein m is selected from 1 and 2.

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

15. The compound or salt of claim 1, wherein Q is selected from C optionally substituted by one or more substituents 5-6 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 .

16. The compound or salt of claim 1, wherein Q is selected from C optionally substituted by one or more substituents 5-6 The one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2.

17. A compound or salt as claimed in claim 1, wherein Q is selected from phenyl and cyclohexyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2.

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

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

20. The compound or salt according to claim 1, wherein R 1 is selected from -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 )2NR 7 .

21. The compound or salt of claim 1, wherein R 1 Selected from -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 )O-.

22. The compound or salt of claim 1, wherein R 1 Selected from -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-.

23. The compound or salt of claim 1, wherein R 1 Selected from -OP(O)(OR 7 )O- and -OP(OR 7 )O-.

24. The compound or salt of claim 1, wherein R 2 is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from halogen, -OR 7 、-OC(O)R 7 、-SR 7 、-N(R 7 )2. -C(O)R 7 and-S(O)R 7 .

25. The compound or salt of claim 1, wherein R 2 is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from -OR 7 、-OC(O)R 7 、-SR 7 and -N(R 7 )2.

26. The compound or salt of claim 1, wherein R 2 is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from -OR 7 and -OC(O)R 7 .

27. The compound or salt of claim 1, wherein R 3 Selected from halogen, -OR 7 、-SR 7 、-N(R 7 )2. -C(O)R 7 、-OC(O)R 7 and-S(O)R 7 .

28. The compound or salt of claim 1, wherein R 3 Select from -OR 7 、-SR 7 、-OC(O)R 7 and -N(R 7 )2.

29. The compound or salt of claim 1, wherein R 3 Select from -OR 7 -and-OC(O)R 7 .

30. The compound or salt of claim 1, wherein R 4 Selected from halogen, -OR 7 、-SR 7 、-N(R 7 )2. -C(O)R 7 、-OC(O)R 7 and-S(O)R 7 .

31. The compound or salt of claim 1, wherein R 4 Select from -OR 7 、-SR 7 、-OC(O)R 7 and -N(R 7 )2.

32. The compound or salt of claim 1, wherein R 4 Select from -OR 7 -and-OC(O)R 7 .

33. The compound or salt of claim 1, wherein R 5 Selected from -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 .

34. The compound or salt of claim 1, wherein R 5 Selected from -OC(O)R 7 and -N(R 7 )C(O)R 7 .

35. The compound or salt of claim 1, wherein each R 7 Independently selected from: Hydrogen; and C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 alkyl), C 3-10 A carbocyclic ring or a 3- to 10-membered heterocyclic ring.

36. A compound or salt as described in any one of claims 1 to 30, wherein each R 7 are independently selected from C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2 and -NH(C 1-6 alkyl).

37. A compound or salt as described in any one of claims 1 to 34, wherein each R 7 are independently selected from C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from halogen, -CN, -OH and -SH.

38. The compound or salt of claim 1, wherein 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 optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2 and C 1-3 alkyl; R 1 selected from -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 a C1 alkyl substituted by -OH or -OC(O)CH3; R 3 is -OH or -OC(O)CH3; R 4 is -OH or -OC(O)CH3; and R 5 It is -NH(O)CH3.

39. The compound or salt of claim 1, wherein the compound comprises:

40. The compound of claim 1, wherein the oligonucleotide (J) is attached at the 5' end or the 3' end of the oligonucleotide.

41. The compound of claim 1, wherein the oligonucleotide comprises DNA.

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

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

44. compounds as claimed in claim 43, wherein the one or more modified internucleoside linkages comprise alkyl phosphonates, phosphorothioates, methyl phosphonates, phosphorodithioates, alkyl phosphonothioates, phosphoramidates, carbamates, carbonates, phosphotriesters, acetimidates or carboxymethyl esters, or a combination thereof.

45. The compound of claim 1, 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 of claim 1, wherein the oligonucleotide comprises one or more modified nucleosides.

