Polyhydroxylated cyclopentane derivatives and methods of use

Abstract

The present disclosure provides linker compounds of formula (I) or (II), their pharma- ceutically acceptable salts, and related scaffolds and conjugates. The present disclosure also relates to the use of the linker compounds, scaffolds, and conjugates, for example, in the delivery of nucleic acids and / or in the treatment or prevention of disease.

Description

[Technical field]

[0001] Related Applications This application claims priority to and the benefit of U.S. Patent Application No. 63 / 252,358, filed October 5, 2021, the entire contents of which are incorporated herein by reference. [Background technology]

[0002] background 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 sugars such as galactose, mannose, mannose-6-phosphate, and peptides and proteins such as transferrin, asialoglycoprotein, vitamin B12, insulin, and epidermal growth factor (EGF). The asialoglycoprotein receptor (ASGP-R) is a highly potent receptor and is highly abundant on hepatocytes. ASGP-R exhibits a higher affinity for N-acetyl-D-galactosylamine (GalNAc) than D-Gal. Recently, certain carbohydrate conjugates have been shown to be valuable alternatives to liposomes for nucleic acid delivery. Furthermore, the stability of nucleic acids within the cellular environment after successful delivery into the cell is important to achieve the desired therapeutic effect.

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

[0004] summary In some aspects, the present disclosure provides a compound of formula (I) or (II): [ka] or a pharma- ceutically acceptable salt thereof, wherein: W is H, C1-C6 alkyl optionally substituted with one or more halogens, or an amino substituent; X is H, halogen, or -OR X and; R X is H, C1-C6 alkyl, or -(C1-C6 alkyl)-(C6-C 10 aryl), where C1-C6 alkyl or -(C1-C6 alkyl)-(C6-C 10 aryl) may be one or more R Xa optionally replaced by; Each R Xa is independently halogen, C1-C6 alkyl, or -O-(C1-C6 alkyl), where C1-C6 alkyl or -O-(C1-C6 alkyl) is optionally substituted with one or more halogens; 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 Yis 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; Alternatively, Y and Z together in formula (I) represent -Si(R L )2-O-Si(R L )2-, where each R L is independently H or C1-C6 alkyl; 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; and Each R 5is independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; and Each R 6 is independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens.

[0005] In some aspects, the disclosure provides a scaffold, or a pharma- ceutically acceptable salt thereof, wherein the scaffold comprises: (i) a ligand; and (ii) comprising a linker unit, wherein the linker unit is: [ka] where the variable R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , X, Y, and Z are as described herein, and # indicates the connection to the ligand.

[0006] In some aspects, the disclosure provides a scaffold or a pharma- ceutically acceptable salt thereof, wherein the scaffold comprises: (i) one or more nucleic acid agents; and (ii) comprises one or more linker units, where each linker unit is independently: [ka] where the variable R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , W, X, Y, and Z are as described herein, and ## indicates the connection to the nucleic acid agent.

[0007] In some aspects, the disclosure provides a conjugate or a pharma- ceutically acceptable salt thereof, wherein the conjugate is: (i) one or more nucleic acid agents; (ii) one or more ligands; and (iii) comprising one or more linker units, wherein each linker unit is independently: [ka] where the variable R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , X, Y, and Z are as described herein, where # indicates the connection to the ligand and ## indicates the connection to the nucleic acid agent.

[0008] In some aspects, the present disclosure provides compounds that are isotopic derivatives of the compounds disclosed herein.

[0009] In some aspects, the disclosure provides pharmaceutical compositions containing the compounds, scaffolds, or conjugates described herein.

[0010] In some aspects, the disclosure provides a method of modulating expression of a target gene in a subject, comprising administering to the subject a conjugate described herein.

[0011] In some aspects, the disclosure provides a method of delivering a nucleic acid agent to a subject, comprising administering to the subject a conjugate described herein.

[0012] 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.

[0013] 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.

[0014] 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.

[0015] 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.

[0016] Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one skilled in the art to which this disclosure belongs. In this specification, the singular form also includes the plural 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 claimed invention. In case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples are merely illustrative 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 will control.

[0017] Other features and advantages of the present disclosure will be apparent from the following detailed description and claims. [Brief description of the drawings]

[0018] BRIEF DESCRIPTION OF THE DRAWINGS [Figure 1] FIG. 1 is a graph showing gene silencing activity of siRNA duplexes in liver 5 days after a single 0.5 mg / kg sc injection of CD-1 female mice, followed by HDI dosing (human gene 1 plasmid, 10 μg) on ​​day 4. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0019] Detailed Description The present disclosure provides 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 disease.

[0020] 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, W is H, C1-C6 alkyl optionally substituted with one or more halogens, or an amino substituent; X is H, halogen, or -OR X and; R X is H, C1-C6 alkyl, or -(C1-C6 alkyl)-(C6-C 10 aryl), where C1-C6 alkyl or -(C1-C6 alkyl)-(C6-C 10 aryl) may be one or more R Xa optionally replaced by; Each R Xa is independently halogen, C1-C6 alkyl, or -O-(C1-C6 alkyl), where C1-C6 alkyl or -O-(C1-C6 alkyl) is optionally substituted with one or more halogens; 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 )RY , -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; Alternatively, Y and Z together in formula (I) represent -Si(R L )2-O-Si(R L )2-, where each R L is independently H or C1-C6 alkyl; 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; and Each R 6 is independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens.

[0021] With respect to the compounds of the present disclosure, the variables W, X, R X , R Xa , Y, R Y , Z, R Z , R L , R 1 , R 2 , R 3 , R 4 , R 5 , and R 6 may each be selected from the groups described herein, where applicable, and the variables W, X, R X , R Xa , Y, R Y , Z, R Z , R L , R 1 , R 2 , R 3 , R 4 , R 5 , and R 6 Any group described herein with respect to any of the variables W, X, R X , R Xa , Y, R Y , Z, R Z , R L , R 1 , R 2 , R 3 , R 4 , R 5 , and R 6 It is understood that one or more of the remaining groups may be combined with any of the groups described herein.

[0022] Variables W, X, R X 、R Xa , Y, R Y , Z, R Z , and R L In some embodiments, W is H.

[0023] 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).

[0024] In some embodiments, W is C1-C6 alkyl (eg, methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0025] In some embodiments, W is methyl, ethyl, or propyl.

[0026] In some embodiments, W is an amino substituent, ie, a group suitable for replacement of a hydrogen on an amino moiety, such as an amino protecting group.

[0027] 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).

[0028] In some embodiments, W is optionally substituted acyl (e.g., —C(═O)(C 30 alkyl), where C1-C 30The alkyl is optionally substituted.

[0029] In some embodiments, W is substituted acyl (e.g., [ka] ).

[0030] In some embodiments, W is trifluoroacetyl (TFA).

[0031] In some embodiments, W is an optionally substituted thioacyl (e.g., —C(═S)(C 30 alkyl), where C1-C 30 The alkyl is optionally substituted.

[0032] In some embodiments, W is a substituted thioacyl (e.g., [ka] ).

[0033] In some embodiments, W is an amino substituent, i.e., a group suitable for replacing a hydrogen on an amino moiety, such as -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), etc., where C1-C 30 Alkyl is optionally substituted. In some embodiments, W is -C(=O)(C-C 25 alkyl), -C(=O)NH(C1-C 25 Alkyl), -C(=S)(C1-C 25 alkyl), or -C(=S)NH(C1-C 25 alkyl), where C1-C 25 The alkyl is optionally substituted.

[0034] In some embodiments, W is -C(=O)NH(C-C 30 alkyl), where C1-C 30 The alkyl is optionally substituted.

[0035] In some embodiments, W is -C(=S)NH(C-C 30 alkyl), where C1-C 30 The alkyl is optionally substituted.

[0036] In some embodiments, X is H.

[0037] In some embodiments, X is not H.

[0038] In some embodiments, X is halogen (eg, F, Cl, Br, or I).

[0039] In some embodiments, X is F or Cl.

[0040] In some embodiments, X is F.

[0041] In some embodiments, X is -OR X It is.

[0042] In some embodiments, X is --OH.

[0043] In some embodiments, X is not --OH.

[0044] In some embodiments, X is one or more R Xa and optionally substituted with -O-(C1-C6 alkyl), for example where C1-C6 alkyl is methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl.

[0045] In some embodiments, X is -O-(C1-C6 alkyl) (e.g., where C1-C6 alkyl is methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0046] In some embodiments, X is -OCH3.

[0047] In some embodiments, X is one or more R Xa and optionally substituted with -O-(C1-C6 alkyl)-O-(C1-C6 alkyl), for example where C1-C6 alkyl is methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl.

[0048] In some embodiments, X is -O-(C1-C6 alkyl)-O-(C1-C6 alkyl) (e.g., where C1-C6 alkyl is methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0049] In some embodiments, X is -OCH2CH2OCH3.

[0050] In some embodiments, X is one or more R Xa -O-(C1-C6 alkyl)-(C6-C 10 aryl).

[0051] In some embodiments, X is -O-(C1-C6 alkyl)-(C6-C 10 aryl).

[0052]

[0053] In some embodiments, X is: [ka] It is.

[0054] In some embodiments, X is one or more R Xa Optionally replaced by: [ka] It is.

[0055] In some embodiments, X is optionally substituted with one or more halogen: [ka] It is.

[0056] In some embodiments, X is optionally substituted with one or more C1-C6 alkyl or -O-(C1-C6 alkyl): [ka] wherein C1-C6 alkyl or -O-(C1-C6 alkyl) is optionally substituted with one or more halogens.

[0057] In some embodiments, R X is H.

[0058] In some embodiments, R X is not H.

[0059] In some embodiments, R X is one or more R Xa and C1-C6 alkyl optionally substituted with (for example, methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0060] In some embodiments, R Xis 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), or -O-(C1-C6 alkyl) (e.g., where C1-C6 alkyl is methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl) optionally substituted with one or more halogens.

[0061] In some embodiments, R X is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0062] In some embodiments, R X is methyl, ethyl, or propyl.

[0063] In some embodiments, R X is methyl.

[0064] In some embodiments, R X 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).

