Multiplexing of targeting ligands via click chemistry at the anomeric sugar moiety

Abstract

The present disclosure relates to monomers and methods for conjugating one or more ligands to an oligonucleotide by click chemistry through attachment of an azide group or a triazolyl moiety at the anomeric site of a pentose or hexose sugar. Another aspect of the invention relates to a method of modulating expression of a target gene in a cell comprising administering to the cell an oligonucleotide and / or a dsRNA molecule as described herein. TIFF2024525713000174.tif105170

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit under 35 U.S.C. § 119(e) of U.S. Provisional Patent Application No. 63 / 236,029, filed August 23, 2021, and U.S. Provisional Patent Application No. 63 / 222,090, filed July 15, 2021, the entire contents of which are incorporated herein by reference in their entirety.

[0002] Technical Field The present disclosure relates generally to monomers and methods for conjugating one or more ligands to oligonucleotides by azide-alkyne cycloaddition (AAC or "click") chemistry at the anomeric position of a sugar, such as a pentose or hexose sugar. [Background technology]

[0003] background There is a need in the art for monomers and methods for conjugating ligands to oligonucleotides. The present disclosure addresses these needs. Summary of the Invention

[0004] overview In one embodiment, a compound of formula (III): Provided herein is a compound of formula TIFF2024525713000002.tif34128, wherein: R 1 is N3 or TIFF2024525713000003.tif18128, During the ceremony, a is 0 or 1, n is 1, 2, 3, 4, or 5; R B is O, N, S, heteroalkyl, branched alkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; Each R C is, independently, TIFF2024525713000004.tif18128, During the ceremony, each b' is independently 0 or 1; each L is independently absent or a linker; Each R L The ligands (e.g., carbohydrates, lipids, vitamins, peptides, proteins, lipoproteins, peptidomimetics, polyamines, nucleosides and nucleotides, oligonucleotides, therapeutic agents, diagnostic agents, detectable labels, antibodies or fragments thereof, optionally substituted C 1-30 Alkyl, optionally substituted C 1-30 Alkenyl, optionally substituted C 1-30 alkynyl, and polyethylene glycol (PEG); R 32 is hydrogen, hydroxy, halogen, protected hydroxy, phosphate group, reactive phosphorus group, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), a solid support, a linker, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—); R 33 is hydrogen, hydroxy, halogen, protected hydroxy, phosphate group, reactive phosphorus group, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), a solid support, a linker, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—), and optionally, R 32 and R 33 is a phosphate group, a reactive phosphorus group, a solid support, a linker, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—); R 4 is hydrogen, optionally substituted C 1-6 Alkyl, optionally substituted C 2-6 Alkenyl, optionally substituted C 2-6 Alkynyl or optionally substituted C 1-6 Is it an alkoxy? or R 4 and R 32 together, 4'-C(R 10 R 11 ) v -Y-2' or 4'-YC(R 10 R 11 ) v -2', Y is -O-, -CH2-, -CH(Me)-, -C(CH3)2-, -S-, -N(R 12 )-, -C(O)-, -C(S)-, -S(O)-, -S(O)2-, -OC(O)-, -C(O)O-, -N(R 12 )C(O)- or -C(O)N(R 12 )- and R 10 and R 11are independently H, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, or optionally substituted C2-C6 alkynyl; R 12 is hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C1-C 30 Alkoxy, C 1-4 Haloalkyl, optionally substituted C 2-4 Alkenyl, optionally substituted C 2-4 Alkynyl, optionally substituted C 1-30 alkyl-COH or nitrogen protecting group, v is 1, 2, or 3; or R 4 and R 33 together with the atoms to which they are attached, may be substituted C 3-8 Cycloalkyl, optionally substituted C 3-8 forming a cycloalkenyl or an optionally substituted 3- to 8-membered heterocyclyl, R 35 is a hydroxyl, a protected hydroxyl, a phosphate group, an optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9), vinyl phosphonate (VP) group, C3-6 cycloalkyl phosphonate (e.g., cyclopropyl phosphonate), monophosphate ((HO)2(O)PO-5'), diphosphate ((HO)2(O)POP(HO)(O)-O-5'), triphosphate ((HO)2(O)PO-(HO)(O)POP(HO)(O)-O-5'), monothiophosphate (phosphorothioate, (HO)2(S)PO-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)PO-5'), phosphorothiolate ((HO)2 (O)PS-5'), alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate ((HO)2(O)P-NH-5', (HO)(NH2)(O)PO-5'), alkyl phosphonate (R(OH)(O)PO-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonate (R(OH)(O)PO-5', R = alkyl ether, e.g., methoxymethyl (CHOMe), ethoxymethyl, etc.), (HO)2(X)PO[-(CH2) a -OP(X)(OH)-O] b -5', or (HO)2(X)PO[-(CH2) a -P(X)(OH)-O] b -5', or (HO)2(X)P-[-(CH2) a -OP(X)(OH)-O] b -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl-terminated phosphates and phosphate mimetics (e.g., HO[-(CH) a -OP(X)(OH)-O] b -5', H2N[-(CH2) a -OP(X)(OH)-O] b -5', H[-(CH2) a -OP(X)(OH)-O] b -5', Me2N[-(CH2) a -OP(X)(OH)-O] b -5', HO[-(CH2) a -P(X)(OH)-O] b-5', H2N[-(CH2) a -P(X)(OH)-O] b -5', H[-(CH2) a -P(X)(OH)-O] b -5', Me2N[-(CH2) a -P(X)(OH)-O] b -5' (wherein X is O or S, and a and b are each independently 1 to 10), Each R 8 and R 9 are independently selected from H, a targeting ligand (e.g., GalNac), a pharmacokinetic modifier, an optionally substituted C 1-30 Alkyl, optionally substituted C 1-30 Alkenyl, or optionally substituted C 1-30 It is alkynyl.

[0005] In some embodiments of any one of the aspects described herein, the compound of Formula (III) has the formula (IIIa): The compound is TIFF2024525713000005.tif35128.

[0006] In some embodiments of any one of the aspects described herein, the compound of Formula (III) has the formula (IIIb): The compound is TIFF2024525713000006.tif35128.

[0007] In another embodiment, the compound of formula (IIIc): Provided herein is a compound of formula TIFF2024525713000007.tif31128, wherein: R 32 is hydrogen, hydroxy, halogen, protected hydroxy, phosphate group, reactive phosphorus group, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), a solid support, a linker, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—); R 33 is hydrogen, hydroxy, halogen, protected hydroxy, phosphate group, reactive phosphorus group, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), a solid support, a linker, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—), and optionally, R 32 and R 33 is a phosphate group, a reactive phosphorus group, a solid support, a linker, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—); R 4 is hydrogen, optionally substituted C 1-6 Alkyl, optionally substituted C 2-6 Alkenyl, optionally substituted C 2-6 Alkynyl or optionally substituted C 1-6 Is it an alkoxy? or R 4 and R 32 together, 4'-C(R 10 R 11 ) v -Y-2' or 4'-YC(R 10 R 11 ) v -2', Y is -O-, -CH2-, -CH(Me)-, -C(CH3)2-, -S-, -N(R 12 )-, -C(O)-, -C(S)-, -S(O)-, -S(O)2-, -OC(O)-, -C(O)O-, -N(R 12 )C(O)- or -C(O)N(R 12 )- and R 10 and R 11 are independently H, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, or optionally substituted C2-C6 alkynyl; R 12 is hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C1-C 30 Alkoxy, C 1-4 Haloalkyl, optionally substituted C 2-4 Alkenyl, optionally substituted C 2-4 Alkynyl, optionally substituted C 1-30 alkyl-COH or nitrogen protecting group, v is 1, 2, or 3; or R 4 and R 33 together with the atoms to which they are attached, may be substituted C 3-8 Cycloalkyl, optionally substituted C 3-8 forming a cycloalkenyl or an optionally substituted 3- to 8-membered heterocyclyl, R 35 is a hydroxyl, a protected hydroxyl, a phosphate group, an optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30Alkynyl, optionally substituted C 1-30 Alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), vinyl phosphonate (VP) group, C3-6 cycloalkyl phosphonate (e.g., cyclopropyl phosphonate), monophosphate ((HO)2(O)PO-5'), diphosphate ((HO)2(O)POP(HO)(O)-O-5'), triphosphate ((HO)2(O)PO-(HO)(O)POP(HO)(O)-O-5'), monothiophosphate (phosphorothioate, (HO)2(S)PO-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)PO-5'), phosphorothiolate ((HO)2 (O)PS-5'), alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate ((HO)2(O)P-NH-5', (HO)(NH2)(O)PO-5'), alkyl phosphonate (R(OH)(O)PO-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonate (R(OH)(O)PO-5', R = alkyl ether, e.g., methoxymethyl (CHOMe), ethoxymethyl, etc.), (HO)2(X)PO[-(CH2) a -OP(X)(OH)-O] b -5', or (HO)2(X)PO[-(CH2) a -P(X)(OH)-O] b -5', or (HO)2(X)P-[-(CH2) a -OP(X)(OH)-O] b -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl-terminated phosphates and phosphate mimetics (e.g., HO[-(CH) a-OP(X)(OH)-O] b -5', H2N[-(CH2) a -OP(X)(OH)-O] b -5', H[-(CH2) a -OP(X)(OH)-O] b -5', Me2N[-(CH2) a -OP(X)(OH)-O] b -5', HO[-(CH2) a -P(X)(OH)-O] b -5', H2N[-(CH2) a -P(X)(OH)-O] b -5', H[-(CH2) a -P(X)(OH)-O] b -5', Me2N[-(CH2) a -P(X)(OH)-O] b -5' (wherein X is O or S, and a and b are each independently 1 to 10), Each R 8 and R 9 are independently selected from H, a targeting ligand (e.g., GalNac), a pharmacokinetic modifier, an optionally substituted C 1-30 Alkyl, optionally substituted C 1-30 Alkenyl, or optionally substituted C 1-30 is alkynyl, Q, Z, and m are defined as one of the sets (i), (ii), or (iii), where: (i) Q is optionally substituted aryl (e.g., phenyl) or optionally substituted heteroaryl; m is an integer selected from 1 to the maximum number of substituents on Q (e.g., when Q is phenyl, m is 1, 2, 3, 4, or 5, e.g., 1, 2, or 3, or 1 or 2); Each Z is -Z 1 , -Z 2 , or -C(R C )3, wherein: R C are aryl (e.g., phenyl or naphthyl) or heteroaryl, each of which is one or more Z1 group or Z 2 substituted with (e.g., 1, 2, or 3) groups; each Z 1 teeth, TIFF2024525713000008.tif15128, wherein R N is hydrogen or C 1-6 is alkyl, Z 2 teeth, TIFF2024525713000009.tif10128 or (ii) m is 1; Q is -CH2O-, -CH2S-, or -CH2N(R N )-, where N, O, or S is attached to Z; Z is TIFF2024525713000010.tif8128, -(CH2) 0-1 -Y-(Z 3 ) p , -C(H)(CHZ 1 )2, -CH2C(H)(CH2Z 1 )2, or -CH2C(CH2Z 1 )3, wherein: Y is an optionally substituted aryl or an optionally substituted heteroaryl; each Z 3 is Z 1 or Z 2 and p is an integer selected from 1 to the maximum number of substituents on Y (e.g., when Y is phenyl, p is 1, 2, 3, 4, or 5, e.g., 1, 2, or 3, or 1 or 2); or (iii) Q is -CHN-; m is 2, Each Z is TIFF2024525713000011.tif9128, -(CH2) 0-1 -Y-(Z 3 ) p , or -CH2C(CH2Z 1 )3.

[0008] In some embodiments of any one of the described aspects, R 35 is a protected hydroxy (e.g., 4,4'-dimethoxytrityl protected) or phosphate group, and R 33 is a hydroxy, or a reactive phosphorus group (e.g., a phosphoramidite, such as 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, or 3'-[(β-thiobenzoylethyl)-(1-pyrrolidinyl)]-thiophosphoramidite).

[0009] In some embodiments of any one of the aspects described herein, R 32 is hydrogen, hydroxy, halogen, protected hydroxy, optionally substituted C alkyl, optionally substituted C alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R8)(CH2R9), -OC 4-30 Alkyl-ON(CH2R8)(CH2R9).

[0010] In some embodiments of any one of the aspects described herein, R 32 is hydrogen, hydroxy, fluoro, chloro, methoxy, ethoxy, 2-methoxyethyl, or C 6-24 Alkyl (e.g., nC 6-24 alkyl).

[0011] In some embodiments of compounds of Formula (IIIc), each Z is independently: TIFF2024525713000012.tif55160.

[0012] In another embodiment, the compound of formula (IV): Provided herein is a compound of formula TIFF2024525713000013.tif46128, wherein: L P is absent or is a linker, R 1 is N3 or TIFF2024525713000014.tif19128, During the ceremony, a is 0 or 1, n is 1, 2, 3, 4, or 5; R B is O, N, S, heteroalkyl, branched alkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; Each R C is, independently, TIFF2024525713000015.tif18128, During the ceremony, each b' is independently 0 or 1; each L is independently absent or a linker; Each R L The ligands (e.g., carbohydrates, lipids, vitamins, peptides, proteins, lipoproteins, peptidomimetics, polyamines, nucleosides and nucleotides, oligonucleotides, therapeutic agents, diagnostic agents, detectable labels, antibodies or fragments thereof, optionally substituted C 1-30 Alkyl, optionally substituted C 1-30 Alkenyl, optionally substituted C 1-30 alkynyl, and polyethylene glycol (PEG); R 42 is a hydroxyl, halogen, protected hydroxyl, phosphate group, reactive phosphorus group, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), a solid support, a linker, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—); R 45 is a hydroxyl, a protected hydroxyl, a phosphate group, an optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9), vinyl phosphonate (VP) group, C3-6 cycloalkyl phosphonate (e.g., cyclopropyl phosphonate), monophosphate ((HO)2(O)PO-5'), diphosphate ((HO)2(O)POP(HO)(O)-O-5'), triphosphate ((HO)2(O)PO-(HO)(O)POP(HO)(O)-O-5'), monothiophosphate (phosphorothioate, (HO)2(S)PO-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)PO-5'), phosphorothiolate ((HO)2 (O)PS-5'), alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate ((HO)2(O)P-NH-5', (HO)(NH2)(O)PO-5'), alkyl phosphonate (R(OH)(O)PO-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonate (R(OH)(O)PO-5', R = alkyl ether, e.g., methoxymethyl (CHOMe), ethoxymethyl, etc.), (HO)2(X)PO[-(CH2) a -OP(X)(OH)-O] b -5', or (HO)2(X)PO[-(CH2) a -P(X)(OH)-O] b -5', or (HO)2(X)P-[-(CH2) a -OP(X)(OH)-O] b -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl-terminated phosphates and phosphate mimetics (e.g., HO[-(CH) a -OP(X)(OH)-O] b -5', H2N[-(CH2) a -OP(X)(OH)-O] b -5', H[-(CH2) a -OP(X)(OH)-O] b -5', Me2N[-(CH2) a -OP(X)(OH)-O] b -5', HO[-(CH2) a -P(X)(OH)-O] b-5', H2N[-(CH2) a -P(X)(OH)-O] b -5', H[-(CH2) a -P(X)(OH)-O] b -5', Me2N[-(CH2) a -P(X)(OH)-O] b -5' (wherein X is O or S, and a and b are each independently 1 to 10), Each R 8 and R 9 are independently selected from H, a targeting ligand (e.g., GalNac), a pharmacokinetic modifier, an optionally substituted C 1-30 Alkyl, optionally substituted C 1-30 Alkenyl, or optionally substituted C 1-30 It is alkynyl.

[0013] In another embodiment, the compound of formula (IVb): Provided herein is a compound of formula TIFF2024525713000016.tif63128, wherein: L P is absent or is a linker, R 42 is a hydroxyl, a halogen-protected hydroxyl, a phosphate group, a reactive phosphorus group, an optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), a solid support, a linker, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—); R45 is a hydroxyl, a protected hydroxyl, a phosphate group, an optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), vinyl phosphonate (VP) group, C3-6 cycloalkyl phosphonate (e.g., cyclopropyl phosphonate), monophosphate ((HO)2(O)PO-5'), diphosphate ((HO)2(O)POP(HO)(O)-O-5'), triphosphate ((HO)2(O)PO-(HO)(O)POP(HO)(O)-O-5'), monothiophosphate (phosphorothioate, (HO)2(S)PO-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)PO-5'), phosphorothiolate ((HO)2 (O)PS-5'), alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate ((HO)2(O)P-NH-5', (HO)(NH2)(O)PO-5'), alkyl phosphonate (R(OH)(O)PO-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonate (R(OH)(O)PO-5', R = alkyl ether, e.g., methoxymethyl (CHOMe), ethoxymethyl, etc.), (HO)2(X)PO[-(CH2) a -OP(X)(OH)-O] b -5', or (HO)2(X)PO[-(CH2) a -P(X)(OH)-O] b -5', or (HO)2(X)P-[-(CH2) a-OP(X)(OH)-O] b -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl-terminated phosphates and phosphate mimetics (e.g., HO[-(CH) a -OP(X)(OH)-O] b -5', H2N[-(CH2) a -OP(X)(OH)-O] b -5', H[-(CH2) a -OP(X)(OH)-O] b -5', Me2N[-(CH2) a -OP(X)(OH)-O] b -5', HO[-(CH2) a -P(X)(OH)-O] b -5', H2N[-(CH2) a -P(X)(OH)-O] b -5', H[-(CH2) a -P(X)(OH)-O] b -5', Me2N[-(CH2) a -P(X)(OH)-O] b -5' (wherein X is O or S, and a and b are each independently 1 to 10), Each R 8 and R 9 are independently selected from H, a targeting ligand (e.g., GalNac), a pharmacokinetic modifier, an optionally substituted C 1-30 Alkyl, optionally substituted C 1-30 Alkenyl, or optionally substituted C 1-30 is alkynyl, Q P , Z P , and m P is defined as one of the set (i), (ii), or (iii), where: (i)Q P is optionally substituted aryl (e.g., phenyl) or optionally substituted heteroaryl; m P From 1 to Q P up to a maximum number of substituents (e.g., Q Pis phenyl, then m is 1, 2, 3, 4, or 5, e.g., 1, 2, or 3, or 1 or 2); each Z P -Z P1 , -Z P2 , or -C(R PC )3, R PC are aryl (e.g., phenyl or naphthyl) or heteroaryl, each of which is one or more Z P1 group or Z P2 substituted with a group (e.g., 1, 2, or 3); each Z P1 teeth, TIFF2024525713000017.tif15128, wherein R N is hydrogen or C 1-6 is alkyl, Z P2 teeth, TIFF2024525713000018.tif10128 or (ii)m P is 1, Q P is -CH2O-, -CH2S-, or -CH2N(R N )-, where N, O, or S is Z P is bound to Z P teeth, TIFF2024525713000019.tif8128, -(CH2) 0-1 -Y-(Z P3 ) pp , -C(H)(CHZ P1 )2, -CH2C(H)(CH2Z P1 )2, or -CH2C(CH2Z P1 )3, wherein: Y P is an optionally substituted aryl or an optionally substituted heteroaryl, each Z P3 is Z P1 or Z P2 and pp is from 1 to Y Pup to a maximum number of substituents (e.g., Y P is phenyl, then pp is 1, 2, 3, 4, or 5, e.g., 1, 2, or 3, or 1 or 2); or or (iii)Q P is -CHN-, m P is 2, each Z P teeth, TIFF2024525713000020.tif9128, -(CH2) 0-1 -Y-(Z P3 ) pp , or -CH2C(CH2Z P1 )3.

[0014] In some embodiments of the compound of Formula (IVb), each Z P is, independently, TIFF2024525713000021.tif50128.

[0015] In some embodiments of any one of the described aspects, R 42 is a hydroxy, protected hydroxy, or phosphate group (e.g., a phosphoramidite, such as 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, or 3'-[(β-thiobenzoylethyl)-(1-pyrrolidinyl)]-thiophosphoramidite).

[0016] In some embodiments of any one of the aspects described herein, R 45is a hydroxy, a protected hydroxy, a vinylphosphonate group, a monophosphate, a diphosphate, a triphosphate, a monothiophosphate (phosphorothioate), a monodithiophosphate (phosphorodithioate), a phosphorothiolate, an alpha-thiotriphosphate, a beta-thiotriphosphate, a gamma-thiotriphosphate, a phosphoramidate, or an alkylphosphonate. 45 is a hydroxy, protected hydroxy, or vinylphosphonate group.

[0017] In some embodiments of any one of the aspects described herein, R 42 is a hydroxy, protected hydroxy, or reactive phosphorus group (e.g., a phosphoramidite, such as 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, or 3'-[(β-thiobenzoylethyl)-(1-pyrrolidinyl)]-thiophosphoramidite), and R 45 is a hydroxy, a protected hydroxy, a vinylphosphonate group, a monophosphate, a diphosphate, a triphosphate, a monothiophosphate (phosphorothioate), a monodithiophosphate (phosphorodithioate), a phosphorothiolate, an alpha-thiotriphosphate, a beta-thiotriphosphate, a gamma-thiotriphosphate, a phosphoramidate, or an alkylphosphonate. 42 is a hydroxy, protected hydroxy, or reactive phosphorus group, and R 45 is a hydroxy, protected hydroxy, or vinylphosphonate group.

[0018] In some embodiments of any one of the aspects described herein, R 42 is a reactive phosphorus group, and R 45 is a protected hydroxy.

[0019] In another embodiment, compounds of formula VI, VII, VIII, or IX: Provided herein is a compound of formula TIFF2024525713000022.tif75128, wherein: R 1 is N3 or TIFF2024525713000023.tif19128, During the ceremony, a is 0 or 1, n is 1, 2, 3, 4, or 5; R B is O, N, S, heteroalkyl, branched alkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; Each R C is, independently, TIFF2024525713000024.tif18128, During the ceremony, each b' is independently 0 or 1; each L is independently absent or a linker; Each R L The ligands (e.g., carbohydrates, lipids, vitamins, peptides, proteins, lipoproteins, peptidomimetics, polyamines, nucleosides and nucleotides, oligonucleotides, therapeutic agents, diagnostic agents, detectable labels, antibodies or fragments thereof, optionally substituted C 1-30 Alkyl, optionally substituted C 1-30 Alkenyl, optionally substituted C 1-30 alkynyl, and polyethylene glycol (PEG); R 62 is a hydroxy, a protected hydroxy, a phosphate group, a reactive phosphorus group, an optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), a solid support, a linker, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—); R 63 and R 64 are independently hydrogen, hydroxy, halogen, protected hydroxy, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), or -O-lipid, R 65 is a hydroxyl, a protected hydroxyl, a phosphate group, an optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9), vinyl phosphonate (VP) group, C3-6 cycloalkyl phosphonate (e.g., cyclopropyl phosphonate), monophosphate ((HO)2(O)PO-5'), diphosphate ((HO)2(O)POP(HO)(O)-O-5'), triphosphate ((HO)2(O)PO-(HO)(O)POP(HO)(O)-O-5'), monothiophosphate (phosphorothioate, (HO)2(S)PO-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)PO-5'), phosphorothiolate ((HO)2 (O)PS-5'), alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate ((HO)2(O)P-NH-5', (HO)(NH2)(O)PO-5'), alkyl phosphonate (R(OH)(O)PO-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonate (R(OH)(O)PO-5', R = alkyl ether, e.g., methoxymethyl (CHOMe), ethoxymethyl, etc.), (HO)2(X)PO[-(CH2) a -OP(X)(OH)-O] b -5', or (HO)2(X)PO[-(CH2) a -P(X)(OH)-O] b -5', or (HO)2(X)P-[-(CH2) a -OP(X)(OH)-O] b -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl-terminated phosphates and phosphate mimetics (e.g., HO[-(CH) a -OP(X)(OH)-O] b -5', H2N[-(CH2) a -OP(X)(OH)-O] b -5', H[-(CH2) a -OP(X)(OH)-O] b -5', Me2N[-(CH2) a -OP(X)(OH)-O] b -5', HO[-(CH2) a -P(X)(OH)-O] b-5', H2N[-(CH2) a -P(X)(OH)-O] b -5', H[-(CH2) a -P(X)(OH)-O] b -5', Me2N[-(CH2) a -P(X)(OH)-O] b -5' (wherein X is O or S, and a and b are each independently 1 to 10), Each R 8 and R 9 are independently selected from H, a targeting ligand (e.g., GalNac), a pharmacokinetic modifier, an optionally substituted C 1-30 Alkyl, optionally substituted C 1-30 Alkenyl, or optionally substituted C 1-30 It is alkynyl.

[0020] In some embodiments of any one of the aspects described herein, R 62 , R 63 , R 64 , and R 65 At least one of R is not hydroxyl. 62 , R 63 , R 64 , and R 65 At least two of R are not simultaneously hydroxyl. 62 and R 63 is not simultaneously hydroxyl. In some embodiments, R 62 and R 64 is not simultaneously hydroxyl. In some embodiments, R 62 and R 65 is not simultaneously hydroxyl. In some embodiments, R 63 and R 64 is not simultaneously hydroxyl. In some embodiments, R 63 and R 65 is not simultaneously hydroxyl. In some embodiments, R 65 is not simultaneously hydroxyl. In some embodiments, R 62 , R 63, R 64 , and R 65 At least three of R are not simultaneously hydroxyl. 62 , R 63 , R 64 , and R 65 All four of these are not simultaneously hydroxyl.

[0021] In another embodiment, the compound of formula VIb, VIIb, VIIIb, or IXb: Provided herein is a compound of formula TIFF2024525713000025.tif177140, wherein: R 62 is a hydroxy, a protected hydroxy, a phosphate group, a reactive phosphorus group, an optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), a solid support, a linker, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—); R 63 and R 64 are independently hydrogen, hydroxy, halogen, protected hydroxy, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), or -O-lipid, R 65 is a hydroxyl, a protected hydroxyl, a phosphate group, an optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9), vinyl phosphonate (VP) group, C3-6 cycloalkyl phosphonate (e.g., cyclopropyl phosphonate), monophosphate ((HO)2(O)PO-5'), diphosphate ((HO)2(O)POP(HO)(O)-O-5'), triphosphate ((HO)2(O)PO-(HO)(O)POP(HO)(O)-O-5'), monothiophosphate (phosphorothioate, (HO)2(S)PO-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)PO-5'), phosphorothiolate ((HO)2 (O)PS-5'), alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate ((HO)2(O)P-NH-5', (HO)(NH2)(O)PO-5'), alkyl phosphonate (R(OH)(O)PO-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonate (R(OH)(O)PO-5', R = alkyl ether, e.g., methoxymethyl (CHOMe), ethoxymethyl, etc.), (HO)2(X)PO[-(CH2) a -OP(X)(OH)-O] b -5', or (HO)2(X)PO[-(CH2) a -P(X)(OH)-O] b -5', or (HO)2(X)P-[-(CH2) a -OP(X)(OH)-O] b -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl-terminated phosphates and phosphate mimetics (e.g., HO[-(CH) a -OP(X)(OH)-O] b -5', H2N[-(CH2) a -OP(X)(OH)-O] b -5', H[-(CH2) a -OP(X)(OH)-O] b -5', Me2N[-(CH2) a -OP(X)(OH)-O] b -5', HO[-(CH2) a -P(X)(OH)-O] b-5', H2N[-(CH2) a -P(X)(OH)-O] b -5', H[-(CH2) a -P(X)(OH)-O] b -5', Me2N[-(CH2) a -P(X)(OH)-O] b -5' (wherein X is O or S, and a and b are each independently 1 to 10), Each R 8 and R 9 are independently selected from H, a targeting ligand (e.g., GalNac), a pharmacokinetic modifier, an optionally substituted C 1-30 Alkyl, optionally substituted C 1-30 Alkenyl, or optionally substituted C 1-30 is alkynyl, Q H , Z H , and m H is defined as one of the set (i), (ii), or (iii), where: (i)Q H is optionally substituted aryl (e.g., phenyl) or optionally substituted heteroaryl; m H From 1 to Q H up to a maximum number of substituents (e.g., Q H is phenyl, then m is 1, 2, 3, 4, or 5, e.g., 1, 2, or 3, or 1 or 2); each Z H -Z H1 , -Z H2 , or -C(R HC )3, wherein: R HC are aryl (e.g., phenyl or naphthyl) or heteroaryl, each of which is one or more Z H1 group or Z H2 substituted with a group (e.g., 1, 2, or 3); each Z H1 teeth, TIFF2024525713000026.tif15128, wherein R N is hydrogen or C 1-6 is alkyl, Z H2 teeth, TIFF2024525713000027.tif10128 or (ii)m H is 1, Q H is -CH2O-, -CH2S-, or -CH2N(R N )-, where N, O, or S is Z H is bound to Z H teeth, TIFF2024525713000028.tif8128, -(CH2) 0-1 -Y-(Z H3 ) hp , -C(H)(CHZ P1 )2, -CH2C(H)(CH2Z H1 )2, or -CH2C(CH2Z H1 )3, wherein: Y H is an optionally substituted aryl or an optionally substituted heteroaryl, each Z H3 is Z H1 or Z H2 and hp is 1 to Y H up to a maximum number of substituents (e.g., Y H is phenyl, then hp is 1, 2, 3, 4, or 5, for example, 1, 2, or 3, or 1 or 2); or or (iii)Q H is -CHN-, m H is 2, each Z H teeth, TIFF2024525713000029.tif9128, -(CH2) 0-1 -Y-(Z H3 ) hp, or -CH2C(CH2Z H1 )3.

[0022] In some embodiments of compounds VIb, VIIb, VIIIb, or H teeth, TIFF2024525713000030.tif50128.

[0023] In some embodiments of any one of the described aspects, R 62 is a hydroxy, protected hydroxy, or phosphate group (e.g., a phosphoramidite, such as 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, or 3'-[(β-thiobenzoylethyl)-(1-pyrrolidinyl)]-thiophosphoramidite).

[0024] In some embodiments of any one of the aspects described herein, R 65 is a hydroxy, a protected hydroxy, a vinylphosphonate group, a monophosphate, a diphosphate, a triphosphate, a monothiophosphate (phosphorothioate), a monodithiophosphate (phosphorodithioate), a phosphorothiolate, an alpha-thiotriphosphate, a beta-thiotriphosphate, a gamma-thiotriphosphate, a phosphoramidate, or an alkylphosphonate. 65 is a hydroxy, protected hydroxy, or vinylphosphonate group.

[0025] In some embodiments of any one of the aspects described herein, R 62 is a hydroxy, protected hydroxy, or reactive phosphorus group (e.g., a phosphoramidite, such as 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, or 3'-[(β-thiobenzoylethyl)-(1-pyrrolidinyl)]-thiophosphoramidite), and R65 is a hydroxy, a protected hydroxy, a vinylphosphonate group, a monophosphate, a diphosphate, a triphosphate, a monothiophosphate (phosphorothioate), a monodithiophosphate (phosphorodithioate), a phosphorothiolate, an alpha-thiotriphosphate, a beta-thiotriphosphate, a gamma-thiotriphosphate, a phosphoramidate, or an alkylphosphonate. 62 is a hydroxy, protected hydroxy, or reactive phosphorus group, and R 65 is a hydroxy, protected hydroxy, or vinylphosphonate group.

[0026] In some embodiments of any one of the aspects described herein, R 62 is a reactive phosphorus group, and R 65 is a protected hydroxy.

[0027] In some embodiments of any one of the aspects described herein, R 62 is a hydroxy, protected hydroxy, or reactive phosphorus group (e.g., a phosphoramidite, such as 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, or 3'-[(β-thiobenzoylethyl)-(1-pyrrolidinyl)]-thiophosphoramidite), and R 63 and R 64 are independently hydrogen, hydroxy, halogen, protected hydroxy, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30Alkyl-ON(CH2R 8 )(CH2R 9 ), or -O-lipid, R 65 is a hydroxy, a protected hydroxy, a vinylphosphonate group, a monophosphate, a diphosphate, a triphosphate, a monothiophosphate (phosphorothioate), a monodithiophosphate (phosphorodithioate), a phosphorothiolate, an alpha-thiotriphosphate, a beta-thiotriphosphate, a gamma-thiotriphosphate, a phosphoramidate, or an alkylphosphonate. 62 is a hydroxy, protected hydroxy, or reactive phosphorus group, and R 63 and R 64 are independently hydroxy, protected hydroxy, optionally substituted C 1-30 alkoxy (e.g., methoxy), alkoxyalkylamine, amino, alkylamino, dialkylamino, or -O-lipid; R 65 is a hydroxy, protected hydroxy, or vinylphosphonate group.

[0028] In some embodiments of any one of the aspects described herein, R 62 is a reactive phosphorus group, and R 63 and R 64 are independently hydroxy, protected hydroxy, optionally substituted C 1-30 alkoxy (e.g., methoxy), alkoxyalkylamine, amino, alkylamino, dialkylamino, or -O-lipid; R 65 is a protected hydroxy.

[0029] The compounds of formula (III), (IIIa), (IIIb), (IIIc), (IV), (IVb), (VI)-(IX), and (VIb)-(IXb) are useful in synthetic oligonucleotides. Thus, in another aspect, oligonucleotides prepared using compounds of formula (III), (IIIa), (IIIb), (IIIc), (IV), (IVb), (VI), (VIb), (VII), (VIIb), (VIII), (VIIIb), (IX), or (IXb) are provided herein. For example, oligonucleotides comprising nucleosides of formula (I): TIFF2024525713000031.tif117134In formula, L p is absent or is a linker, R 1 is N3 or TIFF2024525713000032.tif19128, During the ceremony, a' is 0 or 1; n is 1, 2, 3, 4, or 5; R B is O, N, S, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; Each R C is, independently, TIFF2024525713000033.tif18128, During the ceremony, each b' is independently 0 or 1; each L is independently absent or a linker; Each R L may be carbohydrates, lipids, vitamins, peptides, proteins, lipoproteins, peptidomimetics, polyamines, nucleosides and nucleotides, oligonucleotides, therapeutic agents, diagnostic agents, detectable labels, antibodies or fragments thereof, optionally substituted C 1-30 Alkyl, optionally substituted C 1-30 Alkenyl, optionally substituted C 1-30 independently selected from the group consisting of alkynyl, and polyethylene glycol (PEG); R2 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., 2-methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, 5- to 8-membered heterocyclyl, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), or -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), a bond to an internucleotide bond to a subsequent nucleotide, a 3'-oligonucleotide capping group, a ligand, a linker covalently attached to one or more ligands, a solid support, a linker, or a linker covalently attached to a solid support; R 3 , R 52 , and R 62x independently, a bond to the internucleotide bond to the subsequent nucleotide, hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., 2-methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, 5- to 8-membered heterocyclyl, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9), a 3'-oligonucleotide capping group, a ligand, a linker covalently attached to one or more ligands, a solid support, a linker, or a linker covalently attached to a solid support; R 4 is hydrogen, optionally substituted C 1-6 Alkyl, optionally substituted C 2-6 Alkenyl, optionally substituted C 2-6 Alkynyl or optionally substituted C 1-6 Is it an alkoxy? or R 4 and R 2 together, 4'-C(R 10 R 11 ) v -Y-2' or 4'-YC(R 10 R 11 ) v -2', Y is -O-, -CH2-, -CH(Me)-, -C(CH3)2-, -S-, -N(R 12 )-, -C(O)-, -C(S)-, -S(O)-, -S(O)2-, -OC(O)-, -C(O)O-, -N(R a13 )C(O)- or -C(O)N(R 12 )- and R 10 and R 11 are independently H, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, or optionally substituted C2-C6 alkynyl; R 12 is hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C1-C 30 Alkoxy, C 1-4 Haloalkyl, optionally substituted C 2-4 Alkenyl, optionally substituted C 2-4 Alkynyl, optionally substituted C 1-30 alkyl-COH or a nitrogen protecting group, v is 1, 2, or 3; or R 4 and R 3C, together with the atoms to which they are attached, may be substituted 3-8 Cycloalkyl, optionally substituted C 3-8 forming a cycloalkenyl or an optionally substituted 3- to 8-membered heterocyclyl, R 5 , R 55 , and R 65x are independently a bond to the internucleotide bond to the preceding nucleotide, hydrogen, hydroxy, protected hydroxy, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, optionally substituted 3- to 8-membered heterocyclyl (e.g., morpholin-1-yl, piperidin-1-yl, or pyrrolidin-1-yl), halogen, alkoxyalkyl (e.g., 2-methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), vinylphosphonate (VP) group, C 3-6 Cycloalkylphosphonates (e.g., cyclopropylphosphonate), monophosphates ((HO)2(O)PO-5'), diphosphates ((HO)2(O)POP(HO)(O)-O-5'), triphosphates ((HO)2(O)PO-(HO)(O)POP(HO)(O)-O-5'); monothiophosphates (phosphorothioates, (HO)2(S)PO-5'), monodithiophosphates (phosphorodithioates, (HO)(HS)(S)PO-5'), phosphorothiolates ((HO)2(O)PS-5'); alpha-thiotriphosphates; beta-thiotriphosphates; gamma-thiotriphosphates; phosphoramidates ((HO)2(O)P-NH-5', (HO)(NH2)(O)PO-5'), alkylphosphonates [(RP )(OH)(O)PO-5', R P may be substituted C 1-30 alkyl, for example, methyl, ethyl, isopropyl, or propyl)], alkyl ether phosphonates [(R P1 )(OH)(O)PO-5', R P1 is alkoxyalkyl, for example, methoxymethyl (CH2OMe) or ethoxymethyl], (HO)2(X)PO[—(CH2) a -OP(X)(OH)-O] b -5', or (HO)2(X)PO[-(CH2) a -P(X)(OH)-O] b -5', or (HO)2(X)P-[-(CH2) a -OP(X)(OH)-O] b -5', or optionally substituted alkyl, and dialkyl-terminated phosphates and phosphate mimetics (e.g., HO[-(CH) a -OP(X)(OH)-O] b -5', H2N[-(CH2) a -OP(X)(OH)-O] b -5', H[-(CH2) a -OP(X)(OH)-O] b -5', Me2N[-(CH2) a -OP(X)(OH)-O] b -5', HO[-(CH2) a -P(X)(OH)-O] b -5', H2N[-(CH2) a -P(X)(OH)-O] b -5', H[-(CH2) a -P(X)(OH)-O] b -5', Me2N[-(CH2) a -P(X)(OH)-O] b -5'), wherein X is O or S; a and b each independently represent 1 to 10; R 63 and R 64are independently hydrogen, hydroxy, halogen, protected hydroxy, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), or -O-lipid, Each R 8 and R 9 are independently selected from H, a targeting ligand (e.g., GalNac), a pharmacokinetic modifier, an optionally substituted C 1-30 Alkyl, optionally substituted C 1-30 Alkenyl, or optionally substituted C 1-30 is alkynyl, however, (i)R 2 and R 3 is an internucleotide bond to a subsequent nucleotide, (ii)R 2 and R 3 If neither of the two is an internucleotide bond to the subsequent nucleotide, then R 5 is the bond to the internucleotide bond to the preceding nucleotide, (iii)R 52 and R 55 at least one of which is a bond to an internucleotide bond, (vi)R 62x and R 65x At least one of the bonds is an internucleotide bond This is subject to the condition that:

[0030] In the oligonucleotide containing the nucleoside of formula (I), R 2 , R 3 , and R 5 At least one of the bonds is to an internucleotide bond.

