2'-Alkyl or 3'-Alkyl Modified Ribose Derivatives for Use in In Vivo Delivery of Oligonucleotides

Abstract

The present disclosure relates to linker compounds of formula (I) or (II): JPEG2024534502000192.jpg27128 and its pharma- ceutically acceptable salts, as well as related scaffolds and conjugates. More specifically, compounds of formula (1-A) are disclosed. The present disclosure also relates to the use of linker compounds, scaffolds, and conjugates, for example, in the delivery of nucleic acids and / or in the treatment or prevention of diseases.

Description

[Technical field]

[0001] Related Applications This application claims priority to and the benefit of U.S. Provisional Patent Application No. 63 / 246,870, filed September 22, 2021, the entire contents of which are incorporated herein by reference. [Background technology]

[0002] Efficient delivery of genetic material such as RNA to cells in vivo requires specific targeting and protection from the extracellular environment, especially serum proteins. One way to achieve specific targeting is to conjugate a targeting moiety to the nucleic acid (e.g., an oligonucleotide). The targeting moiety helps direct the nucleic acid to the site of interest. The targeting moiety can improve delivery by receptor-mediated endocytosis. This process is initiated through activation of cell surface or membrane receptors following binding of a specific ligand to the receptor. Many receptor-mediated endocytosis systems are known, including those that recognize sugars such as galactose, mannose, mannose-6-phosphate, peptides and proteins such as transferrin, asialoglycoprotein, vitamin B12, insulin, and epidermal growth factor (EGF). The asialoglycoprotein receptor (ASGP-R) is a high-capacity receptor and is highly abundant on hepatocytes. ASGP-R exhibits a higher affinity for N-acetyl-D-galactosylamine (GalNAc) than for D-Gal. Recently, certain carbohydrate conjugates have been shown to be valuable alternatives to liposomes for nucleic acid delivery. Furthermore, after successful delivery into cells, the stability of the nucleic acid in the cellular environment is important to achieve the desired therapeutic effect.

[0003] Thus, there is a continuing need for new linkers and conjugates for nucleic acid delivery. The present disclosure meets this need. Summary of the Invention

[0004] In some aspects, the present disclosure provides a compound of formula (I) or (II): [ka] or a pharma- ceutically acceptable salt thereof, wherein: B is H or a nucleobase moiety; W is H, C1-C6 alkyl optionally substituted with one or more halogens, or an amino substituent; Y is H, C1-C6 alkyl optionally substituted with one or more halogens, -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y , -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(OR Y )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, -P(=S)(SR Y ) 2, or a hydroxy protecting group; Each R Y is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; Z is H or C1-C6 alkyl optionally substituted with one or more halogens, -P(R Z )2, -P(OR Z )(N(R Z )2), -P(=O)(OR Z )R Z , -P(=S)(OR Z )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z)2, -P(=S)(SR Z ) 2, or a hydroxy protecting group; Each R Z is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; Or Y and Z in formula (I) together represent -Si(R L )2-O-Si(R L )2-, and each R L is independently H or C1-C6 alkyl; Each R a are independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens, or two R a forms a double bond with two adjacent carbon atoms, Each R b is independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 1 is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 2 is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 3 is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 4 is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; Each R 5 is independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; wherein n is an integer ranging from about 0 to about 10.

[0005] In some aspects, the disclosure provides compounds that are isotopic derivatives of the compounds disclosed herein.

[0006] In some aspects, the disclosure provides a scaffold, or a pharma- ceutically acceptable salt thereof, the scaffold comprising: (i) a ligand, and (ii) a linker unit, the linker unit being [ka] and a linker unit, where variables B, R 1 , R 2 , R 3 , R 4 , R 5 , Y, Z, R a , R b and n are as described herein, and # indicates attachment to a ligand.

[0007] In some aspects, the disclosure provides a scaffold, or a pharma- ceutically acceptable salt thereof, the scaffold comprising: (i) one or more nucleic acid agents, and (ii) one or more linker units, each linker unit independently comprising: [ka] and one or more linker units, where variables B, R 1 , R 2 , R 3 , R 4 , R 5 , W, Y, Z, R a , R b , and n are as described herein, and ## indicates attachment to a nucleic acid agent.

[0008] In some aspects, the disclosure provides a conjugate, or a pharma- ceutically acceptable salt thereof, the conjugate comprising: (i) one or more nucleic acid agents, (ii) one or more ligands, and (iii) one or more linker units, each linker unit independently comprising: [ka] and one or more linker units, where variables B, R 1 , R 2 , R 3 , R 4 , R 5 , Y, Z, R a , R b , and n are as described herein, where # indicates attachment to a ligand and ## indicates attachment to a nucleic acid agent.

[0009] In some aspects, the disclosure provides compounds that are isotopic derivatives of the compounds disclosed herein.

[0010] In some aspects, the present disclosure provides pharmaceutical compositions comprising a compound, scaffold, or conjugate described herein.

[0011] In some aspects, the present disclosure provides a method of modulating expression of a target gene in a subject, the method comprising administering to the subject a conjugate described herein.

[0012] In some aspects, the disclosure provides a method of delivering a nucleic acid agent to a subject, the method comprising administering to the subject a conjugate described herein.

[0013] In some aspects, the disclosure provides a method of treating or preventing a disease in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a conjugate described herein.

[0014] In some aspects, the disclosure provides for the use of a conjugate described herein in the manufacture of a medicament for modulating expression of a target gene in a subject.

[0015] In some aspects, the disclosure provides for the use of a conjugate described herein in the manufacture of a medicament for delivering a nucleic acid agent to a subject.

[0016] In some aspects, the disclosure provides for the use of a conjugate described herein in the manufacture of a medicament for the treatment or prevention of a disease in a subject in need thereof.

[0017] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. In this specification, the singular form includes the plural form unless the context clearly indicates otherwise. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of this disclosure, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference. References cited herein are not admitted to be prior art to the invention claimed. In case of conflict, the present specification, including definitions, shall control. Furthermore, the materials, methods, and examples are illustrative only and are not intended to be limiting. In the event of a conflict between the chemical structure and the name of a compound disclosed herein, the chemical structure shall control.

[0018] Other features and advantages of the disclosure will become apparent from the following detailed description, and from the claims. [Brief description of the drawings]

[0019] [Figure 1] FIG. 13 is a graph showing gene silencing activity of siRNA duplexes in liver on day 5 after a single 0.5 mg / kg sc injection of CD-1 female mice followed by HDI administration (human target gene 1 plasmid, 10 μg) on ​​day 4. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0020] The present disclosure provides the compounds, linkers, scaffolds, and conjugates described herein for nucleic acid delivery. The present disclosure also relates to the use of the compounds, linkers, scaffolds, and conjugates, for example, in the delivery of nucleic acids and / or in the treatment or prevention of diseases.

[0021] Linker Compounds of the Present Disclosure In some aspects, the present disclosure provides a compound of formula (I) or (II): [ka] or a pharma- ceutically acceptable salt thereof, wherein: B is H or a nucleobase moiety; W is H, C1-C6 alkyl optionally substituted with one or more halogens, or an amino substituent; Y is H, C1-C6 alkyl optionally substituted with one or more halogens, -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y , -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(OR Y )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, -P(=S)(SR Y ) 2, or a hydroxy protecting group; Each R Y is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; Z is H or C1-C6 alkyl optionally substituted with one or more halogens, -P(R Z )2, -P(OR Z )(N(R Z )2), -P(=O)(OR Z )R Z , -P(=S)(ORZ )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z )2, -P(=S)(SR Z ) 2, or a hydroxy protecting group; Each R Z is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; Or Y and Z in formula (I) together represent -Si(R L )2-O-Si(R L )2-, and each R L is independently H or C1-C6 alkyl; Each R a are independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens, or two R a forms a double bond with two adjacent carbon atoms, Each R b is independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 1 is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 2 is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 3 is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 4 is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; Each R 5 is independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; wherein n is an integer ranging from about 0 to about 10.

[0022] With respect to the compounds of the present disclosure, the variables B, W, Y, Z, R Y , R Z , R L , R a , R b , R 1 , R 2 , R 3 , R 4 , R 5 , and n can each be selected from the groups described herein, where applicable, and the variables B, W, Y, Z, R Y , R Z , R L , R a , R b , R 1 , R 2 , R 3 , R 4 , R 5 Any group described herein for any of the variables B, W, Y, Z, R, Y , R Z , R L , R a , R b , R 1 , R 2 , R 3 , R 4 , R 5 and the remainder of n, one or more of which may be combined with any group described herein.

[0023] Variable B In some embodiments, B is H.

[0024] In some embodiments, B is a nucleobase moiety.

[0025] As used herein, the term "nucleobase moiety" refers to a nucleobase that is attached to the remainder of a compound, e.g., via an atom of the nucleobase or a functional group thereof.

[0026] In some embodiments, the nucleobase moiety is adenine (A), cytosine (C), guanine (G), thymine (T), or uracil (U).

[0027] In some embodiments, the nucleobase moiety is a modified nucleobase.

[0028] In some embodiments, the modified nucleobase is 5-methylcytosine.

[0029] In some embodiments, the modified nucleobase is hypoxanthine, xanthine, or 7-methylguanine.

[0030] In some embodiments, the modified nucleobase is 5,6-dihydrouracil, 5-methylcytosine, or 5-hydroxymethylcytosine.

[0031] In some embodiments, the nucleobase moiety is an artificial nucleobase.

[0032] In some embodiments, the artificial nucleobase is isoguanine, isocytosine, 2-amino-6-(2-thienyl)purine, or pyrrole-2-carbaldehyde.

[0033] Variables W, Y, R Y , Z, R Z , and R L In some embodiments, W is H.

[0034] In some embodiments, W is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0035] In some embodiments, W is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0036] In some embodiments, W is methyl, ethyl, or propyl.

[0037] In some embodiments, W is an amino substituent, ie, a group suitable for replacing a hydrogen of an amino moiety, such as an amino protecting group.

[0038] In some embodiments, W is an amino protecting group, including, but not limited to, fluorenylmethyloxycarbonyl (Fmoc), tert-butyloxycarbonyl (BOC), benzyloxycarbonyl (Cbz), optionally substituted acyl, trifluoroacetyl (TFA), benzyl, triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), or toluenesulfonyl (Ts).

[0039] In some embodiments, W is optionally substituted acyl (e.g., -C(=O)(C-C 30 alkyl), C1-C 30 The alkyl is optionally substituted.

[0040] In some embodiments, W is a substituted acyl (e.g., [ka] ).

[0041] In some embodiments, W is trifluoroacetyl (TFA).

[0042] In some embodiments, W is an optionally substituted thioacyl (e.g., -C(=S)(C-C 30 alkyl), C1-C 30 The alkyl is optionally substituted.

[0043] In some embodiments, W is a substituted thioacyl (e.g., [ka] ).

[0044] In some embodiments, Y is H.

[0045] In some embodiments, Y is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0046] In some embodiments, Y is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0047] In some embodiments, Y is methyl, ethyl, or propyl.

[0048] In some embodiments, Y is -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y , -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(OR Y )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, -P(=S)(SR Y )2.

[0049] In some embodiments, Y is -P(R Y )2.

[0050] In some embodiments, Y is -PH2.

[0051] In some embodiments, Y is -P(OR Y )(N(R Y 2).

[0052] In some embodiments, Y is -P(OH)(NH2).

[0053] In some embodiments, Y is -P(O(C1-C6 alkyl))(N(C1-C6 alkyl)2), wherein C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0054] In some embodiments, Y is -P(=O)(OR Y )R Y It is.

[0055] In some embodiments, Y is -P(=O)(OH)(C1-C6 alkyl), wherein the C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0056] In some embodiments, Y is -P(=S)(OR Y )R Y It is.

[0057] In some embodiments, Y is -P(=S)(OH)(C1-C6 alkyl), wherein the C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0058] In some embodiments, Y is -P(=O)(SR Y )R Y It is.

[0059] In some embodiments, Y is -P(=O)(SH)(C1-C6 alkyl), wherein the C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0060] In some embodiments, Y is -P(=S)(SR Y )R Y It is.

[0061] In some embodiments, Y is -P(=S)(SH)(C1-C6 alkyl), wherein the C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0062] In some embodiments, Y is -P(=O)(OR Y )2.

[0063] In some embodiments, Y is -P(=O)(OH).

[0064] In some embodiments, Y is -P(=S)(OR Y )2.

[0065] In some embodiments, Y is -P(=S)(OH).

[0066] In some embodiments, Y is -P(=O)(SR Y )2.

[0067] In some embodiments, Y is -P(=O)(SH)2.

[0068] In some embodiments, Y is -P(=S)(SR Y )2.

[0069] In some embodiments, Y is -P(=S)(SH)2.

[0070] In some embodiments, Y is a hydroxy protecting group (eg, silyl, Tr, DMTr, acyl, or benzyl).

[0071] In some embodiments, Y is silyl (eg, trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl).

[0072] In some embodiments, Y is triphenylmethyl (Tr) or 4,4'-dimethoxytrityl (DMTr).

[0073] In some embodiments, Y is optionally substituted acyl (eg, optionally substituted acetyl) or benzyl.

[0074] In some embodiments, at least one R Y is H.

[0075] In some embodiments, each R Y is H.

[0076] In some embodiments, at least one R Y is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogen (e.g., F, Cl, Br, or I) or cyano.

[0077] In some embodiments, each R Y is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogen (e.g., F, Cl, Br, or I) or cyano.

[0078] In some embodiments, at least one R Y is H and at least one R Yis C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogen or cyano.

[0079] In some embodiments, Z is H.

[0080] In some embodiments, Z is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0081] In some embodiments, Z is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0082] In some embodiments, Z is methyl, ethyl, or propyl.

[0083] In some embodiments, Z is -P(R Z )2, -P(OR Z )(N(R Z )2), -P(=O)(OR Z )R Z , -P(=S)(OR Z )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z )2, -P(=S)(SR Z )2.

[0084] In some embodiments, Z is -P(R Z )2.

[0085] In some embodiments, Z is -PH2.

[0086] In some embodiments, Z is -P(OR Z )(N(R Z )2).

[0087] In some embodiments, Z is -P(OH)(NH2).

[0088] In some embodiments, Z is -P(O(C1-C6 alkyl))(N(C1-C6 alkyl)2), wherein C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0089] In some embodiments, Z is -P(=O)(OR Z )R Z It is.

[0090] In some embodiments, Z is -P(=O)(OH)(C1-C6 alkyl), wherein the C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0091] In some embodiments, Z is -P(=S)(OR Z )R Z It is.

[0092] In some embodiments, Z is -P(=S)(OH)(C1-C6 alkyl), wherein the C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0093] In some embodiments, Z is -P(=O)(SR Z )R Z It is.

[0094] In some embodiments, Z is -P(=O)(SH)(C1-C6 alkyl), wherein C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0095] In some embodiments, Z is -P(=S)(SR Z )R Z It is.

[0096] In some embodiments, Z is -P(=S)(SH)(C1-C6 alkyl), wherein C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0097] In some embodiments, Z is -P(=O)(OR Z )2.

[0098] In some embodiments, Z is -P(=O)(OH).

[0099] In some embodiments, Z is -P(=S)(OR Z )2.

[0100] In some embodiments, Z is -P(=S)(OH)2.

[0101] In some embodiments, Z is -P(=O)(SR Z )2.

[0102] In some embodiments, Z is -P(=O)(SH)2.

[0103] In some embodiments, Z is -P(=S)(SR Z )2.

[0104] In some embodiments, Z is -P(=S)(SH)2.

[0105] In some embodiments, Z is a hydroxy protecting group (eg, silyl, Tr, DMTr, acyl, or benzyl).

[0106] In some embodiments, Z is silyl (eg, trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl).

[0107] In some embodiments, Z is triphenylmethyl (Tr) or 4,4'-dimethoxytrityl (DMTr).

[0108] In some embodiments, Z is substituted acyl (eg, optionally substituted acetyl) or benzyl.

[0109] In some embodiments, at least one R Z is H.

[0110] In some embodiments, R Z is H.

[0111] In some embodiments, at least one R Z is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogen (e.g., F, Cl, Br, or I) or cyano.

[0112] In some embodiments, each R Z is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogen (e.g., F, Cl, Br, or I) or cyano.

[0113] In some embodiments, at least one R Z is H and at least one R Z is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogen (e.g., F, Cl, Br, or I) or cyano.

[0114] In some embodiments, Y and Z in formula (I) taken together represent -Si(R L )2-O-Si(R L )2- is formed.

[0115] In some embodiments, Y and Z in formula (I) are taken together to form -Si(C1-C6 alkyl)2-O-Si(C1-C6 alkyl)2-.

[0116] In some embodiments, Y and Z in formula (I) together form -Si(iPr)2-O-Si(iPr)2-.

[0117] In some embodiments, at least one R L is H.

[0118] In some embodiments, each R L is independently C1-C6 alkyl.

[0119] In some embodiments, each R L is independently methyl, ethyl, or propyl (e.g., iPr).

[0120] Variable R a , R b , R 1 , R 2 , R 3 , R 4 , R 5 , and n In some embodiments, each R a is H.

[0121] In some embodiments, at least one R a is halogen (e.g., F, Cl, Br, or I) or C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0122] In some embodiments, at least one R a is a halogen (e.g., F, Cl, Br, or I).

[0123] In some embodiments, at least one R a is F or Cl.

[0124] In some embodiments, at least one R a is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0125] In some embodiments, at least one R a is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0126] In some embodiments, at least one R a is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0127] In some embodiments, at least two R on two adjacent carbon atoms a takes together with two adjacent carbon atoms to form a double bond.

[0128] In some embodiments, R b is H.

[0129] In some embodiments, at least one R bis halogen (e.g., F, Cl, Br, or I) or C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0130] In some embodiments, at least one R b is a halogen (e.g., F, Cl, Br, or I).

[0131] In some embodiments, at least one R b is F or Cl.

[0132] In some embodiments, at least one R b is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0133] In some embodiments, at least one R b is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0134] In some embodiments, at least one R b is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0135] In some embodiments, R 1 is H.

[0136] In some embodiments, R 1is a halogen (e.g., F, Cl, Br, or I).

[0137] In some embodiments, R 1 is F or Cl.

[0138] In some embodiments, R 1 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0139] In some embodiments, R 1 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0140] In some embodiments, R 1 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0141] In some embodiments, R 2 is H.

[0142] In some embodiments, R 2 is a halogen (e.g., F, Cl, Br, or I).

[0143] In some embodiments, R 2 is F or Cl.

[0144] In some embodiments, R 2is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0145] In some embodiments, R 2 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0146] In some embodiments, R 2 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0147] In some embodiments, R 3 is H.

[0148] In some embodiments, R 3 is a halogen (e.g., F, Cl, Br, or I).

[0149] In some embodiments, R 3 is F or Cl.

[0150] In some embodiments, R 3 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0151] In some embodiments, R 3 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0152] In some embodiments, R 3 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0153] In some embodiments, R 4 is H.

[0154] In some embodiments, R 4 is a halogen (e.g., F, Cl, Br, or I).

[0155] In some embodiments, R 4 is F or Cl.

[0156] In some embodiments, R 4 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0157] In some embodiments, R 4 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0158] In some embodiments, R 4 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0159] In some embodiments, R 5 is H.

[0160] In some embodiments, at least one R 5 is halogen (e.g., F, Cl, Br, or I) or C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0161] In some embodiments, at least one R 5 is a halogen (e.g., F, Cl, Br, or I).

[0162] In some embodiments, at least one R 5 is F or Cl.

[0163] In some embodiments, at least one R 5 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0164] In some embodiments, at least one R 5 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0165] In some embodiments, at least one R 5 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0166] In some embodiments, R a , R b , R1 , R 2 , R 3 , R 4 , and R 5 Each of is H.

[0167] In some embodiments, n is an integer ranging from about 1 to about 10.

[0168] In some embodiments, n is an integer ranging from about 2 to about 10.

[0169] In some embodiments, n is an integer ranging from about 3 to about 10, from about 4 to about 10, from about 5 to about 10, or from about 6 to about 10.

[0170] In some embodiments, n is an integer ranging from about 1 to about 8, from about 1 to about 7, from about 1 to about 6, from about 1 to about 5, from about 1 to about 4, or from about 1 to about 3.

[0171] In some embodiments, n is an integer ranging from about 2 to about 8, from about 2 to about 7, from about 2 to about 6, from about 2 to about 5, from about 2 to about 4, or from about 2 to about 3.

[0172] In some embodiments, n is 0.

[0173] In some embodiments, n is 1.

[0174] In some embodiments, n is 2.

[0175] In some embodiments, n is 3.

[0176] In some embodiments, n is 4.

[0177] In some embodiments, n is 5.

[0178] In some embodiments, n is 6.

[0179] In some embodiments, n is 7.

[0180] In some embodiments, n is 8.

[0181] In some embodiments, n is 9.

[0182] In some embodiments, n is 10.

[0183] Exemplary embodiments of the compounds In some embodiments, the compound has formula (I') or (II'): [ka] Things, or a pharma- ceutically acceptable salt thereof.

[0184] In some embodiments, the compound has formula (IA) or (II-A): [ka] Things, or a pharma- ceutically acceptable salt thereof.

[0185] In some embodiments, the compound has formula (I'-A) or (II-A): [ka] Things, or a pharma- ceutically acceptable salt thereof.

[0186] In some embodiments, the compound has formula (IB) or (II-B): [ka] Things, or a pharma- ceutically acceptable salt thereof.

[0187] In some embodiments, the compound has formula (I'-B) or (II'-B): [ka] Things, or a pharma- ceutically acceptable salt thereof.

[0188] In some embodiments, Y is a hydroxy protecting group (e.g., silyl, Tr, DMTr, acyl, or benzyl); Z is a hydroxy protecting group (e.g., silyl, Tr, DMTr, acyl, or benzyl); or Y and Z in formula (I), (I'), (IA), (I'-A), (IB), or (I'-B) together represent -Si(R L )2-O-Si(R L )2-, and each R L is independently H or C1-C6 alkyl.

[0189] In some embodiments, Y is a hydroxy protecting group (eg, silyl, Tr, DMTr, acyl, or benzyl) and Z is a hydroxy protecting group (eg, silyl, Tr, DMTr, acyl, or benzyl).

[0190] In some embodiments, Y and Z in formula (I), (I'), (IA), (I'-A), (IB), or (I'-B) taken together represent -Si(R L )2-O-Si(R L )2-, and each R L is independently H or C1-C6 alkyl.

[0191] In some embodiments, the compound is [ka] or a pharma- ceutically acceptable salt thereof, wherein: B is a nucleobase moiety (e.g., adenine (A), cytosine (C), guanine (G), thymine (T), or uracil (U)); Y is -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y, -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(OR Y )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, -P(=S)(SR Y )2, or a hydroxy protecting group (e.g., silyl (e.g., trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl), triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), substituted acyl (e.g., optionally substituted acetyl), or benzyl); Each R Y is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; Z is -P(R Z )2, -P(OR Z )(N(R Z )2), -P(=O)(OR Z )R Z , -P(=S)(OR Z )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z )2, -P(=S)(SR Z )2, or a hydroxy protecting group (e.g., silyl (e.g., trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl), triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), substituted acyl (e.g., optionally substituted acetyl), or benzyl); Each R Zis independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0192] In some embodiments, the compound is [ka] [ka] or a pharma- ceutically acceptable salt thereof, wherein: B is a nucleobase moiety (e.g., adenine (A), cytosine (C), guanine (G), thymine (T), or uracil (U)); Y is -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y , -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(OR Y )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, -P(=S)(SR Y )2, or a hydroxy protecting group (e.g., silyl (e.g., trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl), triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), substituted acyl (e.g., optionally substituted acetyl), or benzyl); Each R Y is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; Z is -P(R Z )2, -P(OR Z )(N(R Z )2), -P(=O)(OR Z )RZ , -P(=S)(OR Z )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z )2, -P(=S)(SR Z )2, or a hydroxy protecting group (e.g., silyl (e.g., trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl), triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), substituted acyl (e.g., optionally substituted acetyl), or benzyl); Each R Z is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; C1-C 30 The alkyl is optionally substituted.

[0193] In some embodiments, the compound is selected from the compounds set forth in Table L and pharma- ceutically acceptable salts thereof. [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5] [Table 1-6] [Table 1-7] [Table 1-8] [Table 1-9] [Table 1-10] [Table 1-11] [Table 1-12] [Table 1-13] [Table 1-14] [Table 1-15] [Table 1-16]

[0194] In some aspects, the disclosure provides compounds that are isotopic derivatives (eg, isotopically labeled compounds) of any one of the compounds of the formulas disclosed herein.

[0195] It will be appreciated that isotopic derivatives can be prepared using any of a variety of art-recognized techniques. For example, isotopic derivatives can generally be prepared by carrying out the procedures disclosed in the schemes and / or examples herein by substituting an isotopically labeled reagent for a non-isotopically labeled reagent.

[0196] In some embodiments, the isotopic derivatives are deuterium-labeled compounds.

[0197] In some embodiments, an isotopic derivative is a deuterium-labeled compound of any one of the compounds of the formulas disclosed herein.

[0198] As used herein, the term "isotopic derivative" refers to a derivative of a compound in which one or more atoms are isotopically enriched or labeled. For example, an isotopic derivative of a compound of formula (I) or (II) is isotopically enriched or labeled with one or more isotopes compared to the corresponding compound of formula (I) or (II). In some embodiments, an isotopic derivative is 2 H, 13 C. 14 C. 15 N, 18 O. 29 Si, 32 P, and 34 S. In some embodiments, the isotopic derivative is a deuterium-labeled compound (i.e., enriched or labeled with one or more atoms selected from S and S). 2 In some embodiments, the compound is 2 H-labeled compound. In some embodiments, the compound is 13 C-labeled compounds or 14 C labeled compound. In some embodiments, the compound is 18 F labeled compound. In some embodiments, the compound is 123 I-labeled compound, 124 I-labeled compound, 125 I-labeled compound, 129 I-labeled compound, 131 I-labeled compound, 135 I-labeled compounds, or any combination thereof. In some embodiments, the compounds are 32 P-labeled compounds or 32 In some embodiments, the compound is 33 S-labeled compound, 34 S-labeled compound, 35 S-labeled compound, 36 S-labeled compounds, or any combination thereof.

[0199] It will be appreciated that isotopic derivatives can be prepared using any of a variety of art-recognized techniques. For example, isotopic derivatives can generally be prepared by carrying out the procedures disclosed in the schemes and / or examples described herein by substituting an isotopically labeled reagent for a non-isotopically labeled reagent.

[0200] It will also be appreciated that isotopic substitution may offer certain therapeutic advantages resulting from greater metabolic stability, for example, increased in vivo half-life or reduced dosage requirements.