47. The compound of claim 46, wherein the one or more modified nucleosides comprise 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.

48. compounds as claimed in claim 46, wherein the one or more modified nucleosides include 2',4'-constrained ethyl nucleosides, 2'-O-methyl nucleosides, 2'-deoxyfluoronucleosides, 2'-ON-methylacetamido (2'-O-NMA) nucleosides, 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.

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

51. The compound of claim 50, wherein the lipid comprises cholesterol, myristoyl, palmitoyl, stearoyl, lithocholyl, behenoyl, docosahexaenoyl, myristyl, palmityl, stearyl, or alpha-tocopherol, or a combination thereof.

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

53. The compound of claim 52, wherein the RGD peptide comprises cyclic (-Arg-Gly-Asp-D-Phe-Cys), cyclic (-Arg-Gly-Asp-D-Phe-Lys), cyclic (-Arg-Gly-Asp-D-Phe-azido), aminobenzoic acid-derivatized RGD, or a combination thereof.

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

55. The compound of claim 54, wherein the sense strand is 12-30 nucleosides in length.

56. The compound of claim 54, wherein the antisense strand is 12-30 nucleosides in length.

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

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

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

60. The compound of claim 59, wherein the 3' overhang comprises 1, 2 or more nucleosides.

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

62. The compound of claim 61, wherein the ASO is 12-30 nucleosides in length.

63. The compound of claim 1, wherein the compound binds to the asialoglycoprotein receptor.

64. The compound of claim 1, wherein the compound is targeted to hepatocytes.

65. A pharmaceutical composition comprising the compound of claim 1 and a pharmaceutically acceptable carrier, excipient or diluent.

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

67. The pharmaceutical composition of claim 65, wherein the pharmaceutical composition comprises a pharmaceutically acceptable carrier.

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

69. 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 target protein by at least 10%.

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

71. The method of claim 70, wherein the effective amount reduces the measured value of the target mRNA or target protein in the subject relative to a baseline target mRNA or target protein measured value.

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

73. The method of claim 72, wherein the effective amount reduces a measured value of a symptom or parameter associated with the condition in the subject relative to a baseline symptom or parameter measured value.

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

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

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

77. A compound represented by formula (A) or (B): 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 from 1 to 20; m is selected from any value from 1 to 20; z is selected from any value from 1 to 3, wherein If z is 3, then Y is C If z is 2, then Y is CR 6 ,or If z is 1, then Y is C(R 6 )2; Q is selected from: C optionally substituted by one or more substituents 3-10 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 Alkyl, wherein the C 1-6 The alkyl group is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2; R 1 Selected from: -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)2R 7 、-OS(O)2R 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; Each R 2 Independently selected from: C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from 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 ; R 3 and R 4 Each independently selected from: -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 ; Each R 5 Independently selected from: -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; Each R 6 Independently selected from: hydrogen; halogen, -CN, -NO2, -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 C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 ; Each R 7 Independently selected from: hydrogen; C 1-6 Alkyl, C 2-6 Alkenyl and C 2-6 Alkynyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 Carbocyclic rings and 3- to 10-membered heterocyclic rings; and C 3-10 carbocyclic and 3- to 10-membered heterocyclic rings, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -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 Halogenated alkyl.

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

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

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

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

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

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

84. The compound or salt of claim 77, wherein n is selected from any value from 1 to 10.

85. The compound or salt of claim 77, wherein n is selected from any value of 1 to 5.

86. The compound or salt of claim 77, wherein n is 2.

87. The compound or salt of claim 77, wherein m is selected from any value of 1 to 10.

88. The compound or salt of claim 77, wherein m is selected from any value of 1 to 5.

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

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

91. The compound or salt of claim 77, wherein Q is selected from C optionally substituted by one or more substituents. 5-6 Carbocyclic ring, wherein the one or more substituents are independently selected from halogen, -CN, -NO2, -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 .