[0031] In the oligonucleotide containing the nucleoside of formula (V), R 42 and R 45 At least one of the bonds is to an internucleotide bond.

[0032] In the oligonucleotides containing a nucleoside of formula (VIx), (VIIx), (VIIIx), or (IXx), R 62x and R 65x At least one of the bonds is to an internucleotide bond.

[0033] In some embodiments of any one of the aspects described herein, the nucleoside of Formula (I) is of Formula (Ia): Nucleoside of TIFF2024525713000034.tif34128.

[0034] In some embodiments of any one of the aspects described herein, the nucleoside of Formula (I) is of Formula (Ib): Nucleoside of TIFF2024525713000035.tif35128.

[0035] In yet another aspect, provided herein is a double-stranded nucleic acid comprising a first strand and a second strand complementary to the first strand, wherein at least one of the first and second strands is an oligonucleotide comprising a nucleotide of Formula (I) described herein.

[0036] In another aspect, provided herein is a method for inhibiting or reducing expression of a target gene in a subject, the method comprising administering to the subject (i) a double-stranded RNA described herein, wherein one strand of the dsRNA is complementary to the target gene, and / or (ii) an oligonucleotide described herein that is complementary to the target gene. [Brief explanation of the drawings]

[0037] The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Patent and Trademark Office upon request and payment of the necessary fee.

[0038] [Figure 1-1]

[0023] Figure 1 is a schematic diagram of the synthesis of multivalent conjugates via CuAAC click chemistry on pentose sugars. Only the β isomer is shown for clarity. [Figure 1-2] See description of Figure 1-1. [Figure 1-3] See description of Figure 1-1. [Figure 2-1]

[0023] Figure 1 is a schematic diagram of the synthesis of multivalent conjugates via CuAAC click chemistry on pentose sugars. Only the β isomer is shown for clarity. [Figure 2-2] See description of Figure 2-1. [Figure 2-3] See description of Figure 2-1. [Figure 3-1]

[0023] Figure 1 is a schematic diagram of the synthesis of multivalent conjugates via CuAAC click chemistry on pentose sugars. Only the β isomer is shown for clarity. [Figure 3-2] See description of Figure 3-1. [Figure 4-1] 1 shows exemplary azide containing ligands suitable for conjugation via click chemistry. [Figure 4-2] See description of Figure 4-1. [Figure 5-1]FIG. 1 is a schematic diagram showing various click chemistry-mediated conjugations. [Figure 5-2] See description of Figure 5-1. [Figure 5-3] See description of Figure 5-1. [Figure 6] Figures 6A-6C show various parameters of multiplexed ligand conjugation via 1' click chemistry: α and β anomers (Figure 6A), valency (Figure 6B), and positional isomers of triazoles (Figure 6C). [Figure 7-1] FIG. 1 is a schematic representation of the diverse regiochemical possibilities with a single ligand R. [Figure 7-2] See description of Figure 7-1. [Figure 7-3] See description of Figure 7-1. [Figure 7-4] See description of Figure 7-1. [Figure 8A-1] 8A and 8B show some exemplary ligands suitable for the present invention. [Figure 8A-2] See legend to Figure 8A-1. [Figure 8A-3] See legend to Figure 8A-1. [Figure 8B-1] 8A and 8B show some exemplary ligands suitable for the present invention. [Figure 8B-2] See legend to Figure 8B-1. [Figure 8B-3] See legend to Figure 8B-1. [Figure 9] The various possible configurations for a single construct are shown, with only the β isomer shown for clarity. [Figure 10-1] 1 illustrates an exemplary building block. [Figure 10-2] See description of Figure 10-1. [Figure 11-1] 1 is a synthetic scheme for the synthesis of exemplary building blocks. [Figure 11-2] See description of Figure 11-1. [Figure 12-1]1 is a synthetic scheme for the synthesis of exemplary building blocks. [Figure 12-2] See description of Figure 12-1. [Figure 13-1] 1 is a synthetic scheme for the synthesis of exemplary building blocks. [Figure 13-2] See description of Figure 13-1. [Figure 14] 1'-deoxy sugar anomer: 1H NMR showing assignment of α-configuration. [Figure 15] 1H NMR showing assignment of 1'-deoxy sugar anomer: β-configuration. [Figure 16-1] 1 is a synthetic scheme showing the synthesis of monovalent and trivalent GalNAc azides. [Figure 16-2] See description of Figure 16-1. [Figure 17-1] Figures 17-19 are synthetic schemes showing solution chemistry of conjugated building blocks for oligonucleotide synthesis - lipid ligand multiplexing (Figure 17), lipid multiplexing (Figure 18), and polyamine multiplexing (Figure 19). [Figure 17-2] See description of Figure 17-1. [Figure 18-1] See description of Figure 17-1. [Figure 18-2] See description of Figure 17-1. [Figure 19] See description of Figure 17-1. [Figure 20] An exemplary dsRNA with an exemplary ligand, GalNAc, is shown. [Figure 21] Another exemplary dsRNA is shown, in which the highlighted (Uhd) nucleoside in the control sense strand is replaced, for example, by the nucleoside structure of one of the boxed nucleotide monomers, where "F" refers to a 2'-deoxy-2'-fluoro modified nucleotide and "OMe" refers to a 2'-methoxy modified nucleotide. [Figure 22-1] Some exemplary azido-sugar building blocks are shown. [Figure 22-2]See description of Figure 22-1. [Figure 23] Some exemplary azido-proline building blocks are shown. [Figure 24] Representative polyalkynes that have been prepared (18) 23 or are commercially available (15-17) are shown. [Figure 25-1] Several exemplary amidites derived from CuAAC between sugar building blocks and polyvalent alkynes are shown: ready for click chemistry on solid support. [Figure 25-2] See description of Figure 25-1. [Figure 26-1] Several exemplary CPGs derived from CuAAC between sugar building blocks and polyvalent alkynes are shown: ready for click chemistry on solid support. [Figure 26-2] See description of Figure 26-1. [Figure 27] Several exemplary amidites derived from RuAAC between sugar building blocks and polyvalent alkynes are shown: ready for click chemistry on solid support. [Figure 28] Several exemplary CPGs derived from RuAAC between sugar building blocks and polyvalent alkynes are shown: ready for click chemistry on solid support. [Figure 29] Some exemplary products derived from CuAAC between alkyne monomers and various azides (Figures 8A and 8B) are shown in Figure 25. All triazoles are 1,4-regioisomers. [Figure 30] Some exemplary products derived from RuAAC between alkyne monomers and various azides (Figures 8A and 8B) are shown in Figure 28. All triazoles are 1,5-regioisomers. [Figure 31] Some exemplary products derived from RuAAC between alkyne monomers and various azides (FIGS. 8A and 8B) are shown in Figure 25. Combinations of 1,4- and 1,5-regioisomers. [Figure 32]Figure 28 shows some exemplary products derived from CuAAC between alkyne monomers and various azides (Figures 8A and 8B). Combinations of 1,4- and 1,5-regioisomers. [Figure 33] Several exemplary compounds derived from CuAAC between GalNA-azide 3 / 4 (shown in Figure 22) and mono-, di-, and trivalent alkyne building blocks (Figure 24) are shown. [Figure 34] Several exemplary compounds derived from CuAAC between FuNA-azide 6 (shown in Figure 22) and mono-, di-, and trivalent alkyne building blocks (Figure 24) are shown. [Figure 35] Several exemplary compounds derived from CuAAC between GluNA-azide 7 / 8 (shown in Figure 22) and mono-, di-, and trivalent alkyne building blocks (Figure 24) are shown. [Figure 36] Some exemplary compounds derived from CuAAC between ManNA-azide 10 / 11 (shown in Figure 22) and mono-, di-, and trivalent alkyne building blocks (Figure 24) are shown. [Figure 37] Figure 37A shows immobilized Cu(I) ions on a solid support, and Figure 37B shows immobilized Ru(III) ions on a polymer support. DETAILED DESCRIPTION OF THE INVENTION

[0039] Detailed Description It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not limiting of the invention as claimed. As used herein, the use of the singular includes the plural unless specifically stated otherwise. As used herein, the use of "or" means "and / or" unless specifically stated otherwise. Furthermore, the use of the term "comprising" and other forms such as "comprises" and "includes" is not limiting. Also, unless specifically stated otherwise, terms such as "element" or "component" encompass both elements and components that constitute a single unit and elements and components that constitute more than one subunit.

[0040] The section headings used herein are for organizational purposes only and should not be construed as limiting the subject matter described. All documents, or portions of documents, cited in this application, including but not limited to patents, patent applications, articles, books, and papers, are expressly incorporated herein by reference in their entirety for all purposes.

[0041] In one embodiment, a compound of formula (III): The compound of TIFF2024525713000036.tif35128 is provided herein.

[0042] In another embodiment, the compound of formula (IIIc): The compound of TIFF2024525713000037.tif32128 is provided herein.

[0043] In another embodiment, a compound of formula (I): Provided herein are oligonucleotides containing the nucleosides of TIFF2024525713000038.tif35128.

[0044] In various embodiments described herein, R 1 is N3 or It could be TIFF2024525713000039.tif16128.

[0045] In some embodiments, and therefore in various aspects described herein, R 1 teeth, TIFF2024525713000040.tif19128, where a' can be 0 or 1. In some embodiments, a' is 0. In some other embodiments, a' is 1.

[0046] -(CH2) a’ R B (R C ) n Note that the group can be attached to the triazole group at the 4 or 5 position. Thus, in some embodiments of any one of the aspects, R 1 teeth, TIFF2024525713000041.tif19128. In some other embodiments of any one of the aspects, R 1 teeth, The file is TIFF2024525713000042.tif23128.

[0047] In various embodiments described herein, R B can be O, N, S, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl. In some embodiments of any one of the aspects, R B is O, N, heteroalkyl, or aryl. For example, R B can be O, N, C(CH2O-)4, or benzyl. In some embodiments, R B is O. In some other embodiments, R B is N. In yet some other embodiments, R B is C(CH2O-)4. In yet some other embodiments, R B is benzyl. In some embodiments, R B teeth, The file is TIFF2024525713000043.tif15128.

[0048] In various aspects described herein, n can be 1, 2, 3, 4, or 5. In some embodiments of any one of the aspects described herein, n is 1. In some other embodiments of any one of the aspects described herein, n is 2. In still some other embodiments of any one of the aspects described herein, n is 3. In still some other embodiments of any one of the aspects described herein, n is 4. In still some other embodiments of any one of the aspects described herein, n is 5.

[0049] In some embodiments of any one of the aspects described herein, n is 1 and R B is O.

[0050] In some embodiments of any one of the aspects described herein, n is 2 and R B is N.

[0051] In some embodiments of any one of the aspects described herein, n is 3 and R B is C(CH2O-)4.

[0052] In some embodiments of any one of the aspects described herein, n is 5 and R B is benzyl.

[0053] In some embodiments, R B is phenyl.

[0054] In various embodiments described herein, each R C is, independently, TIFF2024525713000044.tif18128, or -LR L wherein each b' can independently be 0 or 1. In some embodiments, b' is 0. In some other embodiments, b' is 1.

[0055] In some embodiments, R C teeth, TIFF2024525713000045.tif14128, where b' is 0 or 1.

[0056] In some embodiments, R C teeth, TIFF2024525713000046.tif16128, where b' is 0 or 1. Each R C The triazole group in R is bonded to the 4- or 5-position of the triazole. B Note that R C teeth, It could be TIFF2024525713000047.tif30128.

[0057] In some embodiments, b' is 0. Thus, in some embodiments of any one of the aspects described herein, each R C teeth, TIFF2024525713000048.tif9128. In some other embodiments of any one of the aspects described herein, each R C teeth, TIFF2024525713000049.tif16128. Each R C The triazole group in R is bonded to the 4- or 5-position of the triazole. B Note that R may be attached to R. Thus, in some embodiments of any one of the aspects, C teeth, TIFF2024525713000050.tif15128. In some other embodiments of any one of the aspects, R C teeth, The file is TIFF2024525713000051.tif29128.

[0058] R L Embodiments of the various aspects described herein include a group R L Each R Lmay be independently selected from the group consisting of H, carbohydrates, lipids, vitamins, peptides, proteins, lipoproteins, peptidomimetics, polyamines, nucleosides and nucleotides, oligonucleotides, detectable labels, diagnostic agents (e.g., bitoin), fluorescent dyes, polyethylene glycol (PEG), antibodies, antibody fragments (e.g., nanobodies).

[0059] In some embodiments of any one of the aspects described herein, R L is a ligand. Without wishing to be bound by theory, a ligand modifies one or more properties of a binding molecule (e.g., an oligonucleotide described herein), including, but not limited to, pharmacodynamics, pharmacokinetics, binding, absorption, cellular distribution, cellular uptake, charge, and clearance. Ligands are routinely used in the chemical arts and are attached to the parent compound directly or via an optional linking moiety or group. A preferred list of ligands includes, but is not limited to, intercalators, reporter molecules, polyamines, polyamides, polyethylene glycols, thioethers, polyethers, cholesterol, thiocholesterol, cholic acid moieties, folic acid, lipids, phospholipids, biotin, phenazine, phenanthridine, anthraquinone, adamantane, acridine, fluorescein, rhodamine, coumarin, and dyes.

[0060] Preferred ligands suitable for the present invention include lipid moieties, such as cholesterol moieties (Letsinger et al., Proc. Natl. Acad. Sci. USA, 1989, 86, 6553), cholic acid (Manoharan et al., Bioorg. Med. Chem. Lett., 1994, 4, 1053), thioethers, such as hexyl-S-tritylthiol (Manoharan et al., Ann. NY Acad. Sci., 1992, 660, 306; Manoharan et al., Bioorg. Med. Chem. Lett., 1993, 3, 2765), thiocholesterol (Oberhauser et al., Nucl. Acids Res., 1992, 20, 533), aliphatic chains, such as dodecanediol or undecyl residues (Saison-Behmoaras et al., EMBO J., 1991, 10, 111; Kabanov et al., FEBS Lett., 1990, 259, 327; Svinarchuk et al., Biochimie, 1993, 75, 49), phospholipids, such as di-hexadecyl-rac-glycerol or triethylammonium-1,2-di-O-hexadecyl-rac-glycero-3-H-phosphonate (Manoharan et al., Tetrahedron Lett., 1995, 36, 3651; Shea et al., Nucl. Acids Res., 1990, 18, 3777), polyamines or polyethylene glycol chains (Manoharan et al., Nucleosides & Nucleotides, 1995, 14, 969), adamantaneacetic acid (Manoharan et al., Tetrahedron Lett., 1995, 36, 3651), palmityl moiety (Mishra et al., Biochim. Biophys. Acta, 1995, 1264, 229), or octadecylamine or hexylamino-carbonyl-oxycholesterol moiety (Crooke et al., J. Pharmacol. Exp. Ther., 1996, 277, 923).

[0061] Ligands may include naturally occurring molecules, or recombinant or synthetic molecules. Exemplary ligands include, but are not limited to, polylysine (PLL), poly-L-aspartic acid, poly-L-glutamic acid, styrene-maleic anhydride copolymer, poly(L-lactide-co-glycolide) copolymer, divinyl ether-maleic anhydride copolymer, N-(2-hydroxypropyl) methacrylamide copolymer (HMPA), polyethylene glycol (PEG, e.g., PEG-2K, PEG-5K, PEG-10K, PEG-12K, PEG-15K, PEG-20K, PEG-40K), MPEG, [MPEG]2, polyvinyl alcohol (PVA), polyurethane, poly(2-ethylacrylic acid), N-isopropylacrylamide polymer, polyphosphazine, polyethyleneimine, cationic groups, spermine, spermidine, polyamines, pseudopeptide-polyamines, peptidomimetic polyamines, dendrimeric polyamines, arginine, amidine, protamine, cationic lipids, cationic porphyrins, and quaternary polyamines. Salt, thyrotropin, melanotropin, lectin, glycoprotein, surfactant protein A, mucin, glycosylated polyamino acids, transferrin, bisphosphonates, polyglutamates, polyaspartic acid, aptamers, asialofetuin, hyaluronan, procollagen, immunoglobulins (e.g., antibodies), insulin, transferrin, albumin, sugar-albumin conjugates, intercalating agents (e.g., acridine), crosslinking agents (e.g., psoralens, mitogens), mycin C), porphyrins (e.g., TPPC4, texaphyrin, sapphyrin), polycyclic aromatic hydrocarbons (e.g., phenazine, dihydrophenazine), artificial endonucleases (e.g., EDTA), lipophilic molecules (e.g., steroids, bile acids, cholesterol, cholic acid, adamantaneacetic acid, 1-pyrenebutyric acid, dihydrotestosterone, 1,3-Bis-O(hexadecyl)glycerol, zeranyloxyhexyl group, hexadecylglycerol, borneol, menthol, 1,3-propanediol, heptadecyl group, palmitic acid, myristic acid, O3-(oleoyl)lithocholic acid, O3-(oleoyl)cholenoic acid, dimethoxytrityl, or phenoxazine), peptides (e.g., alpha helical peptides, amphipathic peptides, RGD peptides, cell penetrating peptides, endosomolytic / fusogenic peptides), alkylating agents, phosphate, amino, mercapto, polyamino, alkyl, substituted alkyl, radiolabeled markers, enzymes, haptens (e.g., biotin), transport / absorption enhancers (e.g., naproxen, , aspirin, vitamin E, folic acid), synthetic ribonucleases (e.g., imidazole, bis-imidazole, histamine, imidazole clusters, acridine-imidazole conjugates, Eu3+ complexes of tetraazamacrocycles), dinitrophenyl, HRP, AP, antibodies, hormones and hormone receptors, lectins, carbohydrates, polyvalent carbohydrates, vitamins (e.g., vitamin A, vitamin E, vitamin K, vitamin B, e.g., folic acid, B12, riboflavin, biotin, and pyridoxal), vitamin cofactors, lipopolysaccharides, p38 These include activators of MAP kinase, activators of NF-κB, taxon, vincristine, vinblastine, cytochalasin, nocodazole, japlakinolide, latrunculin A, phalloidin, swinholide A, indanocine, myoservin, tumor necrosis factor alpha (TNF alpha), interleukin-1 beta, gamma interferon, natural or recombinant low-density lipoprotein (LDL), natural or recombinant high-density lipoprotein (HDL), and cell-penetrating agents (e.g., helical cell-penetrating agents).

[0062] Peptide and peptidomimetic ligands include naturally occurring or modified peptides, such as D- or L-peptides, α-, β-, or γ-peptides, N-methylpeptides, azapeptides, peptides with one or more amide bonds, i.e., peptide bonds, replaced with one or more urea, thiourea, carbamate, or sulfonylurea bonds, or cyclic peptides. Peptidomimetics (also referred to herein as oligopeptidomimetics) are molecules capable of folding into defined three-dimensional structures similar to natural peptides. Peptide or peptidomimetic ligands are about 5 to 50 amino acids in length, e.g., about 5, 10, 15, 20, 25, 30, 35, 40, 45, or 50 amino acids in length.

[0063] Exemplary amphipathic peptides include, but are not limited to, cecropin, lycotoxin, paradaxin, buforin, CPF, bombinin-like peptide (BLP), cathelicidin, ceratoxin, S. clava peptide, hagfish intestinal antimicrobial peptide (HFIAP), magainin, brevinin-2, dermaseptin, melittin, pleurocidin, H2A peptide, Xenopus peptide, esculentinis-1, and caerin.

[0064] As used herein, the term "endosomolytic ligand" refers to a molecule having endosomolytic properties. An endosomolytic ligand promotes lysis and / or transport of a composition of the invention or its components from a cellular compartment, such as an endosome, lysosome, endoplasmic reticulum (ER), Golgi apparatus, microtubules, peroxisome, or other intracellular endoplasmic reticulum, into the cytoplasm of a cell. Some exemplary endosomolytic ligands include, but are not limited to, imidazoles, poly- or oligoimidazoles, linear or branched polyethyleneimines (PEI), linear and branched polyamines such as spermine, cationic linear and branched polyamines, polycarboxylates, polycations, masked oligo- or polycations or anions, acetals, polyacetals, ketals / polyketals, orthoesters, linear or branched polymers with masked or unmasked cationic or anionic charge, dendrimers with masked or unmasked cationic or anionic charge, polyanionic peptides, polyanionic peptidomimetics, pH-sensitive peptides, natural and synthetic fusogenic lipids, natural and synthetic cationic lipids.

[0065] Exemplary endosomolytic / fusogenic peptides include, but are not limited to, AALEALAEALAEALEALAEAAAAGGC (GALA), AALEALAEALAEALAEALAEALAAAAGGC (EALA), ALEALEALEALAEA, GLFEAIEGFIENGWEGMIWDYG (INF-7), GLFGAIAGFIENGWEGMIDGWYG (Inf HA-2), GLFEAIEGFIENGWEGMIDGWYGCGLFEAIEGFIENGWEGMID GWYGC (diINF-7), GLFEAIEGFIENGWEGMIDGGCGLFEAIEGFIENGWEGMIDGGC (diINF-3), GLFGALAEALAEALAEHLAEALAEALEALAAGGSC (GLF), GLFEAIEGFIENGWEGLAEALAEALEALAAGGSC (GALA-INF3), GLF EAI EGFI ENGW EGnI DG K GLF EAI EGFI These include ENGW EGnI DG (INF-5, where n is norleucine), LFEALLELLESLWELLLEA (JTS-1), GLFKALLKLLKSLWKLLLKA (ppTG1), GLFRALLRLLRSLWRLLLRA (ppTG20), WEAKLAKALAKALAKHLAKALAKALKACEA (KALA), GLFFEAIAEFIEGGWEGLIEGC (HA), GIGAVLKVLTTGLPALISWIKRKRQQ (Melittin), H5WYG, and CHK6HC.

[0066] Without wishing to be bound by theory, fusogenic lipids fuse with membranes, resulting in membrane destabilization. Fusogenic lipids typically have small head groups and unsaturated acyl chains. Exemplary fusogenic lipids include, but are not limited to, 1,2-dileoyl-sn-3-phosphoethanolamine (DOPE), phosphatidylethanolamine (POPE), palmitoyloleoylphosphatidylcholine (POPC), (6Z,9Z,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-ol (Di-Lin), N-methyl(2,2-di((9Z,12Z)-octadeca-9,12-dienyl)-1,3-dioxolan-4-yl)methanamine (DLin-k-DMA), and N-methyl-2-(2,2-di((9Z,12Z)-octadeca-9,12-dienyl)-1,3-dioxolan-4-yl)ethanamine (also referred to herein as XTC).

[0067] Synthetic polymers with endosomolytic activity suitable for the present invention are described in U.S. Patent Application Publication Nos. 2009 / 0048410, 2009 / 0023890, 2008 / 0287630, 2008 / 0287628, 2008 / 0281044, 2008 / 0281041, 2008 / 0269450, 2007 / 0105804, 2007 / 0036865, and 2004 / 0198687, the contents of which are incorporated herein by reference in their entireties.

[0068] Exemplary cell-penetrating peptides include, but are not limited to, RQIKIWFQNRRMKWKK (penetratin), GRKKRRQRRRPPQC (Tat fragment 48-60), GALFLGWLGAAGSTMGAWSQPKKKRKV (signal sequence-based peptide), LLIILRRRIRKQAHAHSK (PVEC), GWTLNSAGYLLKINLKALAALAKKIL (transportan), KLALKLALKALKAALKLA (amphipathic model peptide), RRRRRRRRR (Arg9), KFFKFFKFFK (bacterial cell wall-penetrating peptide), LLGDFFRKSKEKIGKEFKRIVQRIKDFLRNLVPR These include TES (LL-37), SWLSKTAKKLENSAKKRISEGIAIAIQGGPR (cecropin P1), ACYCRIPACIAGERRYGTCIYQGRLWAFCC (α-defensin), DHYNCVSSGGQCLYSACPIFTKIQGTCYRGKAKCCK (β-defensin), RRRPRPPYLPRPRPPPFFPPRLPPRIPPGFPPRFPPRFPGKR-NH2 (PR-39), ILPWKWPWWPWRR-NH2 (indolicidin), AAVALLPAVLLALLAP (RFGF), AALLPVLLAAP (RFGF analog), and RKCRIVVIRVCR (bactenesin).

[0069] Exemplary cationic groups include, but are not limited to, protonated amino groups derived from, for example, O-amines (amine = NH; alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, or diheteroarylamino, ethylenediamine, polyamino); aminoalkoxy, for example, O(CH) nAmines (e.g., amine = NH; alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, or diheteroarylamino, ethylenediamine, polyamino); amino (e.g., NH; alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, or amino acid), and NH(CHCHNH) n Included are CH2CH2-amine (amine = NH2; alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, or diheteroarylamino).

[0070] As used herein, the term "targeting ligand" refers to any molecule that provides enhanced affinity for a selected target, e.g., a cell, cell type, tissue, organ, body region, or compartment, e.g., a cell, tissue, or organ compartment. Some exemplary targeting ligands include, but are not limited to, antibodies, antigens, folate, receptor ligands, carbohydrates, aptamers, integrin receptor ligands, chemokine receptor ligands, transferrin, biotin, serotonin receptor ligands, PSMA, endothelin, GCPII, somatostatin, LDL, and HDL ligands.

[0071] Carbohydrate targeting ligands include, but are not limited to, D-galactose, multivalent galactose, N-acetyl-D-galactosamine (GalNAc), multivalent GalNAc, such as GalNAc2 and GalNAc3; D-mannose, multivalent mannose, multivalent lactose, N-acetyl-glucosamine, multivalent fucose, glycosylated polyamino acids, and lectins. The term multivalent indicates the presence of more than one monosaccharide unit. These monosaccharide subunits can be linked to each other via glycosidic bonds or can be linked to a scaffold molecule.

[0072] Some folates and folate analogs suitable as ligands for the present invention are described in U.S. Pat. Nos. 2,816,110, 5,552,545, 6,335,434, and 7,128,893, the contents of which are incorporated herein by reference in their entireties.

[0073] As used herein, the terms "PK-modulating ligand" and "PK modulator" refer to a molecule capable of modulating the pharmacokinetics of the oligonucleotides described herein. Some exemplary PK modulators include, but are not limited to, lipophilic molecules, bile acids, sterols, phospholipid analogs, peptides, protein binders, vitamins, fatty acids, phenoxazines, aspirin, naproxen, ibuprofen, suprofen, ketoprofen, (S)-(+)-pranoprofen, carprofen, PEG, biotin, and transthyretia-binding ligands (e.g., tetraiidothyroacetic acid, 2,4,6-triiodophenol, and flufenamic acid). Oligomeric compounds containing multiple phosphorothioate intersugar linkages are also known to bind to serum proteins. Therefore, short oligomeric compounds, e.g., oligonucleotides containing about 5-30 nucleotides (e.g., 5-25 nucleotides, preferably 5-20 nucleotides, e.g., 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides) and containing multiple phosphorothioate linkages in the backbone, are also applicable as ligands (e.g., PK-modulating ligands) in the present invention. PK-modulating oligonucleotides can contain at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or more phosphorothioate and / or phosphorodithioate linkages. In some embodiments, all internucleoside linkages in a PK-modulating oligonucleotide are phosphorothioate and / or phosphorodithioate linkages. Furthermore, aptamers that bind to serum components (e.g., serum proteins) are also suitable as PK-modulating ligands in the present invention. Binding to serum components (e.g., serum proteins) can be predicted from albumin binding assays such as those described in Oravcova, et al., Journal of Chromatography B (1996), 677:1-27.

[0074] When two or more ligands are present, the ligands may all have the same properties, or all have different properties, or some ligands may have the same properties while others have different properties. For example, the ligands may have targeting properties, endosomolytic activity, or PK modulating properties. In a preferred embodiment, all ligands have different properties.

[0075] In some embodiments of any one of the aspects, the ligand has formula (IV)-(VII): TIFF2024525713000052.tif83153, wherein: q 2A , q 2B , q 3A , q 3B , q4 A , q 4B , q 5A , q 5B , and q 5C represents, independently for each occurrence, 0 to 20, and the repeating units may be identical or different; P 2A , P 2B , P 3A , P 3B , P 4A , P 4B , P 5A , P 5B , P 5C , T 2A , T 2B , T 3A , T 3B , T 4A , T 4B , T 5A , T 5B , T 5C is each independently for each occurrence absent, CO, NH, O, S, OC(O), NHC(O), CH, CHNH, or CHO; Q 2A , Q 2B , Q 3A , Q 3B , Q 4A , Q 4B , Q 5A , Q 5B , Q 5Cis, independently for each occurrence, absent, alkylene, or substituted alkylene, wherein one or more methylenes are selected from O, S, S(O), SO, N(R N ), C(R')=C(R''), C≡C or C(O), R 2A , R 2B , R 3A , R 3B , R 4A , R 4B , R 5A , R 5B , R 5C are each independently for each occurrence absent, NH, O, S, CH2, C(O)O, C(O)NH, NHCH(R a )C(O), -C(O)-CH(R a )-NH-, CO, CH=NO, TIFF2024525713000053.tif18156, or heterocyclyl; L 2A , L 2B , L 3A , L 3B , L 4A , L 4B , L 5A , L 5B , and L 5C represents a ligand, i.e., each independently for each occurrence a monosaccharide (such as GalNAc), disaccharide, trisaccharide, tetrasaccharide, oligosaccharide, or polysaccharide; R a is H or an amino acid side chain.

[0076] In some embodiments of any one of the aspects, the ligand has formula (VII): TIFF2024525713000054.tif40128, wherein L 5A , L 5B , and L 5C represents a monosaccharide, such as a GalNAc derivative.

[0077] Exemplary ligands include, but are not limited to, the following: TIFF2024525713000055.tif131128TIFF2024525713000056.tif231147.

[0078] In some embodiments of any one of the aspects described herein, the ligand is a ligand described in U.S. Pat. No. 5,994,517 or U.S. Pat. No. 6,906,182, the contents of each of which are incorporated herein by reference in their entirety.

[0079] In some embodiments, the ligand can be a triantennary ligand as described in Figure 3 of U.S. Patent No. 6,906,182. For example, the ligand is selected from the following triantennary ligands: TIFF2024525713000057.tif192144

[0080] In some embodiments of any one of the aspects described herein, R L is a ligand. More than one R L It should be noted that when present, they may be the same or different. Thus, in some embodiments of any one of the aspects described herein, all R L In some other embodiments of any one of the aspects described herein, R L is different.

[0081] R 2 In some embodiments of any one of the aspects described herein, R 2 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), or -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ) For example, R 2 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 It is an alkoxy, an alkoxyalkyl (eg, methoxyethyl), an alkoxyalkylamine, an alkoxycarboxylate, an amino, an alkylamino, or a dialkylamino.

[0082] In some embodiments of any one of the aspects, R 2 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 alkoxy, or alkoxyalkyl (e.g., methoxyethyl. In some embodiments of any one of the aspects, R 2 is hydrogen, hydroxy, protected hydroxy, fluoro, or methoxy.

[0083] In some embodiments of any one of the aspects, R 2 is a halogen. For example, R 2 can be fluoro, chloro, bromo, or iodo. In some embodiments of any one of the aspects described herein, R 2 is fluoro.

[0084] In some embodiments of any one of the aspects described herein, R 2 and R 4

[0085] In some embodiments of any one of the aspects described herein, R 2 and R 4 together form 4'-C(R 10 R 11 ) v -Y-2' or 4'-YC(R 10 R 11 ) v-2', v is 1, 2, or 3, and Y is -O-, -CH2-, -CH(Me)-, -C(CH3)2-, -S-, -N(R 12 )-, -C(O)-, -C(S)-, -S(O)-, -S(O)2-, -OC(O)-, -C(O)O-, -N(R 12 )C(O)- or -C(O)N(R 12 )- and R 10 and R 11 are independently H, optionally substituted C-C alkyl, optionally substituted C-C alkenyl, or optionally substituted C-C alkynyl; R 12 is hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C1-C 30 Alkoxy, C 1-4 Haloalkyl, optionally substituted C 2-4 Alkenyl, optionally substituted C 2-4 Alkynyl, optionally substituted C 1-30 alkyl-CO2H, or a nitrogen protecting group.

[0086] In some embodiments of any one of the aspects, v is 1. In some other embodiments of any one of the aspects, v is 2.

[0087] In some embodiments, Y is O. For example, R 2 and R 4 together form 4'-C(R 10 R 11 ) v -O-2'.

[0088] R attached to the same carbon 10 and R 11 Note that the can be the same or different. For example, R 10 and R 11 may be H, and one of R 10 and R 11 The other of R can be an optionally substituted C1-C6 alkyl. In one non-limiting example, R 10 and R11 One of the groups can be H and the other can be C1-C6 alkyl, which can be OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8 ) alkyl (i.e., C1-C8 alkoxy), O(C1-C8) haloalkyl, (C2-C8) alkenyl, (C2-C8) alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 10 and R 11 are independently H or C1-C6 optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy. 30 In some embodiments of any one of the aspects, R 10 and R 11 One of R is H and the other is C-C alkyl optionally substituted with C-C alkoxy. For example, R 10 and R 11 One of them is H and the other is —CH or CHOCH.

[0089] In some embodiments of any one of the aspects, R attached to the same C 10 and R 11 For example, R attached to the same C10 and R 11 is H.

[0090] In some embodiments of any one of the aspects, R 2 and R 4 together are 4'-CH2-O-2', 4'-CH(CH3)-O-2', 4'-CH(CH2OCH3)-O-2', or 4'-CH2CH2-O-2'. For example, R 2 and R 4 together form 4'-CH2CH2-O-2'.

[0091] In some embodiments of any one of the aspects described herein, R 2 is the bond to the internucleotide bond to the subsequent nucleotide. 2 and R 3 Note that only one of the may be the bond to the internucleotide linkage to the subsequent nucleotide.

[0092] R 3 In some embodiments of any one of the aspects described herein, R 3 is the bond to the internucleotide bond to the subsequent nucleotide, a hydroxyl, a protected hydroxyl, an optionally substituted C 1-30 It can be an alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), amino, alkylamino, dialkylamino, a 3'-oligonucleotide capping group (e.g., an inverted nucleotide or an inverted abasic nucleotide), a ligand, a linker covalently attached to one or more ligands (e.g., N-acetylgalactosamine (GalNac)), a solid support, or a linker covalently attached to a solid support (e.g., -C(O)CHCHC(O)-).

[0093] In some embodiments of any one of the aspects described herein, R 3 is the bond to the internucleotide bond to the subsequent nucleotide, a hydroxyl, a protected hydroxyl, an optionally substituted C 1-30alkoxy, a 3'-oligonucleotide capping group (e.g., an inverted nucleotide or an inverted abasic nucleotide), a solid support, or a linker covalently attached to a solid support (e.g., -C(O)CHCHC(O)-). For example, R 3 is a bond to an internucleotide bond to a subsequent nucleotide, a hydroxy, a solid support, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—). In some embodiments of any one of the aspects described herein, R 3 is the bond to the internucleotide bond to the subsequent nucleotide, the solid support, or a linker covalently attached to the solid support (e.g., —C(O)CHCHC(O)—).

[0094] In some embodiments of any one of the aspects described herein, R 3 is the bond to the internucleotide bond to the subsequent nucleotide.

[0095] In some embodiments of any one of the aspects described herein, R 3 is a solid support or a linker covalently attached to a solid support.

[0096] In some embodiments of any one of the aspects described herein, R 3 is a hydroxyl.

[0097] In some embodiments of any one of the aspects described herein, R 3 and R 4 C, together with the atoms to which they are attached, may be substituted 3-8 Cycloalkyl, optionally substituted C 3-8 It forms a cycloalkenyl or an optionally substituted 3- to 8-membered heterocyclyl.

[0098] R 4 In some embodiments of any one of the aspects described herein, R 4 is hydrogen, optionally substituted C1-6 Alkyl, optionally substituted C 2-6 Alkenyl, optionally substituted C 2-6 Alkynyl, or optionally substituted C 1-6 For example, R 4 is hydrogen, optionally substituted C 1-6 Alkyl or optionally substituted C 1-6 It may be alkoxy.

[0099] In some embodiments of any one of the aspects described herein, R 4 is H.