[0065] In some embodiments, R Xis 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 -O-(C1-C6 alkyl) (e.g., where C1-C6 alkyl is methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl), where -O-(C1-C6 alkyl) is optionally substituted with one or more halogens.

[0066] In some embodiments, R X is one or more R Xa -(C1-C6 alkyl)-(C6-C 10 aryl).

[0067] In some embodiments, R X is optionally substituted with one or more halogen (e.g., F, Cl, Br, or I), C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl), or -O-(C1-C6 alkyl) (e.g., where C1-C6 alkyl is methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl). 10 aryl), where C1-C6 alkyl or -O-(C1-C6 alkyl) is optionally substituted with one or more halogens.

[0068] In some embodiments, R X is -(C1-C6 alkyl)-(C6-C 10 aryl).

[0069] In some embodiments, at least one R Xa is a halogen (e.g., F, Cl, Br, or I).

[0070] In some embodiments, at least one R Xa is F or Cl.

[0071] In some embodiments, at least one R Xa 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).

[0072] In some embodiments, at least one R Xa is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0073] In some embodiments, at least one R Xa 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).

[0074] In some embodiments, at least one R Xa is -O-(C1-C6 alkyl) optionally substituted with one or more halogen (e.g., F, Cl, Br, or I).

[0075] In some embodiments, at least one R Xa is -O-(C1-C6 alkyl).

[0076] In some embodiments, at least one R Xa is -O-(C1-C6 alkyl) substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0077] In some embodiments, Y is H.

[0078] In some embodiments, Y is not H.

[0079] 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).

[0080] In some embodiments, Y is C1-C6 alkyl (eg, methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0081] In some embodiments, Y is methyl, ethyl, or propyl.

[0082] 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.

[0083] In some embodiments, Y is -P(R Y )2.

[0084] In some embodiments, Y is -PH2.

[0085] In some embodiments, Y is -P(OR Y )(N(RY )2).

[0086] In some embodiments, Y is -P(OH)(NH2).

[0087] In some embodiments, Y is -P(O(C1-C6 alkyl))(N(C1-C6 alkyl)2), where C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0088] In some embodiments, Y is -P(=O)(OR Y )R Y It is.

[0089] In some embodiments, Y is -P(=O)(OH)(C1-C6 alkyl), where C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0090] In some embodiments, Y is -P(=S)(OR Y )R Y It is.

[0091] In some embodiments, Y is -P(=S)(OH)(C1-C6 alkyl), where C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0092] In some embodiments, Y is -P(=O)(SR Y )R Y It is.

[0093] In some embodiments, Y is -P(=O)(SH)(C1-C6 alkyl), where C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0094] In some embodiments, Y is -P(=S)(SR Y )R Y It is.

[0095] In some embodiments, Y is -P(=S)(SH)(C1-C6 alkyl), wherein C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0096] In some embodiments, Y is -P(=O)(OR Y )2.

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

[0098] In some embodiments, Y is -P(=S)(OR Y )2.

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

[0100] In some embodiments, Y is -P(=O)(SR Y )2.

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

[0102] In some embodiments, Y is -P(=S)(SR Y )2.

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

[0104] In some embodiments, Y is a hydroxy protecting group.

[0105] In some embodiments, Y is silyl (eg, trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl).

[0106] In some embodiments, Y is triphenylmethyl (Tr) or 4,4'-dimethoxytrityl (DMTr).

[0107] In some embodiments, Y is optionally substituted acyl (eg, optionally substituted acetyl) or benzyl.

[0108] In some embodiments, Y is not a hydroxy protecting group.

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

[0110] In some embodiments, each R Y is H.

[0111] 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.

[0112] 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.

[0113] In some embodiments, at least one R Y is H, and 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 or cyano.

[0114] In some embodiments, when X is --OH, Y is not H or a hydroxy protecting group.

[0115] In some embodiments, when X is -OH, Y is 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, or -P(=S)(SR Y )2.

[0116] In some embodiments, when Y is H or a hydroxy protecting group, X is not --OH.

[0117] In some embodiments, when Y is H or a hydroxy protecting group, X is H, a halogen, or -OR X and R X is C1-C6 alkyl or -(C1-C6 alkyl)-(C6-C 10 aryl), where C1-C6 alkyl or -(C1-C6 alkyl)-(C6-C 10 aryl) may be one or more R Xa is optionally replaced by

[0118] In some embodiments, Z is H.

[0119] 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).

[0120] In some embodiments, Z is C1-C6 alkyl (eg, methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0121] In some embodiments, Z is methyl, ethyl, or propyl.

[0122] 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.

[0123] In some embodiments, Z is -P(R Z )2.

[0124] In some embodiments, Z is -PH2.

[0125] In some embodiments, Z is -P(OR Z )(N(R Z )2).

[0126] In some embodiments, Z is -P(OH)(NH2).

[0127] In some embodiments, Z is -P(O(C1-C6 alkyl))(N(C1-C6 alkyl)2), where C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0128] In some embodiments, Z is -P(=O)(OR Z )R Z It is.

[0129] In some embodiments, Z is -P(=O)(OH)(C1-C6 alkyl), wherein C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0130] In some embodiments, Z is -P(=S)(OR Z )R Z It is.

[0131] In some embodiments, Z is -P(=S)(OH)(C1-C6 alkyl), where C1-C6 alkyl is optionally substituted with one or more halogen or cyano.

[0132] In some embodiments, Z is -P(=O)(SR Z )R Z It is.

[0133] 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.

[0134] In some embodiments, Z is -P(=S)(SR Z )R Z It is.

[0135] 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.

[0136] In some embodiments, Z is -P(=O)(OR Z )2.

[0137] In some embodiments, Z is -P(=O)(OH)2.

[0138] In some embodiments, Z is -P(=S)(OR Z )2.

[0139] In some embodiments, Z is -P(=S)(OH)2.

[0140] In some embodiments, Z is -P(=O)(SR Z )2.

[0141] In some embodiments, Z is -P(=O)(SH)2.

[0142] In some embodiments, Z is -P(=S)(SR Z )2.

[0143] In some embodiments, Z is -P(=S)(SH)2.

[0144] In some embodiments, Z is a hydroxy protecting group.

[0145] In some embodiments, Z is silyl (eg, trimethylsilyl, triethylsilyl, tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or triisopropylsilyl).

[0146] In some embodiments, Z is triphenylmethyl (Tr) or 4,4'-dimethoxytrityl (DMTr).

[0147] In some embodiments, Z is substituted acyl (eg, optionally substituted acetyl) or benzyl.

[0148] In some embodiments, at least one R Z is H.

[0149] In some embodiments, each R Z is H.

[0150] 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.

[0151] 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.

[0152] 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.

[0153] In some embodiments, Y and Z of formula (I) together represent -Si(R L )2-O-Si(R L )2- is formed.

[0154] In some embodiments, Y and Z of Formula (I) together form -Si(C1-C6 alkyl)2-O-Si(C1-C6 alkyl)2-.

[0155] In some embodiments, Y and Z of Formula (I) together form -Si(C1-C6 alkyl)2-O-SiH(C1-C6 alkyl)-.

[0156] In some embodiments, Y and Z of formula (I) together form -SiH(C1-C6 alkyl)-O-SiH(C1-C6 alkyl)-.

[0157] In some embodiments, Y and Z of Formula (I) together form -Si(iPr)2-O-Si(iPr)2-.

[0158] In some embodiments, at least one R L is H.

[0159] In some embodiments, each R L is independently C1-C6 alkyl.

[0160] In some embodiments, each R L is independently methyl, ethyl, or propyl (e.g., iPr).

[0161] Variable R 1 、R 2 、R 3 、R 4 、R 5 , and R 6 In some embodiments, R 1 is H.

[0162] In some embodiments, R 1 is a halogen (e.g., F, Cl, Br, or I).

[0163] In some embodiments, R 1 is F or Cl.

[0164] In some embodiments, R 1is 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).

[0165] 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).

[0166] 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).

[0167] In some embodiments, R 2 is H.

[0168] In some embodiments, R 2 is a halogen (e.g., F, Cl, Br, or I).

[0169] In some embodiments, R 2 is F or Cl.

[0170] 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) optionally substituted with one or more halogens (e.g., F, Cl, Br, or I).

[0171] 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).

[0172] 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).

[0173] In some embodiments, R 3 is H.

[0174] In some embodiments, R 3 is a halogen (e.g., F, Cl, Br, or I).

[0175] In some embodiments, R 3 is F or Cl.

[0176] 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).

[0177] 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).

[0178] 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).

[0179] In some embodiments, R 4 is H.

[0180] In some embodiments, R 4 is a halogen (e.g., F, Cl, Br, or I).

[0181] In some embodiments, R 4 is F or Cl.

[0182] 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).

[0183] 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).

[0184] 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).

[0185] In some embodiments, each R 5 is H.

[0186] In some embodiments, at least one R 5is 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).

[0187] In some embodiments, at least one R 5 is a halogen (e.g., F, Cl, Br, or I).

[0188] In some embodiments, at least one R 5 is F or Cl.

[0189] 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).

[0190] 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).

[0191] 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).

[0192] In some embodiments, each R 6 is H.

[0193] In some embodiments, at least one R 6is 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).

[0194] In some embodiments, at least one R 6 is a halogen (e.g., F, Cl, Br, or I).

[0195] In some embodiments, at least one R 6 is F or Cl.

[0196] In some embodiments, at least one R 6 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).

[0197] In some embodiments, at least one R 6 is C1-C6 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, pentyl, or hexyl).

[0198] In some embodiments, at least one R 6 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).

[0199] In some embodiments, each R 1 , R 2 , R 3 , R 4 , R 5 , and R 6is H.

[0200] Illustrative Examples of Compounds In some embodiments, the compound has formula (I'-1), (I'-2), (II'-1), or (II'-2): [ka] or a pharma- ceutical acceptable salt thereof.

[0201] In some embodiments, the compound has formula (IA) or (II-A): [ka] or a pharma- ceutical acceptable salt thereof.

[0202] In some embodiments, the compound has formula (I-A'-1), (I-A'-2), (II-A'-1), or (II-A'-2): [ka] or a pharma- ceutical acceptable salt thereof.

[0203] In some embodiments, the compound has formula (IB) or (II-B): [ka] or a pharma- ceutical acceptable salt thereof.