[0201] For the avoidance of doubt, when a group is qualified herein by "as described herein," it is to be understood that the group encompasses the broadest definition occurring first, as well as each and every specific definition of that group.

[0202] It will be understood that the compounds disclosed herein may be presented in one particular configuration. Such a particular configuration should not be interpreted as limiting the present disclosure to one or another isomer, tautomer, positional isomer, or stereoisomer, nor does it exclude mixtures of isomers, tautomers, positional isomers, or stereoisomers. In some embodiments, the presentation of a compound herein in a particular configuration is intended to encompass and refer to each of the available isomers, tautomers, positional isomers, and stereoisomers of the compound, or any mixtures thereof, and the presentation is further intended to refer to the particular configuration of the compound.

[0203] However, it will be understood that the compounds disclosed herein may be presented without a specific stereochemistry (e.g., without a specific stereochemistry). Such presentation is intended to encompass all available isomers, tautomers, positional isomers, and stereoisomers of the compound. In some embodiments, presentation of a compound herein without a specific stereochemistry is intended to refer to each of the available isomers, tautomers, positional isomers, and stereoisomers of the compound, or any mixture thereof.

[0204] As used herein, the term "isomers" refers to compounds that have the same molecular formula but differ in the order of bonding of their atoms or the arrangement of their atoms in space. Compounds that have the same molecular formula but differ in the nature or order of bonding of their atoms or the arrangement of their atoms in space are called "isomers". Isomers that differ in the arrangement of their atoms in space are called "stereoisomers". Stereoisomers that are not mirror images of one another are called "diastereomers" and stereoisomers that are non-superimposable mirror images of one another are called "enantiomers". When a compound has an asymmetric center, for example, when it is bonded to four different groups, a pair of enantiomers is possible. Enantiomers can be characterized by the absolute configuration of their asymmetric center and are described by the R and S ordering rules of Cahn and Prelog or by the way the molecule rotates the plane of polarized light and are designated as dextrorotatory or levorotatory (i.e., as (+) or (-)-isomers, respectively). Chiral compounds can exist as either individual enantiomers or mixtures thereof. A mixture containing equal proportions of the enantiomers is called a "racemic mixture."

[0205] The compounds of the present disclosure may have one or more asymmetric centers, and therefore such compounds can be produced as individual (R)- or (S)-stereoisomers or as mixtures thereof. Unless otherwise indicated, the description or naming of a particular compound in the present specification and claims is intended to include both individual enantiomers and mixtures thereof, racemic or otherwise. Methods for determining stereochemistry and separating stereoisomers are well known in the art (see discussion in Chapter 4 of "Advanced Organic Chemistry", 4th edition J. March, John Wiley and Sons, New York, 2001), for example, by synthesis from optically active starting materials or by resolution of racemic forms. Some of the compounds of the present disclosure may have geometric isomeric centers (E and Z isomers). It is to be understood that the present disclosure encompasses all optical, diastereomeric and geometric isomers and mixtures thereof that have inflammasome inhibitory activity.

[0206] As used herein, the term "chiral center" refers to a carbon atom bonded to four nonidentical substituents.

[0207] As used herein, the term "chiral isomer" refers to a compound having at least one chiral center. Compounds having two or more chiral centers can exist as individual diastereomers or as a mixture of diastereomers called a "diastereomeric mixture." When one chiral center is present, a stereoisomer can be characterized by the absolute configuration (R or S) of that chiral center. Absolute configuration refers to the arrangement in space of the substituents attached to the chiral center. The substituents attached to the chiral center under consideration are ranked according to the Cahn, Ingold and Prelog ranking rules. (Cahn et al.,Angew.Chem.Inter.Edit.1966,5,385;errata 511, Cahn et al.,Angew.Chem.1966,78,413;Cahn and Ingold,J.Chem.Soc.1951(London),612, Cahn et al.,Experientia 1956, 12, 81; Cahn, J. Chem. Educ. 1964, 41, 116).

[0208] As used herein, the term "geometric isomer" refers to diastereomers whose existence is due to hindrance of rotation about a double bond or a cycloalkyl linker (e.g., 1,3-cyclobutyl). These configurations are distinguished in their names by the prefixes cis and trans, or Z and E, indicating that the groups are on the same or opposite sides of a double bond in the molecule, according to the Cahn-Ingold-Prelog rules.

[0209] It is to be understood that the compounds of the present disclosure may be represented as different chiral or geometric isomers, and when a compound has chiral or geometric isomers, all isomers are intended to be included within the scope of the present disclosure, and it should be understood that the naming of the compound does not exclude any isomeric form, and not all isomers may have the same level of activity.

[0210] It is to be understood that the structures and other compounds discussed in this disclosure include all atropisomers thereof. It should also be understood that not all atropisomers may have the same level of activity.

[0211] As used herein, the term "atropisomer" refers to a type of stereoisomer in which the atoms of two isomers are arranged differently in space. Atropisomers owe their existence to rotational constraints caused by hindrance of rotation of large groups around a central bond. Such atropisomers typically exist as mixtures, but as a result of recent advances in chromatographic techniques, it has become possible to separate mixtures of two atropisomers in selected cases.

[0212] As used herein, the term "tautomer" is one of two or more structural isomers that exist in equilibrium and are easily converted from one isomeric form to another. This conversion results in the formal migration of a hydrogen atom accompanied by the switching of adjacent conjugated double bonds. Tautomers exist as a mixture of tautomeric sets in solution. In solutions where tautomerization is possible, a chemical equilibrium of tautomers is reached. The exact ratio of tautomers varies depending on several factors including temperature, solvent and pH. The concept of tautomers that are interconvertible by tautomerization is called tautomerism. Of the various types of tautomerism that are possible, two are commonly observed. In keto-enol tautomerism, a simultaneous shift of electrons and hydrogen atoms occurs. Ring-chain tautomerism occurs as a result of an aldehyde group (-CHO) in a sugar molecule reacting with one of the hydroxyl groups (-OH) in the same molecule to give it a cyclic (ring-shaped) form as exhibited by glucose.

[0213] It should be understood that the compounds of the present disclosure may be represented as different tautomers. When a compound has tautomeric forms, it should also be understood that all tautomeric forms are intended to be included within the scope of the present disclosure, and the naming of the compound does not exclude any tautomeric form. It will be understood that certain tautomers may have a higher level of activity than others.

[0214] It should be understood that the compounds of any formula described herein include the compounds themselves, as well as their salts and solvates, if applicable. Salts can be formed, for example, between an anion and a positively charged group (e.g., amino) on the substituted compounds disclosed herein. Suitable anions include chloride, bromide, iodide, sulfate, bisulfate, sulfamate, nitrate, phosphate, citrate, methanesulfonate, trifluoroacetate, glutamate, glucuronate, glutarate, malate, maleate, succinate, fumarate, tartrate, tosylate, salicylate, lactate, naphthalenesulfonate, and acetate (e.g., trifluoroacetate).

[0215] As used herein, the term "pharmaceutically acceptable anion" refers to an anion suitable for forming a pharmaceutically acceptable salt. Similarly, salts can also be formed between a cation and a negatively charged group (e.g., carboxylate) on the substituted compounds disclosed herein. Suitable cations include sodium, potassium, magnesium, calcium, and ammonium cations such as tetramethylammonium or diethylamine. The substituted compounds disclosed herein also include salts containing quaternary nitrogen atoms.

[0216] It should be understood that the compounds of the present disclosure, for example, salts of the compounds, can exist in either hydrated or non-hydrated (anhydrous) form, or as solvates with other solvent molecules. Non-limiting examples of hydrates include monohydrates, dihydrates, etc. Non-limiting examples of solvates include ethanol solvates, acetone solvates, etc.

[0217] As used herein, the term "solvate" refers to a solvent addition form that contains either stoichiometric or non-stoichiometric amounts of solvent. Some compounds have a tendency to trap a fixed molar ratio of solvent molecules in the crystalline solid state, thereby forming a solvate. When the solvent is water, the solvate formed is a hydrate, and when the solvent is alcohol, the solvate formed is an alcoholate. A hydrate is formed by the combination of one molecule of a substance, where the water retains its molecular state as H2O, with one or more water molecules.

[0218] As used herein, the term "analog" refers to a chemical compound that is structurally similar to another but differs slightly in composition (such as the replacement of one atom with an atom of a different element, or the presence of a particular functional group, or the replacement of one functional group with another functional group). Thus, an analog is a compound that is similar or comparable in function and appearance to a reference compound, but not in structural origin.

[0219] As used herein, the term "derivative" refers to compounds that have a common core structure and are substituted with various groups as described herein.

[0220] As used herein, the term "bioisomer" refers to a compound resulting from the exchange of an atom or group of atoms with another generally similar atom or group of atoms. The purpose of bioisosteric replacement is to create a new compound with similar biological properties as the parent compound. Bioisosteric replacement can be physicochemical or topological based. Examples of carboxylic acid bioisosteres include, but are not limited to, acylsulfonamides, tetrazoles, sulfonates and phosphonates. See, for example, Patani and LaVoie, Chem.Rev.96,3147-3176,1996.

[0221] It should also be understood that any one of the particular compounds of the formulas disclosed herein may exist in unsolvated forms, such as, for example, hydrated forms, in addition to solvated forms. Suitable pharma-ceutically acceptable solvates are, for example, hydrates, such as hemihydrates, monohydrates, dihydrates, or trihydrates. It should be understood that the present disclosure encompasses all such solvated forms that have inflammasome inhibitory activity.

[0222] It should also be understood that any one particular compound of the formula disclosed herein may exhibit polymorphism, and the present disclosure encompasses all such forms or mixtures thereof that have inflammasome inhibitory activity.It is generally known that crystalline materials can be analyzed using conventional techniques such as X-ray powder diffraction analysis, differential scanning calorimetry, thermogravimetry, diffuse reflectance infrared Fourier transform (DRIFT) spectroscopy, near infrared (NIR) spectroscopy, solution and / or solid state nuclear magnetic resonance spectroscopy.The water content of such crystalline materials can be determined by Karl Fischer analysis.

[0223] Compounds of any one of the formulae disclosed herein may exist in a number of different tautomeric forms, and reference to any one of the formulae includes all such forms. For the avoidance of doubt, if a compound can exist in one of several tautomeric forms and only one is specifically described or shown, all others are nevertheless encompassed by the formulae disclosed herein. Examples of tautomeric forms include keto, enol, and enolate forms, such as in the following tautomeric pairs: keto / enol (shown below), imine / enamine, amide / iminoalcohol, amidine / amidine, nitroso / oxime, thioketone / enethiol, and nitro / acintro. [ka]

[0224] Compounds of any one of the formulas disclosed herein that contain an amine function may also form N-oxides. Reference herein to compounds of any one of the formulas disclosed herein that contain an amine function also includes N-oxides. When a compound contains several amine functions, one or more nitrogen atoms may be oxidized to form N-oxides. Particular examples of N-oxides are the N-oxides of tertiary amines or nitrogen atoms of nitrogen-containing heterocycles. N-oxides can be formed by treating the corresponding amine with an oxidizing agent such as hydrogen peroxide or a peracid (e.g., a peroxycarboxylic acid), see, for example, Advanced Organic Chemistry, by Jerry March, 4th Edition, Wiley Interscience, pages. More specifically, N-oxides can be prepared by the procedure of LW Deady (Syn. Comm. 1977, 7, 509-514), in which an amine compound is reacted with meta-chloroperoxybenzoic acid (mCPBA) in an inert solvent such as, for example, dichloromethane.

[0225] The compound of any one of the formulas disclosed herein may be administered in the form of a prodrug that is broken down in the human or animal body to release the compound of the present disclosure.Prodrugs can be used to modify the physical properties and / or pharmacokinetic properties of the compound of the present disclosure.Prodrugs can be formed when the compound of the present disclosure contains a suitable group or substituent to which a property-modifying group can be attached.

[0226] Thus, the present disclosure includes compounds of any one of the formulae disclosed herein above as made available by organic synthesis and by cleavage of a prodrug thereof in the human or animal body. Thus, the present disclosure also includes compounds of any one of the formulae disclosed herein produced by organic synthesis means, and compounds produced in the human or animal body by metabolism of a precursor compound, i.e., compounds of any one of the formulae disclosed herein may be synthetically produced or metabolically produced.

[0227] Suitable pharma- ceutically acceptable prodrugs of a compound of any one of the formulae disclosed herein are those that are based on sound medical judgment, lacking undesirable pharmacological activity, lacking undue toxicity, and are suitable for administration to the human or animal body. Various forms of prodrugs are described, for example, in the following literature: a) Methods in Enzymology, Vol. 42, p. 309-396, edited by K. Widder, et al. (Academic Press, 1985); b) Design of Pro-drugs, edited by H. Bundgaard, (Elsevier, 1985); c) A Textbook of Drug Design and Development, edited by Krogsgaard-Larsen and H. Bundgaard, Chapter 5 "Design and Application of Pro-drugs", by H. Bundgaard p. 113-191 (1991); d) H. Bundgaard, Advanced Drug Delivery Reviews, 8, 1-38 (1992); e) H. Bundgaard, et al., Journal of Pharmaceutical Sciences, 77, 285 (1988); f) N. Kakeya, et al. al., Chem.Pharm.Bull., 32, 692 (1984), g) T. Higuchi and V. Stella, "Pro-Drugs as Novel Delivery Systems", ACS Symposium Series, Volume 14, and h) E. Roche (editor), "Bioreversible Carriers in Drug Design", Pergamon Press, 1987.

[0228] The in vivo effects of the compounds of any one of the formulas disclosed herein may be exerted in part by one or more metabolic products formed in the human or animal body after administration of the compounds of any one of the formulas disclosed herein.As mentioned above, the in vivo effects of the compounds of any one of the formulas disclosed herein may also be exerted by metabolism of precursor compounds (prodrugs).

[0229] Preferably, the present disclosure excludes any individual compound that does not have biological activity as defined herein.

[0230] Linker-Containing Scaffolds and Conjugates As used herein, the term "scaffold" refers to a compound or complex that includes a linker of the present disclosure, where the linker is covalently attached to either a ligand or a nucleic acid agent.

[0231] As used herein, the term "conjugate" refers to a compound or complex that includes a nucleic acid agent covalently attached to a ligand via a linker of the present disclosure.

[0232] In some aspects, the disclosure provides a scaffold, or a pharma- ceutically acceptable salt thereof, the scaffold comprising: (i) a ligand, and (ii) a linker unit, the linker unit being [ka] and a linker unit, where variables B, R 1 , R 2 , R 3 , R 4 , R 5 , Y, Z, R a , R b and n are as described herein, and # indicates attachment to a ligand.

[0233] In some embodiments, the bond "#" is a direct bond to the ligand, ie, without any linking moiety.

[0234] In some embodiments, the bond "#" is an indirect bond to the ligand, i.e., there is a linking moiety between the linker unit and the ligand. In some embodiments, the linking moiety is a C1-C 15 is an alkylene chain, optionally one or more carbon atoms in the alkylene chain may be independently replaced with one or more of -C(O)-, -C(O)0-, -OC(O)-, -C(O)NH-, -NHC(O)-, -NHC(O)NH-, -C(S)-, -C(S)0-, -OC(S)-, -C(S)NH-, -NHC(S)-, or -NHC(S)NH-, e.g., the alkylene chain is optionally substituted with one or more groups independently selected from C1-C6 alkyl, halogen, OH, NH2, C1-C6 alkoxy, CN, and COOH. In some embodiments, the linking moiety is a 2, 3, or more C1-C6 alkyl group. 15 and branched alkylene chains, including alkylene chains, where optionally one or more carbon atoms in each of the alkylene chains may be independently replaced with one or more of -C(O)-, -C(O)0-, -OC(O)-, -C(O)NH-, -NHC(O)-, -NHC(O)NH-, -C(S)-, -C(S)0-, -OC(S)-, -C(S)NH-, -NHC(S)-, or -NHC(S)NH-, where each of the alkylene chains is independently optionally substituted with one or more groups independently selected from, for example, C1-C6 alkyl, halogen, OH, NH2, C1-C6 alkoxy, CN, and COOH. In some embodiments, the linking moiety is a bond between two C1-C 15 In some embodiments, the linking moiety is a branched alkylene chain, including three C-C 15 In some embodiments, the linking moiety is a branched alkylene chain that includes four C-C 15 It is a branched alkylene chain that includes an alkylene chain.

[0235] In some aspects, the disclosure provides a scaffold, or a pharma- ceutically acceptable salt thereof, the scaffold comprising: (i) one or more nucleic acid agents, and (ii) one or more linker units, each linker unit independently comprising: [ka] and one or more linker units, where variables B, R 1 , R 2 , R 3 , R 4 , R 5 , W, Y, Z, R a , R b , and n are as described herein, and ## indicates attachment to a nucleic acid agent.

[0236] In some embodiments, the bond "##" is a direct bond to the nucleic acid agent, i.e., there is no linking moiety.

[0237] In some embodiments, the bond "##" is an indirect bond to the nucleic acid agent, i.e., there is a linking moiety between the linker unit and the nucleic acid agent. In some embodiments, the linking moiety is a radical formed from any of the groups defined herein for Y or Z. For example, the linking moiety is -P(N(CH3)2)(O)-, i.e., a radical formed from -P(N(CH3)2)(OH). In some embodiments, the linking moiety is -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y , -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(OR Y )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, or -P(=S)(SR Y )2 or -P(R Z )2, -P(OR Z )(N(R Z)2), -P(=O)(OR Z )R Z , -P(=S)(OR Z )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z )2, or -P(=S)(SR Z )2.

[0238] In some embodiments, the scaffold comprises double-stranded RNA (eg, double-stranded siRNA).

[0239] In some embodiments, the scaffold comprises a double-stranded RNA (eg, a double-stranded siRNA) and one or more linker units.

[0240] In some embodiments, the scaffold comprises a double-stranded RNA (e.g., a double-stranded siRNA) and 1-10 linker units (e.g., 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, or 1-3 linker units), 2-10 linker units (e.g., 2-10, 2-9, 2-8, 2-7, 2-6, 2-5, 2-4, or 2-3 linker units), 3-10 linker units (e.g., 3-10, 3-9, 3-8, 3-7, 3-6, 3-5, or 3 linker units), 4-10 linker units (e.g., 4-10, 4-9, 4-8, 4-7, 4-6, or 4-5 linker units), 5-10 linker units (e.g., 5-10, 5-9, 5-8, 5-7, or 5-6 linker units), or 6-10 linker units (e.g., 6-10, 6-9, 6-8, or 6-7 linker units).

[0241] In some embodiments, the scaffold comprises a double-stranded RNA (e.g., a double-stranded siRNA) and 1 linker unit, 2 linker units, 3 linker units, 4 linker units, 5 linker units, 6 linker units, 7 linker units, 8 linker units, 9 linker units, or 10 linker units.

[0242] In some embodiments, the scaffold comprises a double-stranded RNA (e.g., a double-stranded siRNA) and one or more linker units; One or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely linked to the sense strand (e.g., at the 3' or 5' end) of a double-stranded RNA (e.g., a double-stranded siRNA); One or more nucleosides or nucleotides at one or more consecutive or discrete internal positions (positions between the 3'-terminal position and the 5'-terminal position) of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA) are replaced with one or more linker units (e.g., 1 to 3 linker units); one or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely attached to the antisense strand (e.g., at the 3'-terminal position or the 5'-terminal position) of a double-stranded RNA (e.g., a double-stranded siRNA); and / or One or more nucleosides or nucleotides at one or more contiguous or discrete internal positions of the antisense strand are replaced with one or more linker units (eg, 1 to 3 linker units).

[0243] In some embodiments, the scaffold comprises a double-stranded RNA (e.g., a double-stranded siRNA) and one or more linker units; One or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely linked to the sense strand (e.g., at the 3' or 5' end) of a double-stranded RNA (e.g., a double-stranded siRNA); One or more nucleosides or nucleotides at one or more contiguous or discrete internal positions (positions between the 3'-terminal position and the 5'-terminal position) of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA) are replaced with one or more linker units (e.g., 1 to 3 linker units).

[0244] In some embodiments, the scaffold comprises a double-stranded RNA (e.g., a double-stranded siRNA) and one or more linker units; one or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely linked to the antisense strand (e.g., the 3'-terminal position or the 5'-terminal position) of a double-stranded RNA (e.g., a double-stranded siRNA); One or more nucleosides or nucleotides at one or more contiguous or discrete internal positions of the antisense strand are replaced with one or more linker units (eg, 1 to 3 linker units).

[0245] In some embodiments, one or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely attached to the sense strand (e.g., at the 3' or 5' end) of a double-stranded RNA (e.g., a double-stranded siRNA).

[0246] In some embodiments, one or more linker units (eg, 1 to 3 linker units) are contiguously or discretely attached to the sense strand at the 3'-end position of the double-stranded RNA (eg, double-stranded siRNA).

[0247] In some embodiments, one or more linker units (eg, 1 to 3 linker units) are contiguously or discretely attached to the sense strand at the 5'-end position of the double-stranded RNA (eg, double-stranded siRNA).

[0248] In some embodiments, one or more nucleosides or nucleotides at one or more contiguous or discrete internal positions of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA) are replaced with one or more linker units (e.g., 1 to 3 linker units).

[0249] In some embodiments, one or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely attached to the antisense strand (e.g., at the 3' or 5' end) of a double-stranded RNA (e.g., a double-stranded siRNA).

[0250] In some embodiments, one or more linker units (eg, 1 to 3 linker units) are contiguously or discretely attached to the antisense strand at the 3'-end position of the double-stranded RNA (eg, double-stranded siRNA).

[0251] In some embodiments, one or more linker units (eg, 1 to 3 linker units) are contiguously or discretely attached to the antisense strand at the 5'-end position of the double-stranded RNA (eg, double-stranded siRNA).

[0252] In some embodiments, one or more nucleosides or nucleotides at one or more contiguous or discrete internal positions of the antisense strand of a double-stranded RNA (e.g., a double-stranded siRNA) are replaced with one or more linker units (e.g., 1 to 3 linker units).

[0253] In some embodiments, the scaffold comprises a (linker unit) p -((nucleic acid agent)-(linker unit) s ) r -(Nucleic acid agents) q and each linker unit is independent of other linker units and each nucleic acid agent is independent of other nucleic acid agents; each r is independently an integer ranging from 0 to 10; each s is independently an integer ranging from 0 to 10; p is an integer ranging from 0 to 10; q is 0 or 1; The scaffold comprises at least one linker unit and at least one nucleic acid agent.

[0254] In some embodiments, the scaffold comprises a (linker unit) p -((nucleic acid agent)-(linker unit) s ) r -(nucleic acid agent).

[0255] In some embodiments, the scaffold comprises a (linker unit) p -((nucleic acid agent)-(linker unit) s )r It is.

[0256] In some embodiments, the scaffold comprises a (linker unit) p -(nucleic acid agent).

[0257] In some embodiments, the scaffold comprises a (nucleic acid agent)-(linker unit) s -(nucleic acid agent).

[0258] In some embodiments, the scaffold comprises: [ka] or a pharma- ceutically acceptable salt thereof, wherein: B is a nucleobase moiety (e.g., adenine (A), cytosine (C), guanine (G), thymine (T), or uracil (U)); Y is -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y , -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(OR Y )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, -P(=S)(SR Y )2, or a hydroxy protecting group (e.g., silyl (e.g., trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl), triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), substituted acyl (e.g., optionally substituted acetyl), or benzyl); Each R Y is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; Z is -P(RZ )2, -P(OR Z )(N(R Z )2), -P(=O)(OR Z )R Z , -P(=S)(OR Z )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z )2, -P(=S)(SR Z )2, or a hydroxy protecting group (e.g., silyl (e.g., trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl), triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), substituted acyl (e.g., optionally substituted acetyl), or benzyl); Each R Z is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; n is an integer ranging from about 0 to about 10.

[0259] In some embodiments, n is an integer ranging from 1-7.

[0260] In some embodiments, n is an integer ranging from 1-6.

[0261] In some embodiments, n is an integer ranging from 2-7.

[0262] In some embodiments, n is an integer ranging from 2-6.

[0263] In some embodiments, n is an integer ranging from 3 to 7.

[0264] In some embodiments, n is an integer ranging from 3 to 6.

[0265] In some embodiments, n is an integer ranging from 4 to 7.

[0266] In some embodiments, n is an integer ranging from 4 to 6.

[0267] In some embodiments, n is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10.

[0268] In some embodiments, n is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8, and 9.

[0269] In some embodiments, n is an integer selected from 1, 2, 3, 4, 5, 6, 7, and 8.

[0270] In some embodiments, n is an integer selected from 1, 2, 3, 4, 5, 6, and 7.

[0271] In some embodiments, n is an integer selected from 2, 3, 4, 5, 6, and 7.

[0272] In some embodiments, n is an integer selected from 2, 3, 4, 5, and 6.

[0273] In some embodiments, n is an integer selected from 3, 4, 5, and 6.

[0274] In some embodiments, the scaffold is formed by linking linker units based on any of the linker compounds described herein with ligands.

[0275] In some embodiments, the scaffold comprises: [ka] [ka] A linker unit based on any of the linker compounds selected from It is formed by linking with a ligand.

[0276] In some embodiments, the scaffold is formed by linking linker units based on any of the linker compounds selected from Table L with the ligands.

[0277] In some embodiments, the ligand is GalNAc.

[0278] In some embodiments, the scaffold is selected from the scaffolds listed in Table S1. [Table 2-1] [Table 2-2] [Table 2-3]

[0279] In some embodiments, the scaffold comprises: [ka] or a pharma- ceutically acceptable salt thereof, wherein: B is a nucleobase moiety (e.g., adenine (A), cytosine (C), guanine (G), thymine (T), or uracil (U)); W is an amino substituent (e.g., fluorenylmethyloxycarbonyl (Fmoc), tert-butyloxycarbonyl (BOC), benzyloxycarbonyl (Cbz), optionally substituted acyl, trifluoroacetyl (TFA), benzyl, triphenylmethyl (Tr), 4,4'-dimethoxytrityl (DMTr), or toluenesulfonyl (Ts), acyl (e.g., -C(=O)(C1-C30 alkyl)), substituted acyl (e.g., [ka] ), trifluoroacetyl (TFA), -C(=O)(C1-C30 alkyl), -C(=O)NH(C1-C30 alkyl), -C(=S)(C1-C30 alkyl), or -C(=S)NH(C1-C30 alkyl), wherein C1-C30 alkyl is optionally substituted; n is an integer ranging from about 0 to about 10.

[0280] In some embodiments, the scaffold is formed by linking linker units based on any of the linker compounds described herein with nucleic acid agents.

[0281] In some embodiments, the scaffold comprises: [ka] [ka] A linker unit based on any of the linker compounds selected from It is formed by linking with a nucleic acid agent.

[0282] In some embodiments, the scaffold is formed by linking a linker unit based on any of the linker compounds selected from Table L with a nucleic acid agent.