[0100] R 5 In some embodiments of any one of the aspects described herein, R 5 is the bond to the internucleotide bond to the preceding nucleotide, hydrogen, hydroxy, protected hydroxy, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9), vinylphosphonate (VP) group, monophosphate ((HO)2(O)PO-5'), diphosphate ((HO)2(O)POP(HO)(O)-O-5'), triphosphate ((HO)2(O)PO-(HO)(O)POP(HO)(O)-O-5'); monothiophosphate (phosphorothioate, (HO)2(S)PO-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)PO-5'), phosphorothiolate ((HO)2(O)PS-5'); alpha-thiotriphosphate Phosphates; beta-thiotriphosphates; gamma-thiotriphosphates; phosphoramidates ((HO)2(O)P-NH-5', (HO)(NH2)(O)PO-5'), alkyl phosphonates (R(OH)(O)PO-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonates (R(OH)(O)PO-5', R = alkyl ether, e.g., methoxymethyl (CHOMe), ethoxymethyl, etc.), (HO)2(X)PO[-(CH2) a -OP(X)(OH)-O] b -5', or (HO)2(X)PO[-(CH2) a -P(X)(OH)-O] b -5', or (HO)2(X)P-[-(CH2) a -OP(X)(OH)-O] b -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl-terminated phosphates and phosphate mimetics (e.g., HO[-(CH) a -OP(X)(OH)-O] b -5', H2N[-(CH2) a -OP(X)(OH)-O] b -5', H[-(CH2) a -OP(X)(OH)-O] b -5', Me2N[-(CH2) a -OP(X)(OH)-O] b -5', HO[-(CH2) a -P(X)(OH)-O] b -5', H2N[-(CH2) a-P(X)(OH)-O] b -5', H[-(CH2) a -P(X)(OH)-O] b -5', Me2N[-(CH2) a -P(X)(OH)-O] b -5' (wherein a and b are each independently 1 to 10).

[0101] In some embodiments of any one of the aspects described herein, R 5 is the bond to the internucleotide bond to the preceding nucleotide, a hydroxy, a protected hydroxy, an optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 It can be an alkoxy, a vinylphosphonate (VP) group, a monophosphate, a diphosphate, a triphosphate, a monothiophosphate (phosphorothioate), a monodithiophosphate (phosphorodithioate), a phosphorothiolate, an alpha-thiotriphosphate, a beta-thiotriphosphate, a gamma-thiotriphosphate, a phosphoramidate, or an alkylphosphonate.

[0102] In some embodiments of any one of the aspects described herein, R 5 is the bond to the internucleotide bond to the preceding nucleotide, a hydroxy, a protected hydroxy, an optionally substituted C 2-30 Alkenyl, optionally substituted C 1-30 It is an alkoxy or vinylphosphonate (VP) group.

[0103] In some embodiments of any one of the aspects described herein, R 5 is the bond to the internucleotide bond to the preceding nucleotide.

[0104] In some embodiments of any one of the aspects described herein, R 5 is a hydroxyl or a protected hydroxyl.

[0105] In some embodiments of any one of the aspects described herein, R 5 may be substituted C 2-30 Alkenyl or optionally substituted C 1-30 It is an alkoxy.

[0106] In some embodiments of any one of the aspects described herein, R 5 is a vinyl phosphonate group.

[0107] In some embodiments of any one of the aspects described herein, R 5 is -CH(R 51 )-X 5 -R 52 wherein X 5 is absent, a bond, or O, and R 51 is hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted -C 2-30 alkenyl, or optionally substituted -C 2-30 alkynyl, and R 52 is the bond to the internucleoside linkage to the preceding nucleotide.

[0108] In some embodiments of any one of the aspects described herein, X 5 is O or a bond. For example, X 5 is O. In some other embodiments of any one of the aspects described herein, X 5 does not exist, i.e., R 5 is -CH(R 51 )R 52 is.

[0109] In some embodiments of the various aspects described herein, R 5 is -CH(R 51 )-R 52 or -C(R 51 )=CHR 52 wherein R 51 is hydrogen, optionally substituted C1-30 Alkyl, optionally substituted -C 2-30 alkenyl, or optionally substituted -C 2-30 alkynyl, and R 52 is the bond to the internucleoside linkage to the preceding nucleotide.

[0110] In some embodiments of the various aspects described herein, R 5 is -CH(R 51 )-X 5 -R 52 For example, R 5 is -CH(R 51 )-X 5 -R 52 where R 51 is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 51 is H. In some other non-limiting examples, R 51is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 It is alkyl.

[0111] In some embodiments of the various aspects described herein, R 5 is -CH(R 51 )-OR 52 where R 51 is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy; 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 51 is H. In some other non-limiting examples, R 51 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 It is alkyl.

[0112] In some embodiments of any one of the aspects described herein, R 5 is -C(R 51 )=CHR 52 -C(R 51 )=CHR 52 It should be noted that the double bond in can be in the cis or trans configuration. Thus, in some embodiments of any one of the aspects, R 5 is -C(R 51 )=CHR 52 and the double bond is in a cis configuration. In some other embodiments of any one of the aspects, R 5 is -C(R 51 )=CHR 52 and the double bond is in a trans configuration. In some embodiments of any one of the aspects described herein, R 5 is -CH=CHR 52 is.

[0113] In some embodiments of any one of the aspects described herein, R 52 is the bond to the internucleoside linkage to the preceding nucleotide.

[0114] In embodiments of the various aspects described herein, R 5 may be substituted C 1-6 Alkyl-R 53 , optionally substituted -C 2-6 Alkenyl-R 53 or optionally substituted -C 2-6 Alkynyl-R 53 In embodiments of the various aspects described herein, R 53 -OR 54 , -SR 55 , -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), or -SP(S)(SR 57 )2, and R 54 is a hydrogen or oxygen protecting group, and R 55 is a hydrogen or sulfur protecting group, and each R 56 are independently hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or oxygen protecting group, and each R 57 are independently hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or sulfur protecting groups.

[0115] In some embodiments of any one of the aspects, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), SP(O)(OR 56 )2, -SP(S)(OR 56 )2, and -SP(S)(SR 57 )(OR 56 ) at least one R 56 is hydrogen.

[0116] In some other embodiments of any one of the aspects, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56), -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), SP(O)(OR 56 )2, -SP(S)(OR 56 )2, or -SP(S)(SR 57 )(OR 56 ) at least one R 56 is not hydrogen. For example, P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), SP(O)(OR 56 )2, -SP(S)(OR 56 )2, and -SP(S)(SR 57 )(OR 56 ) at least one R 56 may be substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or an oxygen protecting group.

[0117] In some embodiments of any one of the aspects, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), SP(O)(OR 56 )2, -SP(S)(OR 56 )2, and -SP(S)(SR 57 )(OR 56 ) at least one R 56 is H and at least one R56 is other than H.

[0118] In some embodiments of any one of the aspects, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) All R in 2 56 is H.

[0119] In some embodiments of any one of the aspects, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) All R in 2 56 is other than H.

[0120] In some embodiments of any one of the aspects, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(S)(OR 56 )2, -OP(S)(SR57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) At least one R in 2 57 is H.

[0121] In some embodiments of any one of the aspects, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) At least one R in 2 57 is other than H. For example, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) At least one R in 2 57 may be substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or sulfur protecting groups.

[0122] In some embodiments of any one of the aspects, -P(S)(SR 57 )2, -OP(S)(SR 57 )2, and SP(S)(SR 57 ) At least one R in 2 57is H and at least one R 57 is other than H.

[0123] In some embodiments, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) All R in 2 57 is H.

[0124] In some embodiments, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) All R in 2 57 is other than H.

[0125] In some embodiments of any one of the aspects described herein, R 5 is optionally substituted -C 2-6 Alkenyl-R 53 For example, R 5 -C 2-6 Alkenyl-R 53 wherein C 2-6Alkenyl includes OH, CN, SC(O)Ph, oxo (=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C 8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p and R is optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH, or CH-aryl-alkoxy, wherein “m” and “p” are independently 1, 2, 3, 4, 5, or 6; 53 is -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), or -SP(S)(SR 57 )2.

[0126] In some embodiments of any one of the aspects, R 5 is -CH=CHR 53Optionally substituted -C 2-6 Alkenyl-R 53 It should be noted that the double bond in can be in the cis or trans configuration. Thus, in some embodiments of any one of the aspects, R 5 is -CH=CHR 53 and the double bond is in a cis configuration. In some other embodiments of any one of the aspects, R 5 is -CH=CHR 53 and the double bond is in the trans configuration.

[0127] In some embodiments of any one of the aspects, R 5 is -CH=CH-P(O)(OR 56 )2, -CH=CH-P(S)(OR 56 )2, -CH=CH-P(S)(SR 57 )(OR 56 ), -CH=CH-P(S)(SR 57 )2, -CH=CH-OP(O)(OR 56 )2, -CH=CH-OP(S)(OR 56 )2, -CH=CH-OP(S)(SR 57 )(OR 56 ), -CH=CH-OP(S)(SR 57 )2, -CH=CH-SP(O)(OR 56 )2, -CH=CH-SP(S)(OR 56 )2, -CH=CH-SP(S)(SR 57 )(OR 56 ), or -CH=CH-SP(S)(SR 57 )2. For example, R 5 is -CH=CH-P(O)(OR 56 )2.

[0128] In some embodiments of any one of the aspects, R 54 is a hydrogen or oxygen protecting group. For example, R 54 is hydrogen or 4,4'-dimethoxytrityl (DMT). In some preferred embodiments, R 54 is H.

[0129] In some embodiments of any one of the aspects described herein, R 5 is optionally substituted -C 1-6 Alkenyl-R 53 For example, R 5 -C 1-6 Alkenyl-R 53 wherein C 1-6 Alkenyl includes OH, CN, SC(O)Ph, oxo (=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C 8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p and R is optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH, or CH-aryl-alkoxy, wherein “m” and “p” are independently 1, 2, 3, 4, 5, or 6; 53 -OR 54 , -SR 55 , -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57)2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), or -SP(S)(SR 57 )2.

[0130] In some embodiments of any one of the aspects described herein, R 5 is -CH(R 58 )-R 53 wherein R 53 -OR 54 , -SR 55 , -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), or -SP(S)(SR 57 )2 and R 58 is H, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 It is alkynyl.

[0131] In some embodiments of any one of the aspects described herein, R 58is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. In one non-limiting example, R 58 is H. In some other non-limiting examples, R 58 is a C1-C6 optionally substituted with a substituent selected from NH2, OH, C(O)NH2, COOH, halo, SH, and C1-C6 alkoxy; 30 It is alkyl.

[0132] In some embodiments of any one of the aspects described herein, R 5 is -CH(R 58 )-OR 59 where R 59 H, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56)2. For example, R 5 is -CH(R 58 )-OR 59 where R 58 is H or optionally substituted C1-C 30 alkyl, and R 59 is H or -P(O)(OR 56 )2.

[0133] In some embodiments of any one of the aspects described herein, R 5 is -CH(R 58 )-SR 60 where R 60 H, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56 )2.

[0134] R 32 In some embodiments of any one of the aspects described herein, R 32 is hydrogen, halogen, -OR 322 , -SR 323 , optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 Alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, -O(CH2CH2O) r CH2CH2OR 324 , cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, heteroaryl, -NH(CH2CH2NH) s CH2CH2-R 325 , NHC(O)R 326, a lipid, a linker covalently attached to a lipid, a ligand, a linker covalently attached to a ligand, a solid support, a linker covalently attached to a solid support, or a reactive phosphorus group.

[0135] R 322 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 323 is H, a sulfur protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 324 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 325 is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30R can be alkoxy, amino (NH), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl. 326 is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl.

[0136] In some embodiments of any one of the aspects described herein, R 32 is R 32 is hydrogen, halogen, -OR 322 , -SR 323 , optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 Alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, -O(CH2CH2O) r CH2CH2OR 324 , cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, heteroaryl, -NH(CH2CH2NH) s CH2CH2-R 325 , NHC(O)R 324 is.

[0137] In some embodiments of any one of the aspects described herein, R 32 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), or -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ) For example, R 32 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 It is an alkoxy, an alkoxyalkyl (eg, methoxyethyl), an alkoxyalkylamine, an alkoxycarboxylate, an amino, an alkylamino, or a dialkylamino.

[0138] In some embodiments of any one of the aspects, R 32 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 In some embodiments of any one of the aspects, R is alkoxy, or alkoxyalkyl (e.g., methoxyethyl). 32 is hydrogen, hydroxy, protected hydroxy, fluoro, or methoxy.

[0139] In some embodiments of any one of the aspects, R 32 is a halogen. For example, R 32 can be fluoro, chloro, bromo, or iodo. In some embodiments of any one of the aspects described herein, R 32 is fluoro.

[0140] In some embodiments of any one of the aspects described herein, R 32 and R 4 together form 4'-C(R 10 R 11 ) v -Y-2' or 4'-YC(R 10 R 11 ) v -2', v is 1, 2, or 3, and Y is -O-, -CH2-, -CH(Me)-, -C(CH3)2-, -S-, or -N(R 12 )-, -C(O)-, -C(S)-, -S(O)-, -S(O)2-, -OC(O)-, -C(O)O-, -N(R 12 )C(O)- or -C(O)N(R 12 )- and R 10 and R 11 are independently H, optionally substituted C-C alkyl, optionally substituted C-C alkenyl, or optionally substituted C-C alkynyl; R 12 is hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C1-C 30 Alkoxy, C 1-4 Haloalkyl, optionally substituted C 2-4 Alkenyl, optionally substituted C 2-4 Alkynyl, optionally substituted C 1-30 In some embodiments of any one of the aspects, v is 1. In some other embodiments of any one of the aspects, v is 2. In some embodiments, Y is O. For example, R 32 and R 4 together form 4'-C(R 10 R 11 ) v -O-2'.

[0141] R attached to the same carbon 10 and R 11 Note that the can be the same or different. For example, R 10 and R11 may be H, and one of R 10 and R 11 The other of R can be an optionally substituted C1-C6 alkyl. In one non-limiting example, R 10 and R 11 One of the groups can be H and the other can be C1-C6 alkyl, which can be OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8 ) alkyl (i.e., C1-C8 alkoxy), O(C1-C8) haloalkyl, (C2-C8) alkenyl, (C2-C8) alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 10 and R 11 are independently H or C1-C6 optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy. 30 In some embodiments of any one of the aspects, R 10 and R 11 One of R is H and the other is C-C alkyl optionally substituted with C-C alkoxy. For example, R 10 and R 11In some embodiments of any one of the aspects, one of R attached to the same C is H and the other is —CH or —CHOCH. 10 and R 11 For example, R attached to the same C 10 and R 11 is H.

[0142] In some embodiments of any one of the aspects, R 32 and R 4 together are 4'-CH2-O-2', 4'-CH(CH3)-O-2', 4'-CH(CH2OCH3)-O-2', or 4'-CH2CH2-O-2'. For example, R 32 and R 4 together form 4'-CH2CH2-O-2'.

[0143] In some embodiments of any one of the aspects described herein, R 32 is a reactive phosphorus group.

[0144] Without wishing to be bound by theory, reactive phosphorus groups are useful, for example, for forming internucleoside linkages, including phosphodiester and phosphorothioate internucleoside linkages. Such reactive phosphorus groups are known in the art and include, but are not limited to, phosphoramidites, H-phosphonates, phosphate triesters, and phosphorus-containing chiral auxiliaries. III or P V Phosphoramidites (P) contain phosphorus atoms in the valence state. III Reactive phosphorus groups in the form of phosphate groups are preferred reactive phosphorus groups for solid-phase oligonucleotide synthesis. The intermediate phosphite compounds are then oxidized to the Pv state using known methods to generate phosphodiester or phosphorothioate internucleoside linkages.

[0145] In some embodiments of any one of the aspects described herein, the reactive phosphorus group is —OP(OR P )(N(R P2)2), -OP(SR P )(N(R P2 )2), -OP(O)(OR P )(N(R P2 )2), -OP(S)(OR P )(N(R P2 )2), -OP(O)(SR P )(N(R P2 )2), -OP(O)(OR P )H, -OP(S)(OR P )H, -OP(O)(SR P )H, -OP(O)(OR P )R P3 , -OP(S)(OR P )R P3 , or -OP(O)(SR P )R P3 For example, a reactive phosphorus group is -OP(OR P )(N(R P2 )2).

[0146] In some embodiments of any one of the aspects, R P may be substituted C 1-6 alkyl. For example, R P is C 1-6 alkyl, which may be OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C 1-C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m-(CH2) p In some embodiments, R is optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH, or CH-aryl-alkoxy, and “m” and “p” are independently 1, 2, 3, 4, 5, or 6. p is C optionally substituted with CN or -SC(O)Ph 1-6 alkyl. For example, R p is cyanoethyl (-CH2CH2CN).

[0147] In the reactive phosphorus group, each R P2 are independently optionally substituted C 1-6 For example, each R P2 may be independently selected from methyl, ethyl, propyl, isopropyl, n-butyl, iso-butyl, pentyl, or hexyl. P2 It should be noted that when groups are present in the reactive phosphorus group, they may be the same or different. Thus, in some non-limiting examples, two or more R P2 When the R group is present, P2 In some other non-limiting examples, two or more R P2 When the R group is present, P2 In some embodiments of any one of the aspects, each R P2 is isopropyl.

[0148] In some embodiments of any one of the aspects, both R P2taken together with the nitrogen atom to which they are attached form an optionally substituted 3- to 8-membered heterocyclyl. Exemplary heterocyclyls include, but are not limited to, pyrrolidinyl, piperazinyl, dioxanyl, morpholinyl, tetrahydrofuranyl, piperidyl, 4-morpholyl, 4-piperazinyl, pyrrolidinyl, perhydropyrrolidinyl, 1,4-diazaperhydroepynyl, 1,3-dioxanyl, 1,4-dioxanyl, and the like, each of which is selected from the group consisting of OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alky. alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p It may be substituted with 1, 2, or 3 substituents independently selected from -NH2, or CH2-aryl-alkoxy, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6.

[0149] In some embodiments of any one of the aspects, R P and R P2together with the atom to which they are attached form an optionally substituted 4- to 8-membered heterocyclyl. Exemplary heterocyclyls include, but are not limited to, pyrrolidinyl, piperazinyl, dioxanyl, morpholinyl, tetrahydrofuranyl, piperidyl, 4-morpholyl, 4-piperazinyl, pyrrolidinyl, perhydropyrrolidinyl, 1,4-diazaperhydroepynyl, 1,3-dioxanyl, 1,4-dioxanyl, and the like, each of which may be substituted with OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alky. alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p It may be substituted with 1, 2, or 3 substituents independently selected from -NH2, or CH2-aryl-alkoxy, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6.

[0150] In the reactive phosphorus group, each R P3 are independently optionally substituted C 1-6 alkyl. For example, R P3 is C 1-6It can be alkyl, which can be OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C 1-C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R P3 is methyl, ethyl, propyl, isopropyl, n-butyl, iso-butyl, pentyl, or hexyl, each of which can be optionally substituted with NH, OH, C(O)NH, COOH, halo, SH, or C-C alkoxy.

[0151] In some embodiments of any one of the aspects, the reactive phosphorus group is —OP(OR P )(N(R P2 )2). For example, a reactive phosphorus group is -OP(OR P )(N(R P2 )2) and R P is cyanoethyl (-CH2CH2CN), and each R P2 is isopropyl.

[0152] In some embodiments of any one of the aspects described herein, R 32 -OP(ORP )(N(R P2 )2), -OP(SR P )(N(R P2 )2), -OP(O)(OR P )(N(R P2 )2), -OP(S)(OR P )(N(R P2 )2), -OP(O)(SR P )(N(R P2 )2), -OP(O)(OR P )H, -OP(S)(OR P )H, -OP(O)(SR P )H, -OP(O)(OR P )R P3 , -OP(S)(OR P )R P3 , or -OP(O)(SR P )R P3 is.

[0153] In some embodiments of any one of the aspects, R 32 -OP(OR P )(N(R P2 )2), -OP(SR P )(N(R P2 )2), -OP(O)(OR P )(N(R P2 )2), -OP(S)(OR P )(N(R P2 )2), -OP(O)(SR P )(N(R P2 )2), -OP(O)(OR P )H, -OP(S)(OR P ), optionally substituted C 1-6 alkyl, and each R P2 are independently optionally substituted C 1-6 alkyl, and each R P3 are independently optionally substituted C 1-6 It is alkyl.

[0154] In some embodiments of any one of the aspects, R 32 -OP(OR P )(N(R P2)2). For example, R 32 -OP(OR P )(N(R P2 )2), wherein R P is cyanoethyl (-CH2CH2CN), and each R P2 is isopropyl.

[0155] In some embodiments of any one of the aspects described herein, R 32 is a solid support or a linker covalently attached to a solid support. For example, R 32 is —OC(O)CHCHC(O)NH—Z, where Z is a solid support. In some embodiments, R 32 is -OC(O)CH2CH2CO2H.

[0156] In some embodiments of any one of the aspects, R 32 -OR 322 If R 322 can be hydrogen or a hydroxyl protecting group.

[0157] R 32 Ga-SR 323 If R 323 can be hydrogen or a sulfur protecting group. Thus, in some embodiments of any one of the aspects, R 323 is hydrogen.

[0158] R 32 -O(CH2CH2O) r CH2CH2OR 324 where r can be 1 to 50, and R 324 independently for each occurrence, H, C1-C 30 alkyl, cyclyl, heterocyclyl, aryl, heteroaryl, aralkyl, sugar, or R 325 and R 325 is independently for each occurrence amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0159] R 32 -NH(CH2CH2NH) s CH2CH2-R 325 where s can be 1 to 50, and R 325 may be, independently for each occurrence, amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0160] In some embodiments of any one of the aspects described herein, R 32 is hydrogen, halogen, -OR 322 or optionally substituted C1-C 30 Alkoxy. For example, R 32 is halogen, -OR 322 or optionally substituted C1-C 30 In some embodiments of any one of the aspects described herein, R 32 is F, OH, or optionally substituted C1-C 30 It is an alkoxy.

[0161] In some embodiments of any one of the aspects described herein, R 32 are OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C8 alkyl) alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p-OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy; 30 alkoxy, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 32 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 In some embodiments of any one of the aspects described herein, R 32 is -O(CH2) t CH3, where t is 1 to 21. For example, t is 14, 15, 16, 17, or 18. In one non-limiting example, t is 16.

[0162] In some embodiments of any one of the aspects, R 32 is -O(CH2) u R 327 wherein u is 2 to 10, and R 327 is C1-C6 alkoxy, amino (NH2), CO2H, OH, or halo. For example, R 327 is —CH or NH. Thus, in some embodiments of any one of the aspects described herein, R 32 is -O(CH2) u -OMe or R 32 is -O(CH2) u It is NH2.

[0163] In some embodiments of any one of the aspects described herein, u is 2, 3, 4, 5, or 6. For example, u is 2, 3, or 6. In one non-limiting example, u is 2. In another non-limiting example, u is 3 or 6.

[0164] In some embodiments of any one of the aspects described herein, R 32 is C1-C6 haloalkyl. For example, R32 is C-C haloalkyl. In some embodiments of any one of the aspects described herein, R 32 is -CF3, -CF2CF3, -CF2CF2CF3, or -CF2(CF3)2.

[0165] In some embodiments of any one of the aspects described herein, R 32 is -OCH(CH2OR 328 )CH2OR 329 where R 328 and R 329 are independently H, optionally substituted C-C 30 Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 328 and R 329 are independently optionally substituted C-C 30 It is alkyl.

[0166] In some embodiments of any one of the aspects described herein, R 32 is -CH2C(O)NHR 3210 where R 3210 is H, optionally substituted C1-C 30 Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 3210 is H or optionally substituted C1-C 30 In some embodiments, R 3210 is optionally substituted C1-C6 alkyl.

[0167] R 33 In some embodiments of any one of the aspects described herein, R 33 is hydrogen, halogen, -OR 332 , -SR 333 , optionally substituted C 1-30Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 Alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, -O(CH2CH2O) r CH2CH2OR 334 , cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, heteroaryl, -NH(CH2CH2NH) s CH2CH2-R 335 , NHC(O)R 336 , a lipid, a linker covalently attached to a lipid, a ligand, a linker covalently attached to a ligand, a solid support, a linker covalently attached to a solid support, or a reactive phosphorus group.

[0168] R 332 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 333 is H, a sulfur protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 334 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 335 is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 R can be alkoxy, amino (NH), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl. 336 is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl.

[0169] In some embodiments of any one of the aspects described herein, R 33 is a reactive phosphorus group. For example, R 33 -OP(OR P )(N(R P2 )2), -OP(SR P )(N(R P2 )2), -OP(O)(OR P )(N(R P2 )2), -OP(S)(OR P )(N(R P2)2), -OP(O)(SR P )(NR P2 )2, -OP(O)(OR P )H, -OP(S)(OR P )H, -OP(O)(SR P )H, -OP(O)(OR P )R P3 , -OP(S)(OR P )R P3 , or -OP(O)(SR P )R P3 is.

[0170] In some embodiments of any one of the aspects, R 33 -OP(OR P )(N(R P2 )2), -OP(SR P )(N(R P2 )2), -OP(O)(OR P )(N(R P2 )2), -OP(S)(OR P )(N(R P2 )2), -OP(O)(SR P )(N(R P2 )2), -OP(O)(OR P )H, -OP(S)(OR P ), optionally substituted C 1-6 alkyl, and each R P2 are independently optionally substituted C 1-6 alkyl, and each R P3 are independently optionally substituted C 1-6 It is alkyl.

[0171] In some embodiments of any one of the aspects, R 33 -OP(OR P )(N(R P2 )2). For example, R 33 -OP(OR P )(N(R P2 )2), wherein R P is cyanoethyl (-CH2CH2CN), and each R P2 is isopropyl.

[0172] Optionally, R 32 and R 33 Only one of the groups is a reactive phosphorus group.

[0173] In some embodiments of any one of the aspects described herein, R 33 is a solid support or a linker covalently attached to a solid support. For example, R 33 is —OC(O)CHCHC(O)NH—Z, where Z is a solid support.

[0174] Optionally, R 32 and R 33 is a solid support or a linker covalently attached to a solid support.

[0175] In some embodiments of any one of the aspects, R 33 -OR 332 If R 332 can be hydrogen or a hydroxyl protecting group. For example, R 332 In some embodiments of any one of the aspects described herein, R can be hydrogen. 33 is -OC(O)CH2CH2CO2H.

[0176] R 33 Ga-SR 333 If R 333 can be hydrogen or a sulfur protecting group. Thus, in some embodiments of any one of the aspects, R 333 is hydrogen.

[0177] R 33 -O(CH2CH2O) r CH2CH2OR 334 where r can be 1 to 50, and R 334 independently for each occurrence, H, C1-C 30 alkyl, cyclyl, heterocyclyl, aryl, heteroaryl, aralkyl, sugar, or R 335 and R 335is independently for each occurrence amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0178] R 33 -NH(CH2CH2NH) s CH2CH2-R 335 where s can be 1 to 50, and R 335 may be, independently for each occurrence, amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0179] In some embodiments of any one of the aspects described herein, R 33 is hydrogen, halogen, -OR 332 or optionally substituted C1-C 30 Alkoxy. For example, R 33 is halogen, -OR 332 or optionally substituted C1-C 30 In some embodiments of any one of the aspects described herein, R 33 is F, OH, or optionally substituted C1-C 30 It is an alkoxy.

[0180] In some embodiments of any one of the aspects described herein, R 33are OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C8 alkyl) alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy; 30 alkoxy, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 33 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 In some embodiments of any one of the aspects described herein, R 33 is -O(CH2) t CH3, where t is 1 to 21. For example, t is 14, 15, 16, 17, or 18. In one non-limiting example, t is 16.

[0181] In some embodiments of any one of the aspects, R 33 is -O(CH2) u R 337 wherein u is 2 to 10, and R 337 is C1-C6 alkoxy, amino (NH2), CO2H, OH, or halo. For example, R 337is —CH or NH. Thus, in some embodiments of any one of the aspects described herein, R 33 is -O(CH2) u -OMe or R 33 is -O(CH2) u It is NH2.

[0182] In some embodiments of any one of the aspects described herein, u is 2, 3, 4, 5, or 6. For example, u is 2, 3, or 6. In one non-limiting example, u is 2. In another non-limiting example, u is 3 or 6.

[0183] In some embodiments of any one of the aspects described herein, R 33 is C1-C6 haloalkyl. For example, R 33 is C-C haloalkyl. In some embodiments of any one of the aspects described herein, R 33 is -CF3, -CF2CF3, -CF2CF2CF3, or -CF2(CF3)2.

[0184] In some embodiments of any one of the aspects described herein, R 33 is -OCH(CH2OR 338 )CH2OR 339 where R 338 and R 339 are independently H, optionally substituted C-C 30 Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 338 and R 339 are independently optionally substituted C-C 30 It is alkyl.

[0185] In some embodiments of any one of the aspects described herein, R 33 is -CH2C(O)NHR 3310 where R 3310is H, optionally substituted C1-C 30 Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 3310 is H or optionally substituted C1-C 30 In some embodiments, R 3310 is optionally substituted C1-C6 alkyl.

[0186] In some embodiments of any one of the aspects described herein, R 33 and R 4 C, together with the atoms to which they are attached, may be substituted 3-8 Cycloalkyl, optionally substituted C 3-8 It forms a cycloalkenyl or an optionally substituted 3- to 8-membered heterocyclyl.

[0187] R 35 In some embodiments of the various aspects described herein, R 35 is R 551 , optionally substituted C 1-6 Alkyl-R 551 , optionally substituted -C 2-6 Alkenyl-R 551 or optionally substituted -C 2-6 Alkynyl-R 551 where R 551 -OR 552 , -SR 553 , hydrogen, a phosphorus group, a solid support, or a linker to a solid support. 551 -OR 552 If R 552 can be H or a hydroxyl protecting group. Similarly, R 551 Ga-SR 553 If R 553 can be H or a sulfur protecting group.

[0188] In some embodiments of any one of the aspects described herein, R35 -OR 552 or -SR 553 is.

[0189] In some embodiments of any one of the aspects described herein, R 552 is a hydroxyl protecting group. 552 Exemplary hydroxyl protecting groups for include, but are not limited to, benzyl, benzoyl, 2,6-dichlorobenzyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, mesylate, tosylate, 4,4'-dimethoxytrityl (DMT), 9-phenylxanthin-9-yl (pixyl), and 9-(p-methoxyphenyl)xanthin-9-yl (MOX). In some embodiments of any one of the aspects described herein, R 35 -OR 552 and R 552 is 4,4'-dimethoxytrityl (DMT), for example, R 35 is -O-DMT.

[0190] In some embodiments of any one of the aspects described herein, R 35 is -CH(R 554 )-R 551 where R 554 is hydrogen, halogen, optionally substituted C1-C 30 Alkyl, optionally substituted C-C 30 Alkenyl, optionally substituted C-C 30 Alkynyl or optionally substituted C-C 30 It is an alkoxy.

[0191] In some embodiments of any one of the aspects, R 35 -CH(R 554 )-R 551 If R 554is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 554 is H. In some other non-limiting examples, R 554 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 It is alkyl.

[0192] In some embodiments of the various aspects described herein, R 35 is -CH(R 554 )-OR 552 where R 554is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 554 is H. In some other non-limiting examples, R 554 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 It is alkyl.

[0193] In some embodiments of the various aspects described herein, R 35 may be substituted C 1-6 Alkyl-R 551 or optionally substituted -C 2-6 Alkenyl-R 551 is.

[0194] In some embodiments of any one of the aspects described herein, R 35 is -C(R 554 )=CHR 551-C(R 554 )=CHR 551 It should be noted that the double bond in can be in the cis or trans configuration. Thus, in some embodiments of any one of the aspects, R d is -C(R 554 )=CHR 551 and the double bond is in a cis configuration. In some other embodiments of any one of the aspects, R d is -C(R 554 )=CHR 551 and the double bond is in the trans configuration.

[0195] In some embodiments of any one of the aspects described herein, R 35 is -CH=CHR 551 is.

[0196] In some embodiments of any one of the aspects, R 35 -C(R 554 )=CHR 551 If R 554 is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) pC1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, "m" and "p" are independently 1, 2, 3, 4, 5, or 6, and R 551 is a phosphorus group. For example, R 35 is -CH=CHR 551 is.

[0197] In some embodiments of any one of the aspects described herein, R 551 is a reactive phosphorus group.

[0198] In some embodiments of any one of the aspects, R 35 is -CH=CH-P(O)(OR 555 )2, -CH=CH-P(S)(OR 555 )2, -CH=CH-P(S)(SR 556 )(OR 555 ), -CH=CH-P(S)(SR 556 )2, -CH=CH-OP(O)(OR 555 )2, -CH=CH-OP(S)(OR 555 )2, -CH=CH-OP(S)(SR 556 )(OR 555 ), -CH=CH-OP(S)(SR 556 )2, -CH=CH-SP(O)(OR 555 )2, -CH=CH-SP(S)(OR 555 )2, -CH=CH-SP(S)(SR 556 )(OR 555 ), or -CH=CH-SP(S)(SR 556 )2, where each R 555 are independently hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or oxygen protecting group, and each R 556 are independently hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30Alkenyl, or optionally substituted C 2-30 alkynyl, or sulfur protecting groups.

[0199] In some embodiments of any one of the aspects, -P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR 555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), SP(O)(OR 555 )2, -SP(S)(OR 555 )2, and -SP(S)(SR 556 )(OR 555 ) at least one R 555 is hydrogen.

[0200] In some other embodiments of any one of the aspects, -P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR 555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), SP(O)(OR 555 )2, -SP(S)(OR 555 )2, or -SP(S)(SR 556 )(OR 555 ) at least one R 555 is not hydrogen. For example, P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR 555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), SP(O)(OR 555 )2, -SP(S)(OR555 )2, and -SP(S)(SR 556 )(OR 555 ) at least one R 555 may be substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or an oxygen protecting group.

[0201] In some embodiments of any one of the aspects, -P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR 555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), SP(O)(OR 555 )2, -SP(S)(OR 555 )2, and -SP(S)(SR 556 )(OR 555 ) at least one R 555 is H and at least one R 555 is other than H.

[0202] In some embodiments of any one of the aspects, -P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR 555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(O)(OR 555 )2, -SP(S)(OR 555 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) All R in 2 555 is H.

[0203] In some embodiments of any one of the aspects, -P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR 555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(O)(OR 555 )2, -SP(S)(OR 555 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) All R in 2 555 is other than H.

[0204] In some embodiments of any one of the aspects, -P(S)(SR 556 )(OR 555 ), -P(S)(SR 556 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) At least one R in 2 556 is H.

[0205] In some embodiments of any one of the aspects, -P(S)(SR 556 )(OR 555 ), -P(S)(SR 556 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR556 ) At least one R in 2 556 is other than H. For example, -P(S)(SR 556 )(OR 555 ), -P(S)(SR 556 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) At least one R in 2 556 may be substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or sulfur protecting groups.

[0206] In some embodiments of any one of the aspects, -P(S)(SR 556 )2, -OP(S)(SR 556 )2, and -SP(S)(SR 556 ) At least one R in 2 556 is H and at least one R 556 is other than H.

[0207] In some embodiments, -P(S)(SR 556 )(OR 555 ), -P(S)(SR 556 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) All R in 2 556 is H.

[0208] In some embodiments, -P(S)(SR 556 )(OR 555), -P(S)(SR 556 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) All R in 2 556 is other than H.

[0209] In some embodiments of any one of the aspects, R 35 is -CH=CH-P(O)(OR 555 )2, where each R 555 is H or an oxygen protecting group.

[0210] In some embodiments of any one of the aspects, R 33 is a reactive phosphorus group, a solid support, a linker to the solid support, and R 35 is a protected hydroxyl.

[0211] In some other embodiments of any one of the aspects, R 32 is a reactive phosphorus group, a solid support, a linker to the solid support, and R 35 is a protected hydroxyl.

[0212] R 42 In some embodiments of any one of the aspects described herein, R 42 is halogen, -OR 422 , -SR 423 , optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30Alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, -O(CH2CH2O) r CH2CH2OR 424 , cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, heteroaryl, -NH(CH2CH2NH) s CH2CH2-R 425 , NHC(O)R 426 , a lipid, a linker covalently attached to a lipid, a ligand, a linker covalently attached to a ligand, a solid support, a linker covalently attached to a solid support, or a reactive phosphorus group.

[0213] R 422 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 423 is H, a sulfur protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 424 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 425is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 R can be alkoxy, amino (NH), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl. 426 is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl.

[0214] In some embodiments of any one of the aspects described herein, R 42 is a reactive phosphorus group. For example, R 42 -OP(OR P )(N(R P2 )2), -OP(SR P )(N(R P2 )2), -OP(O)(OR P )(N(R P2 )2), -OP(S)(OR P )(N(R P2 )2), -OP(O)(SR P )(NR P2 )2, -OP(O)(OR P )H, -OP(S)(OR P)H, -OP(O)(SR P )H, -OP(O)(OR P )R P3 , -OP(S)(OR P )R P3 , or -OP(O)(SR P )R P3 is.

[0215] In some embodiments of any one of the aspects, R 42 -OP(OR P )(N(R P2 )2), -OP(SR P )(N(R P2 )2), -OP(O)(OR P )(N(R P2 )2), -OP(S)(OR P )(N(R P2 )2), -OP(O)(SR P )(N(R P2 )2), -OP(O)(OR P )H, -OP(S)(OR P ), optionally substituted C 1-6 alkyl, and each R P2 are independently optionally substituted C 1-6 alkyl, and each R P3 are independently optionally substituted C 1-6 It is alkyl.

[0216] In some embodiments of any one of the aspects, R 42 -OP(OR P )(N(R P2 )2). For example, R 42 -OP(OR P )(N(R P2 )2), wherein R P is cyanoethyl (-CH2CH2CN), and each R P2 is isopropyl.

[0217] In some embodiments of any one of the aspects described herein, R 42 is a solid support or a linker covalently attached to a solid support. For example, R42 is —OC(O)CHCHC(O)NH—Z, where Z is a solid support.

[0218] In some embodiments of any one of the aspects, R 42 -OR 422 If R 422 can be hydrogen or a hydroxyl protecting group. For example, R 422 In some embodiments of any one of the aspects described herein, R can be hydrogen. 42 is -OC(O)CH2CH2CO2H.

[0219] R 42 Ga-SR 423 If R 423 can be hydrogen or a sulfur protecting group. Thus, in some embodiments of any one of the aspects, R 423 is hydrogen.

[0220] R 42 -O(CH2CH2O) r CH2CH2OR 424 where r can be 1 to 50, and R 424 independently for each occurrence, H, C1-C 30 alkyl, cyclyl, heterocyclyl, aryl, heteroaryl, aralkyl, sugar, or R 425 and R 425 is independently for each occurrence amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0221] R 42 -NH(CH2CH2NH) s CH2CH2-R 425 where s can be 1 to 50, and R 425 may be, independently for each occurrence, amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0222] In some embodiments of any one of the aspects described herein, R 42 -OR 422 or optionally substituted C1-C 30 Alkoxy. For example, R 42 is halogen, -OR 422 or optionally substituted C1-C 30 In some embodiments of any one of the aspects described herein, R 42 is F, OH, or optionally substituted C1-C 30 It is an alkoxy.