[0204] In some embodiments, the compound has formula (I-B'-1), (I-B'-2), (II-B'-1), or (II-B'-2): [ka] or a pharma- ceutical acceptable salt thereof.

[0205] In some embodiments, Y is a hydroxy protecting group (e.g., silyl, Tr, DMTr, acyl, or benzyl) and Z is a hydroxy protecting group (e.g., silyl, Tr, DMTr, acyl, or benzyl); or Y and Z of formula (I), (I'-1), (I'-2), (IA), (I-A'-1), (I-A'-2), (IB), (I-B'-1), or (I-B'-2) together represent -Si(R L )2-O-Si(R L )2-, where each R L is independently H or C1-C6 alkyl.

[0206] 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).

[0207] In some embodiments, Y and Z of formula (I), (I'-1), (I'-2), (IA), (I-A'-1), (I-A'-2), (IB), (I-B'-1), or (I-B'-2) together represent -Si(R L )2-O-Si(R L )2-, where each R L is independently H or C1-C6 alkyl.

[0208] In some embodiments, the compound has the formula: [ka] or a pharma- ceutically acceptable salt thereof, wherein: 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 )RY , -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 (such as, for example, 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, etc.), triphenylmethyl (Tr), 4,4′-dimethoxytrityl (DMTr), substituted acyl (e.g., optionally substituted acetyl), or benzyl, etc.); and Each R Z is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0209] In some embodiments, the compound has the formula: [ka] [ka] or a pharma- ceutically acceptable salt thereof, wherein: 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 (such as, for example, 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 (such as, for example, 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); and Each R Z is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano; Here, C1-C 30 The alkyl is optionally substituted.

[0210] In some embodiments, the compound is selected from the compounds set forth in Table L and their pharma- ceutically acceptable salts. [Table L-1] [Table L-2] [Table L-3] [Table L-4] [Table L-5] [Table L-6]

[0211] In some aspects, the disclosure provides compounds that are isotopic derivatives (eg, isotopically labeled compounds) of a compound of any one of the formulas disclosed herein.

[0212] It will be understood that isotopic derivatives can be prepared using any of a variety of art-recognized techniques, for example, isotopic derivatives can generally be prepared by substituting an isotopically labeled reagent for a non-isotopically labeled reagent and following the procedures disclosed in the schemes and / or examples herein.

[0213] In some embodiments, the isotopic derivative is a deuterium-labeled compound.

[0214] In some embodiments, the isotopic derivative is a deuterium-labeled compound of any one of the compounds of the formulas disclosed herein.

[0215] 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 isotope elements 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). 2 In some embodiments, the compound is 2 In some embodiments, the compound is 13 C-labeled compounds or 14 C labeled compound. In some embodiments, the compound is 18 In some embodiments, the compound is 123 I-labeled compounds, 124 I-labeled compounds,125 I-labeled compounds, 129 I-labeled compounds, 131 I-labeled compounds, 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 compounds, 34 S-labeled compounds, 35 S-labeled compounds, 36 S-labeled compounds, or any combination thereof.

[0216] It will be understood that isotopic derivatives can be prepared using any of a variety of art-recognized techniques, for example, isotopic derivatives can generally be prepared by substituting an isotopically labeled reagent for a non-isotopically labeled reagent and following the procedures disclosed in the schemes and / or examples herein.

[0217] It will also be appreciated that isotopic substitution may offer certain therapeutic advantages resulting from greater metabolic stability, such as increased in vivo half-life or reduced dosage requirements.

[0218] For the avoidance of doubt, when a group is qualified herein by "as described herein," it is to be understood that the group includes the broadest definition occurring first, as well as each and every of the specific definitions relating to that group.

[0219] It will be understood that when a compound is disclosed herein, it is represented in one specific configuration. Such specific configuration is not interpreted to limit the disclosure to one or another of isomers, tautomers, positional isomers or stereoisomers, nor excludes mixtures of isomers, tautomers, positional isomers or stereoisomers. In some embodiments, the presentation of a compound herein in a specific configuration is intended to include and refer to each of the available isomers, tautomers, positional isomers and stereoisomers of this compound, or any mixtures thereof; on the other hand, this presentation is also intended to refer to a specific configuration of this compound.

[0220] It will be understood that when compounds are disclosed herein, they are presented without a specific configuration (e.g., without a specific stereochemistry). Such presentation is intended to encompass all available isomers, tautomers, positional isomers, and stereoisomers of the compounds. In some embodiments, presentation of a compound herein without a specific configuration is intended to refer to each of the available isomers, tautomers, positional isomers, and stereoisomers of the compound, or mixtures thereof.

[0221] The term "isomerism" as used herein means compounds that have the same molecular formula but differ in the nature or sequence of bonds of their atoms or the arrangement of their atoms in space. Compounds that have the same molecular formula but differ in the nature or sequence of bonds 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 each other are called "diastereomers" and those that are non-superimposable mirror images of each other are called "enantiomers". When a compound has an asymmetric center, for example, it is possible for a pair of enantiomers to be bonded to four different groups. Enantiomers can be characterized by the absolute configuration of their asymmetric center and described by the R- and S-sequence rules of Cahn and Prelog, or by the way the molecule rotates in the plane of 32xidizing light and designated as dextrorotatory or levorotatory (i.e., as (+) or (-)-isomers, respectively). A chiral compound can exist as either individual enantiomer or as a mixture thereof. A mixture containing equal proportions of the enantiomers is called a "racemic mixture."

[0222] The compounds of this disclosure may have one or more asymmetric centers; such compounds may therefore be produced as individual I- or (S)-stereoisomers or as mixtures thereof. Unless otherwise indicated, the description or naming of a particular compound in the specification and claims is intended to include both individual enantiomers and mixtures, racemic or otherwise, thereof. Methods for the determination of stereochemistry and separation of stereoisomers are well known in the art, for example, by synthesis from optically active starting materials or by resolution of racemic forms ("Advanced Organic Chemistry", 4 th(See discussion in Chapter 4 of the American Chemical Society, edition J. March, John Wiley and Sons, New York, 2001). Some of the compounds of the present disclosure may have centers of geometric isomerism (E- and Z-isomers). It is to be understood that the present disclosure encompasses optical, diastereoisomers, and geometric isomers, and mixtures thereof, that possess inflammasome inhibitory activity.

[0223] The term "chiral center" as used herein refers to a carbon atom bonded to four nonidentical substituents.

[0224] As used herein, the term "chiral isomer" refers to a compound having at least one chiral center. Compounds with two or more chiral centers may exist either as individual diastereomers or as a mixture of diastereomers, termed a "diastereomeric mixture." When one chiral center is present, stereoisomers may be distinguished (33xidizing 33ati) 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 order of sequence" (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).

[0225] As used herein, "geometric isomers" refers to diastereomers that owe their existence to restricted 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, which indicate that these groups are on the same or opposite sides of a double bond in the molecule according to the Cahn-Ingold-Prelog sequencing rules.

[0226] It should be understood that the compounds of the present disclosure are depicted as different chiral or geometric isomers. When compounds have chiral or geometric isomeric forms, it should also be understood that all isomeric forms are included within the scope of the present disclosure, and the naming of these compounds is not intended to exclude any isomeric form, and it is understood that not all isomers have the same level of activity.

[0227] It should be understood that the structures and other compounds discussed in this disclosure include their atropisomers, and it should also be understood that not all atropisomers have the same level of activity.

[0228] The term "atropisomer" as used herein is a type of stereoisomer in which the atoms of two isomers are arranged differently in space. Atropisomers owe their existence to the restricted rotation caused by the hindrance of the rotation of large groups around a central bond. Such atropisomers typically exist as mixtures, but as a result of the advancement of modern chromatographic techniques, it has become possible to separate mixtures of two atropisomers in selected cases.

[0229] The term "tautomer" as used herein 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 a formal shift of a hydrogen atom associated with the switch of adjacent conjugated double bonds. Tautomers exist as a mixture of tautomeric sets in solution. In solutions where the conversion is possible, a chemical equilibrium of the tautomers will be reached. The exact ratio of the tautomers depends on several factors, including temperature, solvent, and pH. The concept of tautomers that are interconverted by tautomerization is called tautomerism. Among the various tautomeric forms that are possible, two are commonly recognized. 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, resulting in a cyclic (ring-shaped) form, as demonstrated by glucose.

[0230] It should be understood that the compounds of the present disclosure may be depicted as different tautomers. It should also be understood that, where the compounds have tautomeric forms, all tautomeric forms are intended to be included within the scope of the present disclosure, and the naming of these compounds does not exclude any tautomeric form. It will be understood that certain tautomers may have a higher level of activity than others.

[0231] It should be understood that the compounds of any formula described herein include the compounds themselves, as well as, if applicable, their salts, and their solvates.For example, salts can be formed between anions and positively charged groups (e.g., amino) on the substituted compounds disclosed herein.Suitable anions include chloride, bromide, iodide, sulfate, hydrogen sulfate, sulfamate, nitrate, phosphate, citrate, methanesulfonate, trifluoroacetate, glutamate, glucuronate, glutarate, malate, maleate, succinate, fumarate, tartrate, tosylate, salicylate, lactate, naphthalenesulfonate, and acetate (e.g., trifluoroacetate).

[0232] The term "pharmaceutical acceptable anion" as used herein refers to an anion suitable for forming a pharmaceutical 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.Substituted compounds disclosed herein also include those salts that contain a quaternary nitrogen atom.

[0233] 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.

[0234] The term "solvate" as used herein refers to a solvent addition form that contains a solvent in either stoichiometric or non-stoichiometric amounts. Some compounds have a tendency to trap a fixed molar ratio of solvent molecules in the crystalline solid state, thus 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. Hydrates are formed by the combination of one or more water molecules with a molecule of a substance in which the water remains in its molecular state as H2O.

[0235] 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). Thus, an analog is a compound that is similar or equivalent in function and appearance to the reference compound, but differs in structural origin.

[0236] As used herein, the term "derivative" refers to compounds that have a common core structure and are substituted with various groups as described herein.