[0283] In some embodiments, the scaffold is selected from the scaffolds listed in Table S2. [Table 3-1] [Table 3-2] [Table 3-3] [Table 3-4] [Table 3-5]

Table 3-6

Table 3-7

Table 3-8

Table 3-9

Table 3-10

Table 3-11

Table 3-12

Table 3-13

Table 3-14

Table 3-15

Table 3-16

Table 3-17

Table 3-18

Table 3-19

Table 3-20

Table 3-21

Table 3-22

[0284] In some aspects, the disclosure provides a conjugate, or a pharma- ceutically acceptable salt thereof, the conjugate comprising: (i) one or more nucleic acid agents, (ii) one or more ligands, and (iii) one or more linker units, each linker unit independently comprising: [ka] and one or more linker units, where variable R 1 , R 2 , R 3 , R 4 , R 5 , Y, Z, R a , R b , and n are as described herein, where # indicates attachment to a ligand and ## indicates attachment to a nucleic acid agent.

[0285] In some embodiments, the bond "#" is a direct bond to the ligand, ie, without any linking moiety.

[0286] In some embodiments, the bond "#" is an indirect bond to the ligand, i.e., there is a linking moiety between the linker unit and the ligand. In some embodiments, the linking moiety is a C1-C 15 is an alkylene chain, optionally one or more carbon atoms in the alkylene chain may be independently replaced with one or more of -C(O)-, -C(O)0-, -OC(O)-, -C(O)NH-, -NHC(O)-, -NHC(O)NH-, -C(S)-, -C(S)0-, -OC(S)-, -C(S)NH-, -NHC(S)-, or -NHC(S)NH-, e.g., the alkylene chain is optionally substituted with one or more groups independently selected from C1-C6 alkyl, halogen, OH, NH2, C1-C6 alkoxy, CN, and COOH. In some embodiments, the linking moiety is a 2, 3, or more C1-C6 alkyl group. 15and branched alkylene chains, including alkylene chains, where optionally one or more carbon atoms in each of the alkylene chains may be independently replaced with one or more of -C(O)-, -C(O)0-, -OC(O)-, -C(O)NH-, -NHC(O)-, -NHC(O)NH-, -C(S)-, -C(S)0-, -OC(S)-, -C(S)NH-, -NHC(S)-, or -NHC(S)NH-, where each of the alkylene chains is independently optionally substituted with one or more groups independently selected from, for example, C1-C6 alkyl, halogen, OH, NH2, C1-C6 alkoxy, CN, and COOH. In some embodiments, the linking moiety is a bond between two C1-C 15 In some embodiments, the linking moiety is a branched alkylene chain, including three C-C 15 In some embodiments, the linking moiety is a branched alkylene chain that includes four C-C 15 It is a branched alkylene chain that includes an alkylene chain.

[0287] In some embodiments, the bond "##" is a direct bond to the nucleic acid agent, i.e., there is no linking moiety.

[0288] In some embodiments, the bond "##" is an indirect bond to the nucleic acid agent, i.e., there is a linking moiety between the linker unit and the nucleic acid agent. In some embodiments, the linking moiety is a radical formed from any of the groups defined herein for Y or Z. For example, the linking moiety is -P(N(CH3)2)(O)-, i.e., a radical formed from -P(N(CH3)2)(OH). In some embodiments, the linking moiety is -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y , -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(ORY )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, or -P(=S)(SR Y )2 or -P(R Z )2, -P(OR Z )(N(R Z )2), -P(=O)(OR Z )R Z , -P(=S)(OR Z )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z )2, or -P(=S)(SR Z )2.

[0289] In some embodiments, the conjugate comprises a double-stranded RNA (eg, a double-stranded siRNA), one or more ligands, and one or more linker units.

[0290] In some embodiments, the conjugate comprises a double-stranded RNA (e.g., a double-stranded siRNA) and 1 to 10 linker units (e.g., 1 to 10, 1 to 9, 1 to 8, 1 to 7, 1 to 6, 1 to 5, 1 to 4, or 1 to 3 linker units), 2 to 10 linker units (e.g., 2 to 10, 2 to 9, 2 to 8, 2 to 7, 2 to 6, 2 to 5, 2 to 4, or 2 to 3 linker units), 3 to 10 linker units (e.g., 3 to 1 0, 3-9, 3-8, 3-7, 3-6, 3-5, or 3-4 linker units), 4-10 linker units (e.g., 4-10, 4-9, 4-8, 4-7, 4-6, or 4-5 linker units), 5-10 linker units (e.g., 5-10, 5-9, 5-8, 5-7, or 5-6 linker units), or 6-10 linker units (e.g., 6-10, 6-9, 6-8, or 6-7 linker units).

[0291] In some embodiments, the conjugate comprises a double-stranded RNA (e.g., a double-stranded siRNA) and 1 linker unit, 2 linker units, 3 linker units, 4 linker units, 5 linker units, 6 linker units, 7 linker units, 8 linker units, 9 linker units, or 10 linker units.

[0292] In some embodiments, the conjugate comprises a double-stranded RNA (e.g., a double-stranded siRNA), one or more ligands, and one or more linker units; One or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely linked to the sense strand (e.g., at the 3' or 5' end) of a double-stranded RNA (e.g., a double-stranded siRNA); one or more nucleosides or nucleotides at one or more contiguous or discrete internal positions of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA) are replaced with one or more linker units (e.g., 1 to 3 linker units); one or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely attached to the antisense strand (e.g., at the 3'-terminal position or the 5'-terminal position) of a double-stranded RNA (e.g., a double-stranded siRNA); and / or One or more nucleosides or nucleotides at one or more contiguous or discrete internal positions of the antisense strand are replaced with one or more linker units (eg, 1 to 3 linker units).

[0293] In some embodiments, the conjugate comprises a double-stranded RNA (e.g., a double-stranded siRNA), one or more ligands, and one or more linker units; One or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely linked to the sense strand (e.g., at the 3' or 5' end) of a double-stranded RNA (e.g., a double-stranded siRNA); One or more nucleosides or nucleotides at one or more contiguous or discrete internal positions of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA) are replaced with one or more linker units (e.g., 1 to 3 linker units).

[0294] In some embodiments, the conjugate comprises a double-stranded RNA (e.g., a double-stranded siRNA), one or more ligands, and one or more linker units; one or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely linked to the antisense strand (e.g., the 3'-terminal position or the 5'-terminal position) of a double-stranded RNA (e.g., a double-stranded siRNA); One or more nucleosides or nucleotides at one or more contiguous or discrete internal positions of the antisense strand are replaced with one or more linker units (eg, 1 to 3 linker units).

[0295] In some embodiments, one or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely attached to the sense strand (e.g., at the 3' or 5' end) of a double-stranded RNA (e.g., a double-stranded siRNA).

[0296] In some embodiments, one or more linker units (eg, 1 to 3 linker units) are contiguously or discretely attached to the 3'-end position of the sense strand of a double-stranded RNA (eg, a double-stranded siRNA).

[0297] In some embodiments, one or more linker units (eg, 1 to 3 linker units) are contiguously or discretely attached to the 5'-end position of the sense strand of a double-stranded RNA (eg, a double-stranded siRNA).

[0298] In some embodiments, one or more nucleosides or nucleotides at one or more contiguous or discrete internal positions of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA) are replaced with one or more linker units (e.g., 1 to 3 linker units).

[0299] In some embodiments, one or more linker units (e.g., 1 to 3 linker units) are contiguously or discretely attached to the antisense strand (e.g., at the 3' or 5' end) of a double-stranded RNA (e.g., a double-stranded siRNA).

[0300] In some embodiments, one or more linker units (eg, 1 to 3 linker units) are contiguously or discretely attached to the antisense strand at the 3'-end position of the double-stranded RNA (eg, double-stranded siRNA).

[0301] In some embodiments, one or more linker units (eg, 1 to 3 linker units) are contiguously or discretely attached to the antisense strand at the 5'-end position of the double-stranded RNA (eg, double-stranded siRNA).

[0302] In some embodiments, one or more nucleosides or nucleotides at one or more contiguous or discrete internal positions of the antisense strand of a double-stranded RNA (e.g., a double-stranded siRNA) are replaced with one or more linker units (e.g., 1 to 3 linker units).

[0303] In some embodiments, the conjugate comprises a (linker unit-(ligand) 0-1 ) p -((nucleic acid agent)-(linker unit-(ligand) 0-1 ) s ) r -(Nucleic acid agents) q where: each linker unit is independent of other linker units, each nucleic acid agent is independent of other nucleic acid agents, and each ligand is independent of other ligands; each r is independently an integer ranging from 0 to 10; each s is independently an integer ranging from 0 to 10; p is an integer ranging from 0 to 10; q is 0 or 1; The conjugate comprises at least one linker unit, at least one nucleic acid agent, and at least one ligand.

[0304] In some embodiments, the conjugate comprises a (linker unit-(ligand) 0-1 ) p -((nucleic acid agent)-(linker unit-(ligand) 0-1 ) s ) r -(nucleic acid agent).

[0305] In some embodiments, the conjugate comprises a (linker unit-(ligand) 0-1 ) p -((nucleic acid agent)-(linker unit-(ligand) 0-1 ) s ) r It is.

[0306] In some embodiments, the conjugate comprises a (linker unit-(ligand) 0-1 ) p -(nucleic acid agent).

[0307] In some embodiments, the conjugate comprises a (nucleic acid agent)-(linker unit-(ligand) 0-1 ) s -(nucleic acid agent).

[0308] In some embodiments, the conjugate is selected from the conjugates described in Table C, and the nucleic acid agent is linked by ##, where ## is a direct or indirect linkage described herein. [Table 4-1] [Table 4-2] [Table 4-3] [Table 4-4] [Table 4-5] [Table 4-6]

Table 4-7

Table 4-8

Table 4-9

Table 4-10

Table 4-11

Table 4-12

Table 4-13

Table 4-14

Table 4-15

Table 4-16

Table 4-17

Table 4-18

Table 4-19

Table 4-20

Table 4-21

Table 4-22

Table 4-23

Table 4-24

Table 4-25

Table 4-26

Table 4-27

Table 4-28

Table 4-29

Table 4-30

Table 4-31

Table 4-32

Table 4-33

Table 4-34

Table 4-35

Table 4-36

Table 4-37

Table 4-38

[0309] リンカーUnit As used herein, "linker unit" or "linker unit" refers to the portion corresponding to a linker compound in which W, Y, and / or Z are replaced with bonds to a ligand and / or nucleic acid agent. In some embodiments, the bond, e.g., # or ## as described herein, is a direct or indirect bond as described herein.

[0310] In some embodiments, the linker unit is of formula (I) where W is replaced with a bond to a ligand. In some embodiments, the bond, e.g., # or ## as described herein, is a direct or indirect bond as described herein.

[0311] In some embodiments, the linker unit is of formula (I) where Y and / or Z are replaced with a bond to a nucleic acid agent. In some embodiments, the bond, e.g., # or ## as described herein, is a direct or indirect bond as described herein.

[0312] In some embodiments, the linker unit is of formula (I), wherein: W is replaced by a bond to a ligand; Y and / or Z are replaced with a bond to a nucleic acid agent.

[0313] In some embodiments, the linker unit is of formula (II) where W is replaced with a bond to the ligand.

[0314] In some embodiments, the linker unit is of formula (II) where Y and / or Z is replaced with a bond to a nucleic acid agent.

[0315] In some embodiments, the linker unit is of formula (II), wherein: W is replaced by a bond to a ligand; Y and / or Z are replaced with a bond to a nucleic acid agent.

[0316] In some embodiments, the linker unit is of formula (I') or (II') where W is replaced with a bond to the ligand.

[0317] In some embodiments, the linker unit is of formula (I') or (II') where Y and / or Z are replaced with a bond to a nucleic acid agent.

[0318] In some embodiments, the linker unit is of formula (IA) or (II-A), where W is replaced with a bond to the ligand.

[0319] In some embodiments, the linker unit is of formula (IA) or (II-A), where Y and / or Z are replaced with a bond to a nucleic acid agent.

[0320] In some embodiments, the linker unit is of formula (IA) or (II-A), wherein: W is replaced by a bond to a ligand; Y and / or Z are replaced with a bond to a nucleic acid agent.

[0321] In some embodiments, the linker unit is of formula (I'-A) or (II'-A), where W is replaced with a bond to the ligand.

[0322] In some embodiments, the linker unit is of formula (I'-A) or (II'-A), where Y and / or Z are replaced with a bond to a nucleic acid agent.

[0323] In some embodiments, the linker unit is of formula (I'-A) or (II'-A), wherein: W is replaced by a bond to a ligand; Y and / or Z are replaced with a bond to a nucleic acid agent.

[0324] In some embodiments, the linker unit is of formula (IB) or (II-B), where W is replaced with a bond to the ligand.

[0325] In some embodiments, the linker unit is of formula (IB) or (II-B), where Y and / or Z are replaced with a bond to a nucleic acid agent.

[0326] In some embodiments, the linker unit is of formula (IB) or (II-B), wherein: W is replaced by a bond to a ligand; Y and / or Z are replaced with a bond to a nucleic acid agent.

[0327] In some embodiments, the linker unit is of formula (I'-B) or (II'-B), where W is replaced with a bond to the ligand.

[0328] In some embodiments, the linker unit is of formula (I'-B) or (II'-B), where Y and / or Z are replaced with a bond to a nucleic acid agent.

[0329] In some embodiments, the linker unit is of formula (I'-B) or (II'-B), wherein: W is replaced by a bond to a ligand; Y and / or Z are replaced with a bond to a nucleic acid agent.

[0330] In some embodiments, the linker unit prior to attachment is a linker compound described herein.

[0331] In some embodiments, the linker unit prior to attachment is a compound of formula (I) or a pharma- ceutically acceptable salt thereof.

[0332] In some embodiments, the linker unit prior to attachment is a compound of formula (II) or a pharma- ceutically acceptable salt thereof.

[0333] In some embodiments, the linker unit prior to attachment is a compound of formula (I') or (II'), or a pharma- ceutically acceptable salt thereof.

[0334] In some embodiments, the linker unit prior to attachment is a compound of formula (IA) or (II-A), or a pharma- ceutically acceptable salt thereof.

[0335] In some embodiments, the linker unit prior to attachment is a compound of formula (I'-A) or (II'-A), or a pharma- ceutically acceptable salt thereof.

[0336] In some embodiments, the linker unit prior to attachment is a compound of formula (IB) or (II-B), or a pharma- ceutically acceptable salt thereof.

[0337] In some embodiments, the linker unit prior to attachment is a compound of formula (I'-B) or (II'-B), or a pharma- ceutically acceptable salt thereof.

[0338] In some embodiments, the linker unit prior to attachment is a compound selected from the compounds set forth in Table L and pharma- ceutically acceptable salts thereof.

[0339] In any of the above embodiments, a bond, for example, # or ## as described herein, is a direct or indirect bond as described herein.

[0340] Ligand As used herein, the term "ligand" refers to a moiety that, when covalently attached to a nucleic acid agent (e.g., an oligonucleotide), can mediate its entry into or facilitate its delivery to a target site (e.g., a target cell or tissue). A ligand or ligands together with a linker unit form a scaffold as described herein, or one or more ligands or ligands together with one or more linker units and one or more nucleic acid agents form a conjugate as described herein.

[0341] In some embodiments, the ligand comprises a sugar ligand moiety (eg, N-acetylgalactosamine (GalNAc)) that can direct uptake of the oligonucleotide into the liver.

[0342] In some embodiments, the ligand binds to the asialoglycoprotein receptor (ASGPR). In some embodiments, the ligand binds to the liver (e.g., via the ASGPR), such as to liver parenchymal cells.

[0343] Suitable ligands include, but are not limited to, those disclosed in Winkler (Ther. Deliv., 2013, 4(7):791-809), WO 2016 / 100401, WO 2012 / 089352, and WO 2009 / 082607, and U.S. Patent Application Publication Nos. 2009 / 0239814, 2012 / 0136042, 2013 / 0158824, and 2009 / 0247608, each of which is incorporated by reference.

[0344] In some embodiments, the ligand comprises a carbohydrate moiety.

[0345] As used herein, a "carbohydrate moiety" refers to a moiety that includes one or more monosaccharide units, each having at least six carbon atoms (which may be linear, branched, or cyclic) and having an oxygen, nitrogen, or sulfur atom bonded to each carbon atom. In some embodiments, the carbohydrate moiety includes a monosaccharide, a disaccharide, a trisaccharide, or a tetrasaccharide. In some embodiments, the carbohydrate moiety includes an oligosaccharide containing about 4-9 monosaccharide units. In some embodiments, the carbohydrate moiety includes a polysaccharide (e.g., starch, glycogen, cellulose, or a polysaccharide gum).

[0346] In some embodiments, the carbohydrate moiety comprises a monosaccharide, a disaccharide, a trisaccharide, or a tetrasaccharide.

[0347] In some embodiments, the carbohydrate moiety comprises an oligosaccharide (eg, containing from about 4 to about 9 monosaccharide units).

[0348] In some embodiments, the carbohydrate moiety comprises a polysaccharide (eg, starch, glycogen, cellulose, or a polysaccharide gum).

[0349] In some embodiments, the ligand is capable of binding to a human asialoglycoprotein receptor (ASGPR), for example, human asialoglycoprotein receptor 2 (ASGPR2).

[0350] In some embodiments, the carbohydrate moiety comprises a sugar (eg, 1, 2, or 3 sugars).

[0351] In some embodiments, the carbohydrate moiety comprises galactose or a derivative thereof (eg, 1, 2, or 3 galactose or a derivative thereof).

[0352] In some embodiments, the carbohydrate moiety comprises N-acetylgalactosamine or a derivative thereof (eg, 1, 2, or 3 N-acetylgalactosamine or a derivative thereof).

[0353] In some embodiments, the carbohydrate moiety comprises an N-acetyl-D-galactosylamine or derivative thereof (eg, one, two, or three N-acetyl-D-galactosylamines or derivatives thereof).

[0354] In some embodiments, the carbohydrate moiety comprises an N-acetylgalactosamine (eg, one, two, or three N-acetylgalactosamines).

[0355] In some embodiments, the carbohydrate moiety comprises an N-acetyl-D-galactosylamine (eg, one, two, or three N-acetyl-D-galactosylamines).

[0356] In some embodiments, the carbohydrate moiety comprises mannose or a derivative thereof (eg, mannose-6-phosphate).

[0357] In some embodiments, the carbohydrate moiety further comprises a linking moiety that connects one or more sugars (eg, N-acetyl-D-galactosylamine) with the linker unit.

[0358] In some embodiments, the linking moiety comprises a thioether (eg, a thiosuccinimide, or a hydrolysis analog thereof), a disulfide, a triazole, a phosphorothioate, a phosphodiester, an ester, an amide, or any combination thereof.

[0359] In some embodiments, the linking moiety is a trivalent linking moiety.

[0360] Suitable ligands include, but are not limited to, those disclosed in WO 2015 / 006740, WO 2016 / 100401, WO 2017 / 214112, WO 2018 / 039364, and WO 2018 / 045317, each of which is incorporated by reference in its entirety.

[0361] In some embodiments, the ligand is [ka] (e.g. 1, 2, or 3 [ka] ).

[0362] In some embodiments, the ligand is [ka] (e.g. 1, 2, or 3 [ka] ).

[0363] In some embodiments, the ligand is [ka] (e.g. 1, 2, or 3 [ka] ).

[0364] In some embodiments, the ligand is [ka] (e.g. 1, 2, or 3 [ka] ).

[0365] In some embodiments, the ligand is [ka] (e.g. 1, 2, or 3 [ka] ).

[0366] In some embodiments, the ligand is [ka] (e.g. 1, 2, or 3 [ka] ).

[0367] In some embodiments, the ligand is [ka] (e.g. 1, 2, or 3 [ka] ).

[0368] In some embodiments, the ligand is [ka] (e.g. 1, 2, or 3 [ka] ).

[0369] In some embodiments, the ligand is [ka] Includes.

[0370] In some embodiments, the ligand is [ka] Includes.

[0371] In some embodiments, the ligand is [ka] Includes.

[0372] In some embodiments, the ligand is [ka] Includes.

[0373] In some embodiments, the ligand is [ka] Includes.

[0374] In some embodiments, the ligand is [ka] Includes.

[0375] In some embodiments, the ligand is [ka] Includes.

[0376] In some embodiments, the ligand is [ka] Includes.

[0377] In some embodiments, the ligand comprises a lipid moiety (eg, 1, 2, or 3 lipid moieties).

[0378] In some embodiments, the lipid moiety is a (e.g., one, two, three) C8-C 24 The fatty acids include fatty acids, cholesterol, vitamins, sterols, phospholipids, or any combination thereof.

[0379] In some embodiments, the ligand comprises a peptide moiety (eg, one, two, or three peptide moieties).

[0380] In some embodiments, the peptide moiety comprises (eg, one, two, or three) integrins, insulin, glucagon-like peptides, or any combination thereof.

[0381] In some embodiments, the ligand comprises an antibody moiety (eg, transferrin).

[0382] In some embodiments, the ligand comprises one, two, or three antibody moieties (eg, transferrin).

[0383] In some embodiments, the ligand comprises an oligonucleotide (eg, an aptamer or CpG).

[0384] In some embodiments, the ligand comprises one, two, or three oligonucleotides (eg, an aptamer or CpG).

[0385] In some embodiments, the ligand is one, two, or three sugars (e.g., N-acetyl-D-galactosylamine); 1, 2 or 3 lipid moieties, one, two or three peptide moieties, one, two or three antibody moieties, one, two, or three oligonucleotides, or Any combination thereof is included.

[0386] Nucleic acid agents In some embodiments, the nucleic acid agent comprises an oligonucleotide.

[0387] In some embodiments, a nucleic acid agent (eg, an oligonucleotide) includes one or more phosphate groups or one or more analogs of a phosphate group.

[0388] In some embodiments, a linker unit is attached to a nucleic acid agent (eg, an oligonucleotide) via a phosphate group or an analog of a phosphate group in the nucleic acid agent.

[0389] In some embodiments, the oligonucleotide has a length of 1-40 nucleotides, 10-40 nucleotides, 12-35 nucleotides, 15-30 nucleotides, 18-25 nucleotides, or 20-23 nucleotides. In some embodiments, the oligonucleotide has a length of 18, 19, 20, 21, 22, 23, 24, or 25 nucleotides. In some embodiments, the oligonucleotide has a length of 19, 20, 21, 22, or 23 nucleotides.

[0390] In some embodiments, the nucleic acid agent comprises RNA, DNA, or a mixture thereof.

[0391] In some embodiments, the nucleic acid agent comprises RNA.

[0392] In some embodiments, the oligonucleotide is an siRNA (e.g., a single-stranded siRNA (e.g., a hairpin single-stranded siRNA) or a double-stranded siRNA), a microRNA, an anti-microRNA, a microRNA mimic, an antagomir, dsRNA, ssRNA, an aptamer, an immunostimulatory oligonucleotide, a decoy oligonucleotide, a splice variant oligonucleotide, a triplex forming oligonucleotide, a G-quadruplex, or an antisense oligonucleotide.

[0393] In some embodiments, a nucleic acid agent comprises double-stranded RNA (dsRNA), where the double-stranded RNA comprises a sense strand and an antisense strand, as described herein.

[0394] In some embodiments, a nucleic acid agent comprises a double-stranded siRNA (ds-siRNA), where the double-stranded siRNA comprises a sense strand and an antisense strand, as described herein.

[0395] It is understood that the sense strand is also known as the passenger strand, and the terms "sense strand" and "passenger strand" are used interchangeably herein.

[0396] The antisense strand is also known as the guide strand, and it is understood that the terms "antisense strand" and "guide strand" are used interchangeably herein.

[0397] In some embodiments, the oligonucleotide is an iRNA.

[0398] The term "iRNA" refers to an RNA agent that can downregulate the expression of a target gene (e.g., siRNA), e.g., an endogenous or pathogen target RNA. Without being bound by theory, iRNAs can act by one or more of several mechanisms, including post-transcriptional cleavage of the target mRNA (referred to in the art as RNAi), or pre-transcriptional or pre-translational mechanisms. iRNAs can include a single strand or can include two or more strands, e.g., double-stranded iRNAs. When an iRNA is single-stranded, it can include a 5' modification, including one or more phosphate groups or one or more analogs of a phosphate group. In some embodiments, an iRNA is double-stranded. In some embodiments, one or both strands of a double-stranded iRNA can be modified, e.g., 5' modified.

[0399] An iRNA typically comprises a region of sufficient homology to a target gene and is of sufficient length in terms of nucleotides so that the iRNA or a fragment thereof can mediate downregulation of the target gene. An iRNA is or comprises a region that is at least partially, and in some embodiments completely, complementary to a target RNA. It is not necessary that there is perfect complementarity between the iRNA and the target, but the correspondence may be sufficient to allow the iRNA or its cleavage product to direct sequence-specific silencing, for example, by RNAi cleavage of the target RNA, for example, mRNA.

[0400] Nucleotides in an iRNA can be modified (e.g., one or more nucleotides can include a 2'-F or 2'-OCH3 group or can be a nucleotide substitute). Single- or double-stranded regions of an iRNA can be modified or include nucleotide substitutes, for example, the unpaired region of a hairpin structure, e.g., the region linking two complementary regions, can have modifications or nucleotide substitutes. Modifications to stabilize one or more 3' or 5' ends of an iRNA, for example against exonucleases. Modifications can include C3 (or C6, C7, C12) amino linkers, thiol linkers, carboxyl linkers, non-nucleotide spacers (C3, C6, C9, C12, abasic, triethylene glycol, hexaethylene glycol), certain biotin or fluorescein reagents provided as phosphoramidites and bearing separate DMT-protected hydroxyl groups, allowing multiple couplings during RNA synthesis. Modifications can also include, for example, the use of modifications at the 2'OH group of the ribose sugar, for example, the use of deoxyribonucleotides instead of ribonucleotides, e.g., deoxythymidine, and modifications at the phosphate group, e.g., phosphothioate modifications. In some embodiments, different strands contain different modifications.

[0401] In some embodiments, the strands are selected such that the iRNA comprises a single strand or unpaired region at one or both ends of the molecule. A double-stranded iRNA may have an overhang, e.g., one or two 5' or 3' overhangs (e.g., at least a 3' overhang of 2-3 nucleotides). In some embodiments, the iRNA has an overhang, e.g., a 3' overhang, of 1, 2, or 3 nucleotides in length at each end. The overhangs may be the result of one strand being longer than the other, or may be the result of two strands of the same length being staggered.

[0402] In some embodiments, the length of the double-stranded region between the strands of the iRNA is between 6 and 30 nucleotides in length. In some embodiments, the double-stranded region is between 15 and 30, most preferably 18, 19, 20, 21, 22, and 23 nucleotides in length. In some embodiments, the double-stranded region is between 6 and 20 nucleotides in length, most preferably 6, 7, 8, 9, 10, 11, and 12 nucleotides in length.