92. The compound or salt of claim 77, wherein Q is selected from C optionally substituted with one or more substituents. 5-6 The one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2.

93. compounds or salts as claimed in claim 77, wherein Q is selected from phenyl and cyclohexyl, each of which is optionally substituted by one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2 and -NH2.

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

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

96. The compound or salt of claim 77, wherein R 1 Selected from -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.

97. The compound or salt of claim 77, wherein R 1 Selected from -OP(O)(OR 7 )2 and -OP(OR 7 )N(R 7 )2.

98. The compound or salt of claim 77, wherein R 1 Selected from -OP(O)(OCH2CH3)OH and -OP(OCH2CH2CN)N(CH(CH3)2)2.

99. The compound or salt of claim 77, wherein R 1 It is -OP(OCH2CH2CN)N(CH(CH3)2)2.

100. The compound or salt of claim 77, wherein R 2 is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from halogen, -OR 7 、-OC(O)R 7 、-SR 7 、-N(R 7 )2. -C(O)R 7 and-S(O)R 7 .

101. The compound or salt of claim 77, wherein R 2 is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from -OR 7 、-OC(O)R 7 、-SR 7 and -N(R 7 )2.

102. The compound or salt of claim 77, wherein R 2 is selected from C substituted by one or more substituents 1-3 Alkyl, the one or more substituents are independently selected from -OR 7 and -OC(O)R 7 .

103. The compound or salt of claim 77, wherein R 3 Selected from halogen, -OR 7 、-SR 7 、-N(R 7 )2. -C(O)R 7 、-OC(O)R 7 and-S(O)R 7 .

104. The compound or salt of claim 77, wherein R 3 Select from -OR 7 、-SR 7 、-OC(O)R 7 and -N(R 7 )2.

105. The compound or salt of claim 77, wherein R 3 Select from -OR 7 and -OC(O)R 7 .

106. The compound or salt of claim 77, wherein R 4 Selected from halogen, -OR 7 、-SR 7 、-N(R 7 )2. -C(O)R 7 、-OC(O)R 7 and-S(O)R 7 .

107. The compound or salt of claim 77, wherein R 4 Select from -OR 7 、-SR 7 、-OC(O)R 7 and -N(R 7 )2.

108. The compound or salt of claim 77, wherein R 4 Select from -OR 7 and -OC(O)R 7 .

109. The compound or salt of claim 77, wherein R 5 Selected from -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 .

110. The compound or salt of claim 77, wherein R 5 Selected from -OC(O)R 7 and -N(R 7 )C(O)R 7 .

111. The compound or salt of claim 77, wherein each R 7 Independently selected from: Hydrogen; and C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2, -NH(C 1-6 Alkyl), C 3-10 A carbocyclic ring or a 3- to 10-membered heterocyclic ring.

112. The compound or salt of claim 77, wherein each R 7 are independently selected from C optionally substituted by one or more substituents 1-6 Alkyl, wherein the one or more substituents are independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2, =O, =S, -OC 1-6 Alkyl, -SC 1-6 Alkyl, -N(C 1-6 Alkyl)2 and -NH(C 1-6 alkyl).

113. The compound or salt of claim 77, wherein each R 7 are independently selected from C optionally substituted by one or more substituents 1-6 Alkyl, the one or more substituents are independently selected from halogen, -CN, -OH and -SH.

114. The compound or salt of claim 113, wherein 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 optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, -NH2 and C 1-3 alkyl; R 1 Selected from -OP(O)(OR 7 )2 and -OP(OR 7 )N(R 7 )2; R 2 is a C1 alkyl substituted by -OH or -OC(O)CH3; R 3 is -OH or -OC(O)CH3; R 4 is -OH or -OC(O)CH3; and R 5 It is -NH(O)CH3.

115. The compound or salt of claim 77, wherein the compound comprises:

116. A compound comprising: wherein J comprises an oligonucleotide, or comprises a linkage to said oligonucleotide.

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

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

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

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

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