[0237] The term "bioisostere" as used herein refers to a compound resulting from the exchange of an atom or group of atoms with another broadly similar atom or group of atoms. The purpose of the bioisostere exchange is to create a new compound with similar biological properties as the parent compound. The bioisostere exchange may be physicochemical or topological. 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.

[0238] It should also be understood that a particular compound of any one of the formulas disclosed herein may exist in solvated as well as unsolvated forms, such as hydrated forms. Suitable pharma- ceutically acceptable solvates are hydrates, such as hemihydrates, monohydrates, dihydrates, or trihydrates. It should also be understood that the present disclosure encompasses all such solvated forms that have inflammasome inhibitory activity.

[0239] It should also be understood that any one of the particular compounds of the formula disclosed herein may exist in polymorphic forms, and the present disclosure encompasses all such forms, or mixtures thereof, that possess inflammasome inhibitory activity. It is generally known that crystalline materials may 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, liquid and / or solid state nuclear magnetic resonance spectroscopy, etc. The water content of such crystalline materials may be determined by Karl Fischer analysis.

[0240] Compounds of any one of the formulae disclosed herein may exist in many different tautomeric forms, and reference to any one of the compounds of these formulae includes all such forms. For the avoidance of doubt, compounds may exist in several tautomeric forms, and where 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, for example, as seen in the following tautomeric pairs: keto / enol (illustrated below), imine / enamine, amide / iminoalcohol, amidine / amidine, nitroso / oxime, thioketone / enethiol, and nitro / aci-nitro. [ka]

[0241] Compounds of any one of the formulae disclosed herein that contain an amine functionality can also form N-oxides. Reference herein to compounds of any one of the formulae disclosed herein that contain an amine functionality also includes N-oxides. When a compound contains several amine functionalities, one or more nitrogen atoms can 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 treatment of the corresponding amines with an oxidizing agent, such as hydrogen peroxide or a peracid (e.g., peroxycarboxylic acid), and are described, for example, in Advanced Organic Chemistry by Jerry March, 4 th Edition, Wiley Interscience, pages 1010-1015. More particularly, N-oxides can be made 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 dichloromethane.

[0242] The compounds of any one of the formulas disclosed herein may be administered in the form of a prodrug that breaks down in the human or animal body and releases a compound of the present disclosure. Prodrugs may be used to modify the physical properties and / or pharmacokinetic properties of the compounds of the present disclosure. Prodrugs can be formed when the compounds of the present disclosure contain a suitable group or substituent to which a property-modifying group can be attached.

[0243] The present disclosure therefore includes those compounds of any one of the formulae disclosed herein, as defined above, when made available by organic synthesis and when made available in the human or animal body by cleavage of their prodrugs. The present disclosure therefore includes such compounds of any one of the formulae disclosed herein produced by organic synthetic means, and also such compounds produced in the human or animal body by metabolism of precursor compounds, i.e., the compounds of any one of the formulae disclosed herein may be synthetically produced compounds or metabolically produced compounds.

[0244] Suitable pharma- ceutically acceptable prodrugs of a compound of any one of the formulae disclosed herein are those, based on sound medical judgment, suitable for administration to the human or animal body without undesirable pharmacological activity and without undue toxicity. Various types of prodrugs are described, for example, in the following publications: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., 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.

[0245] The in vivo action of a compound of any one of the formulae disclosed herein may be exerted in part by one or more metabolic products formed in the human or animal body after administration of a compound of any one of the formulae disclosed herein. As mentioned above, the in vivo action of a compound of any one of the formulae disclosed herein may also be exerted by metabolism of a precursor compound (prodrug).

[0246] Preferably, the present disclosure excludes any individual compound that does not have a biological activity as defined herein.

[0247] Scaffolds and conjugates containing linkers The term "scaffold" as used herein 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.

[0248] 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.

[0249] In some aspects, the disclosure provides a scaffold or a pharma- ceutically acceptable salt thereof, wherein the scaffold comprises: (i) a ligand; and (ii) a linker unit: [ka] where the variable R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , X, Y, and Z are as described herein, and # indicates the connection to the ligand.

[0250] In some aspects, the disclosure provides a scaffold or a pharma- ceutically acceptable salt thereof, wherein the scaffold comprises: (i) one or more nucleic acid agents; and (ii) one or more linker units: [ka] where the variable R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , W, X, Y, and Z are as described herein, and # indicates the connection to the nucleic acid agent.

[0251] In some embodiments, the connection "##" is a direct connection to the nucleic acid material, i.e., without any linking moieties.

[0252] In some embodiments, the connection "##" is an indirect connection to the nucleic acid material, i.e., there is a linking moiety between the linker unit and the nucleic acid material. 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.

[0253] In some embodiments, the scaffold comprises double-stranded RNA (eg, double-stranded siRNA).

[0254] In some embodiments, the scaffold comprises a double-stranded RNA (eg, a double-stranded siRNA) and one or more linker units.

[0255] 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 3-4 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).

[0256] 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.

[0257] In some embodiments, the scaffold comprises a double-stranded RNA (e.g., a double-stranded siRNA) and one or more linker units, wherein: one or more linker units (e.g., 1 to 3 linker units) are continuously or discontinuously connected to the sense strand (e.g., the 3'- or 5'-end) of a double-stranded RNA (e.g., a double-stranded siRNA); At one or more continuous or discontinuous internal positions (positions between the 3'-end and the 5'-end) of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA), one or more nucleosides or nucleotides are replaced by 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 continuously or discontinuously connected to the antisense strand (e.g., the 3'- or 5'-end) of a double-stranded RNA (e.g., a double-stranded siRNA); and / or At one or more consecutive or interrupted internal positions of the antisense strand, one or more nucleosides or nucleotides are replaced by one or more linker units (eg, 1 to 3 linker units).

[0258] In some embodiments, the scaffold comprises a double-stranded RNA (e.g., a double-stranded siRNA) and one or more linker units, wherein: One or more linker units (e.g., 1 to 3 linker units) are continuously or discontinuously connected to the sense strand (e.g., the 3'- or 5'-end) of a double-stranded RNA (e.g., a double-stranded siRNA); and At one or more continuous or discontinuous internal positions (positions between the 3'-end and the 5'-end) of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA), one or more nucleosides or nucleotides are replaced by one or more linker units (e.g., 1 to 3 linker units).

[0259] In some embodiments, the scaffold comprises a double-stranded RNA (e.g., a double-stranded siRNA) and one or more linker units, wherein: One or more linker units (e.g., 1 to 3 linker units) are continuously or discontinuously connected to the antisense strand (e.g., the 3'- or 5'-end) of the double-stranded RNA (e.g., the double-stranded siRNA); and At one or more consecutive or interrupted internal positions of the antisense strand, one or more nucleosides or nucleotides are replaced by one or more linker units (eg, 1 to 3 linker units).

[0260] In some embodiments, one or more linker units (e.g., 1 to 3 linker units) are continuously or intermittently attached to the sense strand (e.g., the 3'- or 5'-end) of a double-stranded RNA (e.g., a double-stranded siRNA).

[0261] In some embodiments, one or more linker units (eg, 1 to 3 linker units) are continuously or intermittently attached to the sense strand at the 3'-end of the double-stranded RNA (eg, double-stranded siRNA).

[0262] In some embodiments, one or more linker units (eg, 1 to 3 linker units) are continuously or intermittently attached to the sense strand at the 5'-end of the double-stranded RNA (eg, double-stranded siRNA).

[0263] In some embodiments, one or more nucleosides or nucleotides are replaced by one or more linker units (e.g., 1-3 linker units) at one or more continuous or intermittent internal positions of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA).

[0264] In some embodiments, one or more linker units (e.g., 1 to 3 linker units) are continuously or intermittently attached to the antisense strand (e.g., the 3'- or 5'-end) of a double-stranded RNA (e.g., a double-stranded siRNA).

[0265] In some embodiments, one or more linker units (e.g., 1 to 3 linker units) are continuously or intermittently attached to the antisense strand at the 3'-end of the double-stranded RNA (e.g., double-stranded siRNA).

[0266] In some embodiments, one or more linker units (e.g., 1 to 3 linker units) are continuously or intermittently attached to the antisense strand at the 5'-end of the double-stranded RNA (e.g., double-stranded siRNA).

[0267] In some embodiments, one or more nucleosides or nucleotides are replaced by one or more linker units (e.g., 1 to 3 linker units) at one or more continuous or intermittent internal positions of the antisense strand of a double-stranded RNA (e.g., a double-stranded siRNA).

[0268] In some embodiments, the scaffold comprises a linker unit p -((nucleic acid material)-(linker unit) s ) r -(Nucleic acid material) q where: each linker unit is independent of other linker units and each nucleic acid entity is independent of other nucleic acid entities; 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; and The scaffold comprises at least one linker unit and at least one nucleic acid agent.

[0269] In some embodiments, the scaffold comprises a linker unit p -((nucleic acid material)-(linker unit) s ) r -(nucleic acid substance).

[0270] In some embodiments, the scaffold comprises a linker unit p -((nucleic acid material)-(linker unit) s ) r It is.

[0271] In some embodiments, the scaffold comprises a linker unit p -(nucleic acid substance).

[0272] In some embodiments, the scaffold comprises a (nucleic acid agent)-(linker unit) s -(nucleic acid substance).

[0273] In some embodiments, the scaffold comprises: [ka] or a pharma- ceutically acceptable salt thereof, wherein: 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 (such as, for example, 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); and Each R Z is independently H or C1-C6 alkyl optionally substituted with one or more halogen or cyano.

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

[0275] In some embodiments, the scaffold is formed by linking a linker unit based on any of the following linker compounds to a ligand: [ka] [ka]

[0276] 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 ligand.

[0277] In some embodiments, the scaffold is selected from the scaffolds listed in Table S1. [Table S1-1] [Table S1-2]

[0278] In some embodiments, the scaffold comprises: [ka] or a pharma- ceutically acceptable salt thereof, wherein: W can be 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), where C1-C 30 The alkyl is optionally substituted).

[0279] In some embodiments, the scaffold is formed by linking a linker unit based on any of the linker compounds described herein to a nucleic acid agent.