[0403] The oligonucleotides may be those described in U.S. Patent Publication Nos. 2009 / 0239814, 2012 / 0136042, 2013 / 0158824, or 2009 / 0247608, each of which is incorporated herein by reference.

[0404] In some embodiments, the oligonucleotide is a siRNA.

[0405] In some embodiments, the oligonucleotide is a single stranded siRNA.

[0406] In some embodiments, the oligonucleotide is a double-stranded siRNA, eg, a double-stranded siRNA described herein.

[0407] As used herein, a "single-stranded siRNA" is an siRNA that is composed of a single strand, including a double-stranded region formed by intrastrand pairing, e.g., it may be or include a hairpin or panhandle structure. A single-stranded siRNA may be antisense with respect to a target molecule.

[0408] Single-stranded siRNA may be long enough to enter RISC and participate in RISC-mediated cleavage of target mRNA.Single-stranded siRNA is at least 14 nucleotides long, and in some embodiments, at least 15, 20, 25, 29, 35, 40, or 50 nucleotides long.In some embodiments, single-stranded siRNA is 200, 100, 80, 60, 50, 40, or 30 nucleotides long.

[0409] In some embodiments, the single stranded siRNA has a length of 10-40 nucleotides, 12-35 nucleotides, 15-30 nucleotides, 18-25 nucleotides, or 20-23 nucleotides. In some embodiments, the single stranded siRNA has a length of 18, 19, 20, 21, 22, 23, 24, or 25 nucleotides. In some embodiments, the single stranded siRNA has a length of 20, 21, 22, or 23 nucleotides.

[0410] Hairpin siRNAs may have a double-stranded region corresponding to or at least 17, 18, 19, 20, 21, 22, 23, 24, or 25 nucleotide pairs. The double-stranded region may be up to 200, 100, or 50 nucleotide pairs in length. In some embodiments, ranges for the double-stranded region are 15-30, 17-23, 19-23, and 19-21 nucleotide pairs in length. The hairpin may have a single-stranded overhang or terminal unpaired region. In some embodiments, the oligonucleotide is 2-3 nucleotides in length. In some embodiments, the overhang is on the sense side of the hairpin, and in some embodiments, on the antisense side of the hairpin.

[0411] In some embodiments, the oligonucleotide is a double-stranded siRNA.

[0412] As used herein, a "double-stranded siRNA" is an siRNA that contains two or more, and optionally two, strands, capable of interstrand hybridization to form a region of double-stranded structure.

[0413] In some embodiments, the sense strand of the double-stranded siRNA can be equal to or at least 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 29, 40, or 60 nucleotides in length. The sense strand of the double-stranded siRNA can be up to 200, 100, or 50 nucleotides in length. Ranges can be 17-25, 19-23, 19-21, 21-23, or 20-22 nucleotides in length.

[0414] In some embodiments, the sense strand has a length of 10-40 nucleotides, 12-35 nucleotides, 15-30 nucleotides, 18-25 nucleotides, or 20-23 nucleotides. In some embodiments, the sense strand has a length of 18, 19, 20, 21, 22, 23, 24, or 25 nucleotides. In some embodiments, the sense strand has a length of 20, 21, 22, or 23 nucleotides.

[0415] In some embodiments, the sense strand has a length of 18, 19, 20, 21, or 22 nucleotides.

[0416] In some embodiments, the antisense strand of the double-stranded siRNA can be equal to or at least 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 29, 40, or 60 nucleotides in length. The antisense strand of the double-stranded siRNA can be up to 200, 100, or 50 nucleotides in length. Ranges can be 17-25, 19-23, 19-21, 21-23, or 20-22 nucleotides in length.

[0417] In some embodiments, the antisense strand has a length of 10-40 nucleotides, 12-35 nucleotides, 15-30 nucleotides, 18-25 nucleotides, or 20-23 nucleotides. In some embodiments, the antisense strand has a length of 18, 19, 20, 21, 22, 23, 24, or 25 nucleotides. In some embodiments, the antisense strand has a length of 20, 21, 22, or 23 nucleotides.

[0418] In some embodiments, the antisense strand has a length of 20, 21, 22, 23, or 24 nucleotides.

[0419] In some embodiments, the sense strand has a length of 18, 19, 20, 21, or 22 nucleotides and the antisense strand has a length of 20, 21, 22, 23, or 24 nucleotides.

[0420] In some embodiments, the sense strand has a length of 18 nucleotides and the antisense strand has a length of 20 nucleotides.

[0421] In some embodiments, the sense strand has a length of 19 nucleotides and the antisense strand has a length of 21 nucleotides.

[0422] In some embodiments, the sense strand has a length of 20 nucleotides and the antisense strand has a length of 22 nucleotides.

[0423] In some embodiments, the sense strand has a length of 21 nucleotides and the antisense strand has a length of 23 nucleotides.

[0424] In some embodiments, the sense strand has a length of 22 nucleotides and the antisense strand has a length of 24 nucleotides.

[0425] The double-stranded portion of the double-stranded siRNA can be at least 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 29, 40, or 60 nucleotide pairs in length. It can be up to 200, 100, or 50 nucleotide pairs in length. Ranges can be 15-30, 17-23, 19-23, and 19-21 nucleotide pairs in length.

[0426] In some embodiments, the siRNA is large enough that it can be cleaved by endogenous molecules, eg, Dicer, to produce smaller siRNAs, eg, siRNA agents.

[0427] The sense and antisense strands can be selected such that the double-stranded siRNA contains a single strand or unpaired region at one or both ends of the molecule. Thus, the double-stranded siRNA can contain a paired sense and antisense strand to contain an overhang, for example, one or two 5' or 3' overhangs, or a 3' overhang of 1-3 nucleotides. The overhang can be the result of one strand being longer than the other, or the result of two strands of the same length being staggered. Some embodiments have at least one 3' overhang. In some embodiments, both ends of the siRNA molecule have a 3' overhang. In some embodiments, the overhang is 2 nucleotides.

[0428] In some embodiments, the length of the double-stranded region is 15-30, or 18, 19, 20, 21, 22, and 23 nucleotides long, such as the ssiRNA ranges described above. The ssiRNA may resemble in length and structure the natural Dicer processed product from a long dsiRNA. Also included are embodiments in which the two strands of the ssiRNA are linked, e.g., covalently linked. Hairpins or other single-stranded structures that provide the required double-stranded region and 3' overhang are also contemplated.

[0429] The siRNA described herein, including double-stranded siRNA and single-stranded siRNA, can mediate the silencing of target RNA, for example, mRNA, for example, the transcript of a gene that codes for a protein.For convenience, such mRNA is also referred to herein as the mRNA to be silenced.Such gene is also referred to as target gene.Generally, the RNA to be silenced is an endogenous gene or a pathogen gene.In addition, RNA other than mRNA, for example, tRNA and viral RNA, can also be targeted.

[0430] As used herein, the phrase "mediate RNAi" refers to the ability to silence target RNA in a sequence-specific manner.Without being bound by theory, it is believed that silencing uses the RNAi mechanism or process and guide RNA, for example, 21-23 nucleotide ssiRNA.

[0431] In some embodiments, the siRNA is "sufficiently complementary" to the target RNA, e.g., the target mRNA, so that the siRNA silences the production of the protein encoded by the target mRNA. In another embodiment, the siRNA is "exactly complementary" to the target RNA, e.g., the target RNA and the siRNA anneal and form a hybrid made only of Watson-Crick base pairs in the region of exact complementarity. The "sufficiently complementary" target RNA can include an internal region (e.g., of at least 10 nucleotides) that is exactly complementary to the target RNA. Furthermore, in some embodiments, the siRNA specifically discriminates between single nucleotide differences. In this case, the siRNA mediates RNAi only if exact complementarity is found in the region of the single nucleotide difference (e.g., within 7 nucleotides of it).

[0432] MicroRNA: MicroRNAs (miRNAs) are a class of highly conserved small RNA molecules that are transcribed from DNA in the genomes of plants and animals but are not translated into proteins. Processed miRNAs are single-stranded, approximately 17-25 nucleotide (nt) RNA molecules that are incorporated into the RNA-induced silencing complex (RISC) and have been identified as important regulators of development, cell proliferation, apoptosis, and differentiation. They are thought to play a role in regulating gene expression by binding to the 3'-untranslated region of specific mRNAs. RISC mediates downregulation of gene expression through translational inhibition, transcriptional cleavage, or both. RISC has also been implicated in transcriptional silencing in the nuclei of a wide range of eukaryotic organisms.

[0433] The number of miRNA sequences identified to date is large and continues to grow, examples of which can be found, for example, in "miRBase: microRNA sequences, targets and gene nomenclature" Griffiths-Jones S, Grocock RJ, van Dongen S, Bateman A, Enright A J. NAR, 2006, 34, Database Issue, D140-D144; "The microRNA Registry" Griffiths-Jones S. NAR, 2004, 32, Database Issue, D109-D111.

[0434] Antisense oligonucleotides: In some embodiments, the nucleic acid is an antisense oligonucleotide directed to a target polynucleotide. The term "antisense oligonucleotide" or simply "antisense" is meant to include oligonucleotides that are complementary to a target polynucleotide sequence. An antisense oligonucleotide is a single strand of DNA or RNA that is complementary to a selected sequence, for example, a target gene mRNA. Antisense oligonucleotides are thought to inhibit gene expression by binding to complementary mRNA. Binding to the target mRNA can result in inhibition of gene expression by preventing translation of the complementary mRNA strand by binding to it, or by resulting in degradation of the target mRNA. Antisense DNA can be used to target a specific complementary (coding or non-coding) RNA. If binding occurs, this DNA / RNA hybrid can be degraded by the enzyme RNase H. In some embodiments, the antisense oligonucleotide contains about 10 to about 50 nucleotides, more preferably about 15 to about 30 nucleotides. The term also encompasses antisense oligonucleotides that may not be exactly complementary to the desired target gene. Thus, it is contemplated that non-target specific activity may be found with the antisense, or that an antisense sequence containing one or more mismatches with the target sequence may be most preferred for a particular use.

[0435] Antisense oligonucleotides have been demonstrated to be effective target inhibitors of protein synthesis, and therefore can be used to specifically inhibit the protein synthesis of target genes.The effectiveness of antisense oligonucleotides for inhibiting protein synthesis has been well established.For example, the synthesis of polygalacturonase and muscarinic type 2 acetylcholine receptor is inhibited by antisense oligonucleotides directed to their respective mRNA sequences (US Pat. Nos. 5,739,119 and 5,759,829, each of which is incorporated by reference). Additionally, examples of antisense inhibition have been demonstrated for the nuclear protein cyclin, the multidrug resistance gene (MDG1), ICAM-1, E-selectin, STK-1, striatal GABAA receptors and human EGF (Jaskulski et al., Science. 1988 Jun. 10; 240(4858): 1544-6; Vasanthakumar and Ahmed, Cancer Commun. 1989; 1(4): 225-32; Peris et al., Brain Res Mol Brain Res. 1998 Jun. 15; 57(2): 310-20; U.S. Patent Nos. 5,801,154, 5,789,573, 5,718,709 and 5,610,288, each of which is incorporated by reference). Additionally, antisense constructs that can be used to inhibit and treat various abnormal cell proliferations (e.g., cancer) have also been described (U.S. Pat. Nos. 5,747,470, 5,591,317, and 5,783,683, each of which is incorporated by reference).

[0436] Methods for producing antisense oligonucleotides are known in the art and can be easily adapted to produce antisense oligonucleotides targeting any polynucleotide sequence. Selection of antisense oligonucleotide sequences specific for a given target sequence is based on analysis of the selected target sequence and determination of secondary structure, Tm, binding energy, and relative stability. Antisense oligonucleotides can be selected based on their relative inability to form dimers, hairpins, or other secondary structures that reduce or prevent specific binding to the target mRNA in the host cell. Highly preferred target regions of mRNA include the region at or near the AUG translation initiation codon and sequences that are substantially complementary to the 5' region of the mRNA. These secondary structure analysis and target site selection considerations can be performed, for example, using OLIGO primer analysis software v.4 (Molecular Biology Insights) and / or BLASTN 2.0.5 algorithm software (Altschul et al., Nucleic Acids Res. 1997, 25(17):3389-402).

[0437] Antagomir: Antagomir is an RNA-like oligonucleotide with various modifications for pharmacological properties such as RNase protection and enhanced tissue and cellular uptake. Antagomir differs from normal RNA, for example, by complete 2'-O-methylation of sugars, phosphorothioate backbone, and cholesterol moiety at, for example, the 3' end. Antagomir can be used to efficiently silence endogenous miRNA by forming a duplex containing antagomir and endogenous miRNA, thereby preventing miRNA-induced gene silencing. An example of antagomir-mediated miRNA silencing is the silencing of miR-122 described in Krutzfeldt et al., Nature, 2005, 438:685-689, which is expressly incorporated herein by reference in its entirety. Antagomir RNA can be synthesized using standard solid-phase oligonucleotide synthesis protocols. See U.S. Patent Application Publication Nos. 2007 / 0123482 and 2007 / 0213292, each of which is incorporated herein by reference.

[0438] Antagomir can include ligand-conjugated monomer subunits and monomers for oligonucleotide synthesis. Exemplary monomers are described in US Patent Publication No. 2005 / 0107325, which is incorporated by reference in its entirety. Antagomir can have a ZXY structure, such as those described in WO 2004 / 080406, which is incorporated by reference in its entirety. Antagomir can be conjugated with an amphiphilic moiety. Exemplary amphiphilic moieties for use in oligonucleotide agents are described in WO 2004 / 080406, which is incorporated by reference in its entirety.

[0439] Aptamers: Aptamers are nucleic acid or peptide molecules that bind to a particular molecule of interest with high affinity and specificity (Tuerk and Gold, Science 249:505 (1990); Ellington and Szostak, Nature 346:818 (1990), each of which is incorporated by reference in its entirety). DNA or RNA aptamers have been successfully produced that bind to many different entities, from large proteins to small organic molecules. See Eaton, Curr. Opin. Chem. Biol. 1:10-16 (1997), Famulok, Curr. Opin. Struct., J. Biol. 9:324-9 (1999), and Hermann and Patel, Science 287:820-5 (2000), each of which is incorporated by reference in its entirety). Aptamers may be RNA or DNA based and may include riboswitches. Riboswitches are parts of mRNA molecules that can directly bind to small target molecules, and the binding of the target affects the activity of genes. Thus, the mRNA containing the riboswitch is directly involved in regulating its own activity depending on the presence or absence of its target molecule. Generally, aptamers are engineered through repeated in vitro selection or equivalently SELEX (Systematic Evolution of Ligands by Exponential Enrichment) to bind to various molecular targets, such as small molecules, proteins, nucleic acids, and even cells, tissues, and organisms. Aptamers can be prepared by any known method, including synthetic, recombinant, and purified methods, and can be used alone or in combination with other aptamers specific for the same target. Furthermore, as described more fully herein, the term "aptamer" specifically includes "secondary aptamers" that contain consensus sequences derived from comparing two or more known aptamers to a given target.

[0440] Ribozymes: In another embodiment, the nucleic acid-lipid particle is associated with a ribozyme, which is an RNA molecular complex that contains a specific catalytic domain with endonuclease activity (Kim and Cech, Proc Natl Acad Sci USA. 1987 December; 84(24):8788-92; Forster and Symons, Cell. 1987 April 24; 49(2):211-20). For example, many ribozymes accelerate phosphoester transfer reactions with a high degree of specificity, often cleaving only one of several phosphoesters in an oligonucleotide substrate (Cech et al., Cell. 1981 December;27(3 Pt 2):487-96; Michel and Westhof, J Mol Biol. 1990 Dec.5;216(3):585-610; Reinhold-Hurek and Shub, Nature. 1992 May 14;357(6374):173-6). This specificity results from the requirement that the substrate be bound via specific base-pairing interactions to the ribozyme's internal guide sequence ("IGS") prior to chemical reaction.

[0441] At least six basic types of naturally occurring enzymatic RNAs are currently known. Each is capable of catalyzing the hydrolysis of RNA phosphodiester bonds during transfer (and thus capable of cleaving other RNA molecules) under physiological conditions. Broadly, enzymatic nucleic acids act by first binding to a target RNA. Such binding occurs through a target binding portion of the enzymatic nucleic acid that is held in close proximity to the enzymatic portion of the molecule that acts to cleave the target RNA. Thus, an enzymatic nucleic acid first recognizes a target RNA, then binds to the target RNA through complementary base pairing, and upon binding to the correct site, acts enzymatically to cleave the target RNA. Such strategic cleavage of a target RNA destroys its ability to direct synthesis of an encoded protein. After an enzymatic nucleic acid has bound and cleaved its RNA target, it is released from that RNA to seek another target, and can repeatedly bind and cleave new targets.

[0442] The enzymatic nucleic acid molecule may be formed, for example, in a hammerhead, hairpin, Hepatitis δ virus, group I intron, or RNaseP RNA (associated with an RNA guide sequence) or Neurospora VS RNA motif. Specific examples of hammerhead motifs are described in Rossi et al. Nucleic Acids Res. 1992 Sep. 11; 20(17): 4559-65. Examples of hairpin motifs are described in Hampel et al. (European Patent Publication No. 0360257), Hampel and Tritz, Biochemistry 1989 Jun. 13; 28(12): 4929-33, Hampel et al., Nucleic Acids Res. 1990 Jan. 25; 18(2): 299-304, and U.S. Patent No. 5,631,359. Examples of hepatitis delta virus motifs are described in Perrotta and Been, Biochemistry. 1992 Dec. 1;31(47):11843-52, examples of RNase P motifs are described in Guerrier-Takada et al., Cell. 1983 December;35(3 Pt 2):849-57, a Neurospora VS RNA ribozyme motif is described in Collins (Saville and Collins, Cell. 1990 May 18;61(4):685-96; Saville and Collins, Proc Natl Acad Sci USA. 1991 Oct. 1;88(19):8826-30; Collins and Olive, Biochemistry. 1993 Mar. 23;32(11):2795-9), and examples of group I introns are described in U.S. Pat. No. 4,987,071. The important features of the enzymatic nucleic acid molecules used are that they have a specific substrate binding site that is complementary to one or more of the target gene DNA or RNA regions, and that they have nucleotide sequences within or surrounding the substrate binding site that confer RNA cleavage activity to the molecule. Thus, ribozyme constructs need not be limited to the specific motifs mentioned herein.

[0443] Methods for producing ribozymes targeted to any polynucleotide sequence are known in the art. Ribozymes can be designed as described in WO 93 / 23569 and WO 94 / 02595, each of which is specifically incorporated herein by reference, and can be synthesized and tested in vitro and in vivo as described therein.

[0444] Ribozyme activity can be optimized by altering the length of the ribozyme binding arms, or by chemically synthesizing ribozymes with modifications that prevent their degradation by serum ribonucleases (see, e.g., WO 92 / 07065, WO 93 / 15187, and WO 91 / 03162, European Patent Application No. 92110298.4, U.S. Pat. No. 5,334,711, and WO 94 / 13688, which describe various chemical modifications that can be made to the sugar moiety of enzymatic RNA molecules), modifications that increase efficacy within cells, and removal of stem II bases to shorten RNA synthesis time and reduce chemical requirements.

[0445] Immunostimulatory oligonucleotides: The nucleic acid associated with the lipid particles may be immunostimulatory, such as immunostimulatory oligonucleotides (ISS, single-stranded or double-stranded) that can induce an immune response when administered to a subject, which may be a mammal or other patient. ISSs include, for example, specific palindromes that result in hairpin secondary structures (see Yamamoto S., et al. (1992) J. Immunol. 148:4072-4076, which is incorporated by reference in its entirety), or CpG motifs, as well as other known ISS features (e.g., multi-G domains, see WO 96 / 11266, which is incorporated by reference in its entirety).

[0446] The immune response may be an innate immune response or an adaptive immune response. The immune system is divided into a more innate immune system of vertebrates and an adaptive immune system, which is further divided into a humoral cellular component. In some embodiments, the immune response may be mucosal.

[0447] In some embodiments, the immunostimulatory nucleic acid is only immunostimulatory when administered in combination with a lipid particle, and is not immunostimulatory when administered in its "free form." Such oligonucleotides are considered to be immunostimulatory.

[0448] Immunostimulatory nucleic acids are considered to be non-sequence specific if they are not required to specifically bind to and reduce the expression of a target polynucleotide in order to elicit an immune response. Thus, certain immunostimulatory nucleic acids may contain sequences that correspond to regions of naturally occurring genes or mRNAs, yet still be considered non-sequence specific immunostimulatory nucleic acids.

[0449] In some embodiments, the immunostimulatory nucleic acid or oligonucleotide comprises at least one CpG dinucleotide. The oligonucleotide or CpG dinucleotide may be unmethylated or methylated. In another embodiment, the immunostimulatory nucleic acid comprises at least one CpG dinucleotide with a methylated cytosine. In some embodiments, the nucleic acid comprises a single CpG dinucleotide, and the cytosine in the CpG dinucleotide is methylated. In another embodiment, the nucleic acid comprises at least two CpG dinucleotides, and at least one cytosine in the CpG dinucleotide is methylated. In a further embodiment, each cytosine in the CpG dinucleotide present in the sequence is methylated. In another embodiment, the nucleic acid comprises a plurality of CpG dinucleotides, and at least one of the CpG dinucleotides comprises a methylated cytosine.

[0450] Bonds between the linker unit, the nucleic acid agent, and the ligand In some embodiments, the bond between the linker unit and the nucleic acid agent is a bond.

[0451] In some embodiments, the bond between the linker unit and the nucleic acid agent is a moiety (eg, a moiety that includes a cleavable group).

[0452] In some embodiments, the bond between the linker unit and the ligand is a bond.

[0453] In some embodiments, the bond between the Linker unit and the Ligand is a moiety (eg, a moiety that includes a cleavable group).

[0454] In some embodiments, the bond between a linker unit and a ligand comprises a -C(=O)- connected to the linker unit.

[0455] The group may be cleavable or non-cleavable. Suitable groups include, for example, -NR-, -C(=O)-, -C(=O)NH-, -S(=O)-, -S(=O)2-, -S(=O)2NH, or, but not limited to, alkylene, alkenylene, alkynylene, aryl alkylene, aryl alkenylene, aryl alkynylene, heteroaryl alkylene, heteroaryl alkenylene, heteroaryl alkynylene, heterocyclyl alkylene, heterocyclyl alkenylene, heterocyclyl alkynylene, arylene, heteroarylene, heterocyclylene, cycloalkylene, cycloalkenylene, alkylaryl alkylene, alkylaryl alkenylene, alkylaryl alkynylene, alkenylaryl alkylene, alkenylaryl alkenylene, alkenylaryl alkynylene, alkynylaryl alkylene, alkynylaryl alkenylene, alkynylaryl alkynylene, alkylheteroaryl alkylene, alkylheteroaryl alkenylene. Alkylheteroarylalkynylenes Alkenylheteroarylalkylenes Alkenylheteroarylalkenylenes Alkenylheteroarylalkynylenes Alkynylheteroarylalkylenes Alkynylheteroarylalkenylenes Alkynylheteroarylalkynylenes Alkylheteroarylalkynylenes Alkylheterocyclylalkylenes Alkylheterocyclylalkenylenes Alkylheterocyclylalkynylenes Alkenylheterocyclylalkylenes Alkenylheterocyclylalkenylenes Alkenylheterocyclylalkynylenes Alkylheterocyclylalkylenes Alkynylheterocyclylalkenylenes Alkylheterocyclylalkynylenes Alkylarylenes Alkenylarylenes Alkylheteroarylenes Alkylheteroarylenes Alkylheteroarylenes Included are chains of atoms such as alkynylheteroarylene, each of which may be substituted or unsubstituted, and one or more methylenes may be interrupted or terminated by -O-, -S-, -S(=O)-, -S(=O)2-, -NR-, -C(=O)-, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, or substituted or unsubstituted heterocycle, and R is hydrogen, acyl, aliphatic, or substituted aliphatic.

[0456] A cleavable group is one that is sufficiently stable outside a cell, but that upon entry into a target cell is cleaved to release the two moieties that the group holds together. In preferred embodiments, the cleavable group is cleaved at least 10 times faster, preferably at least 100 times faster, in the target cell or under a first reference condition (which may, for example, be selected to replicate or represent intracellular conditions) or under a second reference condition (which may, for example, be selected to replicate or represent conditions found in blood or serum) than in the target cell or in the subject's blood.

[0457] The cleavable group is sensitive to a cleaving agent (e.g., pH, oxidation-reduction redox potential, or the presence of a degradable molecule). In general, the cleaving agent is more prevalent or found at a higher level or activity within the cell than in serum or blood. Examples of such degrading agents include: redox agents that are selective for a particular substrate or have no substrate specificity (e.g., reducing agents such as oxidases or reductases or mercaptans present within the cell, which can degrade the redox-cleavable group by reduction); esterases; agents that can create an endosome or acidic environment, e.g., those that result in a pH of 5 or less; enzymes that can hydrolyze or degrade the acid-cleavable group by acting as general acids, peptidases (which can be substrate specific), phosphatases.

[0458] The cleavable group, e.g., a disulfide bond, may be sensitive to pH. The pH of human serum is 7.4, while the average intracellular pH is slightly lower, ranging from about 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 about 5.0. Some linkers have a cleavable group that is cleaved at a preferred pH, thereby releasing the cationic lipid from the ligand within the cell or into a desired compartment of the cell.

[0459] The conjugate can include a cleavable group that can be cleaved by a specific enzyme. The type of cleavable group incorporated into the conjugate can vary depending on the cell to be targeted. For example, liver targeting ligand can be linked to cationic lipid via a chemical moiety that includes an ester group. Liver cells are rich in esterases, and therefore this group is cleaved more efficiently in liver cells than in cell types that are not rich in esterases. Other cell types that are rich in esterases include lung, renal cortex, and testicular cells.

[0460] Coupling groups containing peptide bonds can be used when targeting cell types that are rich in peptidases, such as hepatocytes and synovial cells.

[0461] In general, the suitability of a candidate cleavable group can be evaluated by testing the ability (or conditions) of a degrading agent to cleave the candidate group. It is also desirable to test the candidate cleavable group for its ability to resist cleavage in blood or when in contact with other non-target tissues. Thus, the relative susceptibility to cleavage between a first condition and a second condition can be determined, the first condition being selected to indicate cleavage in target cells, and the second condition being selected to indicate 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 a whole animal. It can be useful to perform initial evaluation in a cell-free or cultured condition and confirm by further evaluation in a whole animal. In a preferred embodiment, a useful candidate compound is cleaved at least 2, 4, 10, or 100 times faster in cells (or in vitro conditions selected to mimic intracellular conditions) compared to blood or serum (or in vitro conditions selected to mimic extracellular conditions).