[0223] In some embodiments of any one of the aspects described herein, R 42 are OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C8 alkyl) alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy; 30 alkoxy, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 42is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 In some embodiments of any one of the aspects described herein, R 42 is -O(CH2) t CH3, where t is 1 to 21. For example, t is 14, 15, 16, 17, or 18. In one non-limiting example, t is 16.

[0224] In some embodiments of any one of the aspects, R 42 is -O(CH2) u R 427 wherein u is 2 to 10, and R 427 is C1-C6 alkoxy, amino (NH2), CO2H, OH, or halo. For example, R 427 is —CH or NH. Thus, in some embodiments of any one of the aspects described herein, R 42 is -O(CH2) u -OMe or R 42 is -O(CH2) u It is NH2.

[0225] In some embodiments of any one of the aspects described herein, u is 2, 3, 4, 5, or 6. For example, u is 2, 3, or 6. In one non-limiting example, u is 2. In another non-limiting example, u is 3 or 6.

[0226] In some embodiments of any one of the aspects described herein, R 42 is C1-C6 haloalkyl. For example, R 42 is C-C haloalkyl. In some embodiments of any one of the aspects described herein, R 42 is -CF3, -CF2CF3, -CF2CF2CF3, or -CF2(CF3)2.

[0227] In some embodiments of any one of the aspects described herein, R 42 is -OCH(CH2OR 428 )CH2OR 429 where R 428 and R 429 are independently H, optionally substituted C-C 30 Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 428 and R 429 are independently optionally substituted C-C 30 It is alkyl.

[0228] In some embodiments of any one of the aspects described herein, R 42 is -CH2C(O)NHR 4210 where R 4210 is H, optionally substituted C1-C 30 Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 4210 is H or optionally substituted C1-C 30 In some embodiments, R 4210 is optionally substituted C1-C6 alkyl.

[0229] In some embodiments of any one of the aspects described herein, R 42 and R 4 C, together with the atoms to which they are attached, may be substituted 3-8 Cycloalkyl, optionally substituted C 3-8 It forms a cycloalkenyl or an optionally substituted 3- to 8-membered heterocyclyl.

[0230] R 45 In some embodiments of the various aspects described herein, R 45 is R551 , optionally substituted C 1-6 Alkyl-R 551 , optionally substituted -C 2-6 Alkenyl-R 551 or optionally substituted -C 2-6 Alkynyl-R 551 where R 551 -OR 552 , -SR 553 , hydrogen, a phosphorus group, a solid support, or a linker to a solid support. 551 -OR 552 If R 552 can be H or a hydroxyl protecting group. Similarly, R 551 Ga-SR 553 If R 553 can be H or a sulfur protecting group.

[0231] In some embodiments of any one of the aspects described herein, R 45 -OR 552 or -SR 553 is.

[0232] In some embodiments of any one of the aspects described herein, R 552 is a hydroxyl protecting group. 552 Exemplary hydroxyl protecting groups for include, but are not limited to, benzyl, benzoyl, 2,6-dichlorobenzyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, mesylate, tosylate, 4,4'-dimethoxytrityl (DMT), 9-phenylxanthin-9-yl (pixyl), and 9-(p-methoxyphenyl)xanthin-9-yl (MOX). In some embodiments of any one of the aspects described herein, R 45 -OR 552 and R 552 is 4,4'-dimethoxytrityl (DMT), for example, R 45 is -O-DMT.

[0233] In some embodiments of any one of the aspects described herein, R45 is -CH(R 554 )-R 551 where R 554 is hydrogen, halogen, optionally substituted C1-C 30 Alkyl, optionally substituted C-C 30 Alkenyl, optionally substituted C-C 30 Alkynyl or optionally substituted C-C 30 It is an alkoxy.

[0234] In some embodiments of any one of the aspects, R 45 -CH(R 554 )-R 551 If R 554 is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 554 is H. In some other non-limiting examples, R 554is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 It is alkyl.

[0235] In some embodiments of the various aspects described herein, R 45 is -CH(R 554 )-OR 552 where R 554 is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 554 is H. In some other non-limiting examples, R 554 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 It is alkyl.

[0236] In some embodiments of the various aspects described herein, R 45may be substituted C 1-6 Alkyl-R 551 or optionally substituted -C 2-6 Alkenyl-R 551 is.

[0237] In some embodiments of any one of the aspects described herein, R 45 is -C(R 554 )=CHR 551 -C(R 554 )=CHR 551 It should be noted that the double bond in can be in the cis or trans configuration. Thus, in some embodiments of any one of the aspects, R d is -C(R 554 )=CHR 551 and the double bond is in a cis configuration. In some other embodiments of any one of the aspects, R d is -C(R 554 )=CHR 551 and the double bond is in the trans configuration.

[0238] In some embodiments of any one of the aspects described herein, R 45 is -CH=CHR 551 is.

[0239] In some embodiments of any one of the aspects, R 45 -C(R 554 )=CHR 551 If R 554is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, "m" and "p" are independently 1, 2, 3, 4, 5, or 6, and R 551 is a phosphorus group. For example, R 45 is -CH=CHR 551 is.

[0240] In some embodiments of any one of the aspects described herein, R 551 is a reactive phosphorus group.

[0241] In some embodiments of any one of the aspects, R 45 is -CH=CH-P(O)(OR 555 )2, -CH=CH-P(S)(OR 555 )2, -CH=CH-P(S)(SR 556 )(OR 555 ), -CH=CH-P(S)(SR 556 )2, -CH=CH-OP(O)(OR 555 )2, -CH=CH-OP(S)(OR555 )2, -CH=CH-OP(S)(SR 556 )(OR 555 ), -CH=CH-OP(S)(SR 556 )2, -CH=CH-SP(O)(OR 555 )2, -CH=CH-SP(S)(OR 555 )2, -CH=CH-SP(S)(SR 556 )(OR 555 ), or -CH=CH-SP(S)(SR 556 )2, where each R 555 are independently hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or oxygen protecting group, and each R 556 are independently hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or sulfur protecting groups.

[0242] In some embodiments of any one of the aspects, -P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR 555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), SP(O)(OR 555 )2, -SP(S)(OR 555 )2, and -SP(S)(SR 556 )(OR 555 ) at least one R 555 is hydrogen.

[0243] In some other embodiments of any one of the aspects, -P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), SP(O)(OR 555 )2, -SP(S)(OR 555 )2, or -SP(S)(SR 556 )(OR 555 ) at least one R 555 is not hydrogen. For example, P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR 555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), SP(O)(OR 555 )2, -SP(S)(OR 555 )2, and -SP(S)(SR 556 )(OR 555 ) at least one R 555 may be substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or an oxygen protecting group.

[0244] In some embodiments of any one of the aspects, -P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR 555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), SP(O)(OR 555 )2, -SP(S)(OR 555 )2, and -SP(S)(SR 556 )(OR 555 ) at least one R 555is H and at least one R 555 is other than H.

[0245] In some embodiments of any one of the aspects, -P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR 555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(O)(OR 555 )2, -SP(S)(OR 555 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) All R in 2 555 is H.

[0246] In some embodiments of any one of the aspects, -P(O)(OR 555 )2, -P(S)(OR 555 )2, -P(S)(SR 556 )(OR 555 ), -OP(O)(OR 555 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(O)(OR 555 )2, -SP(S)(OR 555 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) All R in 2 555 is other than H.

[0247] In some embodiments of any one of the aspects, -P(S)(SR 556 )(OR 555 ), -P(S)(SR 556 )2, -OP(S)(OR555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) At least one R in 2 556 is H.

[0248] In some embodiments of any one of the aspects, -P(S)(SR 556 )(OR 555 ), -P(S)(SR 556 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) At least one R in 2 556 is other than H. For example, -P(S)(SR 556 )(OR 555 ), -P(S)(SR 556 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) At least one R in 2 556 may be substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or sulfur protecting groups.

[0249] In some embodiments of any one of the aspects, -P(S)(SR 556 )2, -OP(S)(SR 556 )2, and -SP(S)(SR 556) At least one R in 2 556 is H and at least one R 556 is other than H.

[0250] In some embodiments, -P(S)(SR 556 )(OR 555 ), -P(S)(SR 556 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) All R in 2 556 is H.

[0251] In some embodiments, -P(S)(SR 556 )(OR 555 ), -P(S)(SR 556 )2, -OP(S)(OR 555 )2, -OP(S)(SR 556 )(OR 555 ), -OP(S)(SR 556 )2, -SP(S)(SR 556 )(OR 555 ), and -SP(S)(SR 556 ) All R in 2 556 is other than H.

[0252] In some embodiments of any one of the aspects, R 45 is -CH=CH-P(O)(OR 555 )2, wherein each R 555 is H or an oxygen protecting group.

[0253] In some embodiments of any one of the aspects, R 42 is a reactive phosphorus group, a solid support, a linker to the solid support, and R 45 is a protected hydroxyl.

[0254] R 52 In some embodiments of any one of the aspects described herein, R 52 is the bond to the internucleotide bond to the subsequent nucleotide, a hydroxyl, a protected hydroxyl, an optionally substituted C 1-30 It can be an alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), amino, alkylamino, dialkylamino, a 3'-oligonucleotide capping group (e.g., an inverted nucleotide or an inverted abasic nucleotide), a ligand, a linker covalently attached to one or more ligands (e.g., N-acetylgalactosamine (GalNac)), a solid support, or a linker covalently attached to a solid support (e.g., -C(O)CHCHC(O)-).

[0255] In some embodiments of any one of the aspects described herein, R 52 is the bond to the internucleotide bond to the subsequent nucleotide, a hydroxyl, a protected hydroxyl, an optionally substituted C 1-30 alkoxy, a 3'-oligonucleotide capping group (e.g., an inverted nucleotide or an inverted abasic nucleotide), a solid support, or a linker covalently attached to a solid support (e.g., -C(O)CHCHC(O)-). For example, R 52 is a bond to an internucleotide bond to a subsequent nucleotide, a hydroxy, a solid support, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—). In some embodiments of any one of the aspects described herein, R 52 is the bond to the internucleotide bond to the subsequent nucleotide, the solid support, or a linker covalently attached to the solid support (e.g., —C(O)CHCHC(O)—).

[0256] In some embodiments of any one of the aspects described herein, R 52 is the bond to the internucleotide bond to the subsequent nucleotide.

[0257] In some embodiments of any one of the aspects described herein, R52 is a solid support or a linker covalently attached to a solid support.

[0258] In some embodiments of any one of the aspects described herein, R 52 is a hydroxyl.

[0259] R 55 In some embodiments of any one of the aspects described herein, R 55 is the bond to the internucleotide bond to the preceding nucleotide, hydrogen, hydroxy, protected hydroxy, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9), vinylphosphonate (VP) group, monophosphate ((HO)2(O)PO-5'), diphosphate ((HO)2(O)POP(HO)(O)-O-5'), triphosphate ((HO)2(O)PO-(HO)(O)POP(HO)(O)-O-5'); monothiophosphate (phosphorothioate, (HO)2(S)PO-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)PO-5'), phosphorothiolate ((HO)2(O)PS-5'); alpha-thiotriphosphate Phosphates; beta-thiotriphosphates; gamma-thiotriphosphates; phosphoramidates ((HO)2(O)P-NH-5', (HO)(NH2)(O)PO-5'), alkyl phosphonates (R(OH)(O)PO-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonates (R(OH)(O)PO-5', R = alkyl ether, e.g., methoxymethyl (CHOMe), ethoxymethyl, etc.), (HO)2(X)PO[-(CH2) a -OP(X)(OH)-O] b -5', or (HO)2(X)PO[-(CH2) a -P(X)(OH)-O] b -5', or (HO)2(X)P-[-(CH2) a -OP(X)(OH)-O] b -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl-terminated phosphates and phosphate mimetics (e.g., HO[-(CH) a -OP(X)(OH)-O] b -5', H2N[-(CH2) a -OP(X)(OH)-O] b -5', H[-(CH2) a -OP(X)(OH)-O] b -5', Me2N[-(CH2) a -OP(X)(OH)-O] b -5', HO[-(CH2) a -P(X)(OH)-O] b -5', H2N[-(CH2) a-P(X)(OH)-O] b -5', H[-(CH2) a -P(X)(OH)-O] b -5', Me2N[-(CH2) a -P(X)(OH)-O] b -5' (wherein a and b are each independently 1 to 10).

[0260] In some embodiments of any one of the aspects described herein, R 55 is the bond to the internucleotide bond to the preceding nucleotide, a hydroxy, a protected hydroxy, an optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 It can be an alkoxy, a vinylphosphonate (VP) group, a monophosphate, a diphosphate, a triphosphate, a monothiophosphate (phosphorothioate), a monodithiophosphate (phosphorodithioate), a phosphorothiolate, an alpha-thiotriphosphate, a beta-thiotriphosphate, a gamma-thiotriphosphate, a phosphoramidate, or an alkylphosphonate.

[0261] In some embodiments of any one of the aspects described herein, R 55 is the bond to the internucleotide bond to the preceding nucleotide, a hydroxy, a protected hydroxy, an optionally substituted C 2-30 Alkenyl, optionally substituted C 1-30 It is an alkoxy or vinylphosphonate (VP) group.

[0262] In some embodiments of any one of the aspects described herein, R 55 is the bond to the internucleotide bond to the preceding nucleotide.

[0263] In some embodiments of any one of the aspects described herein, R 55 is a hydroxyl or a protected hydroxyl.

[0264] In some embodiments of any one of the aspects described herein, R 55 may be substituted C 2-30 Alkenyl or optionally substituted C 1-30 It is an alkoxy.

[0265] In some embodiments of any one of the aspects described herein, R 55 is a vinyl phosphonate group.

[0266] In some embodiments of any one of the aspects described herein, R 55 is -CH(R 51 )-X 5 -R 52 wherein X 5 is absent, a bond, or O, and R 51 is hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted -C 2-30 alkenyl, or optionally substituted -C 2-30 alkynyl, and R 52 is the bond to the internucleoside linkage to the preceding nucleotide.

[0267] In some embodiments of the various aspects described herein, R 55 is -CH(R 51 )-R 52 or -C(R 51 )=CHR 52 wherein R 51 is hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted -C 2-30 alkenyl, or optionally substituted -C 2-30 alkynyl, and R 52 is the bond to the internucleoside linkage to the preceding nucleotide.

[0268] In some embodiments of the various aspects described herein, R 55 is -CH(R 51 )-X5 -R 52 For example, R 55 is -CH(R 51 )-X 5 -R 52 where R 51 is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 51 is H. In some other non-limiting examples, R 51 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 It is alkyl.

[0269] In some embodiments of the various aspects described herein, R 55 is -CH(R 51 )-OR 52 where R 51is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy; 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 51 is H. In some other non-limiting examples, R 51 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 It is alkyl.

[0270] In some embodiments of any one of the aspects described herein, R 55 is -C(R 51 )=CHR 52 -C(R 51 )=CHR 52 It should be noted that the double bond in can be in the cis or trans configuration. Thus, in some embodiments of any one of the aspects, R 55 is -C(R 51 )=CHR 52and the double bond is in a cis configuration. In some other embodiments of any one of the aspects, R 55 is -C(R 51 )=CHR 52 and the double bond is in a trans configuration. In some embodiments of any one of the aspects described herein, R 55 is -CH=CHR 52 is.

[0271] In some embodiments of any one of the aspects described herein, R 52 is the bond to the internucleoside linkage to the preceding nucleotide.

[0272] In embodiments of the various aspects described herein, R 55 may be substituted C 1-6 Alkyl-R 53 , optionally substituted -C 2-6 Alkenyl-R 53 or optionally substituted -C 2-6 Alkynyl-R 53 In embodiments of the various aspects described herein, R 53 -OR 54 , -SR 55 , -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), or -SP(S)(SR 57 )2, and R 54 is a hydrogen or oxygen protecting group, and R 55is a hydrogen or sulfur protecting group, and each R 56 are independently hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or oxygen protecting group, and each R 57 are independently hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or sulfur protecting groups.

[0273] In some embodiments of any one of the aspects, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), SP(O)(OR 56 )2, -SP(S)(OR 56 )2, and -SP(S)(SR 57 )(OR 56 ) at least one R 56 is hydrogen.

[0274] In some other embodiments of any one of the aspects, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), SP(O)(OR 56 )2, -SP(S)(OR 56 )2, or -SP(S)(SR 57 )(OR 56 ) at least one R 56is not hydrogen. For example, P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), SP(O)(OR 56 )2, -SP(S)(OR 56 )2, and -SP(S)(SR 57 )(OR 56 ) at least one R 56 may be substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or an oxygen protecting group.

[0275] In some embodiments of any one of the aspects, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), SP(O)(OR 56 )2, -SP(S)(OR 56 )2, and -SP(S)(SR 57 )(OR 56 ) at least one R 56 is H and at least one R 56 is other than H.

[0276] In some embodiments of any one of the aspects, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -OP(O)(OR 56 )2, -OP(S)(OR 56)2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) All R in 2 56 is H.

[0277] In some embodiments of any one of the aspects, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) All R in 2 56 is other than H.

[0278] In some embodiments of any one of the aspects, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) At least one R in 2 57 is H.

[0279] In some embodiments of any one of the aspects, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) At least one R in 2 57 is other than H. For example, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) At least one R in 2 57 may be substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or sulfur protecting groups.

[0280] In some embodiments of any one of the aspects, -P(S)(SR 57 )2, -OP(S)(SR 57 )2, and SP(S)(SR 57 ) At least one R in 2 57 is H and at least one R 57 is other than H.

[0281] In some embodiments, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR56 ), -OP(S)(SR 57 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) All R in 2 57 is H.

[0282] In some embodiments, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(S)(SR 57 )(OR 56 ), and -SP(S)(SR 57 ) All R in 2 57 is other than H.

[0283] In some embodiments of any one of the aspects described herein, R 55 is optionally substituted -C 2-6 Alkenyl-R 53 For example, R 55 -C 2-6 Alkenyl-R 53 wherein C 2-6Alkenyl includes OH, CN, SC(O)Ph, oxo (=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C 8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p and R is optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH, or CH-aryl-alkoxy, wherein “m” and “p” are independently 1, 2, 3, 4, 5, or 6; 53 is -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), or -SP(S)(SR 57 )2.

[0284] In some embodiments of any one of the aspects, R 55 is -CH=CHR 53Optionally substituted -C 2-6 Alkenyl-R 53 It should be noted that the double bond in can be in the cis or trans configuration. Thus, in some embodiments of any one of the aspects, R 55 is -CH=CHR 53 and the double bond is in a cis configuration. In some other embodiments of any one of the aspects, R 55 is -CH=CHR 53 and the double bond is in the trans configuration.

[0285] In some embodiments of any one of the aspects, R 55 is -CH=CH-P(O)(OR 56 )2, -CH=CH-P(S)(OR 56 )2, -CH=CH-P(S)(SR 57 )(OR 56 ), -CH=CH-P(S)(SR 57 )2, -CH=CH-OP(O)(OR 56 )2, -CH=CH-OP(S)(OR 56 )2, -CH=CH-OP(S)(SR 57 )(OR 56 ), -CH=CH-OP(S)(SR 57 )2, -CH=CH-SP(O)(OR 56 )2, -CH=CH-SP(S)(OR 56 )2, -CH=CH-SP(S)(SR 57 )(OR 56 ), or -CH=CH-SP(S)(SR 57 )2. For example, R 55 is -CH=CH-P(O)(OR 56 )2.

[0286] In some embodiments of any one of the aspects, R 54 is a hydrogen or oxygen protecting group. For example, R 54 is hydrogen or 4,4'-dimethoxytrityl (DMT). In some preferred embodiments, R 54 is H.

[0287] In some embodiments of any one of the aspects described herein, R 55 is optionally substituted -C 1-6 Alkenyl-R 53 For example, R 55 -C 1-6 Alkenyl-R 53 wherein C 1-6 Alkenyl includes OH, CN, SC(O)Ph, oxo (=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C 8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p and R is optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH, or CH-aryl-alkoxy, wherein “m” and “p” are independently 1, 2, 3, 4, 5, or 6; 53 -OR 54 , -SR 55 , -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57)2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), or -SP(S)(SR 57 )2.

[0288] In some embodiments of any one of the aspects described herein, R 55 is -CH(R 58 )-R 53 wherein R 53 -OR 54 , -SR 55 , -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56 )2, -OP(S)(OR 56 )2, -OP(S)(SR 57 )(OR 56 ), -OP(S)(SR 57 )2, -SP(O)(OR 56 )2, -SP(S)(OR 56 )2, -SP(S)(SR 57 )(OR 56 ), or -SP(S)(SR 57 )2 and R 58 is H, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 It is alkynyl.

[0289] In some embodiments of any one of the aspects described herein, R 58is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. In one non-limiting example, R 58 is H. In some other non-limiting examples, R 58 is a C1-C6 optionally substituted with a substituent selected from NH2, OH, C(O)NH2, COOH, halo, SH, and C1-C6 alkoxy; 30 It is alkyl.

[0290] In some embodiments of any one of the aspects described herein, R 55 is -CH(R 58 )-OR 59 where R 59 H, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56)2. For example, R 55 is -CH(R 58 )-OR 59 where R 58 is H or optionally substituted C1-C 30 alkyl, and R 59 is H or -P(O)(OR 56 )2.

[0291] In some embodiments of any one of the aspects described herein, R 55 is -CH(R 58 )-SR 60 where R 60 H, -P(O)(OR 56 )2, -P(S)(OR 56 )2, -P(S)(SR 57 )(OR 56 ), -P(S)(SR 57 )2, -OP(O)(OR 56 )2.

[0292] R 62 In some embodiments of any one of the aspects described herein, R 62 -OR 622 , -SR 623 , optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 Alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, -O(CH2CH2O) r CH2CH2OR 624 , cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, heteroaryl, -NH(CH2CH2NH) s CH2CH2-R 625 , NHC(O)R 626, a lipid, a linker covalently attached to a lipid, a ligand, a linker covalently attached to a ligand, a solid support, a linker covalently attached to a solid support, or a reactive phosphorus group.

[0293] R 622 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 623 is H, a sulfur protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 624 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 625 is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30R can be alkoxy, amino (NH), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl. 626 is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl.

[0294] In some embodiments of any one of the aspects described herein, R 62 is a reactive phosphorus group. For example, R 62 -OP(OR P )(N(R P2 )2), -OP(SR P )(N(R P2 )2), -OP(O)(OR P )(N(R P2 )2), -OP(S)(OR P )(N(R P2 )2), -OP(O)(SR P )(NR P2 )2, -OP(O)(OR P )H, -OP(S)(OR P )H, -OP(O)(SR P )H, -OP(O)(OR P )R P3 , -OP(S)(OR P )R P3 , or -OP(O)(SR P )R P3 is.

[0295] In some embodiments of any one of the aspects, R 62 -OP(OR P )(N(R P2 )2), -OP(SR P )(N(R P2 )2), -OP(O)(OR P )(N(R P2 )2), -OP(S)(OR P )(N(R P2 )2), -OP(O)(SR P )(N(R P2 )2), -OP(O)(OR P )H, -OP(S)(OR P ), optionally substituted C 1-6 alkyl, and each R P2 are independently optionally substituted C 1-6 alkyl, and each R P3 are independently optionally substituted C 1-6 It is alkyl.

[0296] In some embodiments of any one of the aspects, R 62 -OP(OR P )(N(R P2 )2). For example, R 62 -OP(OR P )(N(R P2 )2), wherein R P is cyanoethyl (-CH2CH2CN), and each R P2 is isopropyl.

[0297] In some embodiments of any one of the aspects described herein, R 62 is a solid support or a linker covalently attached to a solid support. For example, R 62 is —OC(O)CHCHC(O)NH—Z, where Z is a solid support.

[0298] In some embodiments of any one of the aspects, R 62 -OR 622 If R 622can be hydrogen or a hydroxyl protecting group. For example, R 622 In some embodiments of any one of the aspects described herein, R can be hydrogen. 62 is -OC(O)CH2CH2CO2H.

[0299] R 62 Ga-SR 623 If R 623 can be hydrogen or a sulfur protecting group. Thus, in some embodiments of any one of the aspects, R 623 is hydrogen.

[0300] R 62 -O(CH2CH2O) r CH2CH2OR 624 where r can be 1 to 50, and R 624 independently for each occurrence, H, C1-C 30 alkyl, cyclyl, heterocyclyl, aryl, heteroaryl, aralkyl, sugar, or R 625 and R 625 is independently for each occurrence amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0301] R 62 -NH(CH2CH2NH) s CH2CH2-R 625 where s can be 1 to 50, and R 625 may be, independently for each occurrence, amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0302] In some embodiments of any one of the aspects described herein, R 62 -OR 622 or optionally substituted C1-C 30 Alkoxy. For example, R 62 is halogen, -OR622 or optionally substituted C1-C 30 In some embodiments of any one of the aspects described herein, R 62 is F, OH, or optionally substituted C1-C 30 It is an alkoxy.

[0303] In some embodiments of any one of the aspects described herein, R 62 are OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C8 alkyl) alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy; 30 alkoxy, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 62 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 In some embodiments of any one of the aspects described herein, R 62 is -O(CH2) tCH3, where t is 1 to 21. For example, t is 14, 15, 16, 17, or 18. In one non-limiting example, t is 16.

[0304] In some embodiments of any one of the aspects, R 62 is -O(CH2) u R 627 wherein u is 2 to 10, and R 627 is C1-C6 alkoxy, amino (NH2), CO2H, OH, or halo. For example, R 627 is —CH or NH. Thus, in some embodiments of any one of the aspects described herein, R 62 is -O(CH2) u -OMe or R 62 is -O(CH2) u It is NH2.

[0305] In some embodiments of any one of the aspects described herein, u is 2, 3, 4, 5, or 6. For example, u is 2, 3, or 6. In one non-limiting example, u is 2. In another non-limiting example, u is 3 or 6.

[0306] In some embodiments of any one of the aspects described herein, R 62 is C1-C6 haloalkyl. For example, R 62 is C-C haloalkyl. In some embodiments of any one of the aspects described herein, R 62 is -CF3, -CF2CF3, -CF2CF2CF3, or -CF2(CF3)2.

[0307] In some embodiments of any one of the aspects described herein, R 62 is -OCH(CH2OR 628 )CH2OR 629 where R 628 and R 629 are independently H, optionally substituted C-C 30Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 628 and R 629 are independently optionally substituted C-C 30 It is alkyl.

[0308] In some embodiments of any one of the aspects described herein, R 62 is -CH2C(O)NHR 6210 where R 6210 is H, optionally substituted C1-C 30 Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 6210 is H or optionally substituted C1-C 30 In some embodiments, R 6210 is optionally substituted C1-C6 alkyl.

[0309] R 63 In some embodiments of any one of the aspects described herein, R 63 is hydrogen, halogen, -OR 632 , -SR 633 , optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 Alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, -O(CH2CH2O) r CH2CH2OR 634 , cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, heteroaryl, -NH(CH2CH2NH) sCH2CH2-R 635 , NHC(O)R 636 , a lipid, a linker covalently attached to a lipid, a ligand, a linker covalently attached to a ligand, a solid support, a linker covalently attached to a solid support, or a reactive phosphorus group.

[0310] R 632 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 633 is H, a sulfur protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 634 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 635 is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30R can be alkoxy, amino (NH), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl. 636 is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl.

[0311] In some embodiments of any one of the aspects described herein, R 63 is R 63 is hydrogen, halogen, -OR 632 , -SR 633 , optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 Alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, -O(CH2CH2O) r CH2CH2OR 634 , cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, heteroaryl, -NH(CH2CH2NH) s CH2CH2-R 635 , NHC(O)R 634 is.

[0312] In some embodiments of any one of the aspects described herein, R 63 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), or -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ) For example, R 63 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 It is an alkoxy, an alkoxyalkyl (eg, methoxyethyl), an alkoxyalkylamine, an alkoxycarboxylate, an amino, an alkylamino, or a dialkylamino.

[0313] In some embodiments of any one of the aspects, R 63 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 Alkoxy, or alkoxyalkyl (eg, methoxyethyl).

[0314] In some embodiments of any one of the aspects, R 63 is a halogen. For example, R 63 can be fluoro, chloro, bromo, or iodo. In some embodiments of any one of the aspects described herein, R 63 is fluoro.

[0315] In some embodiments of any one of the aspects, R 63 -OR 632If R 632 can be hydrogen or a hydroxyl protecting group.

[0316] R 63 Ga-SR 633 If R 633 can be hydrogen or a sulfur protecting group. Thus, in some embodiments of any one of the aspects, R 633 is hydrogen.

[0317] R 63 -O(CH2CH2O) r CH2CH2OR 634 where r can be 1 to 50, and R 634 independently for each occurrence, H, C1-C 30 alkyl, cyclyl, heterocyclyl, aryl, heteroaryl, aralkyl, sugar, or R 635 and R 635 is independently for each occurrence amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0318] R 63 -NH(CH2CH2NH) s CH2CH2-R 635 where s can be 1 to 50, and R 635 may be, independently for each occurrence, amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0319] In some embodiments of any one of the aspects described herein, R 63 is hydrogen, halogen, -OR 632 or optionally substituted C1-C 30 Alkoxy. For example, R 63 is halogen, -OR 632 or optionally substituted C1-C 30In some embodiments of any one of the aspects described herein, R 63 is F, OH, or optionally substituted C1-C 30 It is an alkoxy.

[0320] In some embodiments of any one of the aspects described herein, R 63 are OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C8 alkyl) alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy; 30 alkoxy, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 63 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 In some embodiments of any one of the aspects described herein, R 63 is -O(CH2) t CH3, where t is 1 to 21. For example, t is 14, 15, 16, 17, or 18. In one non-limiting example, t is 16.

[0321] In some embodiments of any one of the aspects, R 63 is -O(CH2) u R 637 wherein u is 2 to 10, and R 637 is C1-C6 alkoxy, amino (NH2), CO2H, OH, or halo. For example, R 637 is —CH or NH. Thus, in some embodiments of any one of the aspects described herein, R 63 is -O(CH2) u -OMe or R 63 is -O(CH2) u It is NH2.

[0322] In some embodiments of any one of the aspects described herein, u is 2, 3, 4, 5, or 6. For example, u is 2, 3, or 6. In one non-limiting example, u is 2. In another non-limiting example, u is 3 or 6.

[0323] In some embodiments of any one of the aspects described herein, R 63 is C1-C6 haloalkyl. For example, R 63 is C-C haloalkyl. In some embodiments of any one of the aspects described herein, R 63 is -CF3, -CF2CF3, -CF2CF2CF3, or -CF2(CF3)2.

[0324] In some embodiments of any one of the aspects described herein, R 63 is -OCH(CH2OR 638 )CH2OR 639 where R 638 and R 639 are independently H, optionally substituted C-C 30 Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 638 and R 639are independently optionally substituted C-C 30 It is alkyl.

[0325] In some embodiments of any one of the aspects described herein, R 63 is -CH2C(O)NHR 6310 where R 6310 is H, optionally substituted C1-C 30 Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 6310 is H or optionally substituted C1-C 30 In some embodiments, R 6310 is optionally substituted C1-C6 alkyl.

[0326] In some embodiments of any one of the aspects described herein, R 63 is hydrogen, fluoro, -O-MOE, -O-alkyl (e.g., methoxy or -OC 16 aliphatic), -O-alkene, -O-alkyne, -O-lipid, -O-branched lipid, or aminoalkyl.

[0327] R 64 In some embodiments of any one of the aspects described herein, R 64 is hydrogen, halogen, -OR 642 , -SR 643 , optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 Alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, -O(CH2CH2O) r CH2CH2OR 644, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, heteroaryl, -NH(CH2CH2NH) s CH2CH2-R 645 , NHC(O)R 646 , a lipid, a linker covalently attached to a lipid, a ligand, a linker covalently attached to a ligand, a solid support, a linker covalently attached to a solid support, or a reactive phosphorus group.

[0328] R 642 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 643 is H, a sulfur protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 644 is H, a hydroxyl protecting group, an optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl. R 645 is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C1-30 R can be alkoxy, amino (NH), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl. 646 is hydrogen, halogen, hydroxyl, protected hydroxyl, optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 It can be alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, or heteroaryl.

[0329] In some embodiments of any one of the aspects described herein, R 64 is R 64 is hydrogen, halogen, -OR 642 , -SR 643 , optionally substituted C 1-30 Alkyl, C 1-30 Haloalkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, or optionally substituted C 1-30 Alkoxy, amino (NH2), alkylamino, dialkylamino, heterocyclyl, arylamino, diarylamino, heteroarylamino, diheteroarylamino, amino acid, -O(CH2CH2O) r CH2CH2OR 644 , cyano, alkyl-thio-alkyl, thioalkoxy, cycloalkyl, aryl, heteroaryl, -NH(CH2CH2NH) s CH2CH2-R 645 , NHC(O)R 644 is.

[0330] In some embodiments of any one of the aspects described herein, R 64 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 Alkoxy, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), or -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ) For example, R 64 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 It is an alkoxy, an alkoxyalkyl (eg, methoxyethyl), an alkoxyalkylamine, an alkoxycarboxylate, an amino, an alkylamino, or a dialkylamino.

[0331] In some embodiments of any one of the aspects, R 64 is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C 1-30 Alkoxy, or alkoxyalkyl (eg, methoxyethyl).

[0332] In some embodiments of any one of the aspects, R 64 is a halogen. For example, R 64 can be fluoro, chloro, bromo, or iodo. In some embodiments of any one of the aspects described herein, R 64 is fluoro.

[0333] In some embodiments of any one of the aspects, R 64 -OR642 If R 642 can be hydrogen or a hydroxyl protecting group.

[0334] R 64 Ga-SR 643 If R 643 can be hydrogen or a sulfur protecting group. Thus, in some embodiments of any one of the aspects, R 643 is hydrogen.

[0335] R 64 -O(CH2CH2O) r CH2CH2OR 644 where r can be 1 to 50, and R 644 independently for each occurrence, H, C1-C 30 alkyl, cyclyl, heterocyclyl, aryl, heteroaryl, aralkyl, sugar, or R 645 and R 645 is independently for each occurrence amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0336] R 64 -NH(CH2CH2NH) s CH2CH2-R 645 where s can be 1 to 50, and R 645 may be, independently for each occurrence, amino (NH2), alkylamino, dialkylamino, arylamino, diarylamino, heteroarylamino, or diheteroarylamino.

[0337] In some embodiments of any one of the aspects described herein, R 64 is hydrogen, halogen, -OR 642 or optionally substituted C1-C 30 Alkoxy. For example, R 64 is halogen, -OR 642 or optionally substituted C1-C 30In some embodiments of any one of the aspects described herein, R 64 is F, OH, or optionally substituted C1-C 30 It is an alkoxy.

[0338] In some embodiments of any one of the aspects described herein, R 64 are OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1-C8 alkyl) alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy; 30 alkoxy, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 64 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 In some embodiments of any one of the aspects described herein, R 64 is -O(CH2) t CH3, where t is 1 to 21. For example, t is 14, 15, 16, 17, or 18. In one non-limiting example, t is 16.

[0339] In some embodiments of any one of the aspects, R 64 is -O(CH2) u R 647 wherein u is 2 to 10, and R 647 is C1-C6 alkoxy, amino (NH2), CO2H, OH, or halo. For example, R 647 is —CH or NH. Thus, in some embodiments of any one of the aspects described herein, R 64 is -O(CH2) u -OMe or R 64 is -O(CH2) u It is NH2.

[0340] In some embodiments of any one of the aspects described herein, u is 2, 3, 4, 5, or 6. For example, u is 2, 3, or 6. In one non-limiting example, u is 2. In another non-limiting example, u is 3 or 6.

[0341] In some embodiments of any one of the aspects described herein, R 64 is C1-C6 haloalkyl. For example, R 64 is C-C haloalkyl. In some embodiments of any one of the aspects described herein, R 64 is -CF3, -CF2CF3, -CF2CF2CF3, or -CF2(CF3)2.

[0342] In some embodiments of any one of the aspects described herein, R 64 is -OCH(CH2OR 648 )CH2OR 649 where R 648 and R 649 are independently H, optionally substituted C-C 30 Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 648 and R 649are independently optionally substituted C-C 30 It is alkyl.

[0343] In some embodiments of any one of the aspects described herein, R 64 is -CH2C(O)NHR 6410 where R 6410 is H, optionally substituted C1-C 30 Alkyl, optionally substituted C-C 30 Alkenyl or optionally substituted C-C 30 Alkynyl. For example, R 6410 is H or optionally substituted C1-C 30 In some embodiments, R 6410 is optionally substituted C1-C6 alkyl.

[0344] In some embodiments of any one of the aspects described herein, R 63 is hydrogen, fluoro, -O-MOE, -O-alkyl (e.g., methoxy or -OC 16 aliphatic), -O-alkene, -O-alkyne, -O-lipid, -O-branched lipid, or aminoalkyl.

[0345] In some embodiments of any one of the aspects described herein, R 63 and R 64 One of the is hydroxyl and the other is hydrogen, methoxy, fluoro, -O-MOE, -O-alkyl, -O-alkene, -O-alkyne, --O-C16, -O-lipid, -O-branched lipid, or aminoalkyl.

[0346] R 65 In some embodiments of the various aspects described herein, R 65 is R 651 , optionally substituted C 1-6 Alkyl-R 651 , optionally substituted -C 2-6 Alkenyl-R 651or optionally substituted -C 2-6 Alkynyl-R 651 where R 651 -OR 652 , -SR 653 , hydrogen, a phosphorus group, a solid support, or a linker to a solid support. 651 -OR 652 If R 652 can be H or a hydroxyl protecting group. Similarly, R 651 Ga-SR 653 If R 653 can be H or a sulfur protecting group.

[0347] In some embodiments of any one of the aspects described herein, R 65 -OR 652 or -SR 653 is.