[0280] In some embodiments, the scaffold is formed by linking a linker unit based on any of the following linker compounds to a nucleic acid material: [ka] [ka]

[0281] In some embodiments, the scaffold is selected from the scaffolds listed in Table S2: [Table S2-1] [Table S2-2] [Table S2-3] [Table S2-4] [Table S2-5] [Table S2-6]

Table S2-7

Table S2-8

Table S2-9

Table S2-10

Table S2-11

Table S2-12

Table S2-13

Table S2-14

Table S2-15

Table S2-16

Table S2-17

Table S2-18

Table S2-19

Table S2-20

Table S2-21

Table S2-22

Table S2-23

[0282] In some aspects, the disclosure provides a conjugate or a pharma- ceutically acceptable salt thereof, wherein the conjugate is: (i) one or more nucleic acid agents; (ii) one or more ligands; and (iii) one or more linker units, wherein each linker unit is independently: [ka] where the variable R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , X, Y, and Z are as described herein, where # indicates the connection to the ligand and ## indicates the connection to the nucleic acid agent.

[0283] In some embodiments, the connection "#" is a direct connection to the ligand, ie, without any linking moiety.

[0284] In some embodiments, the connection "#" is an indirect connection 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 an alkylene chain, wherein optionally one or more carbon atoms in the alkylene chain may be independently replaced by one or more of -C(O)-, -C(O)O-, -OC(O)-, -C(O)NH-, -NHC(O)-, -NHC(O)NH-, -C(S)-, -C(S)O-, -OC(S)-, -C(S)NH-, -NHC(S)-, or -NHC(S)NH-, and wherein the alkylene chain is optionally substituted by 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 two, three, or more C1-C6 alkyl. 15 and branched alkylene chains, including alkylene chains, where optionally one or more carbon atoms of each alkylene chain may be independently replaced by one or more of -C(O)-, -C(O)O-, -OC(O)-, -C(O)NH-, -NHC(O)-, -NHC(O)NH-, -C(S)-, -C(S)O-, -OC(S)-, -C(S)NH-, -NHC(S)-, or -NHC(S)NH-, and where each alkylene chain may be independently optionally replaced by 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 linker between two C1-C 15 In some embodiments, the linking moiety is a branched alkylene chain, including an alkylene chain. 15 In some embodiments, the linking moiety is a branched alkylene chain, including an alkylene chain. 15 It is a branched alkylene chain, including an alkylene chain.

[0285] In some embodiments, the connection "##" is a direct connection to the nucleic acid material, i.e., without any linking moieties.

[0286] In some embodiments, the connection "##" is an indirect connection to the nucleic acid material, i.e., there is a linking moiety between the linker unit and the nucleic acid material. 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.

[0287] 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.

[0288] 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

[0026] The linker units may comprise 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).

[0289] 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.

[0290] 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, wherein: one or more linker units (e.g., 1 to 3 linker units) are continuously or discontinuously connected to the sense strand (e.g., at the 3'- or 5'-end) of a double-stranded RNA (e.g., a double-stranded siRNA); At one or more continuous or interrupted internal positions of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA), one or more nucleosides or nucleotides are replaced by 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 continuously or discontinuously connected to the antisense strand (e.g., the 3'- or 5'-end) of a double-stranded RNA (e.g., a double-stranded siRNA); and / or At one or more consecutive or interrupted internal positions of the antisense strand, one or more nucleosides or nucleotides are replaced by one or more linker units (eg, 1 to 3 linker units).

[0291] 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, wherein: One or more linker units (e.g., 1 to 3 linker units) are continuously or discontinuously connected to the sense strand (e.g., at the 3'- or 5'-end) of a double-stranded RNA (e.g., a double-stranded siRNA); and At one or more continuous or intermittent internal positions of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA), one or more nucleosides or nucleotides are replaced by one or more linker units (e.g., 1 to 3 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, wherein: One or more linker units (e.g., 1 to 3 linker units) are continuously or discontinuously connected to the antisense strand (e.g., at the 3'- or 5'-end) of the double-stranded RNA (e.g., double-stranded siRNA); and At one or more consecutive or interrupted internal positions of the antisense strand, one or more nucleosides or nucleotides are replaced by one or more linker units (eg, 1 to 3 linker units).

[0293] In some embodiments, one or more linker units (e.g., 1 to 3 linker units) are continuously or intermittently attached to the sense strand (e.g., the 3'- or 5'-end) of a double-stranded RNA (e.g., a double-stranded siRNA).

[0294] In some embodiments, one or more linker units (eg, 1 to 3 linker units) are continuously or discontinuously attached to the 3'-end of the sense strand of a double-stranded RNA (eg, a double-stranded siRNA).

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

[0296] In some embodiments, at one or more continuous or intermittent internal positions of the sense strand of a double-stranded RNA (e.g., a double-stranded siRNA), one or more nucleosides or nucleotides are replaced by one or more linker units (e.g., 1 to 3 linker units).

[0297] In some embodiments, one or more linker units (e.g., 1 to 3 linker units) are continuously or intermittently attached to the antisense strand (e.g., the 3'- or 5'-end) of a double-stranded RNA (e.g., a double-stranded siRNA).

[0298] In some embodiments, one or more linker units (e.g., 1 to 3 linker units) are continuously or intermittently attached to the antisense strand at the 3'-end of the double-stranded RNA (e.g., double-stranded siRNA).

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

[0300] In some embodiments, at one or more continuous or intermittent internal positions of the antisense strand of a double-stranded RNA (e.g., a double-stranded siRNA), one or more nucleosides or nucleotides are replaced by one or more linker units (e.g., 1 to 3 linker units).

[0301] In some embodiments, the conjugate comprises a (linker unit-(ligand) 0-1 ) p -((nucleic acid material)-(linker unit-(ligand) 0-1 ) s ) r -(Nucleic acid material) 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; and The conjugate comprises at least one linker unit, at least one nucleic acid agent, and at least one ligand.

[0302] In some embodiments, the conjugate comprises a (linker unit-(ligand) 0-1 ) p -((nucleic acid material)-(linker unit-(ligand) 0-1 ) s ) r -(nucleic acid substance).

[0303] In some embodiments, the conjugate comprises a (linker unit-(ligand) 0-1 ) p -((nucleic acid material)-(linker unit-(ligand) 0-1 ) s ) r It is.

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

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

[0306] In some embodiments, the conjugate is selected from the conjugates described in Table C, where the nucleic acid agents are joined by ##, and ## is a direct or indirect connection as described herein. [Table C-1] [Table C-2] [Table C-3] [Table C-4] [Table C-5] [Table C-6] [Table C-7] [Table C-8] [Table C-9] [Table C-10] [Table C-11] [Table C-12] [Table C-13] [Table C-14] [Table C-15] [Table C-16] [Table C-17] [Table C-18]

[0307] Linker Units As used herein, "linker unit" or "linker unit" refers to a moiety corresponding to a linker compound in which W, Y, and / or Z are replaced by connections to a ligand and / or nucleic acid agent.

[0308] In some embodiments, the linker unit is of formula (I) where W is replaced by a connection to a ligand.

[0309] In some embodiments, the linker unit is of formula (I) where Y and / or Z is replaced by a connection to a nucleic acid agent.

[0310] In some embodiments, the linker unit is of formula (I) where W is replaced by a connection to a ligand; and Y and / or Z is replaced by a connection to a nucleic acid agent.

[0311] In some embodiments, the linker unit is of formula (I'-1), (I'-2), (II'-1), or (II'-2), where W is replaced by a connection to a ligand.

[0312] In some embodiments, the linker unit is of formula (I'-1), (I'-2), (II'-1), or (II'-2), where Y and / or Z are replaced by a connection to a nucleic acid agent.

[0313] In some embodiments, the linker unit is of formula (IA) or (II-A), where W is replaced by a connection to a ligand.

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

[0315] In some embodiments, the linker unit is of formula (IA) or (II-A), where W is replaced by a connection to a ligand; and Y and / or Z is replaced by a connection to a nucleic acid agent.

[0316] In some embodiments, the linker unit is of formula (I-A'-1), (I-A'-2), (II-A'-1), or (II-A'-2), where W is replaced by a connection to a ligand.

[0317] In some embodiments, the linker unit is of formula (I-A'-1), (I-A'-2), (II-A'-1), or (II-A'-2), where Y and / or Z are replaced by a connection to a nucleic acid agent.

[0318] In some embodiments, the linker unit is of formula (I-A'-1), (I-A'-2), (II-A'-1), or (II-A'-2), where W is replaced by a connection to a ligand; and Y and / or Z is replaced by a connection to a nucleic acid agent.

[0319] In some embodiments, the linker unit is of formula (IB) or (II-B), where W is replaced by a connection to a ligand.

[0320] In some embodiments, the linker unit is of formula (IB) or (II-B), where Y and / or Z is replaced by a connection to a nucleic acid agent.

[0321] In some embodiments, the linker unit is of formula (IB) or (II-B), where W is replaced by a connection to a ligand; and Y and / or Z is replaced by a connection to a nucleic acid agent.

[0322] In some embodiments, the linker unit is of formula (I-B'-1), (I-B'-2), (II-B'-1), or (II-B'-2), where W is replaced by a connection to a ligand.

[0323] In some embodiments, the linker unit is of formula (I-B'-1), (I-B'-2), (II-B'-1), or (II-B'-2), where Y and / or Z are replaced by a connection to a nucleic acid agent.

[0324] In some embodiments, the linker unit is of formula (I-B'-1), (I-B'-2), (II-B'-1), or (II-B'-2), where W is replaced by a connection to a ligand; and Y and / or Z are replaced by a connection to a nucleic acid agent.

[0325] In some embodiments, the linker unit, prior to attachment, is a linker compound described herein.

[0326] In some embodiments, the linker unit, prior to attachment, is a compound of formula (I) or a pharma- ceutically acceptable salt thereof.

[0327] In some embodiments, the linker unit, prior to attachment, is a compound of formula (I'-1), (I'-2), (II'-1), or (II'-2), or a pharma- ceutically acceptable salt thereof.

[0328] 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.

[0329] In some embodiments, the linker unit, prior to attachment, is a compound of formula (I-A'-1), (I-A'-2), (II-A'-1), or (II-A'-2), or a pharma- ceutically acceptable salt thereof.

[0330] 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.

[0331] In some embodiments, the linker unit, prior to attachment, is a compound of formula (I-B'-1), (I-B'-2), (II-B'-1), or (II-B'-2), or a pharma- ceutically acceptable salt thereof.