[0462] Redox-cleavable groups. One class of cleavable groups is redox-cleavable groups that are cleaved upon reduction or oxidation. An example of a reductively cleavable group is a disulfide linking group (-SS-). To determine whether a candidate cleavable 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, the methods described herein can be considered. For example, candidates can be evaluated by incubation with dithiothreitol (DTT) or other reducing agents using reagents known in the art that reproduce the cleavage rate observed in cells, for example, target cells. Candidates can also be evaluated under conditions selected to reproduce blood or serum conditions. In preferred embodiments, the candidate compound is cleaved at most 10% in blood. In preferred embodiments, useful candidate compounds are degraded at least 2, 4, 10, or 100 times faster in cells (or in vitro conditions selected to reproduce intracellular conditions) compared to blood (or in vitro conditions selected to reproduce extracellular conditions). The rate of cleavage of a 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.

[0463] Phosphate-based cleavable groups. Phosphate-based cleavable groups are cleaved by an agent that decomposes or hydrolyzes the phosphate group. An example of an agent that cleaves a phosphate group within a cell is an enzyme such as an intracellular phosphatase. In some embodiments, the phosphate-based linking group is -OP(=O)(OR k )-O-, -OP(=S)(OR k )-O-, -OP(=S)(SR k )-O-, -SP(=O)(OR k )-O-, -OP(=O)(OR k )-S-, -SP(=O)(OR k )-S-, -OP(=S)(OR k )-s-, -SP(=S)(OR k )-O-, -OP(=O)(R k)-O-, -OP(=S)(R k )-O-, -SP(=O)(R k )-O-, -SP(=S)(R k )-O-, -SP(=O)(R k )-S- or -OP(=S)(R k )-S-. In some embodiments, the phosphate based linking group is -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-, or -OP(=S)(H)-S-. In some embodiments, the phosphate based linking group is -OP(=O)(OH)-O-.

[0464] Acid cleavable groups. Acid cleavable groups are linking groups that are cleaved under acidic conditions. In preferred embodiments, acid cleavable groups are cleaved in an acidic environment having 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. In cells, certain low pH organelles such as endosomes and lysosomes can provide a cleavage environment for acid cleavable linking groups. Examples of acid cleavable 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). A preferred embodiment is where the carbon attached 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.

[0465] Ester-based cleavable groups. Ester-based cleavable groups are cleaved by enzymes such as esterases and amidases in cells. Examples of ester-based cleavable groups include, but are not limited to, esters of alkylene, alkenylene, and alkynylene groups. Ester cleavable linkers have the general formula -C(O)O- or -OC(O)-. These candidates can be evaluated using methods similar to those described above.

[0466] Peptide-based cleavable groups. Peptide-based cleavable groups are cleaved by enzymes such as peptidases and proteases in cells. Peptide-based cleavable groups are peptide bonds formed between amino acids to give rise to 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 specific type of amide bond formed between amino acids to give rise to peptides and proteins. Peptide-based cleavable groups are generally limited to peptide bonds (i.e., amide bonds) formed between amino acids to give rise to peptides and proteins, and do not include the entire amide functionality. Peptide-based cleavable linkers have the general formula -NHCHR A C(O)NHCHR B C(O)—, where R A and R Bare the R groups of two adjacent amino acids. These candidates can be evaluated using methods similar to those described above. As used herein, "carbohydrate" refers to either a compound that is a carbohydrate consisting essentially of one or more monosaccharide units having at least six carbon atoms (which may be linear, branched or cyclic) with an oxygen, nitrogen or sulfur atom bonded to each carbon atom, or a compound having as a portion a carbohydrate moiety consisting of one or more monosaccharide units each having at least six carbon atoms (which may be linear, branched or cyclic) with an oxygen, nitrogen or sulfur atom bonded to each carbon atom. Representative carbohydrates include sugars (monosaccharides, disaccharides, trisaccharides and oligosaccharides containing about 4-9 monosaccharide units) and polysaccharides such as starch, glycogen, cellulose and polysaccharide gums. Specific monosaccharides include C5 and above (preferably C5-C8) sugars, and disaccharides and trisaccharides include sugars having two or three monosaccharide units (preferably C5-C8).

[0467] Synthesis method In some aspects, the disclosure provides methods of preparing the disclosed compounds.

[0468] In some aspects, the disclosure provides compounds obtainable or obtainable by the methods for preparing the compounds described herein.

[0469] In some aspects, the disclosure provides intermediates described herein that are suitable for use in the methods for preparing the compounds described herein.

[0470] The compounds of the present disclosure can be prepared by any suitable technique known in the art. Particular processes for the preparation of these compounds are further described in the accompanying Examples.

[0471] In the descriptions of synthetic methods described herein, and in any reference synthetic methods used to prepare starting materials, it is understood that all proposed reaction conditions, including choice of solvent, reaction atmosphere, reaction temperature, duration of experiments and work-up procedures, can be selected by one of ordinary skill in the art.

[0472] It is understood by one skilled in the art of organic synthesis that the functionality present on various portions of the molecule must be compatible with the reagents and reaction conditions employed.

[0473] It will be understood that during the synthesis of the compounds of the present disclosure in the processes defined herein, or during the synthesis of certain starting materials, it may be desirable to protect certain substituents to prevent their undesired reactions. The skilled chemist will understand when such protection is required and how such protecting groups can be put in place and subsequently removed. For examples of protecting groups, see one of the many general texts on the subject, for example 'Protective Groups in Organic Synthesis' by Theodora Green (publisher: John Wiley & Sons). Protecting groups can be removed by any convenient method described in the literature or known to the skilled chemist as being suitable for the removal of the protecting group in question, such a method being selected to effect the removal of the protecting group with minimal interference with groups elsewhere in the molecule. Thus, when a reactant contains a group such as, for example, amino, carboxy or hydroxy, it may be desirable to protect this group in some of the reactions mentioned herein.

[0474] For example, a suitable protecting group for an amino or alkylamino group is, for example, an acyl group, for example an alkanoyl group such as acetyl, an alkoxycarbonyl group, for example a methoxycarbonyl, ethoxycarbonyl or t-butoxycarbonyl group, an arylmethoxycarbonyl group, for example benzyloxycarbonyl, or an aroyl group, for example benzoyl. Suitable protecting groups for hydroxy or alkylhydroxy groups may be, for example, acetyl (Ac), benzoyl (Bz), benzyl (Bn), β-methoxyethoxymethyl ether (MEM), dimethoxytrityl (DMT), methoxymethyl ether (MOM), methoxytrityl (MMT), p-methoxybenzyl ether (PMB), p-methoxyphenyl ether (PMP), pivaloyl (Piv), tetrahydropyranyl (THP), tetrahydrofuran (THF), trityl (triphenylmethyl, Tr), silyl ether (e.g., trimethylsilyl (TMS), tert-butyldimethylsilyl (TBDMS), tri-iso-propylsilyloxymethyl (TOM), and triisopropylsilyl (TIPS) ether), methyl ether, or ethoxyethyl ether (EE). Suitable protecting groups for 1,2-diols may be, for example, acetals. A suitable protecting group for 1,3-diols can be, for example, tetraisopropyldisiloxanylidene (TIPDS).

[0475] The deprotection conditions of the above protecting groups necessarily vary with the choice of protecting group. Thus, for example, acyl groups such as alkanoyl or alkoxycarbonyl groups or aroyl groups can be removed by hydrolysis with a suitable base such as an alkali metal hydroxide, for example lithium hydroxide or sodium hydroxide. Alternatively, acyl groups such as tert-butoxycarbonyl groups can be removed by treatment with a suitable acid, for example hydrochloric acid, sulfuric acid or phosphoric acid or trifluoroacetic acid, and arylmethoxycarbonyl groups such as benzyloxycarbonyl groups can be removed by hydrogenation over a catalyst such as palladium on carbon, or by treatment with a Lewis acid, for example tris(trifluoroacetate)borate. An alternative protecting group suitable for primary amino groups is, for example, the phthaloyl group, which can be removed by treatment with an alkylamine, for example dimethylaminopropylamine, or hydrazine.

[0476] Suitable protecting groups for hydroxyl groups are, for example, acyl groups, e.g. alkanoyl groups such as acetyl, aroyl groups such as benzoyl, or arylmethyl groups such as benzyl. The deprotection conditions for the above protecting groups necessarily vary with the choice of protecting group. Thus, for example, acyl groups such as alkanoyl or aroyl groups can be removed by hydrolysis with a suitable base, e.g., an alkali metal hydroxide, e.g., lithium hydroxide, sodium hydroxide, or ammonia. Alternatively, arylmethyl groups such as benzyl groups can be removed by hydrogenation, e.g., over a catalyst such as palladium on carbon.

[0477] A suitable protecting group for a carboxy group is, for example, an esterifying group, for example a methyl or ethyl group which may be removed by hydrolysis with a base such as sodium hydroxide, or for example a tert-butyl group which may be removed by treatment with an acid, for example an organic acid, such as trifluoroacetic acid, or for example a benzyl group which may be removed by hydrogenation over a catalyst such as palladium on carbon.

[0478] Conveniently, the reaction of the compounds is carried out in the presence of a suitable solvent, preferably inert under the respective reaction conditions. Examples of suitable solvents include hydrocarbons such as hexane, petroleum ether, benzene, toluene, or xylene; chlorinated hydrocarbons such as trichloroethylene, 1,2-dichloroethane, tetrachloromethane, chloroform, or dichloromethane; alcohols such as methanol, ethanol, isopropanol, n-propanol, n-butanol, or tert-butanol; ethers such as diethyl ether, diisopropyl ether, tetrahydrofuran (THF), 2-methyltetrahydrofuran, cyclopentyl methyl ether (CPME), methyl tert-butyl ether (MTBE), or dioxane; ethylene glycol monoethers, diisopropyl ... Examples of suitable solvents include, but are not limited to, glycol ethers such as ethylene glycol monomethyl ether or ethylene glycol dimethyl ether (diglyme); ketones such as acetone, methyl isobutyl ketone (MIBK), or butanone; amides such as acetamide, dimethylacetamide, dimethylformamide (DMF), N-methylpyrrolidinone (NMP), nitriles such as acetonitrile, sulfoxides such as dimethylsulfoxide (DMSO), nitro compounds such as nitromethane or nitrobenzene; esters such as ethyl acetate or methyl acetate, or mixtures of such solvents or with water.

[0479] The reaction temperature is preferably about -100°C to 300°C, depending on the reaction steps and conditions used.

[0480] The reaction time generally ranges from a few minutes to several days depending on the reactivity of each compound and the respective reaction conditions. A suitable reaction time can be easily determined by methods known in the art (e.g., reaction monitoring). Based on the above reaction temperature, a suitable reaction time is generally in the range of 10 minutes to 48 hours.

[0481] Moreover, by utilizing the procedures described herein, in conjunction with ordinary skills in the art, additional compounds of the present disclosure can be readily prepared. Those skilled in the art will readily understand that known variations of the conditions and processes of the following preparative procedures can be used to prepare these compounds.

[0482] As will be appreciated by those skilled in the art of organic synthesis, the compounds of the present disclosure are readily accessible by various synthetic routes, some of which are illustrated in the accompanying examples. Those skilled in the art will readily recognize what types of reagents and reaction conditions should be used to obtain the compounds of the present disclosure, and how they should be applied and adapted to any particular case (whenever necessary or useful). Furthermore, some of the compounds of the present disclosure can be easily synthesized by reacting other compounds of the present disclosure under suitable conditions, for example by applying standard synthetic methods such as reduction, oxidation, addition or substitution reactions, by converting one specific functional group present in the compounds of the present disclosure or in a suitable precursor molecule thereof into another, which methods are well known to those skilled in the art. Similarly, those skilled in the art will readily recognize that, whenever necessary or useful, synthetic protection (or protective) groups, the application of suitable protective groups, as well as methods for their introduction and removal, are well known to those skilled in the art of chemical synthesis, and are described in more detail, for example, in PGM Wuts, TW Greene, "Greene's Protective Groups in Organic Synthesis", 4th edition (2006) (John Wiley & Sons).

[0483] A general route for the preparation of the compounds of the present application is depicted in Scheme 1 herein. [ka]

[0484] Biological assays Once produced, the compounds, scaffolds or conjugates designed, selected, prepared and / or optimized by the above methods can be characterized using various assays known to those skilled in the art to determine whether the compounds, scaffolds or conjugates have biological activity. For example, the compounds, scaffolds or conjugates can be characterized by conventional assays, including but not limited to the assays described below, to determine whether they have the desired activity, e.g., target binding activity and / or specificity and / or stability.

[0485] Furthermore, high throughput screening can be used to speed up the analysis using such assays. As a result, it may be possible to rapidly screen the molecules described herein for activity using techniques known in the art. General methodologies for performing high throughput screening are described, for example, in Devlin (1998) High Throughput Screening, Marcel Dekker, and U.S. Patent No. 5,763,263. High throughput assays can use one or more different assay techniques, including but not limited to those described below.

[0486] A variety of in vitro or in vivo biological assays may be suitable for detecting the effects of the disclosed compounds, scaffolds, or conjugates. These in vitro or in vivo biological assays may include, but are not limited to, enzyme activity assays, electrophoretic mobility shift assays, reporter gene assays, in vitro cell viability assays, and assays described herein.

[0487] In some embodiments, the biological assays are described in the Examples herein.

[0488] Pharmaceutical Compositions In some aspects, the present disclosure provides pharmaceutical compositions comprising a compound, scaffold, or conjugate of the present disclosure as an active ingredient.

[0489] As used herein, the term "composition" is intended to encompass a product comprising the specified ingredients in the specified amounts, as well as any product that results directly or indirectly from combining the specified ingredients in the specified amounts.

[0490] Pharmaceutical compositions suitable for injectable use include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. For intravenous administration, suitable carriers include physiological saline, bacteriostatic water, Cremophor EL™ (BASF, Parsippany, NJ) or phosphate buffered saline (PBS). In all cases, the composition must be sterile and fluid to the extent that easy syringability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof. The proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants. The action of microorganisms can be prevented by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In many cases, it is preferable to include isotonic agents, for example, sugars, polyalcohols such as mannitol and sorbitol, and sodium chloride in the composition. Prolonged absorption of the injectable compositions can be brought about by including in the composition an agent which delays absorption, for example, aluminum monostearate and gelatin.

[0491] Sterile injection solution can be prepared by incorporating the required amount of active compound into suitable solvent with one or combination of the above-listed components as required, followed by filtration sterilization.Generally, dispersion is prepared by incorporating active compound into a sterile vehicle that contains basic dispersion medium and other components required from above-listed.For the preparation of sterile powder for sterile injection solution, preparation method includes vacuum drying and freeze-drying, and obtains powder of active ingredient and any additional desired components from the solution that has been previously sterile-filtered.

[0492] The formulation of the present disclosure may be in the form of an aqueous solution comprising an aqueous vehicle.The aqueous vehicle component may comprise water and at least one pharma- ceutically acceptable excipient.Suitable acceptable excipients include those selected from the group consisting of dissolution enhancers, chelating agents, preservatives, isotonicity agents, viscosity / suspending agents, buffers, and pH adjusters, and mixtures thereof.

[0493] Any suitable dissolution enhancer may be used. Examples of dissolution enhancers include cyclodextrins such as those selected from the group consisting of hydroxypropyl-β-cyclodextrin, methyl-β-cyclodextrin, randomly methylated-β-cyclodextrin, ethylated-β-cyclodextrin, triacetyl-β-cyclodextrin, fully acetylated-β-cyclodextrin, carboxymethyl-β-cyclodextrin, hydroxyethyl-β-cyclodextrin, 2-hydroxy-3-(trimethylammonio)propyl-β-cyclodextrin, glucosyl-β-cyclodextrin, sulfated β-cyclodextrin (S-β-CD), maltosyl-β-cyclodextrin, β-cyclodextrin sulfobutyl ether, branched-β-cyclodextrin, hydroxypropyl-γ-cyclodextrin, randomly methylated-γ-cyclodextrin, and trimethyl-γ-cyclodextrin, and mixtures thereof.

[0494] Any suitable chelating agent may be used. Examples of suitable chelating agents include those selected from the group consisting of ethylenediaminetetraacetic acid and its metal salts, disodium edetate, trisodium edetate, and tetrasodium edetate, and mixtures thereof.

[0495] Any suitable preservative can be used. Examples of preservatives include quaternary ammonium salts, such as benzalkonium halides (preferably benzalkonium chloride), chlorhexidine gluconate, benzethonium chloride, cetylpyridinium chloride, benzyl bromide, phenylmercuric nitrate, phenylmercuric acetate, phenylmercuric neodecanoate, merthiolate, methylparaben, propylparaben, sorbic acid, potassium sorbate, sodium benzoate, sodium propionate, ethyl p-hydroxybenzoate, propylaminopropyl biguanide, and butyl p-hydroxybenzoate, and sorbic acid, and mixtures thereof.

[0496] The aqueous vehicle may also contain an isotonicity agent to adjust tonicity (osmotic pressure), which may be selected from the group consisting of glycols (such as propylene glycol, diethylene glycol, triethylene glycol), glycerol, dextrose, glycerin, mannitol, potassium chloride, and sodium chloride, and mixtures thereof.

[0497] The formulation may contain a pH adjuster to adjust the formulation to an acceptable pH (typically about 5.0 to about 9.0, more preferably about 5.5 to about 8.5, particularly about 6.0 to about 8.5, about 7.0 to about 8.5, about 7.2 to about 7.7, about 7.1 to about 7.9, or about 7.5 to about 8.0). The pH adjuster is typically a mineral acid or metal hydroxide base selected from the group consisting of potassium hydroxide, sodium hydroxide, and hydrochloric acid, and mixtures thereof, preferably sodium hydroxide and / or hydrochloric acid. These acidic and / or basic pH adjusters are added to adjust the formulation to a target acceptable pH range. Thus, it may not be necessary to use both an acid and a base, and depending on the formulation, the addition of either an acid or a base is sufficient to bring the mixture into the desired pH range.

[0498] The aqueous vehicle may also contain a buffer to stabilize the pH. If used, the buffer is selected from the group consisting of phosphate buffers (such as sodium dihydrogen phosphate and disodium hydrogen phosphate), borate buffers (such as boric acid or its salts including disodium tetraborate), citrate buffers (such as citric acid or its salts including sodium citrate), and ε-aminocaproic acid, and mixtures thereof.

[0499] According to a further aspect of the present disclosure, there is provided a pharmaceutical composition comprising a compound of the present disclosure as defined herein, or a pharma- ceutically acceptable salt, hydrate or solvate thereof, together with a pharma- ceutically acceptable diluent or carrier.

[0500] The compositions of the present disclosure may be in a form suitable for oral use (e.g., as tablets, troches, hard or soft capsules, aqueous or oily suspensions, emulsions, dispersible powders or granules, syrups or elixirs), topical use (e.g., as creams, ointments, gels, or aqueous or oily solutions or suspensions), administration by inhalation (e.g., as a finely divided powder or liquid aerosol), administration by insufflation (e.g., as a finely divided powder) or parenteral administration (e.g., as a sterile aqueous or oily solution for intravenous, subcutaneous, intramuscular, intraperitoneal or intramuscular administration, or as a suppository for rectal administration).

[0501] The compositions of the present disclosure can be obtained by conventional procedures using conventional pharmaceutical excipients well known in the art. Thus, compositions intended for oral use may contain, for example, one or more coloring agents, sweeteners, flavoring agents and / or preservatives.

[0502] An effective amount of a compound of the present disclosure for use in therapy is an amount sufficient to treat or prevent, slow the progression of, and / or reduce the symptoms associated with an inflammasome-associated condition referred to herein.

[0503] An effective amount of a compound of the present disclosure for use in therapy is an amount sufficient to treat, slow the progression of, and / or reduce the symptoms associated with an inflammasome-associated condition referred to herein.

[0504] The magnitude of a dose of a compound of formula (I) or (II) for therapeutic or prophylactic purposes will, of course, vary with the nature and severity of the condition, the age and sex of the animal or patient, and the route of administration, in accordance with well-known principles of medicine.

[0505] How to use In some aspects, the present disclosure provides a method of modulating (e.g., reducing or eliminating) expression of a target gene in a subject, the method comprising administering to the subject a conjugate of the present disclosure.

[0506] In some aspects, the present disclosure provides a method of modulating (e.g., reducing or eliminating) expression of a target gene in a cell or tissue of a subject, the method comprising administering to the subject a conjugate of the present disclosure.

[0507] In some aspects, the present disclosure provides a method of delivering a nucleic acid agent to a subject, the method comprising administering to the subject a conjugate of the present disclosure.

[0508] In some aspects, the present disclosure provides a method of treating or preventing a disease in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a conjugate of the present disclosure.

[0509] In some aspects, the present disclosure provides a conjugate of the present disclosure for modulating (e.g., reducing or eliminating) expression of a target gene in a subject.

[0510] In some aspects, the present disclosure provides a conjugate of the present disclosure for modulating (e.g., reducing or eliminating) expression of a target gene in a cell or tissue of a subject.

[0511] In some aspects, the present disclosure provides a conjugate of the present disclosure for delivery of a nucleic acid agent to a subject.

[0512] In some aspects, the present disclosure provides a conjugate of the present disclosure for treatment or prevention of a disease in a subject in need thereof.

[0513] In some aspects, the disclosure provides for the use of a conjugate of the disclosure in the manufacture of a medicament for modulating (e.g., reducing or eliminating) expression of a target gene in a subject.

[0514] In some aspects, the disclosure provides for the use of a conjugate of the disclosure in the manufacture of a medicament for modulating (e.g., reducing or eliminating) expression of a target gene in a cell or tissue of a subject.

[0515] In some aspects, the disclosure provides for the use of a conjugate of the disclosure in the manufacture of a medicament for delivering a nucleic acid agent to a subject.

[0516] In some aspects, the disclosure provides the use of a conjugate of the disclosure in the manufacture of a medicament for the treatment or prevention of a disease in a subject in need thereof.

[0517] In some embodiments, the subject is a cell.

[0518] In some embodiments, the subject is a tissue.

[0519] In some embodiments, the subject is a human.

[0520] In some embodiments, the target gene is Factor VII, Eg5, PCSK9, TPX2, apoB, SAA, TTR, HBV, HCV, RSV, PDGF beta gene, Erb-B gene, Src gene, CRK gene, GRB2 gene, RAS gene, MEKK gene, JNK gene, RAF gene, Erk1 / 2 gene, PCNA (p21) gene, MYB gene, JUN gene, FOS gene, BCL-2 gene, Cyclin D gene, VEGF gene, EGFR gene, cyclin A gene, cyclin E gene, WNT-1 gene, beta-catenin gene, c-MET gene, PKC gene, NFKB gene, STAT3 gene, survivin gene, Her2 / Neu gene, topoisomerase I gene, topoisomerase II alpha gene, p73 gene, p21(WAF1 / CIP1) gene, p27(KIP1) gene, PPM1D gene, RAS gene, caveolin I gene, MIB I gene, MTAI gene, M68 gene, mutations in tumor suppressor genes, p53 tumor suppressor gene, LDHA, or any combination thereof.

[0521] In some embodiments, the disease is characterized by unwanted expression of the target gene.

[0522] In some embodiments, the administration results in the reduction or elimination of expression of the target gene in the subject.

[0523] In some embodiments, the disease is a viral infection, such as an HCV, HBV, HPV, HSV, or HIV infection.

[0524] In some embodiments, the disease is cancer.

[0525] In some embodiments, the cancer is biliary tract cancer, bladder cancer, transitional cell carcinoma, urothelial carcinoma, brain cancer, glioma, astrocytoma, breast cancer, metaplastic carcinoma, cervical cancer, cervical squamous cell carcinoma, rectal cancer, colorectal cancer, colon cancer, hereditary nonpolyposis colorectal cancer, colorectal adenocarcinoma, gastrointestinal stromal tumor (GIST), endometrial cancer, endometrial stromal sarcoma, esophageal cancer, esophageal squamous cell carcinoma, esophageal adenocarcinoma, ocular melanoma, uveal melanoma, gallbladder cancer, gallbladder adenocarcinoma, renal cell carcinoma, clear cell renal cell carcinoma, transitional cell carcinoma, urothelial carcinoma, Wilms' tumor, leukemia, acute lymphocytic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic (CLL), chronic myelogenous (CML), chronic myelomonocytic leukemia ( CMML), liver cancer, liver cancer, hepatocellular carcinoma, cholangiocarcinoma, hepatoblastoma, lung cancer, non-small cell lung cancer (NSCLC), mesothelioma, B cell lymphoma, non-Hodgkin's lymphoma, diffuse large B cell lymphoma, mantle cell lymphoma, T cell lymphoma, non-Hodgkin's lymphoma, precursor T lymphoblastic lymphoma / leukemia, peripheral T cell lymphoma, multiple myeloma, nasopharyngeal carcinoma (NPC), neuroblastoma, oropharyngeal carcinoma, oral squamous cell carcinoma, osteosarcoma, ovarian cancer, pancreatic cancer, pancreatic ductal adenocarcinoma, pseudopapillary neoplasm, acinar cell carcinoma, prostate cancer, prostate adenocarcinoma, skin cancer, melanoma, malignant melanoma, cutaneous melanoma, small intestine cancer, gastric cancer, gastrointestinal stromal tumor (GIST), uterine cancer, or uterine sarcoma.

[0526] In some embodiments, the cancer is liver cancer, liver cancer, hepatoma, hepatocellular carcinoma, cholangiocarcinoma, or hepatoblastoma.

[0527] In some embodiments, the disease is a proliferative, inflammatory, autoimmune, neurological, ocular, respiratory, metabolic, cutaneous, auditory, hepatic, renal, or infectious disease, hi some embodiments, the disease is a liver disease.

[0528] definition Unless stated otherwise, the following terms used in the specification and claims have the following meanings, as set forth below.

[0529] Without wishing to be limited by this description, it is understood that while various options for variables are described herein, the present disclosure is intended to encompass operable embodiments having combinations of options, and the present disclosure may be interpreted as excluding inoperable embodiments caused by a particular combination of options.

[0530] As used herein, "alkyl", "C1, C2, C3, C4, C5 or C6 alkyl" or "C1-C6 alkyl" is intended to include C1, C2, C3, C4, C5 or C6 straight chain (straight-chain) saturated aliphatic hydrocarbon groups, and C3, C4, C5 or C6 branched saturated aliphatic hydrocarbon groups. For example, C1-C6 alkyl is intended to include C1, C2, C3, C4, C5 and C6 alkyl groups. Examples of alkyl include, but are not limited to, moieties having 1 to 6 carbon atoms, such as methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, n-pentyl, i-pentyl, or n-hexyl. In some embodiments, a straight chain or branched alkyl has 6 or fewer carbon atoms (e.g., C1-C6 for straight chain, C3-C6 for branched chain), and in other embodiments, a straight chain or branched alkyl has 4 or fewer carbon atoms.