[0348] In some embodiments of any one of the aspects described herein, R 652 is a hydroxyl protecting group. 652 Exemplary hydroxyl protecting groups for include, but are not limited to, benzyl, benzoyl, 2,6-dichlorobenzyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, mesylate, tosylate, 4,4'-dimethoxytrityl (DMT), 9-phenylxanthin-9-yl (pixyl), and 9-(p-methoxyphenyl)xanthin-9-yl (MOX). In some embodiments of any one of the aspects described herein, R 65 -OR 652 and R 652 is 4,4'-dimethoxytrityl (DMT), for example, R 65 is -O-DMT.

[0349] In some embodiments of any one of the aspects described herein, R 65 is -CH(R 654 )-R 651 where R 654 is hydrogen, halogen, optionally substituted C1-C30 Alkyl, optionally substituted C-C 30 Alkenyl, optionally substituted C-C 30 Alkynyl or optionally substituted C-C 30 It is an alkoxy.

[0350] In some embodiments of any one of the aspects, R 65 -CH(R 654 )-R 651 If R 654 is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 654 is H. In some other non-limiting examples, R 654 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 It is alkyl.

[0351] In some embodiments of the various aspects described herein, R 65 is -CH(R 654 )-OR 652 where R 654 is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6. For example, R 654 is H. In some other non-limiting examples, R 654 is a C1-C6 alkyl group optionally substituted with NH2, OH, C(O)NH2, COOH, halo, SH, or C1-C6 alkoxy; 30 It is alkyl.

[0352] In some embodiments of the various aspects described herein, R 65 may be substituted C 1-6 Alkyl-R 651 or optionally substituted -C 2-6 Alkenyl-R 651 is.

[0353] In some embodiments of any one of the aspects described herein, R 65 is -C(R 654 )=CHR 651 -C(R 654 )=CHR 651 It should be noted that the double bond in can be in the cis or trans configuration. Thus, in some embodiments of any one of the aspects, R d is -C(R 654 )=CHR 651 and the double bond is in a cis configuration. In some other embodiments of any one of the aspects, R d is -C(R 654 )=CHR 651 and the double bond is in the trans configuration.

[0354] In some embodiments of any one of the aspects described herein, R 65 is -CH=CHR 651 is.

[0355] In some embodiments of any one of the aspects, R 65 -C(R 654 )=CHR 651 If R 654 is H or OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl, O(C1-C8)alkyl (i.e., C1 -C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2)p -OH, CH2-[CH(OH)] m -(CH2) p C1-C optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from —NH2, or CH2-aryl-alkoxy 30 alkyl, "m" and "p" are independently 1, 2, 3, 4, 5, or 6, and R 651 is a phosphorus group. For example, R 65 is -CH=CHR 651 is.

[0356] In some embodiments of any one of the aspects described herein, R 651 is a reactive phosphorus group.

[0357] In some embodiments of any one of the aspects, R 65 is -CH=CH-P(O)(OR 655 )2, -CH=CH-P(S)(OR 655 )2, -CH=CH-P(S)(SR 656 )(OR 655 ), -CH=CH-P(S)(SR 656 )2, -CH=CH-OP(O)(OR 655 )2, -CH=CH-OP(S)(OR 655 )2, -CH=CH-OP(S)(SR 656 )(OR 655 ), -CH=CH-OP(S)(SR 656 )2, -CH=CH-SP(O)(OR 655 )2, -CH=CH-SP(S)(OR 655 )2, -CH=CH-SP(S)(SR 656 )(OR 655 ), or -CH=CH-SP(S)(SR 656 )2, wherein each R 655 are independently hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or oxygen protecting group, and each R 656are independently hydrogen, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or sulfur protecting groups.

[0358] In some embodiments of any one of the aspects, -P(O)(OR 655 )2, -P(S)(OR 655 )2, -P(S)(SR 656 )(OR 655 ), -OP(O)(OR 655 )2, -OP(S)(OR 655 )2, -OP(S)(SR 656 )(OR 655 ), SP(O)(OR 655 )2, -SP(S)(OR 655 )2, and -SP(S)(SR 656 )(OR 655 ) at least one R 655 is hydrogen.

[0359] In some other embodiments of any one of the aspects, -P(O)(OR 655 )2, -P(S)(OR 655 )2, -P(S)(SR 656 )(OR 655 ), -OP(O)(OR 655 )2, -OP(S)(OR 655 )2, -OP(S)(SR 656 )(OR 655 ), SP(O)(OR 655 )2, -SP(S)(OR 655 )2, or -SP(S)(SR 656 )(OR 655 ) at least one R 655 is not hydrogen. For example, P(O)(OR 655 )2, -P(S)(OR 655 )2, -P(S)(SR 656 )(OR 655 ), -OP(O)(OR 655 )2, -OP(S)(OR 655 )2, -OP(S)(SR656 )(OR 655 ), SP(O)(OR 655 )2, -SP(S)(OR 655 )2, and -SP(S)(SR 656 )(OR 655 ) at least one R 655 may be substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or an oxygen protecting group.

[0360] In some embodiments of any one of the aspects, -P(O)(OR 655 )2, -P(S)(OR 655 )2, -P(S)(SR 656 )(OR 655 ), -OP(O)(OR 655 )2, -OP(S)(OR 655 )2, -OP(S)(SR 656 )(OR 655 ), SP(O)(OR 655 )2, -SP(S)(OR 655 )2, and -SP(S)(SR 656 )(OR 655 ) at least one R 655 is H and at least one R 655 is other than H.

[0361] In some embodiments of any one of the aspects, -P(O)(OR 655 )2, -P(S)(OR 655 )2, -P(S)(SR 656 )(OR 655 ), -OP(O)(OR 655 )2, -OP(S)(OR 655 )2, -OP(S)(SR 656 )(OR 655 ), -OP(S)(SR 656 )2, -SP(O)(OR 655 )2, -SP(S)(OR 655 )2, -SP(S)(SR 656 )(OR 655), and -SP(S)(SR 656 ) All R in 2 655 is H.

[0362] In some embodiments of any one of the aspects, -P(O)(OR 655 )2, -P(S)(OR 655 )2, -P(S)(SR 656 )(OR 655 ), -OP(O)(OR 655 )2, -OP(S)(OR 655 )2, -OP(S)(SR 656 )(OR 655 ), -OP(S)(SR 656 )2, -SP(O)(OR 655 )2, -SP(S)(OR 655 )2, -SP(S)(SR 656 )(OR 655 ), and -SP(S)(SR 656 ) All R in 2 655 is other than H.

[0363] In some embodiments of any one of the aspects, -P(S)(SR 656 )(OR 655 ), -P(S)(SR 656 )2, -OP(S)(OR 655 )2, -OP(S)(SR 656 )(OR 655 ), -OP(S)(SR 656 )2, -SP(S)(SR 656 )(OR 655 ), and -SP(S)(SR 656 ) At least one R in 2 656 is H.

[0364] In some embodiments of any one of the aspects, -P(S)(SR 656 )(OR 655 ), -P(S)(SR 656 )2, -OP(S)(OR 655 )2, -OP(S)(SR 656 )(OR 655 ), -OP(S)(SR 656)2, -SP(S)(SR 656 )(OR 655 ), and -SP(S)(SR 656 ) At least one R in 2 656 is other than H. For example, -P(S)(SR 656 )(OR 655 ), -P(S)(SR 656 )2, -OP(S)(OR 655 )2, -OP(S)(SR 656 )(OR 655 ), -OP(S)(SR 656 )2, -SP(S)(SR 656 )(OR 655 ), and -SP(S)(SR 656 ) At least one R in 2 656 may be substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, or optionally substituted C 2-30 alkynyl, or sulfur protecting groups.

[0365] In some embodiments of any one of the aspects, -P(S)(SR 656 )2, -OP(S)(SR 656 )2, and -SP(S)(SR 656 ) At least one R in 2 656 is H and at least one R 656 is other than H.

[0366] In some embodiments, -P(S)(SR 656 )(OR 655 ), -P(S)(SR 656 )2, -OP(S)(OR 655 )2, -OP(S)(SR 656 )(OR 655 ), -OP(S)(SR 656 )2, -SP(S)(SR 656 )(OR 655 ), and -SP(S)(SR 656 ) All R in 2 656 is H.

[0367] In some embodiments, -P(S)(SR 656 )(OR 655 ), -P(S)(SR 656 )2, -OP(S)(OR 655 )2, -OP(S)(SR 656 )(OR 655 ), -OP(S)(SR 656 )2, -SP(S)(SR 656 )(OR 655 ), and -SP(S)(SR 656 ) All R in 2 656 is other than H.

[0368] In some embodiments of any one of the aspects, R 65 is -CH=CH-P(O)(OR 655 )2, wherein each R 655 is H or an oxygen protecting group.

[0369] In some embodiments of any one of the aspects, R 63 is a reactive phosphorus group, a solid support, a linker to the solid support, and R 65 is a protected hydroxyl.

[0370] R 62x In some embodiments of any one of the aspects described herein, R 62x is the bond to the internucleotide bond to the subsequent nucleotide, a hydroxyl, a protected hydroxyl, an optionally substituted C 1-30 It can be an alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), amino, alkylamino, dialkylamino, a 3'-oligonucleotide capping group (e.g., an inverted nucleotide or an inverted abasic nucleotide), a ligand, a linker covalently attached to one or more ligands (e.g., N-acetylgalactosamine (GalNac)), a solid support, or a linker covalently attached to a solid support (e.g., -C(O)CHCHC(O)-).

[0371] In some embodiments of any one of the aspects described herein, R62x is the bond to the internucleotide bond to the subsequent nucleotide, a hydroxyl, a protected hydroxyl, an optionally substituted C 1-30 alkoxy, a 3'-oligonucleotide capping group (e.g., an inverted nucleotide or an inverted abasic nucleotide), a solid support, or a linker covalently attached to a solid support (e.g., -C(O)CHCHC(O)-). For example, R 62x is a bond to an internucleotide bond to a subsequent nucleotide, a hydroxy, a solid support, or a linker covalently attached to a solid support (e.g., —C(O)CHCHC(O)—). In some embodiments of any one of the aspects described herein, R 52 is the bond to the internucleotide bond to the subsequent nucleotide, the solid support, or a linker covalently attached to the solid support (e.g., —C(O)CHCHC(O)—).

[0372] In some embodiments of any one of the aspects described herein, R 62x is the bond to the internucleotide bond to the subsequent nucleotide.

[0373] In some embodiments of any one of the aspects described herein, R 62x is a solid support or a linker covalently attached to a solid support.

[0374] In some embodiments of any one of the aspects described herein, R 62x is a hydroxyl.

[0375] R 65x In some embodiments of any one of the aspects described herein, R 65x is the bond to the internucleotide bond to the preceding nucleotide, hydrogen, hydroxy, protected hydroxy, optionally substituted C 1-30 Alkyl, optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30Alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), -OC 4-30 Alkyl-ON(CH2R 8 )(CH2R 9 ), vinylphosphonate (VP) group, monophosphate ((HO)2(O)PO-5'), diphosphate ((HO)2(O)POP(HO)(O)-O-5'), triphosphate ((HO)2(O)PO-(HO)(O)POP(HO)(O)-O-5'); monothiophosphate (phosphorothioate, (HO)2(S)PO-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)PO-5'), phosphorothiolate ((HO)2(O)PS-5'); alpha-thiotriphosphate Phosphates; beta-thiotriphosphates; gamma-thiotriphosphates; phosphoramidates ((HO)2(O)P-NH-5', (HO)(NH2)(O)PO-5'), alkyl phosphonates (R(OH)(O)PO-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonates (R(OH)(O)PO-5', R = alkyl ether, e.g., methoxymethyl (CHOMe), ethoxymethyl, etc.), (HO)2(X)PO[-(CH2) a -OP(X)(OH)-O] b -5', or (HO)2(X)PO[-(CH2) a -P(X)(OH)-O] b -5', or (HO)2(X)P-[-(CH2) a -OP(X)(OH)-O] b -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl-terminated phosphates and phosphate mimetics (e.g., HO[-(CH) a -OP(X)(OH)-O] b -5', H2N[-(CH2) a-OP(X)(OH)-O] b -5', H[-(CH2) a -OP(X)(OH)-O] b -5', Me2N[-(CH2) a -OP(X)(OH)-O] b -5', HO[-(CH2) a -P(X)(OH)-O] b -5', H2N[-(CH2) a -P(X)(OH)-O] b -5', H[-(CH2) a -P(X)(OH)-O] b -5', Me2N[-(CH2) a -P(X)(OH)-O] b -5' (wherein a and b are each independently 1 to 10).

[0376] In some embodiments of any one of the aspects described herein, R 65x is the bond to the internucleotide bond to the preceding nucleotide, a hydroxy, a protected hydroxy, an optionally substituted C 2-30 Alkenyl, optionally substituted C 2-30 Alkynyl, optionally substituted C 1-30 It can be an alkoxy, a vinylphosphonate (VP) group, a monophosphate, a diphosphate, a triphosphate, a monothiophosphate (phosphorothioate), a monodithiophosphate (phosphorodithioate), a phosphorothiolate, an alpha-thiotriphosphate, a beta-thiotriphosphate, a gamma-thiotriphosphate, a phosphoramidate, or an alkylphosphonate.

[0377] In some embodiments of any one of the aspects described herein, R 65x is the bond to the internucleotide bond to the preceding nucleotide, a hydroxy, a protected hydroxy, an optionally substituted C 2-30 Alkenyl, optionally substituted C 1-30 It is an alkoxy or vinylphosphonate (VP) group.

[0378] In some embodiments of any one of the aspects described herein, R 65x is the bond to the internucleotide bond to the preceding nucleotide.

[0379] In some embodiments of any one of the aspects described herein, R 65x is a hydroxyl or a protected hydroxyl.

[0380] In some embodiments of any one of the aspects described herein, R 65x may be substituted C 2-30 Alkenyl or optionally substituted C 1-30 It is an alkoxy.

[0381] In some embodiments of any one of the aspects described herein, R 65x is a vinyl phosphonate group.

[0382] L In embodiments of the various aspects described herein, L is a linker.

[0383] As used herein, the term "linker" means an organic moiety that connects two parts of a compound. A linker is typically a direct bond or an atom such as oxygen or sulfur, NR 1 , C(O), C(O)O, C(O)NR 1, SO, SO2, SO2NH, or other units or chains of atoms, such as substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, aryl alkyl, aryl alkenyl, aryl alkynyl, heteroaryl alkyl, heteroaryl alkenyl, heteroaryl alkynyl, heterocyclyl alkyl, heterocyclyl alkenyl, heterocyclyl alkynyl, aryl, heteroaryl, heterocyclyl, cycloalkyl, cycloalkenyl, alkylaryl alkyl, alkylaryl alkenyl, alkylaryl alkynyl, alkenylaryl alkyl, alkenylaryl alkenyl, alkenylaryl alkynyl, alkynylaryl alkyl, alkynylaryl alkenyl, alkynylaryl alkynyl, alkylheteroaryl alkyl, alkylheteroaryl alkenyl, alkylheteroaryl alkynyl, alkenyl alkylheteroarylalkyl, alkenylheteroarylalkenyl, alkenylheteroarylalkynyl, alkynylheteroarylalkyl, alkynylheteroarylalkenyl, alkynylheteroarylalkynyl, alkylheterocyclylalkyl, alkylheterocyclylalkenyl, alkylhererocyclylalkynyl, alkenylheterocyclylalkyl, alkenylheterocyclylalkenyl, alkenylheterocyclylalkynyl, alkynylheterocyclylalkyl, alkynylheterocyclylalkenyl, alkynylheterocyclylalkynyl, alkylaryl, alkenylaryl, alkynylaryl, alkylheteroaryl, alkenylheteroaryl, alkynylhereroaryl, wherein one or more methylenes are O, S, S(O), SO, N(R 1 )2, C(O), a cleavable linking group, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted heterocycle, wherein R 1 is hydrogen, acyl, aliphatic, or substituted aliphatic.

[0384] In some embodiments, the linker is a cleavable linker. A cleavable linker is a linker that, depending on the process inside the target cell, releases the two moieties it holds together, such as reduction in the cytoplasm, exposure to acidic conditions in a lysosome or endosome, or cleavage by a specific enzyme (e.g., protease) inside the cell. Thus, the cleavable linker allows the two moieties to be released in their original form after internalization and processing inside the target cell. Cleavable linkers include, but are not limited to, linkers whose bonds can be cleaved by enzymes (e.g., peptide linkers), reducing conditions (e.g., disulfide linkers), or acidic conditions (e.g., hydrazones and carbonates).

[0385] Generally, a cleavable linker comprises at least one cleavable linking group. A cleavable linking group is a group that is sufficiently stable outside a cell, but that, once inside a target cell, is cleaved to release the two moieties that the linker holds together. In a preferred embodiment, the cleavable linking group is cleaved at least 10 times faster, preferably at least 100 times faster, in the target cell or under a first reference condition (e.g., which may be selected to mimic or represent intracellular conditions) than in the subject's blood or serum, or under a second reference condition (e.g., which may be selected to mimic or represent conditions found in blood or serum).

[0386] Cleavable linking groups are susceptible to cleavage factors, such as pH, redox potential, or the presence of degradative molecules. Cleavage factors are generally more widespread or found at higher levels or activity inside cells than in serum or blood. Examples of such degradative agents include redox agents that are selective for specific substrates or have no substrate specificity (e.g., reducing agents such as oxidases or reductases or mercaptans present in cells that can degrade redox-cleavable linking groups by reduction); esterases; endosomes or factors that can create an acidic environment (e.g., those that result in a pH of 5 or less); enzymes that can hydrolyze or degrade acid-cleavable linking groups by acting as general acids, peptidases (which may be substrate-specific), and phosphatases.

[0387] Cleavable linking groups, such as disulfide bonds, can be pH-sensitive. While the pH of human serum is 7.4, the average intracellular pH is slightly lower, ranging from approximately 7.1 to 7.3. Endosomes have a more acidic pH, ranging from 5.5 to 6.0, and lysosomes have an even more acidic pH of approximately 5.0. Some linkers have cleavable linking groups that are cleaved at a preferred pH, thereby releasing the cationic lipid from the ligand into the interior of the cell or into a desired compartment of the cell.

[0388] The linker can contain a cleavable linking group that can be cleaved by a specific enzyme.The type of cleavable linking group incorporated into the linker can depend on the target cell.For example, a liver targeting ligand can be linked to a cationic lipid via a linker containing an ester group.Hepatocytes are rich in esterase, so the linker will be cleaved more efficiently in hepatocytes than in cell types that are not rich in esterase.Other cell types that are rich in esterase include lung, renal cortex and testicular cells.When targeting cell types that are rich in peptidase, such as hepatocytes and synovial cells, a linker containing a peptide bond can be used.

[0389] Generally, the suitability of a candidate cleavable linking group can be evaluated by testing the ability (or conditions) of a degradation agent to cleave the candidate linking group. It may also be desirable to test the candidate cleavable linking group for its ability to resist cleavage in blood or when contacted with other non-target tissues. Thus, the relative susceptibility to cleavage can be determined between a first condition and a second condition, where the first condition is selected to exhibit cleavage in target cells and the second condition is selected to exhibit cleavage in other tissues or biological fluids, such as blood or serum. Evaluation can be performed in a cell-free system, in cells, in cell culture, in organ or tissue culture, or in whole animals. It can be useful to perform initial evaluations in cell-free or culture conditions and confirm by further evaluation in whole animals. In preferred embodiments, useful candidate compounds are cleaved at least 2, 4, 10, or 100 times faster in cells (or under in vitro conditions selected to mimic intracellular conditions) than in blood or serum (or under in vitro conditions selected to mimic extracellular conditions).

[0390] One class of cleavable linking group is a redox-cleavable linking group, which can be used in dsRNA molecules according to the present invention and is cleaved upon reduction or oxidation. An example of a reductively cleavable linking group is a disulfide bond (-SS-). To determine whether a candidate cleavable linking group is a suitable "reductively cleavable linking group," or is suitable for use with, for example, a particular iRNA moiety and a particular targeting agent, one can refer to the methods described herein. For example, candidates can be evaluated by incubating with dithiothreitol (DTT) or other reducing agents using reagents known in the art that mimic the cleavage rate that would be observed in cells, e.g., target cells. Candidates can also be evaluated under conditions selected to mimic blood or serum conditions. In a preferred embodiment, the candidate compound is cleaved at a maximum of 10% in blood. In preferred embodiments, useful candidate compounds are degraded at a rate at least 2, 4, 10, or 100 times faster in cells (or under in vitro conditions selected to mimic intracellular conditions) compared to blood (or under in vitro conditions selected to mimic extracellular conditions). The rate of cleavage of the candidate compound can be determined using standard enzyme kinetic assays under conditions selected to mimic the intracellular medium and compared to conditions selected to mimic the extracellular medium.

[0391] Phosphate-cleavable linker groups that may be used in the dsRNA molecules of the present invention are cleaved by agents that degrade or hydrolyze phosphate groups. Examples of agents that cleave phosphate groups within cells include enzymes such as phosphatases within the cell. Examples of phosphate-based linking groups are -OP(O)(ORk)-O-, -OP(S)(ORk)-O-, -OP(S)(SRk)-O-, -SP(O)(ORk)-O-, -OP(O)(ORk)-S-, -SP(O)(ORk)-S-, -OP(S)(ORk)-S-, -SP(S)(ORk)-O-, -OP(O)(Rk)-O-, -OP(S)(Rk)-O-, -SP(O)(Rk)-O-, -SP(S)(Rk)-O-, -SP(O)(Rk)-S-, -OP(S)(Rk)-S-, where Rk, for each occurrence, can independently be hydrogen, C-C alkyl, C-C haloalkyl, C-C aryl, or C-C aralkyl. Preferred embodiments include -OP(O)(OH)-O-, -OP(S)(OH)-O-, -OP(S)(SH)-O-, -SP(O)(OH)-O-, -OP(O)(OH)-S-, -SP(O)(OH)-S-, -OP(S)(OH)-S-, -SP(S)(OH)-O-, -OP(O)(H)-O-, -OP(S)(H)-O-, -SP(O)(H)-O, -SP(S)(H)-O-, -SP(O)(H)-S-, and -OP(S)(H)-S-. A preferred embodiment is -OP(O)(OH)-O-. These candidates can be evaluated using methods similar to those described above.

[0392] Acid-cleavable linking groups that can be used in dsRNA molecules of the present invention are linking groups that are cleaved under acidic conditions. In a preferred embodiment, the acid-cleavable linking group is cleaved in an acidic environment with a pH of about 6.5 or less (e.g., about 6.0, 5.5, 5.0, or less), or by an agent such as an enzyme that can act as a general acid. Within cells, certain low-pH organelles, such as endosomes and lysosomes, can provide a cleavage environment for the acid-cleavable linking group. Examples of acid-cleavable linking groups include, but are not limited to, hydrazones, esters, and esters of amino acids. Acid-cleavable groups can have the general formula -C=NN-, C(O)O, or -OC(O). In a preferred embodiment, the carbon bonded to the oxygen of the ester (alkoxy group) is an aryl group, a substituted alkyl group, or a tertiary alkyl group, such as dimethylpentyl or t-butyl. These candidates can be evaluated using methods similar to those described above.

[0393] The ester-based cleavable linking group that can be used in the dsRNA molecule of the present invention can be cleaved by enzymes such as intracellular esterase and amidase.Examples of ester-based cleavable linking groups include but are not limited to alkylene, alkenylene and alkynylene ester groups.The cleavable linking group of ester has the general formula -C(O)O- or -OC(O)-.These candidates can be evaluated using a method similar to that described above.

[0394] Peptide-based cleavable linking groups that can be used in the dsRNA molecules of the present invention are cleaved by enzymes such as intracellular peptidases and proteases. Peptide-based cleavable linking groups are peptide bonds formed between amino acids to yield oligopeptides (e.g., dipeptides, tripeptides, etc.) and polypeptides. Peptide-based cleavable groups do not include amide groups (-C(O)NH-). Amide groups can be formed between any alkylene, alkenylene, or alkynylene. A peptide bond is a special type of amide bond formed between amino acids to yield peptides and proteins. Peptide-based cleavable groups are generally limited to peptide bonds (i.e., amide bonds) formed between amino acids to yield peptides and proteins, and do not include all amide functional groups. Peptide-based cleavable linking groups have the general formula -NHCHR A C(O)NHCHR B C(O)—, where R A and R B are the R groups of two adjacent amino acids.

[0395] In some embodiments of any one of the aspects, L is a bond.

[0396] In some embodiments of any one of the aspects, L is absent, e.g., R 6 or R 7 -R L is.

[0397] L P In some embodiments of any one of the aspects, L P is a linker. For example, L P can be a bond.

[0398] In some embodiments of any one of the aspects described herein, L P is optionally substituted C1-C 20 Alkylene (e.g., -(CH2) b-, where b is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20), or an optionally substituted C2-C 20 Alkynylene, and the skeleton of the alkylene or alkynylene is O, S, S(O), SO2, NR 1 , N.R. 1 -C(O), C(O), C(O)O, a cleavable linking group, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted heterocyclic, R N1 is hydrogen, acyl, aliphatic, or substituted aliphatic. For example, L P is an optionally substituted C1-C6 alkylene.

[0399] In some embodiments of any one of the aspects, L P is optionally substituted C1-C 20 It is an alkylene, and the alkylene backbone is interrupted by a heteroaryl (eg, triazole) or NHC(O).

[0400] In some embodiments of any one of the aspects, L P is optionally substituted C2-C 20 alkylene. For example, L P are -(CH2)3-, -(CH2)5-, -(CH2)7-, -(CH2)9-, -(CH2) 10 -, -(CH2) 11 -, -(CH2) 12 -, -(CH2) 13 -, -(CH2) 15 - or -(CH2) 17 -It is.

[0401] In some embodiments of any one of the aspects, L P is polyethylene glycol. For example, L P is -(CH2CH2O) L’-O-CH2-, where L' is an integer selected from 1 to 25. In some embodiments, L' is an integer selected from 1 to 10. For example, L' is 1, 2, 3, 4, 5, or 6.

[0402] In some embodiments of any one of the aspects, L P does not exist.

[0403] internucleoside bond As used herein, "internucleoside linkage" refers to a covalent bond between adjacent nucleosides. Two major classes of internucleoside linkages are defined by the presence or absence of a phosphorus atom. Representative phosphorus-containing linkages include, but are not limited to, phosphodiester (P=O), phosphotriester, methylphosphonate, phosphoramidate, and phosphorothioate (P=S). Representative non-phosphorus-containing linking groups include, but are not limited to, methylenemethylimino (-CH2-N(CH3)-O-CH2-), thiodiester (-OC(O)-S-), thionocarbamate (-OC(O)(NH)-S-), siloxane (-O-Si(H)2-O-), and N,N'-dimethylhydrazine (-CH2-N(CH3)-N(CH3)-). Modified internucleoside linkages can be used to alter, typically increase, the nuclease resistance of oligonucleotide compounds compared to natural phosphodiester linkages. In certain embodiments, linkages with chiral atoms can be prepared as racemic mixtures, as separate enantiomers. Representative chiral linkages include, but are not limited to, alkylphosphonates and phosphorothioates. Methods for preparing phosphorus-containing and non-phosphorus-containing linkages are well known to those skilled in the art.

[0404] The phosphate group in the internucleoside linkage can be modified by replacing one of the oxygens with a different substituent. One result of this modification can be increased resistance of the oligonucleotide to nucleolysis. Examples of modified phosphate groups include phosphorothioates, phosphoroselenates, boranophosphates, boranophosphate esters, hydrogen phosphonates, phosphoramidates, alkyl or aryl phosphonates, and phosphotriesters. In some embodiments, one of the non-bridging phosphate oxygen atoms in a phosphodiester internucleoside linkage can be replaced with any of the following: S, Se, BR3 (R is hydrogen, alkyl, aryl), C (i.e., alkyl group, aryl group, etc.), H, NR2 (R is hydrogen, optionally substituted alkyl, aryl), or OR (R is optionally substituted alkyl or aryl). The phosphorus atom in an unmodified phosphate group is achiral. However, replacing one of the non-bridging oxygens with one of the above atoms or groups of atoms makes the phosphorus atom chiral. In other words, the phosphorus atom in such modified phosphate groups is an asymmetric center. The asymmetric phosphorus atom can have either the "R" configuration (herein Rp) or the "S" configuration (herein Sp).

[0405] Phosphorodithioates have both non-bridging oxygens replaced by sulfur. The phosphorus center in phosphorodithioates is achiral, preventing the formation of oligonucleotide diastereomers. Therefore, without wishing to be bound by theory, modifications to both non-bridging oxygens that eliminate the chiral center, such as phosphorodithioate formation, may be desirable in that they cannot produce diastereomeric mixtures. The non-bridging oxygens can independently be any one of O, S, Se, B, C, H, N, or OR (R is alkyl or aryl).

[0406] Phosphodiester internucleoside linkages can also be modified by replacing the bridging oxygen (i.e., the oxygen that connects the phosphate to the sugar of the nucleoside) with nitrogen (bridging phosphoramidates), sulfur (bridging phosphorothioates), and carbon (bridging methylene phosphonates). Substitution can occur at either one or both of the linking oxygens. When the bridging oxygen is the 3'-oxygen of the nucleoside, substitution at carbon is preferred. When the bridging oxygen is the 5'-oxygen of the nucleoside, substitution at nitrogen is preferred.

[0407] Modified phosphate linkages in which at least one of the oxygens linked to the phosphate is replaced, or the phosphate group is replaced with a non-phosphate group, are also referred to as "non-phosphodiester intersugar linkages" or "non-phosphodiester linkers."

[0408] In certain embodiments, the phosphate group can be replaced by a non-phosphorus-containing connector, such as a dephosphoryl linker. Dephosphoryl linkers are also referred to herein as non-phosphodiester linkers. Without wishing to be bound by theory, it is believed that the charged phosphodiester group is the reactive center in nucleotide degradation, and therefore replacement with a neutral structural mimic improves the stability of the nuclease. Again, without wishing to be bound by theory, in some embodiments, it may be desirable to introduce a modification in which the charged phosphate group is replaced with a neutral moiety.

[0409] Examples of moieties that can replace the phosphate group include amide (e.g., amide-3 (3'-CH2-C(=O)-N(H)-5') and amide-4 (3'-CH2-N(H)-C(=O)-5')), hydroxylamino, siloxane (dialkylsiloxane), carboxamide, carbonate, carboxymethyl, carbamate, carboxylate ester, thioether, ethylene oxide linker, sulfide, sulfonate, sulfonamide, sulfonate ester, thioformacetal (3'-S-CH2-O-5'), formacetal (3'-O-CH2-O-5'), oxime, methyleneimino, methylenecarbonylamino, and the like. Nonionic linkages include, but are not limited to, methylenemethylimino (MMI, 3'-CH2-N(CH3)-O-5'), methylenehydrazo, methylenedimethylhydrazo, methyleneoxymethylimino, ether (C3'-O-C5'), thioether (C3'-S-C5'), thioacetamide (C3'-N(H)-C(=O)-CH2-S-C5', C3'-O-P(O)-O-S-S-C5', C3'-CH2-NH-NH-C5', 3'-NHP(O)(OCH3)-O-5', and 3'-NHP(O)(OCH3)-O-5', as well as nonionic linkages containing mixed N, O, S, and CH2 moieties. For example, Carbohydrate See Modifications in Antisense Research; YS Sanghvi and PDCook Eds. ACS Symposium Series 580; Chapters 3 and 4, (pp. 40-65). Preferred embodiments include methylenemethylimino (MMI), methylenecarbonylamino, amide, carbamate, and ethylene oxide linkers.

[0410] Those skilled in the art will recognize that in certain cases, substitution of a non-bridging oxygen may result in enhanced intersugar bond cleavage by the adjacent 2'-OH, and therefore, in many cases, modification of a non-bridging oxygen may require modification of the 2'-OH, e.g., a modification that does not involve cleavage of the adjacent intersugar bond, e.g., arabinose sugars, 2'-O-alkyl, 2'-F, LNA, and ENA.

[0411] Preferred non-phosphodiester internucleoside linkages include phosphorothioates, phosphorothioates having an enantiomeric excess of at least 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95% or more of the Sp isomer, phosphorothioates having an enantiomeric excess of at least 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95% or more of the Rp isomer, phosphorodithioates, phosphorodithioates, phosphotriesters, aminoalkylphosphotrioesters, alkylphosphonates (e.g., methyl-phosphonates), selenophosphates, phosphoramidates (e.g., N-alkylphosphoramidates), and boranophosphonates.

[0412] Further exemplary non-phosphorus-containing internucleoside linking groups are described in U.S. Pat. Nos. 5,034,506, 5,166,315, 5,185,444, 5,214,134, 5,216,141, 5,235,033, 5,264,562, 5,264,564, 5,405,938, 5,434,257, 5,466,677, 5,470,967, 5,489,677, 5,541,307, 5,561,2 25, 5,596,086, 5,602,240, 5,610,289, 5,602,240, 5,608,046, 5,610,289, 5,618,704, 5,623,070, 5,663,312, 5,633,360, 5,677,437, 5,792,608, 5,646,269, and 5,677,439, the contents of each of which are incorporated herein by reference.

[0413] In some embodiments of any one of the aspects, the oligonucleotides described herein include one or more neutral internucleoside linkages that are non-ionic. Suitable neutral internucleoside linkages include phosphotriester, methylphosphonate, MMI (3'-CH2-N(CH3)-O-5'), amide-3 (3'-CH2-C(=O)-N(H)-5'), amide-4 (3'-CH2-N(H)-C(=O)-5'), formacetal (3'-O-CH2-O-5'), and thioformacetal (3'-S-CH2-O-5'); siloxanes (dialkylsiloxanes), carboxylate esters, carboxamides, sulfides, sulfonate esters, and / or nonionic linkages including amides (e.g., Carbohydrate Modifications in Antisense Research; Y.S. Sanghvi and P.D. Cook Eds. ACS Symposium Series 580; Chapters 3 and 4, (pp. 40-65)); and non-ionic bonds containing mixed N, O, S, and CH2 moieties.

[0414] In one embodiment, the non-phosphodiester backbone linkages are selected from the group consisting of phosphorothioate, phosphorodithioate, alkyl-phosphonate, and phosphoramidate backbone linkages.

[0415] In some embodiments of any one of the aspects described herein, the internucleoside linkage is: TIFF2024525713000058.tif25128, where R IL1 and R IL2 are each independently for each occurrence absent, O, S, CH, NR (where R is hydrogen, alkyl, aryl), or optionally substituted alkylene, where the alkylene backbone may contain one or more of O, S, SS, and NR (where R is hydrogen, alkyl, aryl) internally and / or terminally; R IL3 and R IL4are each independently O, OR (R is hydrogen, alkyl, or aryl), S, Se, BR3 (R is hydrogen, alkyl, or aryl), BH3 - , C (i.e., alkyl, aryl, etc.), H, NR2 (where R is hydrogen, alkyl, aryl), alkyl, or aryl. R IL1 and R IL2 replaces the oxygen attached to the 5' carbon of the first nucleoside sugar, and R IL1 and R IL2 It is understood that the other of replaces the oxygen attached to the 3' (or 2') carbon of the second nucleoside sugar.

[0416] In some embodiments of any one of the aspects, R IL1 , R IL2 , R IL3 , and R IL4 are all O.

[0417] In some embodiments, R IL1 and R IL2 is O and R IL3 and R IL4 At least one of the groups is other than O. For example, R IL3 and R IL4 one of which is S and the other is O, or R IL3 and R IL4 Both are S.

[0418] In some embodiments of any one of the aspects described herein, R 3 or R 5 One of the structures is a modified internucleoside linkage, such as the following structure: TIFF2024525713000059.tif25128 to the internucleoside bond, where R IL1 , R IL2 , R IL3 , and R IL4 At least one of the is not O. For example, R IL3 and R IL4 At least one of is S.

[0419] In some embodiments of any one of the aspects described herein, R 3 and R 5 Both of these are bonds to modified internucleoside linkages.

[0420] In some embodiments of any one of the aspects described herein, R 3 is the bond to the phosphodiester internucleoside linkage.

[0421] In some embodiments of any one of the aspects described herein, R 5 is the bond to the phosphodiester internucleoside linkage.

[0422] In some embodiments of any one of the aspects described herein, R 3 is the bond to the modified internucleoside linkage, and R 5 is the bond to the phosphodiester internucleoside linkage.

[0423] In some embodiments of any one of the aspects described herein, R 5 is the bond to the modified internucleoside linkage, and R 3 is the bond to the phosphodiester internucleoside linkage.

[0424] In some embodiments of any one of the aspects, the oligonucleotide can comprise one or more, e.g., 1, 2, 3, 4, 5, 6, 7, 8, or more, modified internucleoside linkages. For example, the oligonucleotide can comprise 1, 2, 3, 4, 5, or 6 modified internucleoside linkages. For example, the oligonucleotide comprises 1, 2, 3, or 4 modified internucleoside linkages. In some embodiments, the oligonucleotide comprises at least two modified internucleoside linkages between the first five nucleotides counting from the 5' end of the oligonucleotide and further comprises at least two modified internucleoside linkages between the first five nucleotides counting from the 3' end of the oligonucleotide. For example, the oligonucleotide comprises modified internucleoside linkages between nucleotides 1 and 2 and between nucleotides 2 and 3 counting from the 5' end of the oligonucleotide, and between nucleotides 1 and 2 and between nucleotides 2 and 3 counting from the 3' end of the oligonucleotide.

[0425] In some embodiments of any one of the aspects, the modified internucleoside linkage is phosphorothioate. Thus, in some embodiments of any one of the aspects, the oligonucleotide comprises one or more, e.g., 1, 2, 3, 4, 5, 6, 7, 8, or more, phosphorothioate internucleoside linkages. For example, the oligonucleotide comprises 1, 2, 3, 4, 5, or 6 phosphorothioate internucleoside linkages. For example, the oligonucleotide comprises 1, 2, 3, or 4 phosphorothioate internucleoside linkages. In some embodiments, the oligonucleotide comprises at least two phosphorothioate internucleoside linkages among the first five nucleotides counting from the 5' end of the oligonucleotide and further comprises at least two phosphorothioate internucleoside linkages among the first five nucleotides counting from the 3' end of the oligonucleotide. For example, the oligonucleotide contains modified internucleoside linkages between nucleotides 1 and 2 and between nucleotides 2 and 3, counting from the 5' end of the oligonucleotide, and between nucleotides 1 and 2 and between nucleotides 2 and 3, counting from the 3' end of the oligonucleotide.