[0332] In some embodiments, the linker unit, prior to attachment, is a compound selected from the compounds set forth in Table L and their pharma- ceutically acceptable salts.

[0333] Ligand As used herein, the term "ligand" refers to a moiety that, when covalently attached to a nucleic acid agent (e.g., an oligonucleotide), is capable of mediating its entry or facilitating its delivery to a target site (e.g., a target cell or tissue).

[0334] In some embodiments, the ligand comprises a sugar ligand moiety (eg, N-acetylgalactosamine (GalNAc)) that allows for direct uptake of the oligonucleotide into the liver.

[0335] In some embodiments, the ligand binds to the asialoglycoprotein receptor (ASGPR). In some embodiments, the ligand binds (eg, via the ASGPR) to the liver, e.g., to liver parenchymal cells.

[0336] Suitable ligands include, but are not limited to, those disclosed in Winkler (Ther. Deliv., 2013, 4(7): 791-809), PCT Patent Application Publication Nos. 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.

[0337] In some embodiments, the ligand comprises a carbohydrate moiety.

[0338] 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), with 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 of about 4-9 monosaccharide units. In some embodiments, the carbohydrate moiety includes a polysaccharide (e.g., starch, glycogen, cellulose, or a polysaccharide gum).

[0339] In some embodiments, the carbohydrate moiety comprises a monosaccharide, a disaccharide, a trisaccharide, or a tetrasaccharide.

[0340] In some embodiments, the carbohydrate moiety comprises an oligosaccharide (eg, containing from about 4 to about 9 monosaccharide units).

[0341] In some embodiments, the carbohydrate moiety comprises a polysaccharide (eg, starch, glycogen, cellulose, or a polysaccharide gum).

[0342] In some embodiments, the ligand is capable of binding to a human asialoglycoprotein receptor (ASGPR), such as the human asialoglycoprotein receptor 2 (ASGPR2).

[0343] In some embodiments, the carbohydrate moiety comprises a saccharide (eg, 1, 2, or 3 saccharides).

[0344] In some embodiments, the carbohydrate moiety comprises a galactose or derivative thereof (eg, 1, 2, or 3 galactose or a derivative thereof).

[0345] In some embodiments, the carbohydrate moiety comprises an N-acetylgalactosamine or derivative thereof (eg, 1, 2, or 3 N-acetylgalactosamines or derivatives thereof).

[0346] In some embodiments, the carbohydrate moiety comprises an N-acetyl-D-galactosylamine or derivative thereof (eg, 1, 2, or 3 N-acetyl-D-galactosylamines or derivatives thereof).

[0347] In some embodiments, the carbohydrate moiety comprises an N-acetylgalactosamine (eg, 1, 2, or 3 N-acetylgalactosamines).

[0348] In some embodiments, the carbohydrate moiety comprises an N-acetyl-D-galactosylamine (eg, 1, 2, or 3 N-acetyl-D-galactosylamines).

[0349] In some embodiments, the carbohydrate moiety comprises mannose or a derivative thereof (eg, mannose-6-phosphate).

[0350] In some embodiments, the carbohydrate moiety further comprises a linking moiety that joins one or more sugars (eg, N-acetyl-D-galactosylamine) to the linker unit.

[0351] In some embodiments, the linking moiety comprises a thioether (eg, a thiosuccinimide, or a hydrolyzable analog thereof), a disulfide, a triazole, a phosphorothioate, a phosphodiester, an ester, an amide, or any combination thereof.

[0352] In some embodiments, the linking moiety is a triantennary linking moiety.

[0353] Suitable ligands include, but are not limited to, those disclosed in PCT Application Publication Nos. WO / 2015 / 006740, WO / 2016 / 100401, WO / 2017 / 214112, WO / 2018 / 039364, and WO / 2018 / 045317, each of which is incorporated herein by reference.

[0354] In some embodiments, the ligand is: [ka] (For example, 1, 2, or 3 [ka] ).

[0355] In some embodiments, the ligand is: [ka] (For example, 1, 2, or 3 [ka] ).

[0356] In some embodiments, the ligand is: [ka] (For example, 1, 2, or 3 [ka] ).

[0357] In some embodiments, the ligand is: [ka] (For example, 1, 2, or 3 [ka] ).

[0358] In some embodiments, the ligand is: [ka] (For example, 1, 2, or 3 [ka] ).

[0359] In some embodiments, the ligand is: [ka] (For example, 1, 2, or 3 [ka] ).

[0360] In some embodiments, the ligand is: [ka] (For example, 1, 2, or 3 [ka] ).

[0361] In some embodiments, the ligand is: [ka] (For example, 1, 2, or 3 [ka] ).

[0362] In some embodiments, the ligand is: [ka] Includes.

[0363] In some embodiments, the ligand is: [ka] Includes.

[0364] In some embodiments, the ligand is: [ka] Includes.

[0365] In some embodiments, the ligand is: [ka] Includes.

[0366] In some embodiments, the ligand is: [ka] Includes.

[0367] In some embodiments, the ligand is: [ka] Includes.

[0368] In some embodiments, the ligand is: [ka] Includes.

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

[0370] In some embodiments, the ligand comprises a lipid moiety (eg, 1, 2, or 3 lipid moieties).

[0371] In some embodiments, the lipid moiety is a C8-C 24 The fatty acids include fatty acids, cholesterol, vitamins, sterols, phospholipids, or any combination thereof.

[0372] In some embodiments, the ligand comprises a peptide moiety (eg, 1, 2, or 3 peptide moieties).

[0373] In some embodiments, the peptide portion includes (eg, one, two, or three) integrins, insulin, glucagon-like peptides, or any combination thereof.

[0374] In some embodiments, the ligand comprises an antibody moiety (eg, transferrin).

[0375] In some embodiments, the ligand comprises one, two, or three antibody moieties (eg, transferrin).

[0376] In some embodiments, the ligand comprises an oligonucleotide (eg, an aptamer or CpG).

[0377] In some embodiments, the ligand comprises one, two, or three oligonucleotides (eg, an aptamer or CpG).

[0378] In some embodiments, the ligand is: Monosaccharides, disaccharides, or trisaccharides (e.g., N-acetyl-D-galactosylamine); 1, 2, or 3 lipid moieties; 1, 2, or 3 peptide moieties; 1, 2, or 3 antibody moieties; 1, 2, or 3 oligonucleotides; or Any combination thereof is included.

[0379] Nucleic acid material In some embodiments, the nucleic acid agent comprises an oligonucleotide.

[0380] In some embodiments, the nucleic acid agent (eg, an oligonucleotide) includes one or more phosphate groups or one or more phosphate group analogs.

[0381] In some embodiments, the linker unit is attached to a nucleic acid agent (eg, an oligonucleotide) via a phosphate group, or a phosphate group analog, in the nucleic acid agent.

[0382] 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 20, 21, 22, or 23 nucleotides.

[0383] In some embodiments, the nucleic acid material comprises RNA, DNA, or a mixture thereof.

[0384] In some embodiments, the nucleic acid material comprises RNA.

[0385] 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 anti-microRNA (antimiR), an antagomir, a dsRNA, a ssRNA, an aptamer, an immunostimulatory oligonucleotide, a decoy oligonucleotide, a splicing modulation oligonucleotide, a triplex forming oligonucleotide, a G-quadruplex, or an antisense oligonucleotide.

[0386] In some embodiments, the nucleic acid agent comprises double-stranded RNA (dsRNA), where the double-stranded RNA comprises a sense strand and an antisense strand, as described herein.

[0387] In some embodiments, the 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.

[0388] 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.

[0389] It is understood that the antisense strand is also known as the guide strand, and the terms "antisense strand" and "guide strand" are used interchangeably herein.

[0390] In some embodiments, the oligonucleotide is an iRNA.

[0391] The term "iRNA" refers to an RNA agent that can downregulate the expression of a target gene (e.g., siRNA), such as an endogenous or pathogenic target RNA. Without wishing to be bound by theory, iRNA can act by one or more of a number of mechanisms, including post-transcriptional cleavage of target mRNA (referred to in the art as RNAi), or pre-transcriptional or pre-translational mechanisms. iRNA can include a single strand, or can include two or more strands, for example, it can be a double-stranded iRNA. When iRNA is single-stranded, it can include a 5' modification, including one or more phosphate groups or one or more phosphate group analogs. In some embodiments, iRNA is double-stranded. In some embodiments, one or both strands of a double-stranded iRNA can be modified, such as, for example, a 5' modification.

[0392] An iRNA typically comprises a region of sufficient homology with 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. Although it is not necessary that there be perfect complementarity between the iRNA and the target, the correspondence is sufficient that the iRNA or its cleavage product can direct sequence-specific silencing, for example, by cleavage of the target RNA, such as an mRNA, RNAi.

[0393] The nucleotides of the iRNA may be modified (e.g., one or more nucleotides may contain a 2'-F or 2'-OCH3 group or may be a nucleotide surrogate). Single-stranded or double-stranded regions of the iRNA may be modified or contain nucleotide surrogates, e.g., unpaired regions or regions of hairpin structures, e.g., the region linking two complementary regions may have modifications or nucleotide surrogates. Modifications may stabilize one or more 3'- or 5'-ends of the iRNA, e.g., against exonucleases. Modifications may include C3 (or C6, C7, C12) amino linkers, thiol linkers, carboxyl linkers, non-nucleotide spacers (C3, C6, C9, C12, abasic, triethylene glycol, hexaethylene glycol), special biotin or fluorescein reagents that become phosphoramidites and have another DMT-protected hydroxyl group, 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, e.g., the use of deoxyribonucleotides, such as deoxythymidine, instead of ribonucleotides, as well as modifications at the phosphate group, e.g., phosphorothioate modifications. In some embodiments, different strands will contain different modifications.

[0394] In some embodiments, the strands are selected such that the iRNA contains a single-stranded or unpaired region at one or both ends of the molecule. The double-stranded iRNA may have an overhang, such as one or two 5' or 3' overhangs (e.g., a 3' overhang of at least 2-3 nucleotides). In some embodiments, the iRNA has an overhang, such as a 3' overhang of 1, 2, or 3 nucleotides in length on each end. These overhangs can be the result of one strand being longer than the other, or the result of two strands of the same length overhanging.