[0531] As used herein, the term "optionally substituted alkyl" refers to an unsubstituted alkyl or an alkyl having specified substituents replacing one or more hydrogen atoms on one or more carbons of the hydrocarbon backbone. Such substituents may include, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, amino (including alkylamino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or aromatic or heteroaromatic moieties.

[0532] As used herein, the term "alkenyl" includes unsaturated aliphatic groups similar in length and possible substitution to the alkyls described above, but containing at least one double bond. For example, the term "alkenyl" includes straight chain alkenyl groups (e.g., ethenyl, propenyl, butenyl, pentenyl, hexenyl, heptenyl, octenyl, nonenyl, decenyl) and branched alkenyl groups. In some embodiments, a straight chain or branched alkenyl group has 6 or fewer carbon atoms in its backbone (e.g., C2-C6 for straight chain, C3-C6 for branched chain). The term "C2-C6" includes alkenyl groups containing 2 to 6 carbon atoms. The term "C3-C6" includes alkenyl groups containing 3 to 6 carbon atoms.

[0533] As used herein, the term "optionally substituted alkenyl" refers to an unsubstituted alkenyl or an alkenyl having specified substituents replacing one or more hydrogen atoms on one or more hydrocarbon backbone carbon atoms. Such substituents may include, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, amino (including alkylamino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, heterocyclyl, alkylaryl, or aromatic or heteroaromatic moieties.

[0534] As used herein, the term "alkynyl" includes unsaturated aliphatic groups analogous in length and possible substitution to the alkyls described above, but containing at least one triple bond. For example, "alkynyl" includes straight chain alkynyl groups (e.g., ethynyl, propynyl, butynyl, pentynyl, hexynyl, heptynyl, octynyl, nonynyl, decynyl) and branched alkynyl groups. In some embodiments, a straight chain or branched alkynyl group has 6 or fewer carbon atoms in its backbone (e.g., C2-C6 for straight chain, C3-C6 for branched chain). The term "C2-C6" includes alkynyl groups containing 2 to 6 carbon atoms. The term "C3-C6" includes alkynyl groups containing 3 to 6 carbon atoms. As used herein, a "C2-C6 alkenylene linker" or a "C2-C6 alkynylene linker" is intended to include a C2, C3, C4, C5 or C6 chain (linear or branched) divalent unsaturated aliphatic hydrocarbon group. For example, a C2-C6 alkenylene linker is intended to include C2, C3, C4, C5 and C6 alkenylene linker groups.

[0535] As used herein, the term "optionally substituted alkynyl" refers to an unsubstituted alkynyl or an alkynyl having specified substituents replacing one or more hydrogen atoms on one or more hydrocarbon backbone carbon atoms. Such substituents may include, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, amino (including alkylamino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or aromatic or heteroaromatic moieties.

[0536] Other optionally substituted moieties (e.g., optionally substituted cycloalkyl, heterocycloalkyl, aryl, or heteroaryl) include both unsubstituted moieties and moieties with one or more of the specified substituents. For example, substituted heterocycloalkyl includes those substituted with one or more alkyl groups, such as 2,2,6,6-tetramethyl-piperidinyl and 2,2,6,6-tetramethyl-1,2,3,6-tetrahydropyridinyl.

[0537] As used herein, the term "cycloalkyl" refers to a group having 3 to 30 carbon atoms (e.g., C3-C 12 , C3-C 10, or C3-C8) saturated or partially unsaturated hydrocarbon monocyclic or polycyclic (e.g., fused, bridged, or spirocyclic) systems. Examples of cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclopentenyl, cyclohexenyl, cycloheptenyl, 1,2,3,4-tetrahydronaphthalenyl, and adamantyl. In the case of a polycyclic cycloalkyl, only one of the rings in the cycloalkyl must be non-aromatic.

[0538] The term "heterocycloalkyl," as used herein, unless otherwise specified, refers to a saturated or partially unsaturated 3-8 membered monocyclic, 7-12 membered bicyclic (fused, bridged, or spirocyclic), or 11-14 membered tricyclic ring system (fused, bridged, or spirocyclic), having one or more heteroatoms independently selected from the group consisting of nitrogen, oxygen, and sulfur (such as O, N, S, P, or Se), e.g., 1 or 1-2 or 1-3 or 1-4 or 1-5 or 1-6 heteroatoms, or e.g., 1, 2, 3, 4, 5, or 6 heteroatoms.Examples of heterocycloalkyl groups include piperidinyl, piperazinyl, pyrrolidinyl, dioxanyl, tetrahydrofuranyl, isoindolinyl, indolinyl, imidazolidinyl, pyrazolidinyl, oxazolidinyl, isoxazolidinyl, triazolidinyl, oxiranyl, azetidinyl, oxetanyl, thietanyl, 1,2,3,6-tetrahydropyridinyl, tetrahydropyranyl, dihydropyranyl, pyranyl, morpholinyl, tetrahydrothiopyranyl, 1,4-diazepanyl, 1,4-oxa Zepanil, 2-oxa-5-azabicyclo[2.2.1]heptanyl, 2,5-diazabicyclo[2.2.1]heptanyl, 2-oxa-6-azaspiro[3.3]heptanyl, 2,6-diazaspiro[3.3]heptanyl, 1,4-dioxa-8-azaspiro[4.5]decanyl, 1,4-dioxaspiro[4.5]decanyl, 1-oxaspiro[4.5]decanyl, 1-azaspiro[4.5]decanyl, 3'H-spiro[cyclohexane-1,1'-isobenzofuran]-yl, 7'H-spiro[cyclohex San-1,5'-furo[3,4-b]pyridin]-yl, 3'H-spiro[cyclohexane-1,1'-furo[3,4-c]pyridin]-yl, 3-azabicyclo[3.1.0]hexanyl, 3-azabicyclo[3.1.0]hexan-3-yl, 1,4,5,6-tetrahydropyrrolo[3,4-c]pyrazolyl, 3,4,5,6,7,8-hexahydropyrido[4,3-d]pyrimidinyl, 4,5,6,7-tetrahydro-1H-pyrazolo[3,4-c]pyridinyl, 5,6,7,8-tetrahydropyrido[4 ,3-d]pyrimidinyl, 2-azaspiro[3.3]heptanyl, 2-methyl-2-azaspiro[3.3]heptanyl, 2-azaspiro[3.5]nonanyl, 2-methyl-2-azaspiro[3.5]nonanyl, 2-azaspiro[4.5]decanyl, 2-methyl-2-azaspiro[4.5]decanyl, 2-oxa-azaspiro[3.4]octanyl, 2-oxa-azaspiro[3.4]octan-6-yl, 5,6-dihydro-4H-cyclopenta[b]thiophenyl, and the like.In the case of polycyclic heterocycloalkyls, only one of the rings in the heterocycloalkyl need be non-aromatic (eg, 4,5,6,7-tetrahydrobenzo[c]isoxazolyl).

[0539] As used herein, the term "aryl" includes groups with aromatic character, including "conjugated" or polycyclic systems that have one or more aromatic rings and do not contain any heteroatoms in the ring structure. The term aryl includes both monovalent and divalent species. Examples of aryl groups include, but are not limited to, phenyl, biphenyl, naphthyl, and the like. Conveniently, the aryl is phenyl.

[0540] As used herein, the term "heteroaryl" is intended to include a stable 5-, 6-, or 7-membered monocyclic or 7-, 8-, 9-, 10-, 11-, or 12-membered bicyclic heteroaromatic ring consisting of carbon atoms and one or more heteroatoms independently selected from the group consisting of nitrogen, oxygen, and sulfur, e.g., 1 or 1-2 or 1-3 or 1-4 or 1-5 or 1-6 heteroatoms, or, e.g., 1, 2, 3, 4, 5, or 6 heteroatoms. The nitrogen atom may be substituted or unsubstituted (i.e., N or NR, where R is H or other defined substituents). The nitrogen and sulfur heteroatoms may be optionally oxidized (i.e., N→O and S(O)). p(wherein p=1 or 2). It should be noted that the total number of S and O atoms in the aromatic heterocycle is not more than 1. Examples of heteroaryl groups include pyrrole, furan, thiophene, thiazole, isothiazole, imidazole, triazole, tetrazole, pyrazole, oxazole, isoxazole, isothiazole, pyridine, pyrazine, pyridazine, pyrimidine, and the like. Heteroaryl groups can also be fused or bridged with alicyclic or heterocyclic rings that are not aromatic so as to form polycyclic systems (e.g., 4,5,6,7-tetrahydrobenzo[c]isoxazolyl). In some embodiments, heteroaryl is thiophenyl or benzothiophenyl. In some embodiments, heteroaryl is thiophenyl. In some embodiments, heteroaryl is benzothiophenyl.

[0541] Furthermore, the terms "aryl" and "heteroaryl" include polycyclic aryl and heteroaryl groups, e.g., tricyclic, bicyclic, such as naphthalene, benzoxazole, benzodioxazole, benzothiazole, benzimidazole, benzothiophene, quinoline, isoquinoline, naphthyridine, indole, benzofuran, purine, benzofuran, deazapurine, indolizine.

[0542] A cycloalkyl, heterocycloalkyl, aryl, or heteroaryl ring may have at one or more ring positions (e.g., a ring-forming carbon or a heteroatom such as N) a substituent as described above, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkoxy, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, alkylaminocarbonyl, aralkylaminocarbonyl, alkenylaminocarbonyl, alkylcarbonyl, arylcarbonyl, aralkylcarbonyl, alkenylcarbonyl, alkoxycarbonyl, amino The aryl and heteroaryl groups may be substituted with carbonyl, alkylthiocarbonyl, phosphate, phosphonato, phosphinato, amino (including alkylamino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or aromatic or heteroaromatic moieties. The aryl and heteroaryl groups may also be fused or bridged with alicyclic or heterocyclic rings which are not aromatic so as to form polycyclic systems (e.g., tetralin, methylenedioxyphenyl such as benzo[d][1,3]dioxol-5-yl).

[0543] As used herein, the term "substituted" means that any one or more hydrogen atoms on the designated atom are replaced with one selected from the indicated group, provided that the normal valence of the designated atom is not exceeded and the replacement results in a stable compound. If the substituent is oxo or keto (i.e., =O), then two hydrogen atoms on the atom are replaced. Keto substituents are not present in aromatic moieties. A ring double bond, as used herein, is a double bond formed between two adjacent ring atoms (e.g., C=C, C=N, or N=N). "Stable compound" and "stable structure" are meant to indicate a compound that is sufficiently robust to survive isolation to a useful degree of purity from a reaction mixture, and formulation into an effective therapeutic agent.

[0544] When a bond to a substituent is shown to cross a bond connecting two atoms in a ring, such substituent may be bonded to any atom in the ring. When substituents are listed without indicating the atom to which such substituent is bonded to the remainder of the compound of a given formula, such substituent may be bonded through any atom in such formula. Combinations of substituents and / or variables are permissible only if such combinations result in stable compounds.

[0545] When any variable (e.g., R) occurs more than one time in any constituent or formula for a compound, its definition at each occurrence is independent of its definition at every other occurrence. Thus, for example, if a group is shown to be substituted with 0-2 R moieties, that group can be optionally substituted with up to 2 R moieties, and R at each occurrence is independently selected from the definitions of R. Additionally, combinations of substituents and / or variables are permissible only if such combinations result in stable compounds.

[0546] As used herein, the term "hydroxy" or "hydroxyl" refers to -OH or -O - The formula includes groups having the formula:

[0547] As used herein, the term "halo" or "halogen" refers to fluoro, chloro, bromo and iodo.

[0548] The terms "haloalkyl" or "haloalkoxyl" refer to an alkyl or alkoxyl substituted with one or more halogen atoms.

[0549] As used herein, the term "optionally substituted haloalkyl" refers to an unsubstituted haloalkyl having specified substituents replacing one or more hydrogen atoms on one or more hydrocarbon backbone carbon atoms. Such substituents may include, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, amino (including alkylamino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or aromatic or heteroaromatic moieties.

[0550] As used herein, the term "alkoxy" or "alkoxyl" includes substituted and unsubstituted alkyl, alkenyl, and alkynyl groups covalently bonded to an oxygen atom. Examples of alkoxy groups or alkoxyl radicals include, but are not limited to, methoxy, ethoxy, isopropyloxy, propoxy, butoxy, and pentoxy groups. Examples of substituted alkoxy groups include halogenated alkoxy groups. Alkoxy groups may be optionally substituted with groups such as alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, amino (including alkylamino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or aromatic or heteroaromatic moieties. Examples of halogen substituted alkoxy groups include, but are not limited to, fluoromethoxy, difluoromethoxy, trifluoromethoxy, chloromethoxy, dichloromethoxy, and trichloromethoxy.

[0551] As used herein, the expressions "one or more of A, B, or C," "one or more of A, B, or C," "one or more of A, B, and C," "one or more of A, B, and C," "selected from the group consisting of A, B, and C," "selected from A, B, and C," etc. are used interchangeably and all refer to a selection from the group consisting of A, B, and / or C, i.e., one or more A, one or more B, one or more C, or any combination thereof, unless otherwise indicated.

[0552] It should be understood that the present disclosure provides methods for the synthesis of the compounds, scaffolds, and conjugates described herein. The present disclosure also provides detailed methods for the synthesis of the various disclosed compounds, scaffolds, and conjugates according to the schemes herein and those shown in the Examples.

[0553] It should be understood that throughout the description, when a composition is described as having, including, or comprising certain components, it is contemplated that the composition also consists essentially of or consists of the recited components. Similarly, when a method or process is described as having, including, or comprising certain process steps, the process also consists essentially of or consists of the recited process steps. Furthermore, it should be understood that the order of steps, or order of performing certain actions, is not important so long as the invention remains operable. Moreover, two or more steps or actions can be performed simultaneously.

[0554] It is to be understood that the synthetic processes of the present disclosure can tolerate a wide variety of functional groups and therefore can employ a variety of substituted starting materials. Although these processes generally provide the desired final compound at or near the end of the overall process, in certain cases it may be desirable to further convert the compound to its pharma-ceutically acceptable salt.

[0555] It is understood that the compounds, scaffolds, and conjugates of the present disclosure can be prepared in a variety of ways using commercially available starting materials, compounds known in the literature, or from readily prepared intermediates, by employing standard synthetic methods and procedures known to those skilled in the art or apparent to those skilled in the art in light of the teachings herein. Standard synthetic methods and procedures for the preparation of organic molecules and functional group transformations and manipulations can be obtained from the relevant scientific literature or from standard textbooks in the field. Examples include, but are not limited to, any one or several sources, such as Smith, MB, March, J., March's Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 5, incorporated herein by reference. th edition,John Wiley & Sons:New York,2001;Greene,TW,Wuts,PGM,Protective Groups in Organic Synthesis,3 rd Representative texts such as R. Larock, Comprehensive Organic Transformations, VCH Publishers (1989); L. Fieser and M. Fieser, Fieser and Fieser's Reagents for organic synthesis, John Wiley and Sons (1994), and L. Paquette, ed., Encyclopedia of Reagents for organic synthesis, John Wiley and Sons (1995), are useful and recognized reference texts in organic synthesis known to those of skill in the art.

[0556] Those skilled in the art will note that the order of certain steps, such as the introduction and removal of protecting groups, may be altered in the reaction sequences and synthetic schemes described herein. Those skilled in the art will recognize that certain groups may require protection from reaction conditions by the use of protecting groups. Protecting groups may also be used to distinguish similar functional groups in a molecule. A list of protecting groups and methods for introducing and removing these groups can be found in Greene, TW, Wuts, PGM, Protective Groups in Organic Synthesis, 3 rd edition, John Wiley & Sons: New York, 1999.

[0557] Unless otherwise stated, any description of a method of treatment or prevention is understood to include the use of the compounds, scaffolds, and conjugates to provide treatment or prevention as described herein. Unless otherwise stated, any description of a method of treatment or prevention is further understood to include the use of the compounds, scaffolds, and conjugates to prepare a medicament for treating or preventing such a condition. Treatment or prevention includes treatment or prevention in humans or non-human animals, including rodents, and other disease models.

[0558] Unless otherwise specified, any description of a method of treatment should be understood to include the use of the compounds, scaffolds, and conjugates to provide treatment as described herein. Unless otherwise specified, any description of a method of treatment should be further understood to include the use of the compounds, scaffolds, and conjugates to prepare a medicament for treating such a condition. Treatment includes the treatment of humans or non-human animals, including rodents, and other disease models.

[0559] As used herein, the term "subject" is interchangeable with the term "subject in need thereof," both of which refer to a subject having a disease or at high risk of developing a disease. "Subject" includes mammals. The mammal may be, for example, a human or a suitable non-human mammal, such as a primate, mouse, rat, dog, cat, cow, horse, goat, camel, sheep, or pig. The subject may be a bird or poultry. In some embodiments, the mammal is a human. A subject in need may be a subject that has previously been diagnosed or identified as having a disease or disorder disclosed herein. A subject in need may also be a subject suffering from a disease or disorder disclosed herein. Alternatively, a subject in need may be a subject at high risk of developing such a disease or disorder compared to the population as a whole (i.e., a subject that is prone to developing such a disorder compared to the population as a whole). A subject in need may be refractory or resistant to a disease or disorder disclosed herein (i.e., a disease or disorder disclosed herein that does not respond or has not yet responded to treatment). The subject may be resistant at the start of treatment or may become resistant during treatment. In some embodiments, the subject in need has undergone and failed all known effective therapies for the disease or disorder disclosed herein. In some embodiments, the subject in need has undergone at least one prior treatment.

[0560] As used herein, the term "treating" or "treat" describes the management and care of a patient for the purpose of combating a disease, condition, or disorder, and includes the administration of a compound of the present disclosure, or a pharma- ceutically acceptable salt, polymorph, or solvate thereof, to alleviate the symptoms or complications of the disease, condition, or disorder, or to eliminate the disease, condition, or disorder. The term "treat" can also include the treatment of an in vitro cell or animal model. It should be understood that reference to "treat" or "treatment" includes the alleviation of established symptoms of a condition. Thus, "treating" a condition, disorder, or condition or "treatment" of a condition, disorder, or condition includes (1) preventing or delaying the appearance of clinical symptoms of the condition, disorder, or condition that occurs in a human who may be afflicted with or susceptible to the condition, disorder, or condition, but who has not yet undergone or exhibited clinical or preclinical symptoms of the condition, disorder, or condition; (2) inhibiting the condition, disorder, or condition, i.e., arresting, reducing, or delaying the onset of the disease or its recurrence (in the case of maintenance treatment), or at least one clinical or preclinical symptom thereof; or (3) relieving or attenuating the disease, i.e., causing regression of the condition, disorder, or condition, or at least one clinical or preclinical symptom thereof.

[0561] It is to be understood that the compounds, scaffolds, and conjugates of the present disclosure, or pharma- ceutically acceptable salts, polymorphs, or solvates thereof, may also be used to prevent the associated disease, condition, or disorder, or to identify suitable candidates for such purposes.

[0562] As used herein, the terms "preventing," "prevent" or "protecting from" describe reducing or eliminating the onset of symptoms or complications of such a disease, condition, or disorder.

[0563] It should be understood that the present disclosure also provides pharmaceutical compositions comprising any compound, scaffold, or conjugate described herein in combination with at least one pharma- ceutically acceptable excipient or carrier.

[0564] As used herein, the term "pharmaceutical composition" is a formulation containing the disclosed compound, scaffold, or conjugate in a form suitable for administration to a subject. In some embodiments, the pharmaceutical composition is in bulk or unit dosage form. The unit dosage form is any of a variety of forms including, for example, a capsule, an IV bag, a tablet, a single pump on an aerosol inhaler, or a vial. The amount of active ingredient (e.g., a formulation of the disclosed compound or its salt, hydrate, solvate, or isomer) in a unit dose of the composition is an effective amount and varies according to the specific treatment involved. Those skilled in the art will understand that it may be necessary to routinely vary the dosage depending on the age and condition of the patient. The dosage also varies depending on the route of administration. Various routes are contemplated, including oral, pulmonary, rectal, parenteral, transdermal, subcutaneous, intravenous, intramuscular, intraperitoneal, inhalation, buccal, sublingual, intrapleural, intrathecal, intranasal, etc. Dosage forms for topical or transdermal administration of the compounds of the present disclosure include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches, and inhalants. In some embodiments, the active compound is mixed under sterile conditions with a pharma- ceutically acceptable carrier, and any required preservatives, buffers, or propellants.

[0565] As used herein, the term "pharmacologically acceptable" refers to compounds, scaffolds, conjugates, anions, cations, materials, compositions, carriers, and / or dosage forms that are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without undue toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio.

[0566] As used herein, the term "pharmaceutically acceptable excipient" means an excipient that is generally safe, non-toxic, and not biologically or otherwise undesirable, which is useful in preparing pharmaceutical compositions, and includes excipients acceptable for veterinary and human pharmaceutical use. As used in the specification and claims, "pharmaceutically acceptable excipient" includes both one excipient and more than one such excipient.

[0567] It is understood that the pharmaceutical composition of the present disclosure is formulated to suit its intended route of administration. Examples of routes of administration include parenteral, e.g., intravenous, intradermal, subcutaneous, oral (e.g., ingestion), inhalation, transdermal (topical), and transmucosal administration. Solutions or suspensions used for parenteral, intradermal, or subcutaneous application can contain the following components: a sterile diluent, e.g., water for injection, saline, fixed oils, polyethylene glycols, glycerin, propylene glycol, or other synthetic solvents; antibacterial agents, e.g., benzyl alcohol or methylparabens; antioxidants, e.g., ascorbic acid or sodium bisulfite; chelating agents, e.g., ethylenediaminetetraacetic acid; buffers, e.g., acetates, citrates, or phosphates, and isotonicity adjusters, e.g., sodium chloride or dextrose. The pH can be adjusted with acids or bases, such as hydrochloric acid or sodium hydroxide. Parenteral preparations can be enclosed in ampoules, disposable syringes, or multiple dose vials made of glass or plastic.

[0568] It should be understood that the compounds or pharmaceutical compositions of the present disclosure can be administered to a subject in many of the well-known methods currently used for chemotherapy treatment.For example, the compounds of the present disclosure can be injected into the bloodstream or body cavity, or can be taken orally, or can be applied through the skin using a patch.The dose selected should be sufficient to constitute an effective treatment, but not so high as to cause unacceptable side effects.The state of the disease state (e.g., the disease or disorder disclosed herein) and the health of the patient should preferably be closely monitored during and for a reasonable period after treatment.

[0569] As used herein, the term "therapeutically effective amount" refers to an amount of an agent to treat, ameliorate or prevent a specified disease or condition, or to exhibit a detectable therapeutic or inhibitory effect. The effect can be detected by any assay method known in the art. The exact effective amount for a subject will vary depending on the subject's weight, size, and health, the nature and extent of the condition, and the therapeutic agent or combination of therapeutic agents selected for administration. The therapeutically effective amount for a given situation can be determined by routine experimentation that is within the skill and judgment of the clinician.

[0570] As used herein, the term "therapeutically effective amount" refers to an amount of an agent to treat or ameliorate a specified disease or condition, or to exhibit a detectable therapeutic or inhibitory effect. The effect can be detected by any assay method known in the art. The exact effective amount for a subject will vary depending on the subject's weight, size, and health, the nature and extent of the condition, and the therapeutic agent or combination of therapeutic agents selected for administration. The therapeutically effective amount for a given situation can be determined by routine experimentation that is within the skill and judgment of the clinician.

[0571] It is understood that for any compound, the therapeutically effective amount can be estimated initially in cell culture assays, for example, of neoplastic cells, or in animal models, usually rats, mice, rabbits, dogs, or pigs. Animal models can also be used to determine appropriate concentration ranges and routes of administration. Such information can then be used to determine useful doses and routes of administration in humans. Therapeutic / prophylactic efficacy and toxicity can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, for example, ED 50 (the dose that is therapeutically effective in 50% of the population), and LD 50 (the dose lethal to 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index, and the LD 50 / ED 50Pharmaceutical compositions that exhibit large therapeutic indices are preferred. The dosage can vary within this range depending on the dosage form used, sensitivity of the patient, and the route of administration.

[0572] Dosage and administration are adjusted to provide sufficient levels of active agent or to maintain the desired effect.Factors that may be considered include the severity of the disease state, the subject's general health, the subject's age, weight, and sex, diet, time and frequency of administration, drug combinations, reaction sensitivities, and tolerability / response to treatment.Long-acting pharmaceutical compositions can be administered every 3-4 days, every week, or once every two weeks, depending on the half-life and clearance rate of the particular formulation.

[0573] The pharmaceutical composition containing the active compound of the present disclosure can be generally manufactured in a known manner, for example, by conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, encapsulating or lyophilizing process.The pharmaceutical composition can be formulated in a conventional manner using one or more pharma-ceutically acceptable carriers, including excipients and / or auxiliary agents, which facilitate the processing of active compound into medicament-usable preparations.Of course, suitable formulations vary according to the selected administration route.

[0574] Pharmaceutical compositions suitable for injectable use include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. For intravenous administration, suitable carriers include physiological saline, bacteriostatic water, Cremophor EL™ (BASF, Parsippany, NJ) or phosphate buffered saline (PBS). In all cases, the composition must be sterile and fluid to the extent that easy syringability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof. The proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants. The action of microorganisms can be prevented by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In many cases, it is preferable to include isotonic agents, for example, sugars, polyalcohols such as mannitol and sorbitol, and sodium chloride in the composition. Prolonged absorption of the injectable compositions can be brought about by including in the composition an agent which delays absorption, for example, aluminum monostearate and gelatin.

[0575] Sterile injection solution can be prepared by incorporating the required amount of active compound into suitable solvent with one or combination of the above-listed components as required, followed by filtration sterilization.Generally, dispersion is prepared by incorporating active compound into a sterile vehicle that contains basic dispersion medium and other components required from above-listed.For the preparation of sterile powder for sterile injection solution, preparation method includes vacuum drying and freeze-drying, and obtains powder of active ingredient and any additional desired components from the solution that has been previously sterile-filtered.

[0576] Oral compositions generally contain an inert diluent or an edible pharma- ceutically acceptable carrier. They can be enclosed in gelatin capsules or compressed into tablets. For the purpose of oral therapeutic administration, the active compound can be incorporated with excipients and used in the form of tablets, troches, or capsules. Oral compositions can also be prepared using a fluid carrier for use as a mouthwash, where the compound in the fluid carrier is orally applied, swished, expectorated, or swallowed. Pharmaceutically compatible binding agents, and / or adjuvant materials can be included as part of the composition. The tablets, pills, capsules, troches and the like can contain any of the following ingredients, or compounds of a similar nature: a binder such as microcrystalline cellulose, gum tragacanth, or gelatin; an excipient such as starch or lactose, a disintegrating agent such as alginic acid, Primogel, or corn starch; a lubricant such as magnesium stearate or Sterotes; a glidant such as colloidal silicon dioxide; a sweetening agent such as sucrose or saccharin; or a flavoring agent such as peppermint, methyl salicylate, or orange flavoring.