[0426] Oxygen Protecting Groups Some embodiments of the various aspects described herein include oxygen protecting groups (also referred to herein as hydroxyl protecting groups). Oxygen protecting groups include R OP1 , -N(R OP2 )2, -C(=O)SR OP1 , -C(=O)R OP1 , -CO2R OP1 , -C(=O)N(R OP2 )2, -C(=NR OP2 )R OP1 , -C(=NR OP2 ) OR OP1 , -C(=NR OP2 )N(R OP2 )2, -S(=O)R OP1 , -SO + 2nd Round OP1 , -Si(R OP1 )3, -P(R OP3 )2, -P(R OP3 ) + 3X - , -P(OR OP3 )2, -P(OR OP3 )3X - , -P(=O)(R OP1 )2, -P(=O)(OR OP3 )2, and -P(=O)(N(R OP2 )2)2, including, but not limited to, wherein each X - is a counterion, and each R OP1 independently, C 1-10 Alkyl, C 1-10 Perhaloalkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, Hetero C 1-10 Alkyl, Hetero C 2-10 Alkenyl, Hetero C 2-10 Alkynyl, C 3-10 Carbocyclyl, 3- to 14-membered heterocyclyl, C 6-14 aryl, or 5- to 14-membered heteroaryl, or two R OP1 groups combine to form a 3- to 14-membered heterocyclyl or 5- to 14-membered heteroaryl ring, and each R OP2is hydrogen, -OH, -OR OP1 , -N(R OP3 )2, -CN, -C(=O)R OP1 , -C(=O)N(R OP3 )2, -CO2R OP1 , -SO2R OP1 , -C(=NR OP3 ) OR OP1 , -C(=NR OP3 )N(R OP3 )2, -SO2N(R OP3 )2, -SO2R OP3 , -SO2OR OP3 , -SOR OP1 , -C(=S)N(R OP3 )2, -C(=O)SR OP3 , -C(=S)SR OP3 , -P(=O)(R OP1 )2, -P(=O)(OR OP3 )2, -P(=O)(N(R OP3 )2)2, C 1-10 Alkyl, C 1-10 Perhaloalkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, Hetero C 1-10 Alkyl, Hetero C 2-10 Alkenyl, Hetero C 2-10 Alkynyl, C 3-10 Carbocyclyl, 3- to 14-membered heterocyclyl, C 6-14 aryl, and 5- to 14-membered heteroaryl, or two R OP2 groups combine to form a 3- to 14-membered heterocyclyl or 5- to 14-membered heteroaryl ring, and each R OP3 are independently hydrogen, C 1-10 Alkyl, C 1-10 Perhaloalkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, Hetero C 1-10 Alkyl, Hetero C 2-10 Alkenyl, Hetero C 2-10 Alkynyl, C 3-10 Carbocyclyl, 3- to 14-membered heterocyclyl, C 6-14 aryl, and 5- to 14-membered heteroaryl, or two R OP3groups combine to form a 3- to 14-membered heterocyclyl or 5- to 14-membered heteroaryl ring; R OP1 , R OP2 , and R OP3 Each alkyl, alkenyl, alkynyl, carbocyclyl, heterocyclyl, aralkyl, aryl, and heteroaryl is OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl alkyl, O(C1-C8)alkyl (i.e., C1-C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p It can be optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from -NH2, or CH2-aryl-alkoxy, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6.

[0427] Oxygen protecting groups are well known in the art and are described in Greene's Protecting Groups in Organic Synthesis, PGM Wuts, 5, incorporated herein by reference. th Edition, John Wiley & Sons, 2014.

[0428] Exemplary oxygen protecting groups include, but are not limited to, methyl, t-butyloxycarbonyl (BOC or Boc), methoxylmethyl (MOM), methylthiomethyl (MTM), t-butylthiomethyl, (phenyldimethylsilyl)methoxymethyl (SMOM), benzyloxymethyl (BOM), p-methoxybenzyloxymethyl (PMBM), (4-methoxyphenoxy)methyl (p-AOM), guaiacolmethyl (GUM), t-butoxymethyl, 4-pentenyloxymethyl (PO M), siloxymethyl, 2-methoxyethoxymethyl (MEM), 2,2,2-trichloroethoxymethyl, bis(2-chloroethoxy)methyl, 2-(trimethylsilyl)ethoxymethyl (SEMOR), tetrahydropyranyl (THP), 3-bromotetrahydropyranyl, tetrahydrothiopyranyl, 1-methoxycyclohexyl, 4-methoxytetrahydropyranyl (MTHP), 4-methoxytetrahydrothiopyranyl, 4-methoxytetrahydrothiopyranyl S,S-dioxide, 1-[(2 -chloro-4-methyl)phenyl]-4-methoxypiperidin-4-yl (CTMP), 1,4-dioxan-2-yl, tetrahydrofuranyl, tetrahydrothiofuranyl, 2,3,3a,4,5,6,7,7a-octahydro-7,8,8-trimethyl-4,7-methanobenzofuran-2-yl, 1-ethoxyethyl, 1-(2-chloroethoxy)ethyl, 1-methyl-1-methoxyethyl, 1-methyl-1-benzyloxyethyl, 1-methyl-1-benzyloxy-2-fluoroethyl, 2,2 ,2-trichloroethyl, 2-trimethylsilylethyl, 2-(phenylselenyl)ethyl, t-butyl, allyl, p-chlorophenyl, p-methoxyphenyl, 2,4-dinitrophenyl, benzyl (Bn), p-methoxybenzyl, 3,4-dimethoxybenzyl, o-nitrobenzyl, p-nitrobenzyl, p-halobenzyl, 2,6-dichlorobenzyl, p-cyanobenzyl, p-phenylbenzyl, 2-picolyl, 4-picolyl, 3-methyl-2-picolyl N-oxide, diphenylmethyl, p,p'-Dinitrobenzhydryl, 5-dibenzosuberyl, triphenylmethyl, α-naphthyldiphenylmethyl, p-methoxyphenyldiphenylmethyl, di(p-methoxyphenyl)phenylmethyl, tri(p-methoxyphenyl)methyl, 4-(4'-bromophenacyloxyphenyl)diphenylmethyl, 4,4',4''-tris(4,5-dichlorophthalimidophenyl)methyl, 4,4',4''-tris(levulinoyloxyphenyl)methyl, 4,4',4''-tris(benzoyloxyphenyl)methyl (nyl)methyl, 3-(imidazol-1-yl)bis(4',4''-dimethoxyphenyl)methyl, 1,1-bis(4-methoxyphenyl)-1'-pyrenylmethyl, 9-anthryl, 9-(9-phenyl)xanthenyl, 9-(9-phenyl-10-oxo)anthryl, 1,3-benzodisulfuran-2-yl, benzisothiazolyl S,S-dioxide, trimethylsilyl (TMS), triethylsilyl (TES), triisopropylsilyl (TIPS), dimethylisopropylsilyl (IPDMS), diisopropylsilyl Ethyl isopropyl silyl (DEIPS), dimethylthexyl silyl, t-butyl dimethyl silyl (TBDMS), t-butyl diphenyl silyl (TBDPS), tribenzyl silyl, tri-p-xylyl silyl, triphenyl silyl, diphenyl methyl silyl (DPMS), t-butyl methoxyphenyl silyl (TBMPS), formate, acetate, chloroacetate, dichloroacetate, trichloroacetate, trifluoroacetate, methoxyacetate, triphenyl methoxyacetate, pheno oxyacetate, p-chlorophenoxyacetate, 3-phenylpropionate, 4-oxopentanoate (levulinate), 4,4-(ethylenedithio)pentanoate (levulinoyl dithioacetal), adamantoate, crotonate, 4-methoxycrotonate, benzoate, p-phenylbenzoate, 2,4,6-trimethylbenzoate (mesitoate), alkyl methyl carbonate, 9-fluorenylmethyl carbonate (Fmoc), alkyl ethyl carbonate, alkyl 2,2,2-Trichloroethyl carbonate (Troc), 2-(trimethylsilyl)ethyl carbonate (TMSEC), 2-(phenylsulfonyl)ethyl carbonate (Psec), 2-(triphenylphosphonio)ethyl carbonate (Peoc), alkyl isobutyl carbonate, alkyl vinyl carbonate, alkyl allyl carbonate, alkyl p-nitrophenyl carbonate, alkyl benzyl carbonate, alkyl p-methoxybenzyl carbonate, alkyl 3,4-dimethoxybenzyl carbonate, alkyl o-nitrobenzyl carbonate, alkyl p-nitrobenzyl carbonate, alkyl S-benzyl thiocarbonate, 4-ethoxy-1-naphthyl carbonate, methyl dithiocarbonate, 2-iodobenzoate, 4-azidobutyrate, 4-nitro-4-methylpentanoate, o-(dibromomethyl)benzoate, 2-formylbenzenesulfonate nate, 2-(methylthiomethoxy)ethyl, 4-(methylthiomethoxy)butyrate, 2-(methylthiomethoxymethyl)benzoate, 2,6-dichloro-4-methylphenoxyacetate, 2,6-dichloro-4-(1,1,3,3-tetramethylbutyl)phenoxyacetate, 2,4-bis(1,1-dimethylpropyl)phenoxyacetate, chlorodiphenylacetate, isobutyrate, monosodium benzoate SP3inoate), (E)-2-methyl-2-butenoate, o-(methoxyacyl)benzoate, α-naphthoate, nitrate, alkyl N,N,N′,N′-tetramethylphosphorodiamidate, alkyl N-phenylcarbamate, borate, dimethylphosphinothioyl, alkyl 2,4-dinitrophenylsulfonate, methanesulfonate (mesylate), and benzylsulfonate, and tosylate (Ts).

[0429] In some embodiments of any one of the aspects described herein, the oxygen protecting group is benzyl, benzoyl, 2,6-dichlorobenzyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, mesylate, tosylate, 4,4'-dimethoxytrityl (DMT), 9-phenylxanthin-9-yl (pixyl), and 9-(p-methoxyphenyl)xanthin-9-yl (MOX). In certain embodiments, the hydroxyl protecting group is selected from acetyl, benzyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, and dimethoxytrityl, with 4,4'-dimethoxytrityl being a more preferred hydroxyl protecting group.

[0430] As used herein, the terms "protected hydroxyl" and "protected hydroxy" refer to a group of the formula -OR Pro R Pro is an oxygen protecting group as defined herein.

[0431] Nitrogen Protecting Groups Some embodiments of the various aspects described herein include a nitrogen protecting group (also referred to herein as an amino protecting group). Nitrogen protecting groups include OH, -OR NP1 , -N(R NP2 )2, -C(=O)R NP1 , -C(=O)N(R NP2 )2, -CO2R NP1 , -SO2R NP1 , -C(=NR NP2 )R NP1 , -C(=NR NP2 ) OR NP1 , -C(=NR NP2 )N(R NP2 )2, -SO2N(R NP2 )2, -SO2R NP2 , -SO2OR NP2 , -SOR NP1 , -C(=S)N(R NP2 )2, -C(=O)SR NP2 , -C(=S)SR NP2 , C 1-10 Alkyl (e.g., aralkyl, heteroaralkyl), C 2-10 Alkenyl, C 2-10Alkynyl, C 3-10 Carbocyclyl, 3- to 14-membered heterocyclyl, C 6-14 aryl, and 5- to 14-membered heteroaryl groups, where each R NP1 independently, C 1-10 Alkyl, C 1-10 Perhaloalkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, Hetero C 1-10 Alkyl, Hetero C 2-10 Alkenyl, Hetero C 2-10 Alkynyl, C 3-10 Carbocyclyl, 3- to 14-membered heterocyclyl, C 6-14 aryl, or 5- to 14-membered heteroaryl, or two R NP1 groups combine to form a 3- to 14-membered heterocyclyl or 5- to 14-membered heteroaryl ring, and each R NP2 are independently hydrogen, C 1-10 Alkyl, C 1-10 Perhaloalkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, Hetero C 1-10 Alkyl, Hetero C 2-10 Alkenyl, Hetero C 2-10 Alkynyl, C 3-10 Carbocyclyl, 3- to 14-membered heterocyclyl, C 6-14 aryl, and 5- to 14-membered heteroaryl, or two R SP3 groups combine to form a 3- to 14-membered heterocyclyl or 5- to 14-membered heteroaryl ring; R NP1 and R NP2Each alkyl, alkenyl, alkynyl, carbocyclyl, heterocyclyl, aralkyl, aryl, and heteroaryl is OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl alkyl, O(C1-C8)alkyl (i.e., C1-C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) p It can be optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from -NH2, or CH2-aryl-alkoxy, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6.

[0432] Nitrogen protecting groups are well known in the art and are described in Greene's Protecting Groups in Organic Synthesis, PGM Wuts, 5, incorporated herein by reference. th Edition, John Wiley & Sons, 2014.

[0433] Exemplary amides (e.g., —C(═O)R NP1) Nitrogen protecting groups include, but are not limited to, formamide, acetamide, chloroacetamide, trichloroacetamide, trifluoroacetamide, phenylacetamide, 3-phenylpropanamide, picolinamide, 3-pyridylcarboxamide, N-benzoylphenylalanyl derivatives, benzamide, p-phenylbenzamide, o-nitrophenylacetamide, o-nitrophenoxyacetamide, acetoacetamide, (N'-dithiobenzyloxyacylamino)acetamide, 3-(p-hydroxyphenyl)propanamide, 3-(o-nitrophenyl)propanamide, 2-methyl-2-(o-nitrophenoxy)propanamide, 2-methyl-2-(o-phenylazophenoxy)propanamide, 4-chlorobutanamide, 3-methyl-3-nitrobutanamide, o-nitrocinnamide, N-acetylmethionine derivatives, o-nitrobenzamide, and o-(benzoyloxymethyl)benzamide.

[0434] Exemplary carbamates (e.g., —C(═O)OR NP1) Nitrogen protecting groups include, but are not limited to, methyl carbamate, ethyl carbamate, 9-fluorenylmethyl carbamate (Fmoc), 9-(2-sulfo)fluorenylmethyl carbamate, 9-(2,7-dibromo)fluoroenylmethyl carbamate, 2,7-di-t-butyl-[9-(10,10-dioxo-10,10,10,10-tetrahydrothioxanthyl)]methyl carbamate (DBD-Tmoc), 4-methoxyphenacyl carbamate (Phenoc), 2,2,2-trichloroethyl carbamate (Troc), ), 2-trimethylsilylethyl carbamate (Teoc), 2-phenylethyl carbamate (hZ), 1-(1-adamantyl)-1-methylethyl carbamate (Adpoc), 1,1-dimethyl-2-haloethyl carbamate, 1,1-dimethyl-2,2-dibromoethyl carbamate (DB-t-BOC), 1,1-dimethyl-2,2,2-trichloroethyl carbamate (TCBOC), 1-methyl-1-(4-biphenylyl)ethyl carbamate (Bpoc), 1-(3,5-di-t-butylphenyl)-1-methylethyl carbamate t-Bumeoc), 2-(2'- and 4'-pyridyl)ethyl carbamate (Pyoc), 2-(N,N-dicyclohexylcarboxamido)ethyl carbamate, t-butyl carbamate (BOC or Boc), 1-adomantyl carbamate (Adoc), vinyl carbamate (Voc), allyl carbamate (Alloc), 1-isopropylallyl carbamate (Ipaoc), cinnamyl carbamate (Coc), 4-nitrocinnamyl carbamate (Noc), 8-quinolyl carbamate, N-hydroxypiperidinyl carbamate , alkyldithiocarbamate, benzyl carbamate (Cbz), p-methoxybenzyl carbamate (Moz), p-nitrobenzyl carbamate, p-bromobenzyl carbamate, p-chlorobenzyl carbamate, 2,4-dichlorobenzyl carbamate, 4-methylsulfinylbenzyl carbamate (Msz), 9-anthrylmethyl carbamate, diphenylmethyl carbamate, 2-methylthioethyl carbamate, 2-methylsulfonylethyl carbamate, 2-(p-toluenesulfonyl)ethyl carbamate, [2-(1,3-dithianyl)]methyl carbamate (Dmoc), 4-methylthiophenylcarbamate (Mtpc), 2,4-dimethylthiophenylcarbamate (Bmpc), 2-phosphonioethyl carbamate (Peoc), 2-triphenylphosphonioisopropylcarbamate (Ppoc), 1,1-dimethyl-2-cyanoethyl carbamate, m-chloro-p-acyloxybenzyl carbamate, p-(dihydroxyboryl)benzyl carbamate, 5-benzisoxazolylmethyl carbamate, 2-(trifluoromethyl)-6-chloro Monylmethyl carbamate (Tcroc), m-nitrophenyl carbamate, 3,5-dimethoxybenzyl carbamate, o-nitrobenzyl carbamate, 3,4-dimethoxy-6-nitrobenzyl carbamate, phenyl(o-nitrophenyl)methyl carbamate, t-amyl carbamate, S-benzylthiocarbamate, p-cyanobenzyl carbamate, cyclobutyl carbamate, cyclohexyl carbamate, cyclopentyl carbamate, cyclopropylmethyl carbamate, p-decyloxybenzyl carbamate, 2 ,2-Dimethoxyacylvinylcarbamate, o-(N,N-dimethylcarboxamido)benzylcarbamate, 1,1-dimethyl-3-(N,N-dimethylcarboxamido)propylcarbamate, 1,1-dimethylpropynylcarbamate, di(2-pyridyl)methylcarbamate, 2-furanylmethylcarbamate, 2-iodoethylcarbamate, isoborylcarbamate, isobutylcarbamate, isonicotinecarbamate, p-(p'-methoxyphenylazo)benzylcarbamate, 1-methylcyclobutylcarbamate, 1 -methylcyclohexyl carbamate, 1-methyl-1-cyclopropylmethyl carbamate, 1-methyl-1-(3,5-dimethoxyphenyl)ethyl carbamate, 1-methyl-1-(p-phenylazophenyl)ethyl carbamate, 1-methyl-1-phenylethyl carbamate, 1-methyl-1-(4-pyridyl)ethyl carbamate, phenyl carbamate, p-(phenylazo)benzyl carbamate, 2,4,6-tri-t-butylphenyl carbamate, 4-(trimethylammonium)benzyl carbamate, and 2,4,6-trimethylbenzyl carbamate.

[0435] Exemplary sulfonamides (e.g., —S(═O)R NP1 Nitrogen protecting groups include, for example, p-toluenesulfonamide (Ts), benzenesulfonamide, 2,3,6-trimethyl-4-methoxybenzenesulfonamide (Mtr), 2,4,6-trimethoxybenzenesulfonamide (Mtb), 2,6-dimethyl-4-methoxybenzenesulfonamide (Pme), 2,3,5,6-tetramethyl-4-methoxybenzenesulfonamide (Mte), 4-methoxybenzenesulfonamide (Mbs), 2,4,6-trimethylbenzenesulfonamide (Mts). , 2,6-dimethoxy-4-methylbenzenesulfonamide (iMds), 2,2,5,7,8-pentamethylchroman-6-sulfonamide (Pmc), methanesulfonamide (Ms), β-trimethylsilylethanesulfonamide (SES), 9-anthracenesulfonamide, 4-(4′,8′-dimethoxynaphthylmethyl)benzenesulfonamide (DNMBS), benzylsulfonamide, trifluoromethylsulfonamide, and phenacylsulfonamide.

[0436] Additional exemplary nitrogen protecting groups include, but are not limited to, phenothiazinyl-(10)-acyl derivatives, N'-p-toluenesulfonylaminoacyl derivatives, N'-phenylaminothioacyl derivatives, N-benzoylphenylalanine derivatives, N-acetylmethionine derivatives, 4,5-diphenyl-3-oxazolin-2-one, N-phthalimide, N-dithiasulfinimide (Dts), N-2,3-diphenylmaleimide, N-2,5-dimethylpyrrole, N-1, 1,4,4-Tetramethyldisilylazacyclopentane adduct (STABASE), 5-substituted 1,3-dimethyl-1,3,5-triazacyclohexan-2-one, 5-substituted 1,3-dibenzyl-1,3,5-triazacyclohexan-2-one, 1-substituted 3,5-dinitro-4-pyridone, N-methylamine, N-allylamine, N-[2-(trimethylsilyl)ethoxy]methylamine (SEM), N-3-acetoxypropylamine, N-(1-isopropyl-4-nitro-2-oxo-3 -pyrroolin-3-yl)amine, quaternary ammonium salts, N-benzylamine, N-di(4-methoxyphenyl)methylamine, N-5-dibenzosuberylamine, N-triphenylmethylamine (Tr), N-[(4-methoxyphenyl)diphenylmethyl]amine (MMTr), N-9-phenylfluorenylamine (PhF), N-2,7-dichloro-9-fluorenylmethyleneamine, N-ferrocenylmethylamino (Fcm), N-2-picolylamino N'-oxide, N-1,1-di Methylthiomethyleneamine, N-benzylideneamine, Np-methoxybenzylideneamine, N-diphenylmethyleneamine, N-[(2-pyridyl)mesityl]methyleneamine, N-(N',N'-dimethylaminomethylene)amine, N,N'-isopropyllidenediamine, Np-nitrobenzylideneamine, N-salicylideneamine, N-5-chlorosalicylideneamine, N-(5-chloro-2-hydroxyphenyl)phenylmethyleneamine, N-cyclohexylideneamine, N-(5,5-dimethyl-3-oxo-1-cyclohexenyl)amine, N-borane and N-diphenylborinic acid derivatives, N-[phenyl(pentachloro- or tungsten)acyl]amine, N-copper chelate, N-zinc chelate, N-nitroamine, N-nitrosamine, amine N-oxide, diphenylphosphinamide (Dpp), dimethylthiophosphinamide (Mpt), diphenylthiophosphinamide (Ppt), dialkyl phosphoramidate, dibenzyl phosphoramidate, diphenyl phosphoramidate, benzenesulfenamide, o-nitrobenzenesulfenamide (Nps), 2,4-dinitrobenzenesulfenamide, pentachlorobenzenesulfenamide, 2-nitro-4-methoxybenzenesulfenamide, triphenylmethylsulfenamide, and 3-nitropyridine sulfenamide (Npys).

[0437] Sulfur Protecting Groups Some embodiments of the various aspects described herein include sulfur protecting groups (also referred to herein as thiol protecting groups). Sulfur protecting groups include -R SP1 , -N(R SP2 )2, -C(=O)SR SP1 , -C(=O)R SP1 , -CO2R SP1 , -C(=O)N(R SP2 )2, -C(=NR SP2 )R SP1 , -C(=NR SP2 ) OR SP1 , -C(=NR SP2 )N(R SP2 )2, -S(=O)R SP1 , -SO2R SP1 , -Si(R SP1 )3, -P(R SP3 )2, -P(R SP3 ) + 3X - , -P(OR SP3 )2, -P(OR SP3 ) + 3X - , -P(=O)(R SP1 )2, -P(=O)(OR SP3)2, and -P(=O)(N(R SP2 )2)2, including, but not limited to, wherein: X - is a counterion, and each R SP1 independently, C 1-10 Alkyl, C 1-10 Perhaloalkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, Hetero C 1-10 Alkyl, Hetero C 2-10 Alkenyl, Hetero C 2-10 Alkynyl, C 3-10 Carbocyclyl, 3- to 14-membered heterocyclyl, C 6-14 aryl, or 5- to 14-membered heteroaryl, or two R SP1 groups combine to form a 3- to 14-membered heterocyclyl or 5- to 14-membered heteroaryl ring, and each R SP2 is hydrogen, -OH, -OR SP1 , -N(R SP3 )2, -CN, -C(=O)R SP1 , -C(=O)N(R SP3 )2, -CO2R SP1 , -SO2R SP1 , -C(=NR SP3 ) OR SP1 , -C(=NR SP3 )N(R SP3 )2, -SO2N(R SP3 )2, -SO2R SP3 , -SO2OR SP3 , -SOR SP1 , -C(=S)N(R SP3 )2, -C(=O)SR SP3 , -C(=S)SR SP3 , -P(=O)(R SP1 )2, -P(=O)(OR SP3 )2, -P(=O)(N(R SP3 )2)2, C 1-10 Alkyl, C 1-10 Perhaloalkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, Hetero C 1-10 Alkyl, Hetero C 2-10 Alkenyl, Hetero C 2-10 Alkynyl, C3~10 Carbocyclyl, 3- to 14-membered heterocyclyl, C 6-14 aryl, and 5- to 14-membered heteroaryl, or two R SP2 groups combine to form a 3- to 14-membered heterocyclyl or 5- to 14-membered heteroaryl ring, and each R SP3 are independently hydrogen, C 1-10 Alkyl, C 1-10 Perhaloalkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, Hetero C 1-10 Alkyl, Hetero C 2-10 Alkenyl, Hetero C 2-10 Alkynyl, C 3-10 Carbocyclyl, 3- to 14-membered heterocyclyl, C 6-14 aryl, and 5- to 14-membered heteroaryl, or two R SP3 groups combine to form a 3- to 14-membered heterocyclyl or 5- to 14-membered heteroaryl ring; R SP1 , R SP2 , and R SP3 Each alkyl, alkenyl, alkynyl, carbocyclyl, heterocyclyl, aralkyl, aryl, and heteroaryl is OH, CN, SC(O)Ph, oxo(=O), SH, SO2NH2, SO2(C1-C4)alkyl, SO2NH(C1-C4)alkyl, halogen, carbonyl, thiol, cyano, NH2, NH(C1-C4)alkyl, N[(C1-C4)alkyl]2, C(O)NH2, COOH, COOMe, acetyl, (C1-C8)alkyl alkyl, O(C1-C8)alkyl (i.e., C1-C8 alkoxy), O(C1-C8)haloalkyl, (C2-C8)alkenyl, (C2-C8)alkynyl, haloalkyl, thioalkyl, cyanomethylene, alkylaminyl, aryl, heteroaryl, substituted aryl, NH2-C(O)-alkylene, NH(Me)-C(O)-alkylene, CH2-C(O)-alkyl, C(O)-alkyl, alkylcarbonylaminyl, CH2-[CH(OH)] m -(CH2) p -OH, CH2-[CH(OH)] m -(CH2) pIt can be optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from -NH2, or CH2-aryl-alkoxy, and "m" and "p" are independently 1, 2, 3, 4, 5, or 6.

[0438] Sulfur protecting groups are well known in the art and are described in Greene's Protecting Groups in Organic Synthesis, PGM Wuts, 5, incorporated herein by reference. th Edition, John Wiley & Sons, 2014.

[0439] It should be noted that the nucleoside of formula (I) can be located anywhere in the oligonucleotide. In some embodiments, the nucleoside of formula (I) is located at the 5'-end or 3'-end of the oligonucleotide. In some embodiments, the nucleoside of formula (I) is located at an internal position of the oligonucleotide.

[0440] In some embodiments of any one of the aspects described herein, the oligonucleotide further comprises a nucleoside having a modified sugar, i.e., in addition to a nucleotide of Formula (I). By "modified sugar" is meant a sugar or moiety that is not a 2'-deoxy (i.e., 2'-H) ribose sugar or a 2'-OH ribose sugar. Some exemplary nucleotides containing modified sugars are 2'-F ribose, 2'-OMe ribose, 2'-O,4'-C-methylene ribose (locked nucleic acid, LNA), anhydrohexitol (1,5-anhydrohexitol nucleic acid, HNA), cyclohexene (cyclohexene nucleic acid, CeNA), 2'-methoxyethyl ribose, 2'-O-allyl ribose, 2'-C-allyl ribose, 2'-ON-methylacetamido (2'-O-NMA) ribose, 2'-O-dimethylaminoethoxyethyl (2'-O-DMAEOE) ribose, 2'-O-aminopropyl (2'-O-AP) ribose, 2'-F arabinose (2'-ara-F), threose (threose nucleic acid, TNA), and 2,3-dihydroxypropyl (glycol nucleic acid, GNA). It should be noted that the nucleoside having a modified sugar can be present at any position in the oligonucleotide.

[0441] In some embodiments, the oligonucleotide further comprises at least one, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more 2'-fluoro (2'-F) nucleotides. For example, the oligonucleotide can comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 2'-F nucleotides. Note that the 2'-F nucleotides can be present at any position in the oligonucleotide.

[0442] In some embodiments, an oligonucleotide comprises a 2'-nucleoside and a 2'-F nucleoside of Formula (I), eg, alone.

[0443] In some embodiments, the oligonucleotide further comprises at least one, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more, 2'-OMe nucleotides. For example, the oligonucleotide can comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 2'-OMe nucleotides. Note that the 2'-OMe nucleotides can be present at any position in the oligonucleotide.

[0444] In some embodiments, the oligonucleotide comprises, e.g., alone, 2'-nucleosides and 2'-OMe nucleosides of Formula (I). In some other embodiments, the oligonucleotide comprises, e.g., alone, 2'-nucleosides, 2'-OMe nucleosides, and 2'-F nucleosides of Formula (I).

[0445] In some embodiments, the oligonucleotide further comprises at least one, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more 2'-deoxy, e.g., 2'-H nucleotides. For example, the oligonucleotide can comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 2'-deoxy, e.g., 2'-H nucleotides. Note that the 2'-deoxy, e.g., 2'-H nucleotides can be present at any position in the oligonucleotide. For example, the oligonucleotide can comprise 2'-deoxy, e.g., 2'-H nucleotides at 1, 2, 3, 4, 5, or 6 of the following positions, counting from the 5' end of the oligonucleotide: 2, 5, 7, 12, 14, and 16. In some embodiments, the oligonucleotide comprises 2'-deoxy nucleotides at positions 5 and 7, counting from the 5' end of the oligonucleotide.

[0446] In some embodiments, an oligonucleotide comprises a nucleoside of Formula (I) and a 2'-deoxy (2'-H) nucleotide, e.g., alone. In some embodiments, an oligonucleotide comprises a nucleoside of Formula (I), a 2'-OMe nucleoside, and a 2'-deoxy (2'-H) nucleotide, e.g., alone. In some embodiments, an oligonucleotide comprises a nucleoside of Formula (I), a 2'-F nucleoside, and a 2'-deoxy (2'-H) nucleotide, e.g., alone. In some embodiments, an oligonucleotide comprises a nucleoside of Formula (I), a 2'-OMe nucleoside, a 2'-F nucleoside, and a 2'-deoxy (2'-H) nucleotide, e.g., alone.

[0447] In some embodiments of any one of the aspects described herein, the oligonucleotide further comprises an unnatural nucleobase, i.e., in addition to the nucleotide of Formula (I). In some embodiments, the oligonucleotide can comprise one or more, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more, nucleotides comprising independently selected unnatural nucleobases. When present, nucleotides comprising unnatural nucleobases can be present anywhere in the oligonucleotide.

[0448] "Unnatural nucleobase" refers to a nucleobase that is not adenine, guanine, cytosine, uracil, or thymine. Exemplary unnatural nucleobases include inosine, xanthine, hypoxanthine, nubularine, isoguanisine, thurcidin, and substituted or modified analogs of adenine, guanine, cytosine, and uracil, such as 2-aminoadenine and other alkyl derivatives of adenine and guanine, 2-propyl and other alkyl derivatives of adenine and guanine, 5-halouracil and cytosine, 5-propynyluracil and cytosine, 6-azouracil ... Uracil, cytosine, and thymine, 5-uracil (pseudouracil), 4-thiouracil, 5-halouracil, 5-(2-aminopropyl)uracil, 5-aminoallyluracil, 8-halo, amino, thiol, thioalkyl, hydroxyl, and other 8-substituted adenines and guanines, 5-trifluoromethyl and other 5-substituted uracils and cytosines, 7-methylguanine, 5-substituted pyrimidines, 6-azapyrimidines, and N-2, N-6, and O-6 substituted purines include 2-aminopropyladenine, 5-propynyluracil and 5-propynylcytosine, dihydrouracil, 3-deaza-5-azacytosine, 2-aminopurine, 5-alkyluracil, 7-alkylguanine, 5-alkylcytosine, 7-deazaadenine, N6, N6-dimethyladenine, 2,6-diaminopurine, 5-amino-allyl-uracil, N3-methyluracil, Substituted 1,2,4-triazole, 2-pyridinone, 5-nitroindole, 3-nitropyrrole, 5-methoxyuracil, uracil-5-oxyacetic acid, 5-methoxycarbonylmethyluracil, 5-methyl-2-thiouracil, 5-methoxycarbonylmethyl-2-thiouracil, 5-methylaminomethyl-2-thiouracil, 3-(3-amino-3-carboxypropyl)uracil, 3-methylcytosine, 5-methylcytosine, N 4

[0033] Additional purines and pyrimidines include, but are not limited to, acetylcytosine, 2-thiocytosine, N6-methyladenine, N6-isopentenyladenine, 2-methylthio-N6-isopentenyladenine, N-methylguanine, or O-alkylated bases. Additional purines and pyrimidines include those disclosed in U.S. Pat. No. 3,687,808, those disclosed in the Concise Encyclopedia of Polymer Science and Engineering, pages 858-859, Kroschwitz, JI, ed. John Wiley & Sons, 1990, and those disclosed by Englisch et al., Angewandte Chemie, International Edition, 1991, 30, 613, the contents of all of which are incorporated herein by reference.

[0449] In some embodiments, the unnatural nucleobase is inosine, xanthine, hypoxanthine, nubularine, isoguanisine, thurcidin, 2-(halo)adenine, 2-(alkyl)adenine, 2-(propyl)adenine, 2-(amino)adenine, 2-(aminoalkyl)adenine, 2-(aminopropyl)adenine, 2-(methylthio)-N 6 -(Isopentenyl)adenine, 7-(Deaza)adenine, 8-(Alkenyl)adenine, 8-(Alkyl)adenine, 8-(Alkynyl)adenine, 8-(Amino)adenine, 8-(Halo)adenine, 8-(Hydroxyl)adenine, 8-(Thioalkyl)adenine, 8-(Thiol)adenine, N 6 -(Isopentyl)adenine, N 6 -(methyl)adenine, N 6 , N 6-(Dimethyl)adenine, 2-(alkyl)guanine, 2-(propyl)guanine, 6-(alkyl)guanine, 6-(methyl)guanine, 7-(alkyl)guanine, 7-(methyl)guanine, 7-(deaza)guanine, 8-(alkyl)guanine, 8-(alkenyl)guanine, 8-(alkynyl)guanine, 8-(amino)guanine, 8-(halo)guanine, 8-(hydroxyl)guanine, 8-(thioalkyl)guanine cytosine, 8-(thiol)guanine, N-(methyl)guanine, 2-(thio)cytosine, 3-(deaza)-5-(aza)cytosine, 3-(alkyl)cytosine, 3-(methyl)cytosine, 5-(alkyl)cytosine, 5-(alkynyl)cytosine, 5-(halo)cytosine, 5-(methyl)cytosine, 5-(propynyl)cytosine, 5-(trifluoromethyl)cytosine, 6-(azo)cytosine, N 4 -(Acetyl)cytosine, 3-(3-amino-3-carboxypropyl)uracil, 2-(thio)uracil, 5-(methyl)-2-(thio)uracil, 5-(methylaminomethyl)-2-(thio)uracil, 4-(thio)uracil, 5-(methyl)-4-(thio)uracil, 5-(methylaminomethyl)-4-(thio)uracil, 5-(methyl)-2,4-(dithio)uracil, 5-(methylaminomethyl)-2,4-(dithio)uracil, 5-(2-aminopropyl)uracil, 5-(alkyl)uracil, 5-(alkynyl)uracil, 5-(allylamino)uracil, 5-(aminoallyl)uracil , 5-(aminoalkyl)uracil, 5-(guanidiniumalkyl)uracil, 5-(1,3-diazole-1-alkyl)uracil, 5-(cyanoalkyl)uracil, 5-(dialkylaminoalkyl)uracil, 5-(dimethylaminoalkyl)uracil, 5-(halo)uracil, 5-(methoxy)uracil, uracil-5-oxyacetic acid, 5-(methoxycarbonylmethyl)-2-(thio)uracil, 5-(methoxycarbonylmethyl)uracil, 5-(propynyl)uracil, 5-(propynyl)uracil, 5-(trifluoromethyl)uracil, 6-(azo)uracil, dihydrouracil, N 3-(methyl)uracil, 5-uracil (i.e., pseudouracil), 2-(thio)pseudouracil, 4-(thio)pseudouracil, 2,4-(dithio)pseudouracil, 5-(alkyl)pseudouracil, 5-(methyl)pseudouracil, 5-(alkyl)-2-(thio)pseudouracil, 5-(methyl)-2-(thio)pseudouracil, 5-(alkyl)-4-(thio)pseudouracil, 5-(methyl)-4-(thio)pseudouracil, 5-(alkyl)-2,4-(dithio)pseudouracil pseudouracil, 5-(methyl)-2,4-(dithio)pseudouracil, 1-substituted pseudouracil, 1-substituted 2(thio)-pseudouracil, 1-substituted 4-(thio)pseudouracil, 1-substituted 2,4-(dithio)pseudouracil, 1-(aminocarbonylethylenyl)-pseudouracil, 1-(aminocarbonylethylenyl)-2(thio)-pseudouracil, 1-(aminocarbonylethylenyl)-4-(thio)pseudouracil, 1-(aminocarbonylethylenyl)-2,4-(dithio)pseudouracil Uracil, 1-(aminoalkylaminocarbonylethylenyl)-pseudouracil, 1-(aminoalkylamino-carbonylethylenyl)-2(thio)-pseudouracil, 1-(aminoalkylaminocarbonylethylenyl)-4-(thio)pseudouracil, 1-(aminoalkylaminocarbonylethylenyl)-2,4-(dithio)pseudouracil, 1,3-(diaza)-2-(oxo)-phenoxazin-1-yl, 1-(aza)-2-(thio)-3-(aza)-phenoxazin-1-yl, 1 ,3-(diaza)-2-(oxo)-phenthiazin-1-yl, 1-(aza)-2-(thio)-3-(aza)-phenthiazin-1-yl, 7-substituted 1,3-(diaza)-2-(oxo)-phenoxazin-1-yl, 7-substituted 1-(aza)-2-(thio)-3-(aza)-phenoxazin-1-yl, 7-substituted 1,3-(diaza)-2-(oxo)-phenthiazin-1-yl, 7-substituted 1-(aza)-2-(thio)-3-(aza)-phenoxazin-1-yl, 7-(aminoalkylhydroxy)-1,3-(diaza)-2-(oxo)-phenoxazin-1-yl, 7-(aminoalkylhydroxy)-1-(aza)-2-(thio)-3-(aza)-phenoxazin-1-yl, 7-(aminoalkylhydroxy)-1,3-(diaza)-2-(oxo)-phenthiazin-1-yl, 7-(aminoalkylhydroxy)-1-(aza)-2-(thio)-3-(aza)-phenthiazin-1-yl, 7-(guanidiniumalkylhydroxy)-1,3-(diaza)-2-(oxo)-phenoxazin-1-yl, 7-(guanidiniumalkylhydroxy)-1,3-(diaza)-2-(oxo)-phenoxazin-1-yl (hydroxy)-1-(aza)-2-(thio)-3-(aza)-phenoxazin-1-yl, 7-(guanidiniumalkyl-hydroxy)-1,3-(diaza)-2-(oxo)-phenthiazin-1-yl, 7-(guanidiniumalkyl-hydroxy)-1-(aza)-2-(thio)-3-(aza)-phenthiazin-1-yl, 1,3,5-(triaza)-2,6-(dioxa-naphthalene, inosine, xanthine, hypoxanthine, nubularine, thurcidin, isoguanisine, inosinyl, 2-aza-inosinyl, 7-deaza-inosinyl Nyl, nitroimidazolyl, nitropyrazolyl, nitrobenzimidazolyl, nitroindazolyl, aminoindolyl, pyrrolopyrimidinyl, 3-(methyl)isocarbostyrilyl, 5-(methyl)isocarbostyrilyl, 3-(methyl)-7-(propynyl)isocarbostyrilyl, 7-(aza)indolyl, 6-(methyl)-7-(aza)indolyl, imidizopyridinyl, 9-(methyl)-imidizopyridinyl, pyrrolopyridinyl, isocarbostyrilyl, 7-(propynyl)isocarbostyrilyl, propynyl-7-(aza)indolyl phenyl, 2,4,5-(trimethyl)phenyl, 4-(methyl)indolyl, 4,6-(dimethyl)indolyl, phenyl, naphthalenyl, anthracenyl, phenanthracenyl, pyrenyl, stilbenyl, tetracenyl, pentacenyl, difluorotolyl, 4-(fluoro)-6-(methyl)benzimidazole, 4-(methyl)benzimidazole, 6-(azo)thymine, 2-pyridinone, 5-nitroindole, 3-nitropyrrole, 6-(aza)pyrimidine, 2-(amino)purine, 2,6-(diamino)purine, 5-substituted pyrimidines, N, 2 -substituted purines, N6 -substituted purines, O 6 -substituted purines, substituted 1,2,4-triazoles, and any O- or N-alkylated derivatives thereof.