[0395] In some embodiments, the length of the duplex region between the strands of the iRNA is between 6 and 30 nucleotides in length. In some embodiments, the duplex region is between 15 and 30, most preferably 18, 19, 20, 21, 22, and 23 nucleotides in length. In some embodiments, the duplex region is between 6 and 20 nucleotides in length, most preferably 6, 7, 8, 9, 10, 11, and 12 nucleotides in length.

[0396] The oligonucleotides may be those described in U.S. Patent Publication Nos. 2009 / 0239814, 2012 / 0136042, 2013 / 0158824, or 2009 / 0247608, which are incorporated herein by reference.

[0397] In some embodiments, the oligonucleotide is a siRNA.

[0398] In some embodiments, the oligonucleotide is a single stranded siRNA.

[0399] In some embodiments, the oligonucleotide is a double-stranded siRNA, such as a double-stranded siRNA described herein.

[0400] As used herein, a "single-stranded siRNA" is an siRNA made up of a single strand that includes a duplex region formed by intrastrand pairing, which may be, for example, a hairpin or pan-handle structure. A single-stranded siRNA may be antisense with respect to its target molecule.

[0401] Single-stranded siRNA is long enough so that it can enter RISC and participate in the RISC-mediated cleavage of target mRNA.Single-stranded siRNA is at least 14, and in some embodiments at least 15, 20, 25, 29, 35, 40 or 50 nucleotides in length.In some embodiments, it is less than 200, 100, 80, 60, 50, 40 or 30 nucleotides in length.

[0402] In some embodiments, the single-stranded siRNA has a length of 10 to 40 nucleotides, 12 to 35 nucleotides, 15 to 30 nucleotides, 18 to 25 nucleotides, or 20 to 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.

[0403] The hairpin siRNA may have a duplex region equal to or at least 17, 18, 19, 20, 21, 22, 23, 24, or 25 nucleotide pairs. The duplex region may be equal to or less than 200, 100, or 50 nucleotide pairs in length. In some embodiments, the duplex region ranges from 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 overhang 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.

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

[0405] As used herein, a "double-stranded siRNA" is an siRNA that contains more than one strand, and optionally two strands, in which interstrand hybridization is capable of forming a region of duplex structure.

[0406] In some embodiments, the sense strand of the double stranded siRNA is equal to or at least 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 29, 40, or 60 nucleotides in length. It is equal to or less than 200, 100, or 50 nucleotides in length. Ranges may be 17-25, 19-23, 19-21, 21-23, or 20-22 nucleotides in length.

[0407] 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.

[0408] In some embodiments, the sense strand has a length of 18, 19, 20, 21, or 22 nucleotides.

[0409] In some embodiments, the antisense strand of the double stranded siRNA is equal to or at least 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 29, 40, or 60 nucleotides in length. It is equal to or less than 200, 100, or 50 nucleotides in length. Ranges may be 17-25, 19-23, 19-21, 21-23, or 20-22 nucleotides in length.

[0410] 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.

[0411] In some embodiments, the antisense strand has a length of 20, 21, 22, 23, or 24 nucleotides.

[0412] 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.

[0413] In some embodiments, the sense strand has a length of 18 nucleotides and the antisense strand has a length of 20 nucleotides.

[0414] In some embodiments, the sense strand has a length of 19 nucleotides and the antisense strand has a length of 21 nucleotides.

[0415] In some embodiments, the sense strand has a length of 20 nucleotides and the antisense strand has a length of 22 nucleotides.

[0416] In some embodiments, the sense strand has a length of 21 nucleotides and the antisense strand has a length of 23 nucleotides.

[0417] In some embodiments, the sense strand has a length of 22 nucleotides and the antisense strand has a length of 24 nucleotides.

[0418] The double-stranded portion 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 nucleotide pairs in length. It can be equal to or less than 200, 100, or 50 nucleotide pairs in length. Ranges can be 15-30, 17-23, 19-23, and 19-21 nucleotide pairs in length.

[0419] In some embodiments, the siRNA is large enough to be cleaved by an endogenous molecule, such as Dicer, to generate smaller siRNAs, such as siRNA agents.

[0420] These sense and antisense strands may be selected so that the double-stranded siRNA comprises a single-stranded or unpaired region at one or both ends of the molecule. Thus, the double-stranded siRNA may comprise a sense strand and an antisense strand that are paired to include an overhang, such as, for example, one or two 5' or 3' overhangs, or a 3' overhang of 1-3 nucleotides. These overhangs can be the result of one strand being longer than the other, or the result of two strands of the same length overhanging. Some embodiments will have at least one 3' overhang. In some embodiments, both ends of the siRNA molecule will have a 3' overhang. In some embodiments, the overhang is 2 nucleotides.

[0421] In some embodiments, the length of the double-stranded region is within the range of 15-30, or 18, 19, 20, 21, 22, and 23 nucleotides in length, such as the ssiRNAs discussed above. The ssiRNA can be similar in length and structure to the natural Dicer-processed product from a long dsiRNA. Also included are embodiments in which the two strands of the ssiRNA are connected, such as covalently linked. Hairpins or other single-stranded structures that provide the necessary double-stranded region and 3' overhang are also contemplated.

[0422] The siRNA described herein, including double-stranded siRNA and single-stranded siRNA, can mediate the silencing of target RNA, such as mRNA, such as the transcription product 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 pathogenic gene.In addition, non-mRNA RNA, such as tRNA and viral RNA, can also be targeted.

[0423] As used herein, the phrase "mediate RNAi" refers to the ability to silence target RNA in a sequence-specific manner.Without wishing to be bound by theory, it is believed that silencing uses the RNAi mechanism or process and guide RNA, for example, 21-23 nucleotide ssiRNA.

[0424] In some embodiments, siRNA has "sufficient complementarity" with target RNA, e.g., target mRNA, so that siRNA silences the production of protein encoded by target mRNA. In another embodiment, siRNA has "strict complementarity" with target RNA, e.g., target RNA and siRNA anneal, e.g., to form a hybrid exclusively made by Watson-Crick base pairing in the region of strict complementarity. Target RNA with "sufficient complementarity" can include an internal region (e.g., of at least 10 nucleotides) that has strict complementarity with target RNA. In some embodiments, siRNA specifically discriminates one nucleotide difference. In this case, siRNA mediates RNAi only when strict complementarity is found within the region of 1-nucleotide difference (e.g., within 7 nucleotides).

[0425] MicroRNA:MicroRNAs (miRNAs) are a highly conserved class of small RNA molecules that are transcribed from DNA in plant and animal genomes but are not translated into proteins. Processed miRNAs are single-stranded, ~17-25 nucleotide (nt) RNA molecules that are incorporated into RNA-induced silencing complexes (RISCs) and have been identified as important regulators of development, cell proliferation, apoptosis, and differentiation. They are believed to play a role in regulating gene expression by binding to the 3′-untranslated regions of specific mRNAs. RISCs mediate downregulation of gene expression through translational inhibition, transcript cleavage, or both. RISCs are also involved in transcriptional silencing in the nuclei of a wide range of eukaryotic organisms.

[0426] The number of miRNA sequences identified to date is large and growing, illustrative 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.

[0427] Antisense oligonucleotidesIn 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 targeted polynucleotide sequence. Antisense oligonucleotides are single-stranded DNA or RNA that are complementary to a selected sequence, such as a target gene mRNA. Antisense oligonucleotides are believed to inhibit gene expression by binding to complementary mRNA. Binding to the target mRNA can lead to inhibition of gene expression either by binding to it and thereby preventing translation of the complementary mRNA strand, or by leading to degradation of the target mRNA. Antisense DNA can be used to target 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, antisense oligonucleotides comprise about 10 to about 50 nucleotides, more preferably about 15 to about 30 nucleotides. The term also encompasses antisense oligonucleotides that may not be strictly complementary to the desired target gene. Thus, it is contemplated that cases may occur in which non-target specific activity is observed with the antisense, or where an antisense sequence containing one or more mismatches with the target sequence may be most preferred for a particular application.

[0428] Antisense oligonucleotides are clearly effective and targeted inhibitors of protein synthesis, and can be used to specifically inhibit protein synthesis by targeted genes.The effectiveness of antisense oligonucleotides in 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). Further 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 have also been described that can be used to inhibit and treat various abnormal cell growth, such as cancer (U.S. Pat. Nos. 5,747,470; 5,591,317 and 5,783,683, each of which is incorporated by reference).

[0429] Methods for making antisense oligonucleotides are known in the art and can be easily adapted to make antisense oligonucleotides that target any polynucleotide sequence. Selection of antisense oligonucleotide sequences specific to a given target sequence is based on the analysis of the selected target sequence and the determination of secondary structure, Tm, binding energy, and relative stability. Antisense oligonucleotides may be selected based on their relative inability to form dimers, hairpins, or other secondary structures that reduce or inhibit specific binding to target mRNA in host cells. Highly preferred target regions of mRNA include those regions at or near the AUG translation initiation codon and those sequences that are substantially complementary to the 5' region of mRNA. These secondary structure analyses 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).

[0430] Antagomir:Antagomir is an RNA-like oligonucleotide with various modifications related to pharmacological properties such as RNAse protection and enhanced tissue and cellular uptake. They differ from normal RNA, for example, by complete 2'-O-methylation of sugars, phosphorothioate backbone, and cholesterol-moiety, for example, at the 3'-terminus. Antagomir can be used to efficiently silence endogenous miRNA by forming a duplex containing antagomir and endogenous miRNA, thereby preventing miRNA-induced 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.

[0431] Antagomir can include ligand-conjugated monomer subunits and monomers for oligonucleotide synthesis. Exemplary monomers are described in U.S. Patent Application Publication No. 2005 / 0107325, which is incorporated by reference in its entirety. Antagomir can have a ZXY structure as described in WO2004 / 080406, which is incorporated by reference in its entirety. Antagomir can be conjugated with an amphiphilic moiety. Examples of amphiphilic moieties for use with oligonucleotide materials are described in WO2004 / 080406, which is incorporated by reference in its entirety.