[0577] For administration by inhalation, the compounds are delivered in the form of an aerosol spray from pressured container or dispenser which contains a suitable propellant, e.g., a gas such as carbon dioxide, or a nebulizer.

[0578] For intranasal administration, the compound is delivered in a solution or solid formulation. In some embodiments, the compound is delivered in a solution as a mist, drops, or swab. In some embodiments, the compound is delivered as a powder. In some embodiments, the compound is included in a kit that further includes an intranasal applicator.

[0579] Systemic administration may be by transmucosal or transdermal means. For transmucosal or transdermal administration, a penetrant suitable for the barrier to be permeated is used in the formulation. Such penetrants are generally known in the art, and include, for example, for transmucosal administration, detergents, bile salts, and fusidic acid derivatives. Transmucosal administration can be achieved by using nasal sprays or suppositories. For transdermal administration, the active compound is formulated into ointments, salves, gels, or creams, as generally known in the art.

[0580] The active compounds can be prepared with pharma- ceutically acceptable carriers that protect the compounds from rapid elimination from the body, such as controlled release formulations, including implants and microencapsulated delivery systems. Biodegradable, biocompatible polymers, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid, can be used. Methods for preparing such formulations will be clear to those skilled in the art. Materials can also be obtained commercially from Alza Corporation and Nova Pharmaceuticals, Inc. Liposomal suspensions, including liposomes targeted to infected cells with monoclonal antibodies against viral antigens, can also be used as pharma-ceutically acceptable carriers. These can be prepared according to methods known to those skilled in the art, for example, as described in U.S. Pat. No. 4,522,811.

[0581] For ease of administration and uniformity of dosage, it is particularly advantageous to formulate oral or parenteral compositions in unit dosage form.Unit dosage form as used herein refers to a physically separate unit suitable as a unit dose for the subject to be treated, each unit containing a predetermined amount of active compound calculated to produce desired therapeutic effect in association with necessary pharmaceutical carrier.The specification of unit dosage form of the present disclosure is determined by and directly depends on the unique characteristics of active compound and the specific therapeutic effect to be achieved.

[0582] In therapeutic applications, dosages of pharmaceutical compositions used in accordance with the present disclosure will vary depending on the agent, the age, weight, and clinical condition of the recipient patient, and the experience and judgment of the clinician or practitioner administering the treatment, among other factors that will affect the dosage selected. In general, the dosage should be sufficient to slow, preferably regress, and preferably cause complete regression of the symptoms of the disease or disorder disclosed herein. Dosages may range from about 0.01 mg / kg / day to about 5000 mg / kg / day. An effective amount of a pharmaceutical agent is an amount that provides an objectively identifiable improvement as noted by a clinician or other qualified observer. Improved survival and growth indicate regression. As used herein, the term "dosage effective manner" refers to an amount of an active compound that produces a desired biological effect in a subject or cell.

[0583] It should be understood that the pharmaceutical compositions can be included in a container, pack, or dispenser together with instructions for administration.

[0584] For compounds, scaffolds, or conjugates of the present disclosure that are capable of further forming salts, it is understood that all of these forms are also contemplated within the scope of the claimed disclosure.

[0585] As used herein, the term "pharmaceutical acceptable salts" refers to derivatives of the disclosed compounds, where the parent compound is modified by making its acid or base salt. Examples of pharmaceutical acceptable salts include, but are not limited to, mineral acid organic acid salts of basic residues such as amines, alkali organic salts of acidic residues such as carboxylic acids, and the like. Pharmaceutically acceptable salts include the conventional non-toxic salts or quaternary ammonium salts of the parent compound, for example, formed from non-toxic inorganic and organic acids. For example, such conventional non-toxic salts include 2-acetoxybenzoic acid, 2-hydroxyethanesulfonic acid, acetic acid, ascorbic acid, benzenesulfonic acid, benzoic acid, bicarbonate, carbonic acid, citric acid, edetic acid, ethanedisulfonic acid, 1,2-ethanesulfonic acid, fumaric acid, glucoheptonic acid, gluconic acid, glutamic acid, glycolic acid, glycollyarsanilic acid, hexylresorcinic acid, hydrabamic acid, hydrobromic acid, hydrochloric acid, hydroiodic acid, hydroxymaleic acid, hydroxyisobutyl acid, ethyl ... These include, but are not limited to, those derived from inorganic and organic acids selected from naphthoic, isethionic, lactic, lactobionic, laurylsulfonic, maleic, malic, mandelic, methanesulfonic, napsylic, nitric, oxalic, pamoic, pantothenic, phenylacetic, phosphoric, polygalacturonic, propionic, salicylic, stearic, hypoacetic, succinic, sulfamic, sulfanilic, sulfuric, tannic, tartaric, toluenesulfonic, and commonly occurring amino acids such as glycine, alanine, phenylalanine, arginine, and the like.

[0586] In some embodiments, the pharma- ceutically acceptable salt is a sodium salt, a potassium salt, a calcium salt, a magnesium salt, a diethylamine salt, a choline salt, a meglumine salt, a benzathine salt, a tromethamine salt, an ammonia salt, an arginine salt, or a lysine salt.

[0587] Other examples of pharma- ceutically acceptable salts include hexanoic acid, cyclopentanepropionic acid, pyruvic acid, malonic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, 4-methylbicyclo-[2.2.2]-oct-2-ene-1-carboxylic acid, 3-phenylpropionic acid, trimethylacetic acid, tert-butylacetic acid, muconic acid, etc. The present disclosure also encompasses salts formed when acidic protons present in the parent compound are either replaced by metal ions, e.g., alkali metal ions, alkaline earth ions, or aluminum ions, or when coordinated with organic bases, such as ethanolamine, diethanolamine, triethanolamine, tromethamine, N-methylglucamine, and the like. It is understood that in the salt form, the ratio of the compound to the cation or anion of the salt can be 1:1, or any ratio other than 1:1, for example, 3:1, 2:1, 1:2, or 1:3.

[0588] It should be understood that all references to pharma- ceutically acceptable salts include the solvent addition forms (solvates) or crystal forms (polymorphs), as defined herein, of the same salt.

[0589] The compound or a pharma- ceutically acceptable salt thereof may be administered orally, nasally, transdermally, pulmonary, inhalation, buccal, sublingually, intraperitoneally, subcutaneously, intramuscularly, intravenously, rectally, intrapleurally, intrathecally, and parenterally. In some embodiments, the compound is administered orally. Those skilled in the art will recognize the advantages of certain routes of administration.

[0590] The dosage regimen utilizing the compound is selected according to a variety of factors, including the type, species, age, weight, sex, and medical condition of the patient, the severity of the condition to be treated, the route of administration, the renal and hepatic function of the patient, and the particular compound or salt thereof used. A physician or veterinarian of ordinary skill can readily determine and prescribe the effective amount of the drug required to prevent, counter, or halt the progression of the condition. A physician or veterinarian of ordinary skill can readily determine and prescribe the effective amount of the drug required to counter or halt the progression of the condition.

[0591] Techniques for formulation and administration of the disclosed compounds of this disclosure can be found in Remington: The Science and Practice of Pharmacy, 1999. th edition, Mack Publishing Co., Easton, PA (1995). In one embodiment, the compounds described herein and their pharma- ceutically acceptable salts are used in pharmaceutical preparations in combination with a pharma- ceutically acceptable carrier or diluent. Suitable pharma- ceutically acceptable carriers include inert solid fillers or diluents and sterile organic aqueous solutions. The compounds are present in such pharmaceutical compositions in an amount sufficient to provide the desired dosage within the range described herein.

[0592] All percentages and ratios used herein are by weight unless otherwise indicated. Other features and advantages of the present disclosure are apparent from the various examples. The examples provided illustrate different components and methodologies useful in implementing the present disclosure. The examples do not limit the claimed disclosure. Based on the present disclosure, one skilled in the art can identify and use other components and methods useful in implementing the present disclosure.

[0593] In the synthesis schemes described herein, compounds may be drawn in one specific configuration for simplicity. Such specific configurations should not be interpreted as limiting the present disclosure to one or another isomer, tautomer, positional isomer, or stereoisomer, nor do they exclude mixtures of isomers, tautomers, positional isomers, or stereoisomers. However, it will be understood that a given isomer, tautomer, positional isomer, or stereoisomer may have a higher level of activity than another isomer, tautomer, positional isomer, or stereoisomer.

[0594] All publications and patent documents are incorporated herein by reference to the same extent as if each such publication or patent document was specifically and individually indicated to be incorporated herein by reference. Citation of publications and patent documents is not intended as an admission that any is relevant prior art, and does not constitute any admission as to its contents or date. Although the present invention has been described by way of written description, those skilled in the art will recognize that the present invention can be practiced in various embodiments, and that the foregoing description and the following examples are intended to be illustrative and not limiting of the scope of the following claims.

[0595] Exemplary embodiments Exemplary embodiment 1. A compound of formula (I) or (II), [ka] or a pharma- ceutically acceptable salt thereof, wherein: B is H or a nucleobase moiety; W is H, C1-C6 alkyl optionally substituted with one or more halogens, or an amino substituent; Y is H, C1-C6 alkyl optionally substituted with one or more halogens, -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y , -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(OR Y )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, -P(=S)(SR Y ) 2, or a hydroxy protecting group; Each R Y is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; Z is H or C1-C6 alkyl optionally substituted with one or more halogens, -P(R Z )2, -P(OR Z )(N(R Z )2), -P(=O)(OR Z )R Z , -P(=S)(OR Z )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z )2, -P(=S)(SR Z ) 2, or a hydroxy protecting group; Each R Z is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; or Y and Z in formula (I) together represent -Si(R L )2-O-Si(R L )2-, and each R L is independently H or C1-C6 alkyl; Each R a are independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens, or two R a forms a double bond with two adjacent carbon atoms, Each R b is independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 1 is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 2 is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 3is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; R 4 is H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; Each R 5 is independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; A compound wherein n is an integer ranging from about 0 to about 10.

[0596] Exemplary embodiment 2. A scaffold or a pharma- ceutically acceptable salt thereof, comprising: (i) a ligand, and (ii) a linker unit, the linker unit being [ka] and a linker unit, where variables B, R 1 , R 2 , R 3 , R 4 , R 5 , Y, Z, R a , R b and n is as described herein and # indicates attachment to a ligand, or a pharma- ceutically acceptable salt thereof.

[0597] Exemplary embodiment 3. A scaffold or a pharma- ceutically acceptable salt thereof, comprising: (i) one or more nucleic acid agents, and (ii) one or more linker units, each linker unit independently comprising: [ka] and one or more linker units, where variables B, R 1 , R 2 , R 3 , R 4 , R 5 , W, Y, Z, R a , R band n is described herein and ## indicates attachment to a nucleic acid agent, or a pharma- ceutically acceptable salt thereof.

[0598] Exemplary embodiment 4. A conjugate or a pharma- ceutically acceptable salt thereof, wherein the scaffold comprises: (i) one or more nucleic acid agents, (ii) one or more ligands, and (iii) one or more linker units, each linker unit independently comprising: [ka] and one or more linker units, where variables B, R 1 , R 2 , R 3 , R 4 , R 5 , Y, Z, R a , R b and n are described herein, # indicates attachment to a ligand, and ## indicates attachment to a nucleic acid agent, or a pharma- ceutically acceptable salt thereof.

[0599] Exemplary Embodiment 5. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein B is H.

[0600] Exemplary Embodiment 6. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein B is a nucleobase moiety.

[0601] Exemplary Embodiment 7. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein the nucleobase moiety is adenine (A), cytosine (C), guanine (G), thymine (T), or uracil (U).

[0602] Exemplary Embodiment 8 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein the nucleobase moiety is a modified nucleobase.

[0603] Exemplary Embodiment 9 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein the nucleobase moiety is an artificial nucleobase.

[0604] Exemplary Embodiment 10. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein W is H.

[0605] Exemplary Embodiment 11. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein W is C1-C6 alkyl.

[0606] Exemplary Embodiment 12. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein W is an amino substituent.

[0607] Exemplary embodiment 13. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein W is fluorenylmethyloxycarbonyl (Fmoc), tert-butyloxycarbonyl (BOC), benzyloxycarbonyl (Cbz), optionally substituted acyl, trifluoroacetyl (TFA), benzyl, triphenylmethyl (Tr), 4,4'-dimethoxytrityl (DMTr), or toluenesulfonyl (Ts).

[0608] Exemplary Embodiment 14. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein W is optionally substituted acyl.

[0609] Exemplary Embodiment 15. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein W is trifluoroacetyl (TFA).

[0610] Exemplary Embodiment 16. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein Y is C1-C6 alkyl optionally substituted with one or more halogens.

[0611] Exemplary embodiment 17. Y is -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y , -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(OR Y )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, -P(=S)(SR Y )2. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments.

[0612] Exemplary Embodiment 18. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein Y is a hydroxy protecting group.

[0613] Exemplary Embodiment 19. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein Y is silyl.

[0614] Exemplary Embodiment 20. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein Y is triphenylmethyl (Tr) or 4,4'-dimethoxytrityl (DMTr).

[0615] Exemplary Embodiment 21. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein Y is optionally substituted acyl or benzyl.

[0616] Exemplary embodiment 22. At least one R Y The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0617] Exemplary embodiment 23. At least one R YThe compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0618] Exemplary embodiment 24. At least one R Y is H and at least one R Y The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0619] Exemplary Embodiment 25. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein Z is H.

[0620] Exemplary Embodiment 26. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein Z is C1-C6 alkyl optionally substituted with one or more halogens.

[0621] Exemplary embodiment 27. Z is -P(R Z )2, -P(OR Z )(N(R Z )2), -P(=O)(OR Z )R Z , -P(=S)(OR Z )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z )2, -P(=S)(SR Z )2. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments.

[0622] Exemplary Embodiment 28. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein Z is a hydroxy protecting group.

[0623] Exemplary Embodiment 29. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein Z is silyl.

[0624] Exemplary Embodiment 30. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein Z is triphenylmethyl (Tr) or 4,4'-dimethoxytrityl (DMTr).

[0625] Exemplary Embodiment 31. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein Z is substituted acyl or benzyl.

[0626] Exemplary embodiment 32. At least one R Z The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0627] Exemplary embodiment 33. At least one R Z The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0628] Exemplary embodiment 34. At least one R Z is H and at least one R Z The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0629] Exemplary embodiment 35. Y and Z in formula (I) together represent -Si(R L )2-O-Si(R L )2-forming the compound, scaffold, or conjugate of any one of the preceding exemplary embodiments.

[0630] Exemplary embodiment 36. At least one R LThe compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0631] Exemplary embodiment 37. Each R L is independently C1-C6 alkyl.

[0632] Exemplary embodiment 38. Each R a The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0633] Exemplary embodiment 39. At least one R a The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is halogen, or C1-C6 alkyl optionally substituted with one or more halogens.

[0634] Exemplary embodiment 40. Each R b The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0635] Exemplary embodiment 41. At least one R b The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is halogen, or C1-C6 alkyl optionally substituted with one or more halogens.

[0636] Exemplary embodiment 42.R 1 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0637] Exemplary embodiment 43.R 1 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is a halogen.

[0638] Exemplary embodiment 44.R 1The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens.

[0639] Exemplary embodiment 45.R 2 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0640] Exemplary embodiment 46.R 2 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0641] Exemplary embodiment 47.R 2 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens.

[0642] Exemplary embodiment 48.R 3 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0643] Exemplary embodiment 49.R 3 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0644] Exemplary embodiment 50.R 3 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens.

[0645] Exemplary embodiment 51.R 4 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0646] Exemplary embodiment 52.R 4 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0647] Exemplary embodiment 53.R 4 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens.

[0648] Exemplary embodiment 54.R 5 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0649] Exemplary embodiment 55.R 5 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein

[0650] Exemplary embodiment 56.R 5 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens.

[0651] Exemplary embodiment 57.R a , R b , R 1 , R 2 , R 3 , R 4 , and R 5 The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein each of is H.

[0652] Exemplary embodiment 58. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein n is an integer in the range of about 1 to about 10, about 2 to about 10, about 3 to about 10, about 4 to about 10, about 5 to about 10, or about 6 to about 10.

[0653] Exemplary Embodiment 59. The compound, scaffold, or conjugate of any one of the preceding exemplary embodiments, wherein n is an integer in the range of about 2 to about 8, about 2 to about 7, about 2 to about 6, about 2 to about 5, about 2 to about 4, or about 2 to about 3.

[0654] Exemplary embodiment 60. The compound has formula (I') or (II'): [ka] Things, or a pharma- ceutically acceptable salt thereof.

[0655] Exemplary embodiment 61. The compound has formula (IA) or (II-A): [ka] Things, or a pharma- ceutically acceptable salt thereof.

[0656] Exemplary embodiment 62. The compound has formula (I'-A) or (II'-A): [ka] Things, or a pharma- ceutically acceptable salt thereof.

[0657] Exemplary embodiment 63. The compound has formula (IB) or (II-B): [ka] Things, or a pharma- ceutically acceptable salt thereof.

[0658] Exemplary embodiment 64. The compound has formula (I'-B) or (II'-B): [ka] Things, or a pharma- ceutically acceptable salt thereof.

[0659] Exemplary embodiment 65. Y is a hydroxy protecting group and Z is a hydroxy protecting group; or In formula (I), (I'), (IA), (I'-A), (IB), or (I'-B), Y and Z together represent -Si(R L )2-O-Si(R L )2-, and each R L is independently H or C1-C6 alkyl.

[0660] Exemplary embodiment 66. The compound comprises: [ka] or a pharma- ceutically acceptable salt thereof, wherein: B is a nucleobase moiety (e.g., adenine (A), cytosine (C), guanine (G), thymine (T), or uracil (U)); Y is -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y , -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(OR Y )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, -P(=S)(SR Y )2, or a hydroxy protecting group (e.g., silyl (e.g., trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl), triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), substituted acyl (e.g., optionally substituted acetyl), or benzyl); Each R Yis independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; Z is -P(R Z )2, -P(OR Z )(N(R Z )2), -P(=O)(OR Z )R Z , -P(=S)(OR Z )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z )2, -P(=S)(SR Z )2, or a hydroxy protecting group (e.g., silyl (e.g., trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl), triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), substituted acyl (e.g., optionally substituted acetyl), or benzyl); Each R Z The compound of any one of the preceding exemplary embodiments, wherein is independently H, or C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0661] Exemplary Embodiment 67. The compound of any one of the preceding exemplary embodiments, wherein the compound is selected from the compounds set forth in Table L and pharma- ceutically acceptable salts thereof.

[0662] Exemplary embodiment 68. A compound that is an isotopic derivative of the compound of any one of the preceding exemplary embodiments.

[0663] Exemplary embodiment 69. The scaffold comprises (linker unit) p -((nucleic acid agent)-(linker unit) s ) r -(Nucleic acid agents) q and each linker unit is independent of another linker unit, and each nucleic acid agent is independent of another nucleic acid agent; each r is independently an integer ranging from 0 to 10; each s is independently an integer ranging from 0 to 10; p is an integer ranging from 0 to 10; q is 0 or 1; The scaffold of any one of the preceding exemplary embodiments, wherein the scaffold comprises at least one linker unit and at least one nucleic acid agent.

[0664] Exemplary embodiment 70. The scaffold comprises: [ka] or a pharma- ceutically acceptable salt thereof, wherein: B is a nucleobase moiety (e.g., adenine (A), cytosine (C), guanine (G), thymine (T), or uracil (U)); Y is -P(R Y )2, -P(OR Y )(N(R Y )2), -P(=O)(OR Y )R Y , -P(=S)(OR Y )R Y , -P(=O)(SR Y )R Y , -P(=S)(SR Y )R Y , -P(=O)(OR Y )2, -P(=S)(OR Y )2, -P(=O)(SR Y )2, -P(=S)(SR Y )2, or a hydroxy protecting group (e.g., silyl (e.g., trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl), triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), substituted acyl (e.g., optionally substituted acetyl), or benzyl); Each R Yis independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; Z is -P(R Z )2, -P(OR Z )(N(R Z )2), -P(=O)(OR Z )R Z , -P(=S)(OR Z )R Z , -P(=O)(SR Z )R Z , -P(=S)(SR Z )R Z , -P(=O)(OR Z )2, -P(=S)(OR Z )2, -P(=O)(SR Z )2, -P(=S)(SR Z )2, or a hydroxy protecting group (e.g., silyl (e.g., trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl), triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), substituted acyl (e.g., optionally substituted acetyl), or benzyl); Each R Z is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; The scaffold of any one of the preceding exemplary embodiments, wherein n is an integer ranging from about 0 to about 10.

[0665] Exemplary Embodiment 71. The scaffold of any one of the preceding exemplary embodiments, wherein the scaffold is selected from the scaffolds listed in Table S1.

[0666] Exemplary embodiment 72. The scaffold comprises: [ka] or a pharma- ceutically acceptable salt thereof, wherein: B is a nucleobase moiety (e.g., adenine (A), cytosine (C), guanine (G), thymine (T), or uracil (U)); W is an amino substituent (e.g., fluorenylmethyloxycarbonyl (Fmoc), tert-butyloxycarbonyl (BOC), benzyloxycarbonyl (Cbz), optionally substituted acyl, trifluoroacetyl (TFA), benzyl, triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), or toluenesulfonyl (Ts), acyl (e.g., —C(═O)(C1-C 30 alkyl), substituted acyls (e.g., [ka] ), trifluoroacetyl (TFA), -C(=O)(C1-C 30 alkyl), -C(=O)NH(C1-C 30 Alkyl), -C(=S)(C1-C 30 alkyl), or -C(=S)NH(C1-C 30 alkyl) (wherein C1-C 30 alkyl is optionally substituted) The scaffold of any one of the preceding exemplary embodiments, wherein n is an integer ranging from about 0 to about 10.

[0667] Exemplary Embodiment 73. The scaffold of any one of the preceding exemplary embodiments, wherein the scaffold is selected from the scaffolds listed in Table S2.

[0668] Exemplary embodiment 74. The conjugate comprises (linker unit - (ligand) 0-1 ) p -((nucleic acid agent)-(linker unit-(ligand) 0-1 ) s ) r -(Nucleic acid agents) q where: each linker unit is independent of other linker units, each nucleic acid agent is independent of other nucleic acid agents, and each ligand is independent of other ligands; each r is independently an integer ranging from 0 to 10; each s is independently an integer ranging from 0 to 10; p is an integer ranging from 0 to 10; q is 0 or 1; The conjugate of any one of the preceding exemplary embodiments, wherein the conjugate comprises at least one linker unit, at least one nucleic acid agent, and at least one ligand.

[0669] Exemplary Embodiment 75. The conjugate of any one of the preceding exemplary embodiments, wherein the conjugate is selected from the conjugates listed in Table C.

[0670] Exemplary Embodiment 76. The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the linker unit is of formula (I), wherein W is replaced with a bond to a ligand.

[0671] Exemplary Embodiment 77. The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the linker unit is of formula (I), wherein Y and / or Z is replaced with a bond to a nucleic acid agent.

[0672] Exemplary Embodiment 78. The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the linker unit is of formula (II), wherein W is replaced with a bond to a ligand.

[0673] Exemplary Embodiment 79. The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the linker unit is of formula (II), wherein Y and / or Z is replaced with a bond to a nucleic acid agent.

[0674] Exemplary Embodiment 80 The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the ligand comprises a carbohydrate moiety.

[0675] Exemplary Embodiment 81 The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the carbohydrate moiety comprises a monosaccharide, a disaccharide, a trisaccharide, or a tetrasaccharide.

[0676] Exemplary Embodiment 82 The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the carbohydrate moiety comprises galactose or a derivative thereof.

[0677] Exemplary embodiment 83. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0678] Exemplary embodiment 84. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0679] Exemplary embodiment 85. The ligand comprises: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0680] Exemplary embodiment 86. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0681] Exemplary embodiment 87. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0682] Exemplary embodiment 88. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0683] Exemplary embodiment 89. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0684] Exemplary embodiment 90. The ligand comprises: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0685] Exemplary embodiment 91. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0686] Exemplary embodiment 92. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0687] Exemplary embodiment 93. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0688] Exemplary embodiment 94. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0689] Exemplary embodiment 95. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0690] Exemplary embodiment 96. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0691] Exemplary embodiment 97. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0692] Exemplary embodiment 98. The ligand is: [ka] The scaffold or conjugate of any one of the preceding exemplary embodiments, comprising:

[0693] Exemplary Embodiment 99. The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the ligand comprises a lipid.

[0694] Exemplary Embodiment 100. The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the ligand comprises a peptide moiety.

[0695] Exemplary Embodiment 101. The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the ligand comprises an antibody moiety.

[0696] Exemplary Embodiment 102 The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the nucleic acid agent comprises an oligonucleotide.

[0697] Exemplary Embodiment 103. The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the nucleic acid agent comprises one or more phosphate groups or one or more analogs of phosphate groups.

[0698] Exemplary Embodiment 104. The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the linker unit is attached to the nucleic acid agent via a phosphate group or an analog of a phosphate group in the nucleic acid agent.

[0699] Exemplary Embodiment 105 The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the nucleic acid agent comprises RNA.

[0700] Exemplary embodiment 106. The scaffold or conjugate of any one of the preceding exemplary embodiments, wherein the oligonucleotide is an siRNA, a microRNA, an anti-microRNA, a microRNA mimic, an antagomir, a dsRNA, a ssRNA, an aptamer, an immunostimulatory oligonucleotide, a decoy oligonucleotide, a splice variant oligonucleotide, a triplex forming oligonucleotide, a G-quadruplex, or an antisense oligonucleotide.

[0701] Exemplary embodiment 107. A pharmaceutical composition comprising a compound, scaffold, or conjugate of any one of the preceding exemplary embodiments.

[0702] Exemplary embodiment 108. A method for regulating expression of a target gene in a subject, comprising administering to the subject a conjugate of any one of the preceding exemplary embodiments.

[0703] Exemplary embodiment 109. A method of delivering a nucleic acid agent to a subject, comprising administering to the subject a conjugate of any one of the preceding exemplary embodiments.

[0704] Exemplary embodiment 110. A method of treating or preventing a disease in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a conjugate of any one of the preceding exemplary embodiments.

[0705] Exemplary embodiment 111. A conjugate of any one of the preceding exemplary embodiments for modulating expression of a target gene in a subject.

[0706] Exemplary embodiment 112. The conjugate of any one of the preceding exemplary embodiments for delivering a nucleic acid agent to a subject.