[0450] In some embodiments, the non-natural nucleobase is a modified nucleobase, i.e., the nucleobase comprises a nucleobase modification described herein, e.g., the nucleobase is a substituted or modified analog of any of the naturally occurring nucleobases. Examples of nucleobase modifications include C-5 pyrimidines with alkyl or aminoalkyl groups and other cationic groups such as guanidinium and amidine functional groups, alkyl or aminoalkyl groups, and N-type nucleobases with other cationic groups such as purines, G-clamps, guanidinium G-clamps, and guanidinium and amidine functional groups of pseudouridines, as known in the art. 2 - and N 6 -Includes, but is not limited to:

[0451] In some embodiments of any one of the aspects, the unnatural nucleobase is a universal nucleobase. As used herein, a universal nucleobase is any modified or unmodified natural or unnatural nucleobase that can base pair with all of adenine, cytosine, guanine, and uracil without substantially affecting the melting behavior, recognition by intracellular enzymes, or activity of an oligonucleotide comprising the universal nucleobase. Some exemplary universal nucleobases include, but are not limited to, 2,4-difluorotoluene, nitropyrrolyl, nitroindolyl, 8-aza-7-deazaadenine, 4-fluoro-6-methylbenzimidazole, 4-methylbenzimidazole, 3-methylisocarbostyrilyl, 5-methylisocarbostyrilyl, 3-methyl-7-propynylisocarbostyrilyl, 7-azaindolyl, 6-methyl-7-azaindolyl, imidizopyridinyl, 9-methyl-imidizopyridinyl, pyrrolopyridinyl, isocarbostyrilyl, 7-propynylisocarbostyrilyl, propynyl-7-azaindolyl, 2,4,5-trimethylphenyl, 4-methylinolyl, 4,6-dimethylindolyl, phenyl, napthalenyl, anthracenyl, phenanthracenyl, pyrenyl, stilbenyl, tetracenyl, pentacenyl, and structural derivatives thereof.

[0452] In some embodiments of any one of the aspects described herein, the non-mature nucleobase is a protected nucleobase. As used herein, "protected nucleobase" refers to a nucleobase that includes a nitrogen-protecting group, an oxygen-protecting group, and / or a sulfur-protecting group.

[0453] In some embodiments of any one of the aspects described herein, the unnatural nucleobase is a modified, protected, or substituted analog of a nucleobase selected from adenine, cytosine, guanine, thymine, and uracil.

[0454] In some embodiments, the oligonucleotide further comprises a solid support attached thereto.

[0455] The oligonucleotides described herein can range in length from a few nucleotides (e.g., 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides) to hundreds of nucleotides in length. For example, oligonucleotides can be 5 nucleotides to 100 nucleotides in length. In some embodiments, oligonucleotides are 10 nucleotides to 50 nucleotides in length. For example, oligonucleotides are 15 to 35, more commonly 18 to 25, even more commonly 19 to 24, and most commonly 19 to 21 base pairs in length. In some embodiments, longer oligonucleotides, 25 to 30 nucleotides in length, are preferred. In some embodiments, shorter oligonucleotides, 10 to 15 nucleotides in length, are preferred. In another embodiment, the oligonucleotide is at least 21 nucleotides in length.

[0456] In some embodiments, the oligonucleotides described herein comprise a pattern of backbone chiral centers. In some embodiments, the regular pattern of backbone chiral centers comprises at least five internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least six internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least seven internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least eight internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least nine internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least ten internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least 11 internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least 12 internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least 13 internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least 14 internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least 15 internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least 16 internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least 17 internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least 18 internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises at least 19 internucleotide linkages in the Sp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises no more than 8 internucleotide linkages in the Rp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises no more than 7 internucleotide linkages in the Rp configuration.In some embodiments, the regular pattern of backbone chiral centers comprises six or fewer internucleotide linkages in the Rp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises five or fewer internucleotide linkages in the Rp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises four or fewer internucleotide linkages in the Rp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises three or fewer internucleotide linkages in the Rp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises two or fewer internucleotide linkages in the Rp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises one or fewer internucleotide linkages in the Rp configuration. In some embodiments, the regular pattern of backbone chiral centers comprises eight or fewer non-chiral internucleotide linkages (phosphodiesters as a non-limiting example). In some embodiments, the regular pattern of backbone chiral centers comprises seven or fewer non-chiral internucleotide linkages. In some embodiments, the regular pattern of backbone chiral centers comprises six or fewer non-chiral internucleotide linkages. In some embodiments, the regular pattern of backbone chiral centers comprises five or fewer non-chiral internucleotide linkages. In some embodiments, the regular pattern of backbone chiral centers comprises four or fewer non-chiral internucleotide linkages. In some embodiments, the regular pattern of backbone chiral centers comprises three or fewer non-chiral internucleotide linkages. In some embodiments, the regular pattern of backbone chiral centers comprises two or fewer non-chiral internucleotide linkages. In some embodiments, the regular pattern of backbone chiral centers comprises one or fewer non-chiral internucleotide linkages. In some embodiments, the regular pattern of backbone chiral centers comprises at least 10 internucleotide linkages in the Sp configuration and eight or fewer non-chiral internucleotide linkages. In some embodiments, the regular pattern of backbone chiral centers comprises at least 11 internucleotide linkages in the Sp configuration and seven or fewer non-chiral internucleotide linkages. In some embodiments, the regular pattern of backbone chiral centers comprises at least 12 internucleotide linkages in the Sp configuration and six or fewer non-chiral internucleotide linkages.In some embodiments, the regular pattern of backbone chiral centers comprises at least 13 internucleotide linkages in the Sp configuration and no more than 6 internucleotide linkages that are not chiral. In some embodiments, the regular pattern of backbone chiral centers comprises at least 14 internucleotide linkages in the Sp configuration and no more than 5 internucleotide linkages that are not chiral. In some embodiments, the regular pattern of backbone chiral centers comprises at least 15 internucleotide linkages in the Sp configuration and no more than 4 internucleotide linkages that are not chiral. In some embodiments, the internucleotide linkages in the Sp configuration may optionally be contiguous or non-contiguous. In some embodiments, the internucleotide linkages in the Rp configuration may optionally be contiguous or non-contiguous. In some embodiments, the non-chiral internucleotide linkages may optionally be contiguous or non-contiguous.

[0457] In some embodiments, the oligonucleotides described herein comprise stereochemical blocks. In some embodiments, the blocks are Rp blocks, in that each internucleotide linkage of the block is Rp. In some embodiments, the 5'-block is an Rp block. In some embodiments, the 3'-block is an Rp block. In some embodiments, the blocks are Sp blocks, in that each internucleotide linkage of the block is Sp. In some embodiments, the 5'-block is an Sp block. In some embodiments, the 3'-block is an Sp block. In some embodiments, provided oligonucleotides comprise both Rp and Sp blocks. In some embodiments, provided oligonucleotides comprise one or more Rp blocks but no Sp blocks. In some embodiments, provided oligonucleotides comprise one or more Sp blocks but no Rp blocks. In some embodiments, provided oligonucleotides comprise one or more PO blocks, in which each internucleotide linkage is a natural phosphate linkage.

[0458] In some embodiments, the oligonucleotides described herein comprise a 5'-block that is an Sp block, in which each sugar moiety comprises a 2'-fluoro modification. In some embodiments, the 5'-block is an Sp block, in which each internucleotide linkage is a modified internucleotide linkage and each sugar moiety comprises a 2'-fluoro modification. In some embodiments, the 5'-block is an Sp block, in which each internucleoside linkage is a phosphorothioate linkage and each sugar moiety comprises a 2'-fluoro modification. In some embodiments, the 5'-block comprises four or more nucleoside units. In some embodiments, the 5'-block comprises five or more nucleoside units. In some embodiments, the 5'-block comprises six or more nucleoside units. In some embodiments, the 5'-block comprises seven or more nucleoside units. In some embodiments, the 3'-block is an Sp block, in which each sugar moiety comprises a 2'-fluoro modification. In some embodiments, the 3'-block is an Sp block, in which each internucleotide linkage is a modified internucleotide linkage and each sugar moiety comprises a 2'-fluoro modification. In some embodiments, the 3'-block is an Sp block, in which each of the internucleotide linkages is a phosphorothioate linkage and each sugar moiety comprises a 2'-fluoro modification. In some embodiments, the 3'-block comprises 4 or more nucleoside units. In some embodiments, the 3'-block comprises 5 or more nucleoside units. In some embodiments, the 3'-block comprises 6 or more nucleoside units. In some embodiments, the 3'-block comprises 7 or more nucleoside units.

[0459] In some embodiments, the oligonucleotides described herein comprise certain types of nucleosides in certain regions, or are followed by a particular type of internucleotide linkage, e.g., a natural phosphate linkage, a modified internucleotide linkage, an Rp chiral internucleotide linkage, an Sp chiral internucleotide linkage, etc. In some embodiments, A is followed by Sp. In some embodiments, A is followed by Rp. In some embodiments, A is followed by a natural phosphate linkage (PO). In some embodiments, U is followed by Sp. In some embodiments, U is followed by Rp. In some embodiments, U is followed by a natural phosphate linkage (PO). In some embodiments, C is followed by Sp. In some embodiments, C is followed by Rp. In some embodiments, C is followed by a natural phosphate linkage (PO). In some embodiments, G is followed by Sp. In some embodiments, G is followed by Rp. In some embodiments, G is followed by a natural phosphate linkage (PO). In some embodiments, C and U are followed by Sp. In some embodiments, C and U are followed by Rp. In some embodiments, C and U are followed by a natural phosphate bond (PO). In some embodiments, A and G are followed by Sp. In some embodiments, A and G are followed by Rp.

[0460] In some embodiments of any one of the aspects described herein, the oligonucleotides described herein are 5' phosphorylated or comprise a phosphoryl analog at the 5' prime end. 5'-phosphate modifications include modifications that are compatible with RISC-mediated gene silencing. Suitable modifications include 5'-monophosphate ((HO)2(O)PO-5'), 5'-diphosphate ((HO)2(O)POP(HO)(O)-O-5'), 5'-triphosphate ((HO)2(O)PO-(HO)(O)POP(HO)(O)-O-5'), 5'-guanosine cap (7-methylated or unmethylated) (7m-GO-5'-(HO)(O)PO-(HO)(O)POP(HO)(O)-O-5'). '), 5'-adenosine cap (Appp), and any modified or unmodified nucleotide cap structure (NO-5'-(HO)(O)PO-(HO)(O)POP(HO)(O)-O-5'), 5'-monothiophosphate (phosphorothioate, (HO)2(S)PO-5'), 5'-monodithiophosphate (phosphorodithioate, (HO)(HS)(S)PO-5'), 5'-phosphorothiolate ((HO )2(O)PS-5'), any additional combination of oxygen / sulfur substituted monophosphates, diphosphates, and triphosphates (e.g., 5'-alpha-thiotriphosphate, 5'-gamma-thiotriphosphate, etc.), 5'-phosphoramidates ((HO)2(O)P-NH-5', (HO)(NH2)(O)PO-5'), 5'-alkyl phosphonates (e.g., R-P(OH)(O)-O-5'-, R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), 5'-alkenyl phosphonates (i.e., vinyl, substituted vinyl, e.g., OH)2(O)P-5'-CH= or (OH)2(O)P-5'-CH2-), 5'-alkyl ether phosphonates (e.g., R(OH)(O)PO-5', R = alkyl ether, e.g., methoxymethyl (MeOCH2-), ethoxymethyl, etc.). Other exemplary 5'-modifications include when Z is alkyl which may be substituted at least once, for example, ((HO)2(X)PO[-(CH2) a -OP(X)(OH)-O] b-5', ((HO)2(X)PO[-(CH2) a -P(X)(OH)-O] b -5', ((HO)2(X)P-[-(CH2) a -OP(X)(OH)-O] b -5', Dialkyl Terminal Phosphates and Phosphate Mimics: HO[-(CH2) a -OP(X)(OH)-O] b -5', H2N[-(CH2) a -OP(X)(OH)-O] b -5', H[-(CH2) a -OP(X)(OH)-O] b -5', Me2N[-(CH2) a -OP(X)(OH)-O] b -5', HO[-(CH2) a -P(X)(OH)-O] b -5', H2N[-(CH2) a -P(X)(OH)-O] b -5', H[-(CH2) a -P(X)(OH)-O] b -5', Me2N[-(CH2) a -P(X)(OH)-O] b -5', where a and b are each independently 1 to 10. Other embodiments include BH3, BH3 - and / or Se substitution.

[0461] In some embodiments of any one of the aspects described herein, the oligonucleotide comprises a 5'-vinyl phosphonate group. For example, the oligonucleotide comprises a 5'-E-vinyl phosphonate group. In some other non-limiting examples, the oligonucleotide comprises a 5'-Z-vinyl phosphonate group.

[0462] In some embodiments of any one of the aspects, the oligonucleotides described herein comprise a 5'-morpholino, 5'-dimethylamino, 5'-deoxy, reverse abasic, or reverse abasic locked nucleic acid modification at the 5' end.

[0463] In some embodiments of any one of the aspects, the oligonucleotides described herein may comprise a thermally destabilizing modification. For example, the oligonucleotide may comprise at least one thermally destabilizing modification of the duplex within the first nine nucleotide positions, counting from the 5' end of the oligonucleotide. In some embodiments, the thermally destabilizing modification is located at positions 2, 3, 4, 5, 6, 7, 8, or 9, counting from the 5' end of the antisense strand. In some embodiments, the thermally destabilizing modification is located at positions 2-9, or preferably 4-8, counting from the 5' end of the oligonucleotide. In some further embodiments, the thermally destabilizing modification is located at positions 5, 6, 7, or 8, counting from the 5' end of the oligonucleotide. In some further embodiments, the thermally destabilizing modification is located at position 7, counting from the 5' end of the oligonucleotide.

[0464] The term "thermally destabilizing modification" includes modifications that result in a dsRNA having a lower overall melting temperature (Tm), preferably 1, 2, 3, or 4 degrees lower than the melting temperature (Tm) of a dsRNA not having such a modification. In some embodiments, the thermally destabilizing modification is located at position 2, 3, 4, 5, 6, 7, 8, or 9, counting from the 5' end of the antisense strand.

[0465] Thermally destabilizing modifications can include, but are not limited to, abasic modifications, mismatches with the opposing nucleotide in the opposing strand, and sugar modifications, such as 2'-deoxy modifications or acyclic nucleotides, such as unlocked nucleic acids (UNAs) or glycol nucleic acids (GNAs). For example, thermally destabilizing modifications can include, but are not limited to, mUNA and GNA components such as: TIFF2024525713000060.tif106162TIFF2024525713000061.tif197158TIFF2024525713000062.tif96164

[0466] In some embodiments, the destabilizing modification is selected from the group consisting of GNA-isoC, GNA-isoG, 5'-mUNA, 4'-mUNA, 3'-mUNA, and 2'-mUNA.

[0467] In some embodiments, the destabilizing modified mUNA is selected from the group consisting of: TIFF2024525713000063.tif87152R = H, OH, OMe, Cl, F, OH, O-(CH2)2OMe, SMe, NMe2, NH2, Me, CCH(alkyne), O-nPr, O-alkyl, O-alkylamino, R' = H, Me, B=A, C, 5-Me-C, G, I, U, T, Y, 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, C2-modified purine, N8-modified purine, phenoxazine, G-clamp, non-standard mono-, bi-, and tricyclic heterocycle, pseudouracil, isoC, isoG, 2,6-diamninopurine, pseudocytosine, 2-aminopurine, xanthosine, N6-alkyl-A, O6-alkyl-G, 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, C2-modified purine, N8-modified purine, 7-deazapurine, phenoxazine, G-clamp, non-standard mono-, bi-, and tricyclic heterocycle. The stereochemistry is R or S, with a combination of R and S for any unspecified chiral center.

[0468] In some embodiments, the destabilizing modified mUNA is selected from the group consisting of: TIFF2024525713000064.tif52147R = H, OH, OMe, Cl, F, OH, O-(CH2)2OMe, SMe, NMe2, NH2, Me, CCH(alkyne), O-nPr, O-alkyl, O-alkylamino, R' = H, Me, B=A, C, 5-Me-C, G, I, U, T, Y, 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, C2-modified purine, N8-modified purine, phenoxazine, G-clamp, non-standard mono-, bi-, and tricyclic heterocycle, pseudouracil, isoC, isoG, 2,6-diamninopurine, pseudocytosine, 2-aminopurine, xanthosine, N6-alkyl-A, O6-alkyl-G, 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, C2-modified purine, N8-modified purine, 7-deazapurine, phenoxazine, G-clamp, non-standard mono-, bi-, and tricyclic heterocycle. The stereochemistry is R or S, with a combination of R and S for any unspecified chiral center.

[0469] In some embodiments, the destabilizing modified mUNA is selected from the group consisting of: TIFF2024525713000065.tif85157R = H, OMe, F, OH, O-(CH2)2OMe, SMe, NMe2, NH2, Me, O-nPr, O-alkyl, O-alkylamino, R' = H, Me, B = A, C, 5-Me-C, G, I, U, T, Y, 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, C2-modified purine, N8-modified purine, phenoxazine, G-clamp, non-standard mono-, bi-, and tricyclic heterocycle, pseudouracil, isoC, isoG, 2,6-diamninopurine, pseudocytosine, 2-aminopurine, xanthosine, N6-alkyl-A, O6-alkyl-G, 7-deazapurine, The stereochemistry is R or S, with a combination of R and S for any unspecified chiral center.

[0470] In some embodiments, the destabilizing modified mUNA is selected from the group consisting of: TIFF2024525713000066.tif86151R = H, OH, OMe, Cl, F, OH, O-(CH2)2OMe, SMe, NMe2, NH2, Me, CCH(alkyne), O-nPr, O-alkyl, O-alkylamino, R' = H, Me, B=A, C, 5-Me-C, G, I, U, T, Y, 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, C2-modified purine, N8-modified purine, phenoxazine, G-clamp, non-standard mono-, bi-, and tricyclic heterocycle, pseudouracil, isoC, isoG, 2,6-diamninopurine, pseudocytosine, 2-aminopurine, xanthosine, N6-alkyl-A, O6-alkyl-G, 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, C2-modified purine, N8-modified purine, 7-deazapurine, phenoxazine, G-clamp, non-standard mono-, bi-, and tricyclic heterocycle. The stereochemistry is R or S, with a combination of R and S for any unspecified chiral center.

[0471] In some embodiments, the destabilizing modified mUNA is selected from the group consisting of: TIFF2024525713000067.tif51147R = H, OH, OMe, Cl, F, OH, O-(CH2)2OMe, SMe, NMe2, NH2, Me, CCH(alkyne), O-nPr, O-alkyl, O-alkylamino, R' = H, Me, B=A, C, 5-Me-C, G, I, U, T, Y, 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, C2-modified purine, N8-modified purine, phenoxazine, G-clamp, non-standard mono-, bi-, and tricyclic heterocycle, pseudouracil, isoC, isoG, 2,6-diamninopurine, pseudocytosine, 2-aminopurine, xanthosine, N6-alkyl-A, O6-alkyl-G, 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, C2-modified purine, N8-modified purine, 7-deazapurine, phenoxazine, G-clamp, non-standard mono-, bi-, and tricyclic heterocycle. The stereochemistry is R or S, with a combination of R and S for any unspecified chiral center.

[0472] In some embodiments, the modified mUNA is selected from the group consisting of: TIFF2024525713000068.tif84156R = H, OMe, F, OH, O-(CH2)2OMe, SMe, NMe2, NH2, Me, O-nPr, O-alkyl, O-alkylamino, R' = H, Me, B = A, C, 5-Me-C, G, I, U, T, Y, 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, C2-modified purine, N8-modified purine, phenoxazine, G-clamp, non-standard mono-, bi-, and tricyclic heterocycle, pseudouracil, isoC, isoG, 2,6-diamninopurine, pseudocytosine, 2-aminopurine, xanthosine, N6-alkyl-A, O6-alkyl-G, 7-deazapurine, The stereochemistry is R or S, with a combination of R and S for any unspecified chiral center.

[0473] Exemplary abasic modifications include, but are not limited to, the following: TIFF2024525713000069.tif60128In the formula, R=H, Me, Et or OMe, R'=H, Me, Et or OMe, R"=H, Me, Et or OMe. TIFF2024525713000070.tif45134 where B is a modified or unmodified nucleobase and the asterisk on each structure represents either R, S or racemic.

[0474] Exemplary sugar modifications include, but are not limited to, the following: TIFF2024525713000071.tif81132 where B is a modified or unmodified nucleobase and the asterisk on each structure represents either R, S or racemic.

[0475] In some embodiments, the thermally destabilizing modification of the duplex is selected from the mUNA and GNA components described herein in Examples 1-3. In some embodiments, the destabilizing modification is selected from the group consisting of GNA-isoC, GNA-isoG, 5'-mUNA, 4'-mUNA, 3'-mUNA, and 2'-mUNA. In some further embodiments, the dsRNA molecule further comprises at least one thermally destabilizing modification selected from the group consisting of GNA, 2'-OMe, 3'-OMe, 5'-Me, Hyp-spacer, SNA, hGNA, hhGNA, mGNA, TNA, and h'GNA (Mod A-Mod K).

[0476] The term "acyclic nucleotide" refers to any nucleotide having an acyclic ribose sugar, e.g., where any of the bonds between the ribose carbons (e.g., C1'-C2', C2'-C3', C3'-C4', C4'-O4', or C1'-O4') are absent, and / or at least one of the ribose carbons or oxygens (e.g., C1', C2', C3', C4', or O4'), independently or in combination, is absent from the nucleotide. In some embodiments, an acyclic nucleotide is TIFF2024525713000072.tif35151, where B is a modified or unmodified nucleobase, R1 and R2 are independently H, halogen, OR3, or alkyl, and R3 is H, alkyl, cycloalkyl, aryl, aralkyl, heteroaryl, or sugar. The term "UNA" refers to an acyclic, unlocked nucleic acid in which one of the sugar bonds has been removed to form an unlocked "sugar" residue. In one example, UNA also encompasses monomers in which the C1'-C4' bond has been removed (i.e., a carbon-oxygen-carbon covalent bond between the C1' and C4' carbons). In another example, the C2'-C3' bond of the sugar (i.e., the carbon-carbon covalent bond between the C2' and C3' carbons) is removed (see Mikhailov et al., Tetrahedron Letters, 26(17):2059 (1985), and Fluiter et al., Mol. Biosyst., 10:1039 (2009), which are incorporated herein by reference in their entireties). Acyclic derivatives offer greater backbone flexibility without affecting Watson-Crick pairing. Acyclic nucleotides can be attached via a 2'-5' or 3'-5' linkage.

[0477] The term "GNA" refers to glycol nucleic acid, which is a polymer similar to DNA or RNA, but differs in the composition of its "backbone" in that it is made up of repeating glycerol units joined by phosphodiester bonds. TIFF2024525713000073.tif57128

[0478] The thermally destabilizing modification of the duplex can be a mismatch (i.e., non-complementary base pair) between the thermally destabilizing nucleotide and the opposing nucleotide in the opposing strand of the dsRNA duplex. Exemplary mismatch base pairs include G:G, G:A, G:U, G:T, A:A, A:C, C:C, C:U, C:T, U:U, T:T, U:T, or combinations thereof. Other mismatch base pairings known in the art are also contemplated by the present invention. Mismatches can occur between nucleotides that are either naturally occurring or modified nucleotides; that is, mismatch base pairing can occur between the nucleobases derived from each nucleotide, independently of the modification on the ribose sugar of the nucleotide. In certain embodiments, the dsRNA molecule contains at least one nucleobase in the mismatch pairing that is a 2'-deoxynucleobase, for example, the 2'-deoxynucleobase is in the sense strand.

[0479] In some embodiments, the duplex thermodestabilizing modifications in the seed region of the antisense strand include nucleotides that impair WCH bonding with complementary bases on the target mRNA, such as: TIFF2024525713000074.tif72135.

[0480] Many examples of abasic nucleotides, acyclic nucleotide modifications (including UNA and GNA) and mismatch modifications are described in detail in WO2011 / 133876, which is incorporated herein by reference in its entirety.

[0481] Thermally destabilizing modifications can also include universal base and phosphate modifications that have reduced or eliminated ability to form hydrogen bonds with opposing bases.

[0482] In some embodiments, thermal destabilizing modifications include nucleotides with non-standard bases, for example, but not limited to, nucleobase modifications that impair or completely eliminate the ability to form hydrogen bonds with the base in the opposite strand.These nucleobase modifications have been evaluated for destabilizing the central region of dsRNA duplexes, as described in WO2010 / 0011895, the entire contents of which are incorporated herein by reference.Exemplary nucleobase modifications include: TIFF2024525713000075.tif63147.

[0483] In some embodiments, the thermally destabilizing modification of the duplex in the seed region of the antisense strand is TIFF2024525713000076.tif20146, where R is H, OH, OCH3, F, NH2, NHMe, NMe2, or O-alkyl.

[0484] Exemplary phosphate modifications that have been shown to reduce the thermal stability of dsRNA duplexes compared to natural phosphodiester bonds include the following: TIFF2024525713000077.tif31139.

[0485] The alkyl R group can be a C1-C6 alkyl. Specific alkyl R groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, pentyl, and hexyl.

[0486] In some embodiments of any one of the aspects described herein, the oligonucleotide can include one or more stabilizing modifications. For example, the oligonucleotide can include at least two (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10 or more) stabilizing modifications.

[0487] In some embodiments, the oligonucleotide comprises at least two (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10 or more) stabilizing modifications. Without limitation, the stabilizing modifications in the oligonucleotide can be present at any position. In some embodiments, the oligonucleotide comprises stabilizing modifications at positions 2, 6, 8, 9, 14, and 16, counting from the 5' end. In some other embodiments, the oligonucleotide comprises stabilizing modifications at positions 2, 6, 14, and 16, counting from the 5' end. In yet some other embodiments, the oligonucleotide comprises stabilizing modifications at positions 2, 14, and 16, counting from the 5' end. In some embodiments, the oligonucleotide comprises stabilizing modifications at positions 7, 10, and 11, counting from the 5' end. In some other embodiments, the oligonucleotide comprises stabilizing modifications at positions 7, 9, 10, and 11, counting from the 5' end.

[0488] In some embodiments, the oligonucleotide comprises at least one stabilizing modification adjacent to a destabilizing modification. For example, the stabilizing modification can be a nucleotide at the 5' or 3' end of the destabilizing modification, i.e., at the -1 or +1 position from the position of the destabilizing modification. In some embodiments, the oligonucleotide comprises a stabilizing modification at each of the 5' and 3' ends of the destabilizing modification, i.e., at the -1 and +1 positions from the position of the destabilizing modification.

[0489] In some embodiments, the oligonucleotide comprises at least two stabilizing modifications at the 3' end of the destabilizing modification, ie, at positions +1 and +2 from the position of the destabilizing modification.

[0490] Exemplary thermally stabilizing modifications include, but are not limited to, 2'-fluoro modifications. Other thermally stabilizing modifications include, but are not limited to, LNA.

[0491] double stranded RNA Those skilled in the art are well aware that double-stranded RNA with a double-stranded structure of 20-23 base pairs, specifically 21 base pairs, has been evaluated as being particularly effective in inducing RNA interference (Elbashir et al., EMBO 2001, 20:6877-6888).However, it has also been found that shorter or longer double-stranded oligonucleotides can be equally effective.

[0492] Therefore, in one aspect, the present invention provides a double-stranded RNA (dsRNA) comprising a first strand (also referred to as antisense strand or guide strand) and a second strand (also referred to as sense strand or passenger strand), wherein at least one of the first strand (i.e., antisense strand) or the second strand (i.e., sense strand) is the oligonucleotide described herein.In other words, at least one of the first strand (i.e., antisense strand) or the second strand (i.e., sense strand) comprises at least one nucleotide of formula (I).

[0493] In some embodiments of any one of the aspects described herein, the sense strand is an oligonucleotide described herein. In other words, the sense strand comprises at least one nucleotide of formula (I).

[0494] In some embodiments of any one of the aspects described herein, the antisense strand is an oligonucleotide described herein. In other words, the antisense strand comprises at least one nucleotide of formula (I).

[0495] In some embodiments of the various aspects described herein, the antisense strand is substantially complementary to a target nucleic acid, e.g., a target gene or mRNA gene, and the dsRNA is capable of directing targeted cleavage of the target nucleic acid.

[0496] Each strand of the dsRNA molecule can be in the range of 15 to 35 nucleotides in length. For example, each strand can be 17 to 35 nucleotides, 17 to 30 nucleotides, 25 to 35 nucleotides, 27 to 30 nucleotides, 17 to 23 nucleotides, 17 to 21 nucleotides, 17 to 19 nucleotides, 19 to 25 nucleotides, 19 to 23 nucleotides, 19 to 21 nucleotides, 21 to 25 nucleotides, or 21 to 23 nucleotides in length. Without limitation, the sense and antisense strands can be equal or unequal in length. For example, the sense and antisense strands can independently be 18, 19, 20, 21, 22, 23, 24, or 25 nucleotides in length.

[0497] In some embodiments, the antisense strand is 15-35 nucleotides in length. In some embodiments, the antisense strand is 15-35, 17-35, 17-30, 25-35, 27-30, 17-23, 17-21, 17-19, 19-25, 19-23, 19-21, 21-25, 21-25, or 21-23 nucleotides in length. For example, the antisense strand can be 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, or 35 nucleotides in length. In some embodiments, the antisense strand is 19, 20, 21, 22, 23, 24, or 25 nucleotides in length. For example, the antisense strand is 21, 22, 23, 24, or 25 nucleotides in length. In some embodiments, the antisense strand is 22, 23, or 24 nucleotides in length. For example, the antisense strand is 23 nucleotides in length.

[0498] Like the antisense strand, the sense strand can be 15 to 35 nucleotides in length in some embodiments. In some embodiments, the sense strand is 15 to 35, 17 to 35, 17 to 30, 25 to 35, 27 to 30, 17 to 23, 17 to 21, 17 to 19, 19 to 25, 19 to 23, 19 to 21, 21 to 25, 21 to 25, or 21 to 23 nucleotides in length. For example, the sense strand can be 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, or 35 nucleotides in length. In some embodiments, the sense strand is 17, 18, 19, 20, 21, 22, 23, 24, or 25 nucleotides in length. For example, the sense strand is 19, 20, 21, 22, or 23 nucleotides in length. In some embodiments, the sense strand is 20, 21, or 22 nucleotides in length. For example, the sense strand is 21 nucleotides in length.

[0499] In some embodiments, the sense strand can be 15-35 nucleotides in length, and the antisense strand, independently of the sense strand, can be 15-35 nucleotides in length. In some embodiments, the sense strand is 15-35, 17-35, 17-30, 25-35, 27-30, 17-23, 17-21, 17-19, 19-25, 19-23, 19-21, 21-...

Claims

1. A compound of formula (III), (IV), (VI), (VII), (VIII), or (IX): wherein, L Ｐ is absent or is a linker, R １ is is either or N3, wherein, a' is 0 or 1, n is 1, 2, 3, 4, or 5, R Ｂ is O, N, S, heteroalkyl, branched alkyl, cycloalkyl, heterocyclyl, aryl (e.g., phenyl), or heteroaryl, Each R Ｃ is independently and, wherein, each b' is independently 0 or 1, each L is independently absent or a linker, Each R Ｌ is independently selected from the group consisting of a ligand (e.g., a carbohydrate, lipid, vitamin, peptide, protein, lipoprotein, peptidomimetic, polyamine, nucleoside and nucleotide, oligonucleotide, therapeutic agent, diagnostic agent, detectable label, antibody or fragment thereof, optionally substituted C １－３０ alkyl, optionally substituted C １－３０ alkenyl, optionally substituted C １－３０ alkynyl, and polyethylene glycol (PEG)). R ３２ is hydrogen, hydroxy, halogen, protected hydroxy, phosphate group, reactive phosphorus group, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxylate, amino, alkylamino, dialkylamino, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), a solid support, a linker, or a linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), and R ３３ is a reactive phosphorus group, hydrogen, hydroxy, halogen, protected hydroxy, phosphate group, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxocarboxylate, amino, alkylamino, dialkylamino, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), a solid support, a linker, or a linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), and optionally, only one of R ３２ and R ３３ is a phosphate group, a reactive phosphorus group, a solid support, a linker, or a linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), R ４ is hydrogen, optionally substituted C １－６ alkyl, optionally substituted C ２－６ alkenyl, optionally substituted C ２－６ alkynyl, or optionally substituted C １－６ alkoxy, or or R ４ and R ３２ are joined together to form 4'-C(R １０ R １１ ) ｖ -Y-2' or 4'-Y-C(R １０ R １１ ) ｖ -2', and Y is —O—, —CH ２ —, —CH(Me)—, —C(CH ３ ) ２ —, —S—, —N(R １２ )—, —C(O)—, —C(S)—, —S(O)—, —S(O) ２ —, —OC(O)—, —C(O)O—, —N(R １２ )C(O)—, or —C(O)N(R １２ )—, and R １０ and R １１ each independently is H, optionally substituted C １ -C ６ alkyl, optionally substituted C ２ -C ６ alkenyl, or optionally substituted C ２ -C ６ alkynyl, and R １２ is hydrogen, optionally substituted C １－３０ alkyl, optionally substituted C １ -C ３０ alkoxy, C １－４ haloalkyl, optionally substituted C ２－４ alkenyl, optionally substituted C ２－４ alkynyl, optionally substituted C １－３０ alkyl-CO ２ H, or a nitrogen protecting group, v is 1, 2, or 3, or or R ４ and R ３３ together with the atoms to which they are attached form an optionally substituted C ３－８ cycloalkyl, an optionally substituted C ３－８ cycloalkenyl, or an optionally substituted 3- to 8-membered heterocyclyl, R ３５ is a protected hydroxy, hydroxy, phosphate group, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxylate, amino, alkylamino, dialkylamino, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), vinylphosphonate (VP) group, C3-6 cycloalkylphosphonate (e.g., cyclopropylphosphonate), monophosphate ((HO) ２ (O)P-O-5'), diphosphate ((HO) ２ (O)P-O-P(HO)(O)-O-5'), triphosphate ((HO) ２ (O)P-O-(HO)(O)P-O-P(HO)(O)-O-5'), monothiophosphate (phosphorothioate, (HO)2(S)P-O-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)P-O-5'), phosphorothiolate ((HO)2(O)P-S-5'), alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate ((HO) ２ (O)P-NH-5', (HO)(NH ２ )(O)P-O-5'), alkylphosphonate (R(OH)(O)P-O-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonate (R(OH)(O)P-O-5', R = alkyl ether, e.g., methoxymethyl (CH ２ OMe), ethoxymethyl, etc.), (HO) ２ (X)P-O[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', or (HO)2(X)P-O[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', or (HO)2(X)P-[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl terminal phosphate and phosphate mimics (e.g., HO[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', H ２ N[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', H[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', Me ２ N[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', HO[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', H ２ N[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', H[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', Me ２ N[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5' (wherein X is O or S, and a and b are each independently 1 to 10)), and R ４２ is hydroxy, halogen, protected hydroxy, phosphate group, reactive phosphorus group, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxylate, amino, alkylamino, dialkylamino, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), solid support, linker, or linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), and R ４５ is hydroxy, protected hydroxy, a phosphate group, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxycarboxylate, amino, alkylamino, dialkylamino, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), vinylphosphonate (VP) group, C3-6 cycloalkylphosphonate (e.g., cyclopropylphosphonate), monophosphate ((HO) ２ (O)P-O-5'), diphosphate ((HO) ２ (O)P-O-P(HO)(O)-O-5'), triphosphate ((HO) ２ (O)P-O-(HO)(O)P-O-P(HO)(O)-O-5'), monothiophosphate (phosphorothioate, (HO)2(S)P-O-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)P-O-5'), phosphorothiolate ((HO)2(O)P-S-5'), alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate ((HO) ２ (O)P-NH-5', (HO)(NH ２ ) (O)P-O-5'), alkylphosphonate (R(OH)(O)P-O-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonate (R(OH)(O)P-O-5', R = alkyl ether, e.g., methoxymethyl (CH ２ OMe), ethoxymethyl, etc.), (HO) ２ (X)P-O[- (CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', or (HO)2(X)P-O[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', or (HO)2(X)P-[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl terminal phosphate and phosphate mimics (e.g., HO[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', H ２ N[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', H[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', Me ２ N[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', HO[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', H ２ N[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', H[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', Me ２ N[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5' (wherein X is O or S, and a and b are each independently 1 to 10)), and R ６２ is hydroxy, protected hydroxy, a phosphate group, a reactive phosphorus group, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxylate, amino, alkylamino, dialkylamino, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), a solid support, a linker, or a linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), and R ６３ and R ６４ are, independently, hydrogen, hydroxy, halogen, protected hydroxy, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxylate, amino, alkylamino, dialkylamino, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), or -O-lipid, R ６５ is hydroxy, protected hydroxy, phosphate group, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy, halogen, alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxocarboxylate, amino, alkylamino, dialkylamino, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), vinylphosphonate (VP) group, C3-6 cycloalkylphosphonate (e.g., cyclopropylphosphonate), monophosphate ((HO) ２ (O)P-O-5'), diphosphate ((HO) ２ (O)P-O-P(HO)(O)-O-5'), triphosphate ((HO) ２ (O)P-O-(HO)(O)P-O-P(HO)(O)-O-5'), monothiophosphate (phosphorothioate, (HO)2(S)P-O-5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)P-O-5'), phosphorothiolate ((HO)2(O)P-S-5'), alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate ((HO) ２ (O)P-NH-5', (HO)(NH ２ )(O)P-O-5'), alkylphosphonate (R(OH)(O)P-O-5', R = alkyl, e.g., methyl, ethyl, isopropyl, propyl, etc.), alkyl ether phosphonate (R(OH)(O)P-O-5', R = alkyl ether, e.g., methoxymethyl (CH ２ OMe), ethoxymethyl, etc.), (HO) ２ (X)P-O[- (CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', or (HO)2(X)P-O[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', or (HO)2(X)P-[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5' (wherein X is O, S, or optionally substituted alkyl), and dialkyl terminal phosphate and phosphate mimics (e.g., HO[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', H ２ N[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', H[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', Me ２ N[-(CH ２ ) ａ -O-P(X)(OH)-O] ｂ -5', HO[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', H ２ N[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', H[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5', Me ２ N[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5' (wherein X is O or S, and a and b are each independently 1 to 10)), Each R ８ and R ９ is, independently, H, a targeting ligand (e.g., GalNAc), a pharmacokinetic modifying factor, an optionally substituted C １－３０ alkyl, an optionally substituted C １－３０ alkenyl, or an optionally substituted C １－３０ alkynyl. said compound.