[0707] Exemplary embodiment 113. A conjugate of any one of the preceding exemplary embodiments for the treatment or prevention of a disease in a subject in need thereof.

[0708] Exemplary embodiment 114. Use of a conjugate of any one of the preceding exemplary embodiments in the manufacture of a medicament for modulating expression of a target gene in a subject.

[0709] Exemplary embodiment 115. Use of a conjugate of any one of the preceding exemplary embodiments in the manufacture of a medicament for delivering a nucleic acid agent to a subject.

[0710] Exemplary embodiment 116. Use of a conjugate of any one of the preceding exemplary embodiments in the manufacture of a medicament for the treatment or prevention of a disease in a subject in need thereof.

[0711] Exemplary embodiment 117. The method, conjugate, or use of any one of the preceding exemplary embodiments, wherein the subject is a human. EXAMPLES

[0712] Example 1. Chemical structure and synthesis scheme of 2'-C-alkyl-GalNAc (GalNAc2) [ka]

[0713] Synthesis of 1-((6aR,8R,9R,9aS)-9-hydroxy-2,2,4,4-tetraisopropyltetrahydro-6H-furo[3,2-f][1,3,5,2,4]trioxadisilosin-8-yl)pyrimidine-2.4(1H,3H)-dione (1-2). To a solution of compound 1-1 (44.6 g, 182.64 mmol) in pyridine (446 mL) was added TIPSCl2 (63.4 g, 200.90 mmol) at 0° C., and the mixture was stirred at 25° C. for 16 h. The reaction was quenched with MeOH and concentrated in vacuum. The residue was dissolved in EtOAc (500 mL), washed with aqueous citric acid (500 mL×2) and brine (500 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuum. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 3 / 1 to 1 / 2) to give compound 1-2 (75.6 g, 155.33 mmol, yield 85.05%) as a white solid. 1 H NMR: 400MHz, DMSO-d 6, δ 11.37(s,1H),7.68(d,J=8.0Hz,1H),5.59(d,J=4.8Hz,1H),5.53-5.51(m,2H),4.15-4.03(m,3H),3.98-3.93(m,2H),1.17-0.96(m,28H).

[0714] Synthesis of O-((6aR,8R,9R,9aR)-8-(2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-2,2,4,4-tetraisopropyltetrahydro-6H-furo[3,2-f][1,3,5,2,4]trioxadisilosin-9-yl)O-phenylcarbonothioate (1-3). To a solution of compound 1-2 (37.5 g, 77.05 mmol) in ACN (150 mL) was added a solution of DMAP (18.83 g, 154.10 mmol) and PhOCSCl (16.0 g, 92.46 mmol, 12.77 mL, 1.2 equiv.) in ACN (150 mL) at 0° C. The mixture was stirred at 15° C. for 1 h. The reaction mixture was then dissolved in DCM (500 mL), washed with brine (500 mL), dried over anhydrous Na2SO4, filtered and concentrated under vacuum. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 10 / 1 to 1 / 1) to give compound 1-3 (64.0 g, 102.75 mmol, yield 66.7%) as a white solid. 1 H NMR: 400MHz, CDCl 3, δ 8.11(s,1H),7.75(d,J=4.0Hz,1H),7.72-7.44(m,2H),7.41-7.31(m,1 H),7.26-7.25(m,1H),7.14-7.12(m,2H),6.85-6.83(m,1H),6.02(d,J =4.8Hz,1H),5.75(s,1H),5.75-5.72(m,1H),4.58-4.55(m,1H),4.28- 4.25(m,1H),4.14-4.11(m,2H),4.06-4.05(m,1H),1.12-1.00(m,28H).

[0715] Synthesis of 1-((6aR,8R,9R,9aS)-9-allyl-2,2,4,4-tetraisopropyltetrahydro-6H-furo[3,2-f][1,3,5,2,4]trioxadicyclosin-8-yl)pyrimidine-2.4(1H,3H)-dione (1-4). To a solution of compound 1-3 (67.0 g, 107.57 mmol) in toluene (670 mL) was added allyl(tributyl)stannane (180.4 g, 544.81 mmol). Then, a solution of BPO (6.70 g, 27.66 mmol) in toluene (670 mL) was added in small portions at 120° C. for 1 h. The mixture was stirred at 120° C. for 15 h and then concentrated under vacuum. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 10 / 1 to 3 / 1) to give compound 1-4 (22.0 g, 43.07 mmol, yield 40.0%) as a yellow oil. 1 H NMR: 400MHz, CDCl 3, δ 8.14(s,1H),7.73(d,J=8.4Hz,1H),5.96-5.81(m,1H),5.80(s,1H),5.70-5.68(m,1H),5.18-5.10(m,2H),4.51( t,J=7.6Hz,1H),4.16-4.10(m,2H),4.03-3.95(m,2H),2.65-2.61(m,1H),2.32-2.22(m,2H),1.10-1.03(m,28H).

[0716] Synthesis of 1-((6aR,8R,9R,9aS)-9-(3-hydroxypropyl)-2,2,4,4-tetraisopropyltetrahydro-6H-furo[3,2-f][1,3,5,2,4]trioxadisilosin-8-yl)pyrimidine-2.4(1H,3H)-dione (1-5). To a solution of compound 1-4 (6.00 g, 11.75 mmol) in THF (30 mL) was added 9-BBN (0.5 M in THF, 140.96 mL) at 15° C. and the mixture was stirred for 1 h. NaBO3.4(HO) (10.8 g, 70.48 mmol, 13.56 mL) and HO (24 mL) were added to the reaction and the mixture was stirred for an additional 1 h. The mixture was diluted with EtOAc (200 mL), washed with brine (200 mL), dried over anhydrous Na2SO4, filtered and concentrated in vacuo. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 10 / 1 to 1 / 2) to give compound 1-5 (4.20 g, 7.94 mmol, yield 67.6%) as a colorless oil. 1 H NMR: 400MHz, CDCl 3, δ 9.80(s,1H),7.89(d,J=8.0Hz,1H),5.80-5.70(m,2H),4.43(t,J=8.0Hz,1H),4.20-4.17(m,1H),4.00- 3.89(m,2H),3.71-3.65(m,2H),2.28-2.23(m,2H),1.95-1.93(m,4H),1.54(m,1H),1.11-0.95(m,28H).

[0717] Synthesis of 3-((6aR,8R,9R,9aS)-8(2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-2,2,4,4-tetraisopropyltetrahydro-6H-furo[3,2-f][1,3,5,2,4]trioxadisilosin-9-yl)propyl methanesulfonate (1-6). To a solution of compound 1-5 (11.0 g, 20.80 mmol) in DCM (110 mL) was added TEA (4.21 g, 41.60 mmol) and MsCl (3.12 g, 27.24 mmol, 2.11 mL) at 0° C. The mixture was stirred at 15° C. for 1 h. The reaction mixture was then quenched with water (10 mL) at 0° C. and extracted with DCM (3×100 mL). The combined organic layers were washed with brine, dried, filtered and concentrated under reduced pressure to give compound 1-6 (12.6 g, crude) as a yellow oil, which was used in the next step without further purification.

[0718] Synthesis of 1-((6aR,8R,9R,9aS)-9-(3-azidopropyl)-2,2,4,4-tetraisopropyltetrahydro-6H-furo[3,2-f][1,3,5,2,4]trioxadicyclosin-8-yl)pyrimidine-2.4(1H,3H)-dione (1-7). To a solution of compound 1-6 (12.6 g, 20.80 mmol) in DMF (120 mL) was added NaN3 (2.68 g, 41.22 mmol) at 15 °C. The mixture was stirred at 50 °C for 1 h. The reaction mixture was then quenched with aqueous NaHCO3 (600 mL) and extracted with EtOAc (300 mL). The organic layer was washed with brine (500 mL), dried over Na2SO4, filtered and concentrated in vacuo to give compound 1-7 (11.5 g, crude) as a yellow oil, which was used in the next step without further purification.

[0719] Synthesis of 1-((2R,3R,4S,5R)-3-(3-azidopropyl)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (1-8). To a solution of compound 1-7 (11.5 g, 20.80 mmol) in MeOH (110 mL) was added NHF (7.70 g, 208.02 mmol). The mixture was stirred at 60 °C for 2 h and concentrated under vacuum. The residue was purified by column chromatography (SiO, DCM:MeOH = 50:1 to 10:1) to give compound 1-8 (3.00 g, 9.64 mmol, 46.3% yield) as a white solid. 1 H NMR: 400MHz, DMSO-d 6, δ 7.84(d,J=8.0Hz,1H),5.86(d,J=9.2Hz,1H),5.69-5.67(m,1H),5.28(d,J=5.2Hz,1H),5.05(t,J=5.2Hz,1H),4.12(t,J=5.2Hz,1H), 3.85-3.83(m,1H),3.55-3.53(m,2H),3.33-3.29(m,2H),2.18-2.12(m,1H),1.61-1.55(m,2H),1.47-1.44(m,1H),1.25-1.23(m,1H).

[0720] Synthesis of 1-((2R,3R,4S,5R)-3-(3-azidopropyl)-5-((bis(4-methoxyphenyl)(phenyl)-methoxy)methyl)-4-hydroxytetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (1-9). To a solution of compound 1-8 (3.00 g, 9.64 mmol) in pyridine (30 mL) was added DMTCl (3.59 g, 10.60 mmol). The mixture was stirred at 15° C. for 1 h, quenched with MeOH (3 mL) and concentrated in vacuo. The residue was redissolved in EtOAc (100 mL) and washed with aqueous citric acid (100 mL) and brine (100 mL). The organic layer was dried over Na2SO4, filtered and concentrated in vacuo. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 10 / 1 to 1 / 1, 0.1% TEA) to give compound 1-9 (5.00 g, 8.15 mmol, yield 84.55%) as a yellow solid. 1 H NMR: 400MHz, DMSO-d 6, δ 7.57(d,J=8.0Hz,1H),7.39-7.25(m,9H),6.91(d,J=8.8Hz,4H),5.87(d,J=8.8Hz,1H),5.46(d,J=8.0Hz,1H),5.37(d,J=4.8Hz,1H),4.12( t,J=4.6Hz,1H),3.95(s,1H),3.74(s,6H),3.33-3.26(m,3H),3.17-3.16(m,1H),2.21-2.19(m,1H),1.65-1.49(m,3H),1.27-1.26(m,1H).

[0721] Synthesis of 1-((2R,3R,4S,5R)-3-(3-aminopropyl)-5-((bis(4-methoxyphenyl)(phenyl)-methoxy)methyl)-4-hydroxytetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (1-10). To a solution of compound 1-9 (3.90 g, 6.36 mmol) in EtOAc (39 mL) was added Pd / C (1.95 g, 6.36 mmol, 10% on carbon). The suspension was stirred under H2 (15 psi) for 1 h, filtered and concentrated in vacuo to give compound 1-10 (3.50 g, crude) as a brown solid. The crude was used in the next step without further purification.

[0722] Synthesis of (2S,3S,4S,5S,6S)-5-acetamido-2-(acetoxymethyl)-6-((5-((3-((2R,3R,4R,5R)-5-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-2-(2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-4-hydroxytetrahydrofuran-3-yl)propyl)amino)-5-oxopentyl)oxy)tetrahydro-2H-pyran-3,4-diyl diacetate (1-11). To a solution of compound 1-10 (3.50 g, 5.96 mmol) and GalNAc pentanoic acid (2.66 g, 5.96 mmol) in DMF (35 mL) was added HCTU (3.70 g, 8.93 mmol, 1.5 equiv.) and NMM (1.81 g, 17.87 mmol, 1.96 mL, 3 equiv.). The reaction was stirred at 15 °C for 1 h. The mixture was then quenched with aqueous NH4Cl (200 mL), extracted with EtOAc (200 mL), and washed with brine (200 mL). The organic layer was dried over Na2SO4, filtered, and concentrated under vacuum. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 2 to EtOAc / acetone = 1 / 2, 0.1% TEA) to give compound 1-11 (4.40 g, 53% yield) as a yellow oil. 1H NMR:400MHz DMSO-d6,δ 11.36(s,1H),7.81(d,J=9.6Hz,1H),7.56-7.38(m,1H),7.35(d,J=7.6Hz,1H),7.32-7.30(m,4H),7.25-7.23(m,5H),6.90(d,J=8.8Hz,4H),5.85(d,J=9.2Hz,1H),5.43(d,J=8.0Hz,1H),5.33(d,J=5.2Hz,1H),5.21(d,J=3.2Hz,1H),4.95-4.94(m,1H),4.47(d,J=8.8Hz,1H),4.12(m,1H),4.05-4.03(m,5H),4.02-4.01(m,2H),4.00-3.74(m,7H),3.25-3.24(m,4H),3.02-2.99(m,3H),2.10-2.08(m,4H),2.03-2.02(m,1H),1.99(s,6H),1.89(s,3H),1.76(s,3H),1.47-1.44(m,7H)。

[0723] Synthesis of (2S,3S,4S,5S,6S)-5-acetamido-2-(acetoxymethyl)-6-((5-((3-((2R,3R,4S,5R)-5-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-(((2-cyanoethoxy)(diisopropylamino)-phosphanyl)oxy)-2-(2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)tetrahydrofuran-3-yl)propyl)amino)-5-oxopentyl)oxy)tetrahydro-2H-pyran-3,4-diyl diacetate (GalNAc2). To a solution of compound 1-11 (4.00 g, 3.93 mmol) in DCM (40 mL) was added DCI (696.7 mg, 5.90 mmol) and 2-cyanoethyl-N,N,N',N'-tetraisopropylphosphorodiamidite (2.37 g, 7.87 mmol, 2.50 mL). The mixture was stirred at 15 °C for 1 h, diluted with DCM (100 mL), washed with aqueous NaHCO3 (100 mL) and brine (100 mL). The organic layer was dried over anhydrous Na2SO4, filtered and concentrated under vacuum. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 3 / 1 to 0 / 1, 0.1% TEA) to give GalNAc2 (3.20 g, 2.63 mmol, 66.84% yield) as a yellow solid. 1 H NMR:400MHz CD3CN, δ 9.17(s,1H),7.58-7.42(M,1H),7.33-7.22(m,2H),7.31-7.30(m,7H),6.91-6.87(m,4H),6.55-6.35(m,2H) ),5.92-5.88(m,1H),5.45-5.42(m,1H),5.28-5.27(m,1H),5.01(m,1H),4.53-4.51(m,2H),4.11-4.05(m, 5H),3.96-3.94(m,9H),3.80-3.77(m,3H),3.77-3.61(m,1H),3.60-3.33(m,5H),2.46-2.17(m,2H),2.09( s,5H),2.07(s,5H),1.91(s,3H),1.83(s,3H),1.55-1.51(m,9H).1.19-1.16(m,9H),1.10(d,J=6.8Hz,3H).

[0724] Example 2. mRNA knockdown activity of siRNA duplexes conjugated with GalNAc G2 to target gene 1. Gene silencing activity was tested using the exemplary siRNA duplexes listed in Table 1. These siRNA duplexes were conjugated with either GalNAc L96 or GalNAc G2 for liver delivery to target gene 1. As shown in Figure 1, GalNAc G2 provides comparable or better delivery efficiency and KD activity than GalNAc L96.

[0725] CD-1 female mice were subcutaneously administered 0.5 mg / kg of siRNA duplexes conjugated with GalNAc. The control group received phosphate-buffered saline (PBS). After 4 days of treatment, the animals were then hydrodynamically injected (HDI) with 10 μg of human gene 1 in pcDNA3.1(+) via the tail vein. Mice were sacrificed 1 day after treatment. Liver tissue was collected and stored in RNAlater® at 4°C overnight and transferred to -80°C after RNAlater removal for mRNA analysis. Target mRNA reduction was measured by qPCR using a CFX384 TOUCH™ Real-Time PCR Detection System (BioRad Laboratories, Inc., Hercules, CA). All samples were normalized to PBS-treated control animals and plotted using GraphPad Prism software (GraphPad Software Inc., La Jolla, CA). [Table 5] The lowercase letters "f" and "m" indicate 2'-deoxy-2'-fluoro (2'-F) and 2'-O-methyl (2'-OMe) sugar modifications to adenosine, cytidine, guanosine and uridine, respectively. The letter "s" indicates a phosphorothioate (PS) linkage, "EP" indicates an ethylphosphonate modification at the 5' end, and L96 and G2 indicate the GalNAc structure shown below: [ka]

[0726] Equivalent

[0727] Details of one or more embodiments of the present disclosure are described in the accompanying specification above. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present disclosure, the preferred methods and materials are described herein. Other features, objects, and advantages of the present disclosure will be apparent from the specification and claims. In this specification and the appended claims, the singular form "a," "an," or "the" includes plural referents unless the context clearly indicates otherwise. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this disclosure belongs. All patents and publications cited herein are incorporated by reference.

[0728] The foregoing description has been presented for purposes of illustration only and is not intended to limit the disclosure to the precise form disclosed, but rather to be limited by the scope of the claims appended hereto.

Claims

1. A compound of formula (I) or (II), 【Chemical 1】 or a pharmaceutically acceptable salt thereof, wherein: B is H or a nucleobase moiety; W is H, C optionally substituted with one or more halogens １ -C ６ alkyl or amino substituents, Y is H, C optionally substituted with one or more halogens １ -C ６ Alkyl, -P(R Ｙ ) ２ , -P(OR Ｙ ) (N (R Ｙ ) ２ ), -P(=O)(OR Ｙ ) R Ｙ , -P(=S)(OR Ｙ ) R Ｙ , -P(=O)(SR Ｙ ) R Ｙ , -P(=S)(SR Ｙ ) R Ｙ , -P(=O)(OR Ｙ ) ２ , -P(=S)(OR Ｙ ) ２ , -P(=O)(SR Ｙ ) ２ , -P(=S)(SR Ｙ ) ２ or a hydroxy protecting group, Each R Ｙ are independently H or C optionally substituted with one or more halogens or cyanos １ -C ６ is alkyl, Z is H or C optionally substituted with one or more halogens １ -C ６ Alkyl, -P(R Ｚ ) ２ , -P(OR Ｚ ) (N (R Ｚ ) ２ ), -P(=O)(OR Ｚ ) R Ｚ , -P(=S)(OR Ｚ ) R Ｚ , -P(=O)(SR Ｚ ) R Ｚ , -P(=S)(SR Ｚ ) R Ｚ , -P(=O)(OR Ｚ ) ２ , -P(=S)(OR Ｚ ) ２ , -P(=O)(SR Ｚ ) ２ , -P(=S)(SR Ｚ ) ２ or a hydroxy protecting group, Each R Ｚ are independently H or C optionally substituted with one or more halogens or cyanos １ -C ６ Is it alkyl? Or Y and Z in formula (I) together represent —Si(R Ｌ ) ２ —O—Si(R Ｌ ) ２ -, and each R Ｌ are independently H or C １ -C ６ is alkyl, Each R ａ are independently H, halogen, or C optionally substituted with one or more halogens １ -C ６ alkyl or two R on two adjacent carbon atoms ａ together with the two adjacent carbon atoms form a double bond, Each R ｂ are independently H, halogen, or C optionally substituted with one or more halogens １ -C ６ is alkyl, R １ is H, halogen, or C optionally substituted with one or more halogens １ -C ６ is alkyl, R ２ is H, halogen, or C optionally substituted with one or more halogens １ -C ６ is alkyl, R ３ is H, halogen, or C optionally substituted with one or more halogens １ -C ６ is alkyl, R ４ is H, halogen, or C optionally substituted with one or more halogens １ -C ６ is alkyl, Each R ５ are independently H, halogen, or C optionally substituted with one or more halogens １ -C ６ is alkyl, A compound wherein n is an integer ranging from about 0 to about 10.

2. 1. A scaffold, or a pharmaceutically acceptable salt thereof, comprising: (i) one or more ligands, or one or more nucleic acid agents, and (ii) one or more linker units, each linker unit independently comprising: 【Chemistry 2-1】 【Chemistry 2-2】 and During the ceremony, B is H or a nucleobase moiety; W is H, C 1 -C 6 alkyl optionally substituted with one or more halogens, or an amino substituent; Y is H, C 1 -C 6 alkyl optionally substituted with one or more halogens, —P(R Y ) 2 , —P(OR Y )(N(R Y ) 2 , —P(═O)(OR Y )R Y , —P(═S)(OR Y )R Y , —P(═O)(SR Y )R Y , —P(═S)(SR Y )R Y , —P(═O)(OR Y ) 2 , —P(═S)(OR Y ) 2 , —P(═O)(SR Y ) 2 , —P(═S)(SR Y ) 2 , or a hydroxy protecting group; each R Y is independently H or C 1 -C 6 alkyl optionally substituted with one or more halogen or cyano; Z is H, C 1 -C 6 alkyl optionally substituted with one or more halogens, —P(R Z ) 2 , —P(OR Z )(N(R Z ) 2 , —P(═O)(OR Z )R Z , —P(═S)(OR Z )R Z , —P(═O)(SR Z )R Z , —P(═S)(SR Z )R Z , —P(═O)(OR Z ) 2 , —P(═S)(OR Z ) 2 , —P(═O)(SR Z ) 2 , —P(═S)(SR Z ) 2 , or a hydroxy protecting group; each R Z is independently H or C 1 -C 6 alkyl optionally substituted with one or more halogen or cyano; or Y and Z in formula (I) together form -Si(R L ) 2 -O-Si(R L ) 2 -, where each R L is independently H or C 1 -C 6 alkyl; each R a is independently H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens, or two R a on two adjacent carbon atoms together with said two adjacent carbon atoms form a double bond; each R b is independently H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; R 1 is H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; R 2 is H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; R 3 is H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; R 4 is H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; each R 5 is independently H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; n is an integer ranging from about 0 to about 10; A scaffold or a pharmaceutically acceptable salt thereof, wherein # indicates attachment to a ligand and ## indicates attachment to a nucleic acid agent.

3. A conjugate or a pharmaceutically acceptable salt thereof, wherein the conjugate comprises: (i) one or more nucleic acid agents; (ii) one or more ligands, and (iii) one or more linker units, each linker unit independently comprising: 【Chemistry 3】 and one or more linker units, B is H or a nucleobase moiety; Y is H, C 1 -C 6 alkyl optionally substituted with one or more halogens, —P(R Y ) 2 , —P(OR Y )(N(R Y ) 2 , —P(═O)(OR Y )R Y , —P(═S)(OR Y )R Y , —P(═O)(SR Y )R Y , —P(═S)(SR Y )R Y , —P(═O)(OR Y ) 2 , —P(═S)(OR Y ) 2 , —P(═O)(SR Y ) 2 , —P(═S)(SR Y ) 2 , or a hydroxy protecting group; each R Y is independently H or C 1 -C 6 alkyl optionally substituted with one or more halogen or cyano; Z is H, C 1 -C 6 alkyl optionally substituted with one or more halogens, —P(R Z ) 2 , —P(OR Z )(N(R Z ) 2 , —P(═O)(OR Z )R Z , —P(═S)(OR Z )R Z , —P(═O)(SR Z )R Z , —P(═S)(SR Z )R Z , —P(═O)(OR Z ) 2 , —P(═S)(OR Z ) 2 , —P(═O)(SR Z ) 2 , —P(═S)(SR Z ) 2 , or a hydroxy protecting group; each R Z is independently H or C 1 -C 6 alkyl optionally substituted with one or more halogen or cyano; or Y and Z in formula (I) together form -Si(R L ) 2 -O-Si(R L ) 2 -, where each R L is independently H or C 1 -C 6 alkyl; each R a is independently H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens, or two R a on two adjacent carbon atoms together with said two adjacent carbon atoms form a double bond; each R b is independently H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; R 1 is H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; R 2 is H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; R 3 is H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; R 4 is H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; each R 5 is independently H, halogen, or C 1 -C 6 alkyl optionally substituted with one or more halogens; n is an integer ranging from about 0 to about 10; A conjugate or a pharmaceutically acceptable salt thereof, wherein # indicates attachment to a ligand and ## indicates attachment to a nucleic acid agent.

4. 4. The conjugate of claim 3, wherein B is H.

5. 4. The conjugate of claim 3, wherein B is a nucleobase moiety.

6. 4. The conjugate of claim 3, wherein the nucleobase moiety is adenine (A), cytosine (C), guanine (G), thymine (T), or uracil (U).

7. Y is C optionally substituted with one or more halogens １ -C ６ 4. The compound, scaffold or conjugate of claim 3, wherein said compound is alkyl.

8. Y is -P(R Ｙ ) ２ , -P(OR Ｙ ) (N (R Ｙ ) ２ ), -P(=O)(OR Ｙ ) R Ｙ , -P(=S)(OR Ｙ ) R Ｙ , -P(=O)(SR Ｙ ) R Ｙ , -P(=S)(SR Ｙ ) R Ｙ , -P(=O)(OR Ｙ ) ２ , -P(=S)(OR Ｙ ) ２ , -P(=O)(SR Ｙ ) ２ , -P(=S)(SR Ｙ ) ２ 4. The compound, scaffold, or conjugate of claim 3, wherein:

9. 4. The conjugate of claim 3, wherein Y is a hydroxy protecting group.

10. 4. The conjugate of claim 3, wherein Z is H.

11. Z is C optionally substituted with one or more halogens １ -C ６ The conjugate of claim 3 , wherein the alkyl is alkyl.

12. Z is -P(R Ｚ ) ２ , -P(OR Ｚ ) (N (R Ｚ ) ２ ), -P(=O)(OR Ｚ ) R Ｚ , -P(=S)(OR Ｚ ) R Ｚ , -P(=O)(SR Ｚ ) R Ｚ , -P(=S)(SR Ｚ ) R Ｚ , -P(=O)(OR Ｚ ) ２ , -P(=S)(OR Ｚ ) ２ , -P(=O)(SR Ｚ ) ２ , -P(=S)(SR Ｚ ) ２ The conjugate of claim 3, wherein

13. 4. The conjugate of claim 3, wherein Z is a hydroxy protecting group.

14. In formula (I), Y and Z together represent —Si(R Ｌ ) ２ —O—Si(R Ｌ ) ２ The conjugate of claim 3, which forms

15. 4. The conjugate of claim 3, wherein the conjugate is selected from the conjugates set forth in Table C.

16. The ligand is 【Chemistry 4】 16. The scaffold or conjugate of any one of claims 3 to 15, comprising:

17. 16. The scaffold or conjugate of any one of claims 3 to 15, wherein the ligand comprises a lipid, a peptide moiety, or an antibody moiety.

18. 16. The scaffold or conjugate of any one of claims 3 to 15, wherein the nucleic acid agent comprises an oligonucleotide.

19. A pharmaceutical composition comprising the conjugate of any one of claims 3 to 15.

20. 16. The conjugate of any one of claims 3 to 15 for modulating expression of a target gene in a subject, for delivering a nucleic acid agent to a subject, or for treating or preventing a disease in a subject in need thereof.

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