2. A compound of formula (IIIa): or a compound of formula (IIIb): The compound according to claim 1, which is a compound of formula (IIIa) or (IIIb).

3. R １ is The compound according to claim 1, which is a compound of formula (IIIc).

4. R Ｃ is The compound according to claim 1, which is a compound of formula (IV).

5. R ｃ is The compound according to claim 1, which is a compound of formula (V).

6. The compound according to claim 1, wherein at least one L is a linker.

7. At least one R Ｌ is independently selected from the group consisting of carbohydrates, lipids, vitamins, peptides, proteins, lipoproteins, peptidomimetics, polyamines, nucleosides and nucleotides, oligonucleotides, therapeutic agents, diagnostic agents, detectable labels, antibodies or fragments thereof, the compound according to any one of claims 1 to 6.

8. At least one R Ｌ The compound according to claim 1, wherein at least one R is selected from the group consisting of a targeting ligand, an endosome-lytic ligand, and a PK-modulating ligand.

9. The compound according to claim 1, wherein n is 1, 2, 3, or 5.

10. R Ｂ is O, N, C(CH ２ O−) ４ , benzyl, or The compound according to claim 1, which is a compound of formula (VI).

11. R ３２ is hydrogen, hydroxyl, protected hydroxyl, halogen, optionally substituted C １－３０ alkoxy (e.g., methoxy), halogen, alkoxyalkyl (e.g., 2-methoxyethyl), amino, alkylamino, dialkylamino, a reactive phosphorus group, a solid support, a linker, or a linker covalently attached to a solid support, or R ３２ and R ４ together form 4’-C(R １０ R １１ ), ｖ -Y-2’ or 4’-Y-C(R １０ R １１ ), ｖ -2’, the compound according to claim 1.

12. R ３２ is hydrogen, hydroxyl, protected hydroxyl, fluoro, methoxy, or 2 - methoxyethoxy, or R ３２ and R ４ together form 4’ - C(R １０ R １１ ) ｖ -Y-2, the compound according to claim 11.

13. R ３２ The compound according to claim 12, wherein R is hydrogen.

14. R ４ The compound according to claim 1, wherein R is H.

15. R ３３ The compound according to claim 14, wherein R is a reactive phosphorus group, hydrogen, hydroxyl, protected hydroxyl, solid support, linker, or a linker covalently attached to the solid support.

16. R ３３ The compound according to claim 1, wherein R is a reactive phosphorus group, a solid support, a linker, or a linker covalently attached to a solid support.

17. R ３３ The compound according to claim 16, wherein R is a reactive phosphorus or a linker covalently bonded to a solid support.

18. R ３５ is a protected hydroxy, hydroxy, optionally substituted C １－３０ alkoxy, vinyl phosphonate (VP) group, monophosphate, diphosphate, triphosphate, monothiophosphate (phosphorothioate), monodithiophosphate, phosphorothiolate, alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate, alkyl phosphonate, alkyl ether phosphonate, dialkyl terminal phosphate, and phosphate mimetic, the compound according to claim 17.

19. R ３５ The compound according to claim 18, wherein R is a protected hydroxyl or a hydroxyl.

20. R ３２ is hydrogen, hydroxyl, protected hydroxyl, fluoro, methoxy, or 2 - methoxyethoxy, and R ３３ is a reactive phosphorus group, hydrogen, hydroxyl, protected hydroxyl, a solid support, a linker, or a linker covalently attached to a solid support, and R ４ is H, and R ３５ is protected hydroxyl or hydroxyl, the compound according to claim 1.

21. R ３２ is a reactive phosphorus group, hydrogen, hydroxyl, protected hydroxyl, solid support, linker, or a linker covalently attached to the solid support, R ３３ is hydrogen, hydroxyl, protected hydroxyl, fluoro, methoxy, or 2 - methoxyethoxy, R ４ is H, R ３５ is a protected hydroxyl or hydroxyl, the compound according to claim 1.

22. R ３２ and R ４ together form 4'-C(R １０ R １１ ) ｖ -Y-2, and R ３３ is a reactive phosphorus group, hydrogen, hydroxyl, protected hydroxyl, solid support, linker, or a linker covalently attached to a solid support, and R ３５ is a protected hydroxyl or hydroxyl, the compound according to claim 1.

23. R ３５ is a vinylphosphonate (VP) group, cyclopropylphosphonate, monophosphate, diphosphate, triphosphate, monothiophosphate (phosphorothioate), monodithiophosphate, phosphorothiolate, alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate, alkylphosphonate, alkyl ether phosphonate, dialkyl terminal phosphate, or a phosphate mimetic, and R ３３ is a reactive phosphorus group, the compound according to claim 1.

24. R ３５ is a vinylphosphonate (VP) group, cyclopropylphosphonate, or phosphate mimetic, and R ３３ is a reactive phosphorus group, the compound according to claim 1.

25. R ３５ is a vinylphosphonate (VP) group (e.g., E-vinylphosphonate), cyclopropylphosphonate, or phosphate mimetic, and R ３３ is a phosphoramidite group, the compound according to claim 1.

26. R ３５ is a triphosphate group, R ３３ is allyloxy, azidomethoxy, or aminooxy, the compound according to claim 1.

27. The compound according to claim 1, wherein the compound is a compound of formula (IV).

28. L Ｐ The compound according to claim 27, wherein L is a linker.

29. R １ is N ３ The compound according to claim 27, wherein

30. R ４２ is a reactive phosphorus group, hydroxyl, protected hydroxyl, phosphate group, optionally substituted C １－３０ alkoxy (e.g., methoxy), solid support, linker, or a linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), the compound according to claim 27.

31. R ４２ wherein R is a reactive phosphorus group, hydroxyl, protected hydroxyl, phosphate group, solid support, linker, or a linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), the compound according to claim 30.

32. R ４２ The compound according to claim 31, wherein R is a hydroxyl group or a protected hydroxyl group.

33. R ４２ is a solid support, a linker, or a linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), the compound according to claim 31.

34. R ４２ The compound according to claim 31, wherein R is a reactive phosphorus group.

35. R ４５ is hydroxy, protected hydroxy, optionally substituted C １－３０ alkoxy, vinylphosphonate (VP) group, monophosphate, diphosphate, triphosphate, monothiophosphate (phosphorothioate), monodithiophosphate, phosphorothiolate, alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate, alkylphosphonate, alkyl ether phosphonate, dialkyl terminal phosphate, and a phosphate mimetic, the compound according to claim 27.

36. R ４５ The compound according to claim 35, wherein R is hydroxyl or protected hydroxyl.

37. R ４５ The compound according to claim 35, wherein R is a vinylphosphonate (VP) group, cyclopropylphosphonate, monophosphate, diphosphate, triphosphate, monothiophosphate (phosphorothioate), monodithiophosphate, phosphorothiolate, alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate, alkylphosphonate, alkyl ether phosphonate, or dialkyl terminal phosphate.

38. R ４５ The compound according to claim 37, wherein R is a vinylphosphonate (VP) group (e.g., E-vinylphosphonate), cyclopropylphosphonate, or a phosphate mimetic.

39. R ４５ is a protecting hydroxy (e.g., 4,4'-dimethoxytrityl protection) or a phosphate group, R ４２ is a reactive phosphorus group (e.g., phosphoramidite, e.g., 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, or 3'-[(β-thiobenzoylethyl)-(1-pyrrolidinyl)]-thiophosphoramidite), the compound according to claim 27.

40. R ４５ is a protecting hydroxy (e.g., 4,4'-dimethoxytrityl protection) or a phosphate group, and R ４２ is a solid support, a linker, or a linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), the compound according to claim 27.

41. The compound according to claim 1, which is a compound of formula (V), (VII), (VIII), or (IX).

42. R ６２ is hydroxyl, protected hydroxyl, a phosphate group, a reactive phosphorus group, optionally substituted C １－３０ alkoxy (e.g., methoxy), a solid support, a linker, or a linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), the compound according to claim 41.

43. R ６２ wherein R is hydroxyl, protected hydroxyl, a phosphate group, a reactive phosphorus group, a solid support, a linker, or a linker covalently attached to the solid support (e.g., -C(O)CH ２ CH ２ C(O)-), the compound according to claim 42.

44. R ６２ The compound according to claim 43, wherein R is a hydroxyl group or a protected hydroxyl group.

45. R ６２ wherein R is a solid support, a linker, or a linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), the compound according to claim 43.

46. R ６２ The compound according to claim 43, wherein R is a reactive phosphorus group.

47. R ６５ is hydroxy, protected hydroxy, optionally substituted C １－３０ alkoxy, vinyl phosphonate (VP) group, monophosphate, diphosphate, triphosphate, monothiophosphate (phosphorothioate), monodithiophosphate, phosphorothiolate, alpha-thiotriphosphate, beta-thiotriphosphate, gamma-thiotriphosphate, phosphoramidate, alkyl phosphonate, alkyl ether phosphonate, dialkyl terminal phosphate, and phosphate mimetic, the compound according to claim 41.

48. R ６５ The compound according to claim 47, wherein R is hydroxyl or protected hydroxyl.

49. R ６５ The compound according to claim 48, wherein R is a vinylphosphonate (VP) group, cyclopropylphosphonate, monophosphate, diphosphate, triphosphate, monothiophosphate (phosphorothioate), monodithiophosphate, phosphorothiolate, alpha - thiotriphosphate, beta - thiotriphosphate, gamma - thiotriphosphate, phosphoramidate, alkylphosphonate, alkyl ether phosphonate, or dialkyl terminal phosphate.

50. R ６５ The compound according to claim 49, wherein R is a vinylphosphonate (VP) group (e.g., E-vinylphosphonate), cyclopropylphosphonate, or phosphate mimetic.

51. R ６５ is a protecting hydroxy (e.g., 4,4'-dimethoxytrityl protection) or a phosphate group, and R ６２ is a reactive phosphorus group (e.g., phosphoramidite, e.g., 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, or 3'-[(β-thiobenzoylethyl)-(1 The compound according to claim 49, which is a compound of formula (VII).

52. R ６５ is a protecting hydroxy (e.g., 4,4'-dimethoxytrityl protection) or a phosphate group, R ６２ is a solid support, a linker, or a linker covalently attached to a solid support (e.g., -C(O)CH ２ CH ２ C(O)-), the compound according to claim 41.

53. A compound of formula (IIIc): wherein, Q, Z, and m are defined as one of the sets (i), (ii), or (iii), where (i) Q is optionally substituted aryl (e.g., phenyl) or optionally substituted heteroaryl, m is an integer selected from 1 to the maximum number of substituents of Q (e.g., when Q is phenyl, m is 1, 2, 3, 4, or 5, e.g., 1, 2, or 3, or 1 or 2), Each Z is -Z １ , -Z ２ , or -C(R Ｃ ) ３ , wherein R Ｃ is aryl (e.g., phenyl or naphthyl) or heteroaryl, each substituted with one or more Z １ groups or Z ２ groups (e.g., 1, 2, or 3), Each Z １ is selected from the group consisting of, In the formula, R Ｎ is hydrogen or C １－６ alkyl, and Z ２ is or Alternatively (ii) m is 1, Q is -CH ２ O-, -CH ２ S-, or -CH ２ N(R Ｎ ), where N, O, or S is bonded to Z, Z is , -(CH ２ ), ０－１ -Y-(Z ３ ), ｐ , -C(H)(CH ２ Z １ ), ２ , -CH ２ C(H)(CH ２ Z １ ), ２ , or -CH ２ C(CH ２ Z １ ), ３ wherein, Y is optionally substituted aryl or optionally substituted heteroaryl, Each Z ３ is Z １ or Z ２ and p is an integer selected from 1 to the maximum number of substituents of Y (for example, when Y is phenyl, p is 1, 2, 3, 4, or 5, for example, 1, 2, or 3, or 1 or 2), or Alternatively (iii) Q is -CH ２ N-, and m is 2, each Z is , -(CH ２ ) ０－１ -Y-(Z ３ ) ｐ , or -CH ２ C(CH ２ Z １ ) ３ is the compound according to claim 1.

54. R ３５ is a protecting hydroxy (e.g., 4,4'-dimethoxytrityl protection) or a phosphate group, and R ３３ is hydroxy or a reactive phosphorus group (e.g., phosphoramidite, e.g., 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, or 3'-[(β-thiobenzoylethyl)-(1-pyrrolidinyl)]-thiophosphoramidite), the compound according to claim 41.

55. R ３２ is hydrogen, hydroxy, halogen, protected hydroxy, optionally substituted C1-30 alkyl, optionally substituted C2-30 alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxylate, amino, alkylamino, dialkylamino, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), and is the compound according to claim 42.

56. R ３２ is hydrogen, hydroxy, fluoro, chloro, methoxy, ethoxy, 2-methoxyethyl, or C ６－２４ alkyl (e.g., n-C ６－２４ alkyl) and the compound according to claim 43.

57. Formula (IVb): is a compound of formula: Q Ｐ , Z Ｐ , and m Ｐ is defined as one of the sets (i), (ii), or (iii), where (i) Q Ｐ is an optionally substituted aryl (e.g., phenyl) or an optionally substituted heteroaryl, m Ｐ is an integer selected from 1 to the maximum number of substituents of Q Ｐ (for example, when Q Ｐ is phenyl, m is 1, 2, 3, 4, or 5, for example, 1, 2, or 3, or 1 or 2). Each Z Ｐ is -Z Ｐ１ , -Z Ｐ２ , or -C(R ＰＣ ) ３ and R ＰＣ is aryl (e.g., phenyl or naphthyl) or heteroaryl, each being substituted with one or more Z Ｐ１ groups or Z Ｐ２ groups (e.g., 1, 2, or 3), Each Z Ｐ１ is selected from the group consisting of, wherein R Ｎ is hydrogen or C １－６ alkyl, and Z Ｐ２ is or (ii) m Ｐ is 1, Q Ｐ is -CH ２ O-, -CH ２ S-, or -CH ２ N(R Ｎ ), where N, O, or S is attached to Z Ｐ and Z Ｐ is , -(CH ２ ) ０－１ -Y-(Z Ｐ３ ) ｐｐ , -C(H)(CH ２ Z Ｐ１ ) ２ , -CH ２ C(H)(CH ２ Z Ｐ１ ) ２ , or -CH ２ C(CH ２ Z Ｐ１ ) ３ , wherein Y Ｐ is an aryl which may be substituted or a heteroaryl which may be substituted, Each Z Ｐ３ is Z Ｐ１ or Z Ｐ２ and pp is an integer selected from 1 to the maximum number of substituents of Y Ｐ (for example, when Y Ｐ is phenyl, pp is 1, 2, 3, 4, or 5, for example, 1, 2, or 3, or 1 or 2), or Alternatively (iii) Q Ｐ is -CH ２ N-, and m Ｐ is 2, Each Z Ｐ is , -(CH ２ ), ０－１ -Y-(Z Ｐ３ ), ｐｐ or -CH ２ C(CH ２ Z Ｐ１ ), ３ wherein. the compound according to claim 1.

58. R ４２ is hydroxy, a linker to a solid support, or a reactive phosphorus group (e.g., phosphoramidite, e.g., 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, or 3'-[(β-thiobenzoylethyl)-(1-pyrrolidinyl)]-thiophosphoramidite), and R ４５ is protected hydroxy (e.g., 4,4'-dimethoxytrityl protected) or a phosphate group, the compound according to claim 57.

59. Formula VIb, VIIb, VIIIb, or IXb: is a compound of formula: Q Ｈ 、Z Ｈ 、and m Ｈ is defined as one of the sets (i), (ii), or (iii), where (i) Q Ｈ is an optionally substituted aryl (for example, phenyl) or an optionally substituted heteroaryl, m Ｈ is an integer selected from 1 to the maximum number of substituents of Q Ｈ (for example, when Q Ｈ is phenyl, m is 1, 2, 3, 4, or 5, for example, 1, 2, or 3, or 1 or 2), Each Z Ｈ is -Z Ｈ１ , -Z Ｈ２ , or -C(R ＨＣ ) ３ wherein R ＨＣ is aryl (e.g., phenyl or naphthyl) or heteroaryl, each being substituted with one or more Z Ｈ１ groups or Z Ｈ２ groups (e.g., 1, 2, or 3), Each Z Ｈ１ is selected from the group consisting of, wherein R Ｎ is hydrogen or C １－６ alkyl, and Z Ｈ２ is or (ii) m Ｈ is 1, Q Ｈ is -CH ２ O-, -CH ２ S-, or -CH ２ N(R Ｎ ), where N, O, or S is bonded to Z Ｈ and Z Ｈ is , -(CH ２ ), ０－１ -Y-(Z Ｈ３ ), ｈｐ , -C(H)(CH ２ Z Ｐ１ ), ２ , -CH ２ C(H)(CH ２ Z Ｈ１ ), ２ , or -CH ２ C(CH ２ Z Ｈ１ ), ３ wherein, Y Ｈ is an aryl which may be substituted or a heteroaryl which may be substituted, Each Z Ｈ３ is Z Ｈ１ or Z Ｈ２ and hp is an integer selected from 1 to the maximum number of substituents of Y Ｈ (for example, when Y Ｈ is phenyl, hp is 1, 2, 3, 4, or 5, for example, 1, 2, or 3, or 1 or 2), or Alternatively (iii) Q Ｈ is -CH ２ N-, and m Ｈ is 2, and Each Z Ｈ is , -(CH ２ ) ０－１ -Y-(Z Ｈ３ ) ｈｐ , or -CH ２ C(CH ２ Z Ｈ１ ) ３ is the compound according to claim 1.

60. R ６２ is hydroxy, a linker to a solid support, or a reactive phosphorus group (e.g., a phosphoramidite, e.g., 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, 3'-[(2-cyanoethyl)-(N,N-diisopropyl)]-phosphoramidite, or 3'-[(β-thiobenzoylethyl)-(1-pyrrolidinyl)]-thio-phosphoramidite), and R ６５ is a protected hydroxy (e.g., 4,4'-dimethoxytrityl protected) or a phosphate group, the compound according to claim 59.

61. The first compound according to any one of claims 53 to 60, wherein L is a linker and R Ｌ is a ligand, of the formula R Ｌ -L-N ３ A composition which is an azide-alkyne cycloaddition (AAC) reaction product with a second compound of.

62. The composition according to claim 61, wherein all ethynyl groups in the first compound have reacted with the second compound.

63. The composition according to claim 61, wherein not all but at least one ethynyl group of the first compound has reacted with the second compound.

64. The composition according to claim 61, wherein the second compound is an azide selected from the group consisting of azide compounds 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 108, 117, and 121.

65. Formula (I), (V), (VIx), (VIIx), (VIIIx), or (IXx): is an oligonucleotide containing a nucleoside of formula: L Ｐ either does not exist or is a linker, R １ is or N 3, wherein: a' is 0 or 1, n is 1, 2, 3, 4, or 5, R Ｂ is O, N, S, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl, Each R Ｃ is, independently, and wherein: each b' is 0 or 1, each L is independently absent or a linker, Each R Ｌ is independently selected from the group consisting of carbohydrates, lipids, vitamins, peptides, proteins, lipoproteins, peptidomimetics, polyamines, nucleosides and nucleotides, oligonucleotides, therapeutic agents, diagnostic agents, detectable labels, antibodies or fragments thereof, optionally substituted C １－３０ alkyl, optionally substituted C １－３０ alkenyl, optionally substituted C １－３０ alkynyl, and polyethylene glycol (PEG). R ２ is hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy (e.g., methoxy), alkoxyalkyl (e.g., 2-methoxyethyl), alkoxyalkylamine, alkoxyoxylate, amino, alkylamino, dialkylamino, 5- to 8-membered heterocyclyl, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), or -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), a bond to the internucleotide linkage to the subsequent nucleotide, a 3'-oligonucleotide capping group, a ligand, a linker covalently attached to one or more ligands, a solid support, a linker, or a linker covalently attached to a solid support, R ３ 、R ５２ 、and R ６３ｘ are, independently, a bond to an internucleotide bond to a subsequent nucleotide, hydrogen, hydroxy, protected hydroxy, halogen, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy (e.g., methoxy), alkoxyalkyl (e.g., 2-methoxyethyl), alkoxyalkylamine, alkoxyoxylate, amino, alkylamino, dialkylamino, 5- to 8-membered heterocyclyl, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), a 3'-oligonucleotide capping group, a ligand, a linker covalently attached to one or more ligands, a solid support, a linker, or a linker covalently attached to a solid support, R ４ is hydrogen, optionally substituted C １－６ alkyl, optionally substituted C ２－６ alkenyl, optionally substituted C ２－６ alkynyl, or optionally substituted C １－６ alkoxy, or or R ４ and R ２ together form 4'-C(R １０ R １１ ) ｖ -Y-2' or 4'-Y-C(R １０ R １１ ) ｖ -2', and Y is -O-, -CH ２ -, -CH(Me)-, -C(CH ３ ) ２ -, -S-, -N(R １２ ), -C(O)-, -C(S)-, -S(O)-, -S(O) ２ -, -OC(O)-, -C(O)O-, -N(R ａ１３ ), -C(O)-, or -C(O)N(R １２ ), and R １０ and R １１ each independently is H, optionally substituted C １ -C ６ alkyl, optionally substituted C ２ -C ６ alkenyl, or optionally substituted C ２ -C ６ alkynyl, R １２ is hydrogen, optionally substituted C １－３０ alkyl, optionally substituted C １ -C ３０ alkoxy, C １－４ haloalkyl, optionally substituted C ２－４ alkenyl, optionally substituted C ２－４ alkynyl, optionally substituted C １－３０ alkyl-CO ２ H, or a nitrogen protecting group, v is 1, 2, or 3, or or R ４ and R ３ together with the atoms to which they are attached form an optionally substituted C ３－８ cycloalkyl, an optionally substituted C ３－８ cycloalkenyl, or an optionally substituted 3- to 8-membered heterocyclyl, R ５ 、 R ５５ 、 and R ６５ｘ are, independently, a bond to a nucleotide - nucleotide bond to a preceding nucleotide, hydrogen, hydroxy, protected hydroxy, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy, optionally substituted 3 - 8 - membered heterocyclyl (e.g., morpholin - 1 - yl, piperidin - 1 - yl, or pyrrolidin - 1 - yl), halogen, alkoxyalkyl (e.g., 2 - methoxyethyl), alkoxyalkylamine, alkoxyoxocarboxylate, amino, alkylamino, dialkylamino, - O - C ４－３０ alkyl - ON(CH ２ R ８ )(CH ２ R ９ ), - O - C ４－３０ alkyl - ON(CH ２ R ８ )(CH ２ R ９ ), vinylphosphonate (VP) group, C ３－６ cycloalkylphosphonate (e.g., cyclopropylphosphonate), monophosphate ((HO) ２ )(O)P - O - 5'), diphosphate ((HO) ２ )(O)P - O - P(HO)(O) - O - 5'), triphosphate ((HO) ２ )(O)P - O - (HO)(O)P - O - P(HO)(O) - O - 5'); monothiophosphate (phosphorothioate, (HO)2(S)P - O - 5'), monodithiophosphate (phosphorodithioate, (HO)(HS)(S)P - O - 5'), phosphorothiolate ((HO)2(O)P - S - 5'); alpha - thiotriphosphate; beta - thiotriphosphate; gamma - thiotriphosphate; phosphoramidate ((HO) ２ )(O)P - NH - 5', (HO)(NH ２ )(O)P - O - 5'), alkylphosphonate [(R Ｐ )(OH)(O)P−O−5’,R Ｐ is optionally substituted C １－３０ alkyl, such as methyl, ethyl, isopropyl, or propyl)], alkyl ether phosphonate [(R Ｐ１ )(OH)(O)P−O−5’,R Ｐ１ is alkoxyalkyl, such as methoxymethyl (CH ２ OMe) or ethoxymethyl], (HO) ２ (X)P−O[−(CH ２ ) ａ −O−P(X)(OH)−O] ｂ −5’, or (HO) ２ (X)P−O[−(CH ２ ) ａ −P(X)(OH)−O] ｂ −5’, or (HO) ２ (X)P−[−(CH ２ ) ａ −O−P(X)(OH)−O] ｂ −5’, or optionally substituted alkyl, and dialkyl terminal phosphate and phosphate mimics (e.g., HO[−(CH ２ ) ａ −O−P(X)(OH)−O] ｂ −5’,H ２ N[−(CH ２ ) ａ −O−P(X)(OH)−O] ｂ −5’,H[−(CH ２ ) ａ −O−P(X)(OH)−O] ｂ −5’,Me ２ N[−(CH ２ ) ａ −O−P(X)(OH)−O] ｂ −5’,HO[−(CH ２ ) ａ −P(X)(OH)−O] ｂ −5’,H ２ N[−(CH ２ ) ａ −P(X)(OH)−O] ｂ −5’,H[−(CH ２ ) ａ −P(X)(OH)−O] ｂ -5', Me ２ N[-(CH ２ ) ａ -P(X)(OH)-O] ｂ -5') represents, wherein X is O or S, a and b are each independently 1 to 10, R ６３ and R ６４ are, independently, hydrogen, hydroxy, halogen, protected hydroxy, optionally substituted C １－３０ alkyl, optionally substituted C ２－３０ alkenyl, optionally substituted C ２－３０ alkynyl, optionally substituted C １－３０ alkoxy (e.g., methoxy), alkoxyalkyl (e.g., methoxyethyl), alkoxyalkylamine, alkoxyoxylate, amino, alkylamino, dialkylamino, -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), -O-C ４－３０ alkyl-ON(CH ２ R ８ )(CH ２ R ９ ), or -O-lipid, Each R ８ and R ９ is independently H, a targeting ligand (e.g., GalNAc), a pharmacokinetic modifying factor, an optionally substituted C １－３０ alkyl, an optionally substituted C １－３０ alkenyl, or an optionally substituted C １－３０ alkynyl, provided that (i) R ２ and R ３ one or less of which is a bond to a nucleotide bond to a subsequent nucleotide, (ii) R ２ and R ３ If both of and R are not linked to the internucleotide bond to the subsequent nucleotide, R ５ is linked to the internucleotide bond to the preceding nucleotide, (iii) R ５２ wherein, when R is not bonded to the internucleotide linkage to a subsequent nucleotide, ５５ R is bonded to the internucleotide linkage to a preceding nucleotide, (iv) R ５５ is not a bond to the internucleotide bond to the preceding nucleotide, R ５２ is a bond to the internucleotide bond to the subsequent nucleotide on the condition that the oligonucleotide.

66. The nucleoside of formula (I) is a nucleoside of formula (Ia): or formula (Ib): The oligonucleotide according to claim 65.

67. R １ is as defined in claim 65.

68. R Ｃ is as defined in claim 65.

69. R １ is as defined in claim 65.

70. The oligonucleotide according to claim 65, wherein at least one L is a linker.

71. At least one R Ｌ is independently selected from the group consisting of carbohydrates, lipids, vitamins, peptides, proteins, lipoproteins, peptidomimetics, polyamines, nucleosides and nucleotides, oligonucleotides, therapeutic agents, diagnostic agents, detectable labels, antibodies or fragments thereof, the oligonucleotide according to claim 65.

72. At least one R Ｌ The oligonucleotide according to claim 65, wherein the at least one R is selected from the group consisting of a targeting ligand, an endosome-lytic ligand, and a PK-modulating ligand.

73. The oligonucleotide according to claim 65, wherein n is 1, 2, 3, or 5.

74. R Ｂ is O, N, C(CH ２ O−) ４ , benzyl, or The oligonucleotide according to claim 65, which is as follows.

75. R ２ is hydrogen, hydroxyl, protected hydroxyl, halogen, optionally substituted C １－３０ alkoxy (e.g., methoxy), halogen, alkoxyalkyl (e.g., 2-methoxyethyl), amino, alkylamino, dialkylamino, a reactive phosphorus group, a solid support, a linker, or a linker covalently attached to a solid support, or R ２ and R ４ together form 4’-C(R １０ R １１ )) ｖ -Y-2’ or 4’-Y-C(R １０ R １１ )) ｖ -2’, the oligonucleotide according to claim 65.

76. R ２ is hydrogen, hydroxyl, protected hydroxyl, fluoro, methoxy, or 2 - methoxyethoxy, or R ３２ and R ４ together form 4’ - C(R １０ R １１ ) ｖ -Y-2, the oligonucleotide according to claim 75.

77. R ２ The oligonucleotide according to claim 76, wherein R is hydrogen.

78. R ４ The oligonucleotide according to claim 65, wherein R is H.

79. R ３ is a linkage to an internucleotide linkage to a subsequent nucleotide, hydroxy, optionally substituted C １－３０ alkoxy, 3'-oligonucleotide capping group, solid support, linker, or a linker covalently attached to a solid support, the oligonucleotide according to claim 65.

80. R ３ The oligonucleotide according to claim 79, wherein R is a linkage to an internucleotide linkage to a subsequent nucleotide.

81. R ３ The oligonucleotide according to claim 65, wherein R is hydroxyl.

82. R ５ is a bond to a nucleotide - nucleotide bond to a preceding nucleotide, hydroxy, optionally substituted C １－３０ alkoxy, vinylphosphonate (VP) group, monophosphate, diphosphate, triphosphate, monothiophosphate (phosphorothioate), monodithiophosphate, phosphorothiolate, alpha - thiotriphosphate, beta - thiotriphosphate, gamma - thiotriphosphate, phosphoramidate, alkylphosphonate, alkyl ether phosphonate, dialkyl terminal phosphate, and phosphate mimetic, the oligonucleotide according to claim 65.

83. R ５ The oligonucleotide according to claim 82, wherein R is a bond to an internucleotide bond to a preceding nucleotide.

84. R ５ is hydroxy, optionally substituted C １－３０ alkoxy, vinyl phosphonate (VP) group, monophosphate, diphosphate, triphosphate, monothiophosphate (phosphorothioate), monodithiophosphate, phosphorothiolate, alpha-thiotriphosphate, beta-thiotriphosphate, or gamma-thiotriphosphate, the oligonucleotide according to claim 65.

85. The oligonucleotide according to claim 65, wherein the nucleoside is a nucleoside of formula (V).

86. R ５２ is a bond to an internucleotide bond to a subsequent nucleotide, hydroxy, optionally substituted C １－３０ alkoxy, 3'-oligonucleotide capping group, solid support, linker, or linker covalently attached to a solid support, the oligonucleotide according to claim 85.

87. R ５２ The oligonucleotide according to claim 86, wherein R is a bond to an internucleotide bond to a subsequent nucleotide.

88. R ５２ The oligonucleotide according to claim 86, wherein R is hydroxyl.

89. R ５５ The oligonucleotide according to claim 85, wherein R is a bond to an internucleotide bond to a preceding nucleotide.

90. R ５５ is hydroxy, optionally substituted C １－３０ alkoxy, vinylphosphonate (VP) group, monophosphate, diphosphate, triphosphate, monothiophosphate (phosphorothioate), monodithiophosphate, phosphorothiolate, alpha-thiotriphosphate, beta-thiotriphosphate, or gamma-thiotriphosphate, the oligonucleotide according to claim 85.

91. The oligonucleotide according to claim 65, wherein the nucleoside is a nucleoside of formula (VIx), (VIIx), (VIIIx), or (Ixx).

92. R ６２ｘ is a bond to an internucleotide bond to a subsequent nucleotide, hydroxy, optionally substituted C １－３０ alkoxy, 3'-oligonucleotide capping group, solid support, linker, or a linker covalently attached to a solid support, the oligonucleotide according to claim 91.

93. R ６２ｘ The oligonucleotide according to claim 91, wherein R is a bond to a nucleotide - nucleotide bond to a subsequent nucleotide.

94. R ６２ｘ The oligonucleotide according to claim 91, wherein R is hydroxyl.

95. R ６５ｘ The oligonucleotide according to claim 91, wherein R is a bond to an internucleotide bond to a preceding nucleotide.

96. R ６５ｘ is hydroxy, optionally substituted C １－３０ alkoxy, vinylphosphonate (VP) group, monophosphate, diphosphate, triphosphate, monothiophosphate (phosphorothioate), monodithiophosphate, phosphorothiolate, alpha-thiotriphosphate, beta-thiotriphosphate, or gamma-thiotriphosphate, the oligonucleotide according to claim 91.

97. The oligonucleotide according to claim 65, comprising 3 to 50 nucleotides.

98. The oligonucleotide according to claim 65, comprising at least one ribonucleotide.

99. The oligonucleotide according to claim 65, comprising at least one 2'-deoxyribonucleotide.

100. The oligonucleotide according to claim 65, comprising at least one nucleotide having a modified nucleobase or a non-natural nucleobase.

101. The oligonucleotide according to claim 65, comprising at least one nucleotide having a modified ribose sugar.

102. The oligonucleotide according to claim 65, comprising at least one nucleotide containing a group that is neither H nor OH at the 2'-position of the ribose sugar.

103. The oligonucleotide according to claim 65, comprising at least one nucleotide having 2'-F ribose.

104. The oligonucleotide according to claim 65, comprising at least one nucleotide having 2'-OMe ribose.

105. The oligonucleotide according to claim 65, comprising at least one nucleotide containing a moiety other than ribose sugar.

106. The oligonucleotide according to claim 65, comprising at least one modified nucleotide internucleoside linkage.

107. The oligonucleotide according to claim 65, comprising at least two, for example, three, four, or five consecutively and independently selected monomers of formula (I), formula (V), formula (VIx), formula (VIIx), formula (VIIIx), and / or (IXx).

108. The oligonucleotide according to claim 65, which is attached to a solid support.

109. The oligonucleotide according to claim 65, comprising at least one hydroxyl, phosphate, or amino protecting group.

110. A double-stranded nucleic acid comprising a first oligonucleotide strand and a second oligonucleotide strand substantially complementary to the first strand, wherein the first or second strand is the oligonucleotide according to claim 65, said double-stranded nucleic acid.

111. The double-stranded nucleic acid according to claim 110, wherein the first and second strands are independently 15 to 25 nucleotides in length.

112. The double-stranded nucleic acid according to claim 110, wherein the double-stranded nucleic acid is capable of inducing RNA interference.

113. The double-stranded nucleic acid according to claim 110, wherein one or both strands have an overhang of 1 to 5 nucleotides at their respective 5'- or 3'-ends.

114. The double-stranded nucleic acid according to claim 113, wherein only one strand has an overhang of 2 nucleotides at its 5'- or 3'-end.

115. The double-stranded nucleic acid according to claim 113, wherein only one strand has an overhang of 2 nucleotides at its 3'-end.

116. A method for reducing the expression of a target gene in a subject, comprising administering to the subject (i) the double-stranded RNA according to claim 110, wherein the first strand or the second strand is complementary to the target gene, or (ii) the oligonucleotide according to claim 65, which is complementary to the target gene in either case, said method.

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