[0432] AptamersAptamers 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 engineered to 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. 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 can be RNA or DNA based and can include riboswitches. Riboswitches are parts of mRNA molecules that can directly bind to small target molecules, and binding to the target affects the activity of genes. Thus, the mRNA that contains 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 trials of 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 even organisms. Aptamers may be prepared by any known method, including synthetic, recombinant, and purification methods, and may be used alone or in combination with other aptamers that are specific for the same target. As further described more fully herein, the term "aptamer" specifically includes "secondary aptamers" that include consensus sequences derived from comparing two or more known aptamers to a given target.

[0433] RibozymesIn another embodiment, the nucleic acid-lipid particles are related to ribozymes, which are RNA molecular complexes that contain specific catalytic domains 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 catalyze phosphoester transfer reactions with a high degree of specificity, sometimes 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 bind via base-pairing interactions to the ribozyme's internal guide sequence ("IGS") prior to the specific chemical reaction.

[0434] At least six basic types of naturally occurring enzymatic RNAs are currently known. Each is capable of catalyzing the hydrolysis of RNA phosphodiester bonds in trans under physiological conditions (and thus capable of cleaving other RNA molecules). Generally, enzymatic nucleic acids act by first binding to a target RNA. Such binding occurs through the target binding portion of the enzymatic nucleic acid, which is held in close proximity to the enzymatic portion of the molecule that acts to cleave the target RNA. Thus, the enzymatic nucleic acid first recognizes and then binds the target RNA through complementary base-pairing, and once bound to the correct site, acts enzymatically to cleave the target RNA. Such strategic cleavage of a target RNA would destroy 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, as well as repeatedly bind and cleave new targets.

[0435] The enzymatic nucleic acid molecule may be formed, for example, in a hammerhead, hairpin, hepatitis delta 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 by Rossi et al. Nucleic Acids Res. 1992 Sep. 11; 20(17):4559-65. Examples of hairpin motifs are described by Hampel et al. (European Patent Application Publication No. EP 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. An example of a hepatitis delta virus motif is described by Perrotta and Been, Biochemistry. 1992 Dec. 1; 31(47):11843-52; an example of an RNase P motif is described by Guerrier-Takada et al., Cell. 1983 Dec; 35(3 Pt 2):849-57; and a Neurospora VS RNA ribozyme motif is described by 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. Patent No. 4,987,071. 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 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, the ribozyme constructs required are not limited to the specific motifs mentioned herein.

[0436] Methods for making ribozymes targeted to any polynucleotide sequence are known in the art. Ribozymes may be designed as described in International Patent Application Publication Nos. WO 93 / 23569 and WO 94 / 02595, each of which is specifically incorporated herein by reference, and synthesized for testing in vitro and in vivo as described therein.

[0437] 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., International Patent Application Publication Nos. WO 92 / 07065, WO 93 / 15187, and WO 91 / 03162; European Patent Application Publication No. 92110298.4; U.S. Pat. No. 5,334,711; and International Patent Application Publication No. WO 94 / 13688, which describe various chemical modifications that can be made to the sugar portion of enzymatic RNA molecules), by removing stem II bases to shorten RNA synthesis time and reduce chemical requirements.

[0438] Immunostimulatory Oligonucleotides The nucleic acid associated with the lipid particle includes an immunostimulatory oligonucleotide (ISS; single-stranded or double-stranded) that is immunostimulatory and capable of inducing an immune response when administered to a subject, which may be a mammal or other patient. The ISS may contain, for example, specific palindromes that lead to hairpin-type 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 (such as multi-G domains, see WO96 / 11266, which is incorporated by reference in its entirety).

[0439] The immune response may be an innate or adaptive immune response. The immune system is divided into a more innate immune system and an acquired adaptive immune system in vertebrates, the latter being further divided into humoral and cellular components. In some embodiments, the immune response may be a mucosal system.

[0440] 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.

[0441] Immunostimulatory nucleic acids are considered to be non-sequence specific if they are not required to specifically bind to and reduce expression of a target polynucleotide in order to elicit an immune response. Thus, certain immunostimulatory nucleic acids can contain sequences that correspond to regions of a naturally occurring gene or mRNA, yet still be considered non-sequence specific immunostimulatory nucleic acids.

[0442] 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, wherein the cytosine in the CpG dinucleotide is methylated. In an alternative embodiment, the nucleic acid comprises at least two CpG dinucleotides, wherein 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, wherein at least one of the CpG dinucleotides comprises a methylated cytosine.

[0443] Connections between linker units, nucleic acid entities, and ligands In some embodiments, the connection between the linker unit and the nucleic acid material is a bond.

[0444] In some embodiments, the connection between the linker unit and the nucleic acid agent is a moiety (eg, a moiety that includes a cleavable group).

[0445] In some embodiments, the connection between the Linker unit and the Ligand is a bond.

[0446] In some embodiments, the connection between the Linker unit and the Ligand is a moiety (eg, a moiety that includes a cleavable group).

[0447] In some embodiments, the connection between the Linker unit and the Ligand comprises a -C(=O)- attached to the Linker unit.

[0448] The group can 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 a chain of atoms, such as, 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, alkylarylalkylene, alkylarylalkenylene, alkylarylalkynylene, alkenylarylalkylene, alkenylarylalkenylene, alkenylarylalkynylene, alkynylarylalkylene, alkynylarylalkenylene, alkynylarylalkynylene, alkylheteroarylalkylene, alkylheteroarylalkenylene, alkylheteroarylalkynylene, alkenylheteroarylalkylene, alkenylheteroarylalkylene, alkenylheteroarylalkylene, Heteroarylalkenylene, alkenylheteroarylalkynylene, alkynylheteroarylalkylene, alkynylheteroarylalkenylene, alkynylheteroarylalkynylene, alkylheterocyclylalkylene, alkylheterocyclylalkenylene, alkylheterocyclylalkynylene, alkenylheterocyclylalkylene, alkenylheterocyclylalkenylene, alkenylheterocyclylalkynylene, alkynylheterocyclylalkylene, alkynylheterocyclylalkenylene, a includes alkynylheterocyclylalkynylene, alkylarylene, alkenylarylene, alkynylarylene, alkylheteroarylene, alkenylheteroarylene, alkynylheteroarylene, and the like, each of which may be substituted or unsubstituted, and one or more methylenes thereof 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, where R is hydrogen, acyl, aliphatic or substituted aliphatic.

[0449] The cleavable group is sufficiently stable outside a cell, but is cleaved upon entry into a target cell to release the two moieties that hold the group together. In a preferred embodiment, 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 can, for example, be selected to mimic or represent intracellular conditions), or under a second reference condition (which can, for example, be selected to mimic or represent conditions found in blood or serum), than in the subject's blood.

[0450] Cleavable groups are sensitive to cleaving agents, such as pH, redox potential, or the presence of degradable molecules. In general, cleaving agents are more prevalent or found at higher levels or activity in cells than in serum or blood. Examples of such degradable agents include: redox agents, selected for a particular substrate or with no substrate specificity, including, for example, oxidases or reductases, or reducing agents such as mercaptans present in cells, which can degrade redox-cleavable groups by reduction; esterases; agents that can create an endosomal or acidic environment, such as those that result in a pH of 5 or less; enzymes that can hydrolyze or degrade acid cleavable groups by acting as general acids, peptidases (which can be substrate specific), and phosphatases.

[0451] Cleavable groups, such as disulfide bonds, can 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 approximately 5.0. Some linkers have cleavable groups that are cleaved at a preferred pH, thereby releasing the cationic lipid from the ligand inside the cell or to a desired compartment of the cell.

[0452] The conjugate can contain a cleavable group that can be cleaved by a specific enzyme. The type of cleavable group incorporated into the conjugate can depend on the cell to be targeted. For example, a ligand that targets the liver can be connected to a cationic lipid through a chemical moiety that contains an ester group. Hepatocytes are rich in esterases, and therefore this group will be cleaved more effectively in hepatocytes 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.

[0453] Coupling groups that include peptide bonds can be used when targeting cell types that are rich in peptidases, such as hepatocytes and synovial cells.

[0454] In general, the suitability of a candidate cleavable group can be evaluated by testing the ability of a degradable agent (or condition) to cleave the candidate group. It may also be 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 and a second condition can be determined, where the first condition is selected to be indicative of cleavage in target cells, and the second condition is selected to be indicative of cleavage in other tissues or biological fluids, such as blood or serum. These evaluations can be performed in cell-free systems, cells, cell cultures, organ or tissue cultures, or in whole animals. It may be useful to perform initial evaluations in cell-free or culture conditions and confirm with further evaluations in whole animals. In preferred embodiments, useful candidate compounds are cleaved at least 2, 4, 10, or 100 times faster in cells (or under in vitro conditions selected to mimic intracellular conditions) than in blood or serum (or under in vitro conditions selected to mimic extracellular conditions).

[0455] Redox-cleavable groups:One class of cleavable groups are redox cleavable groups that are cleaved as soon as they are reduced or oxidized. 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 tried. For example, candidates can be evaluated by using reagents known in the art that mimic the cleavage rate observed in cells, for example target cells, and incubation with dithiothreitol (DTT) or other reducing agents. These candidates can also be evaluated under conditions selected to mimic the conditions of blood or serum. In a preferred embodiment, the candidate compound is cleaved in blood by a maximum of 10%. In a preferred embodiment, a useful candidate compound is degraded at least 2, 4, 10, or 100 times faster in cells (or under in vitro conditions selected to mimic intracellular conditions) than in blood (or under in vitro conditions selected to mimic 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, compared to conditions selected to mimic the extracellular medium.

[0456] Phosphate-Based Cleavable Groups Phosphate-based cleavable groups are cleaved by substances that degrade or hydrolyze phosphate groups. Examples of substances that cleave phosphate groups within a cell are enzymes such as intracellular phosphatases. 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-.

[0457] Acid-Cleavable Groups : Acid cleavable groups are linking groups that are cleaved under acidic conditions. In a preferred embodiment, the acid cleavable groups are cleaved in an acidic environment with a pH of about 6.5 or lower (e.g., about 6.0, 5.5, 5.0, or lower) or by substances such as enzymes that can act as general acids. In cells, certain low pH organelles such as endosomes and lysosomes can provide a cleavage environment for the 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 connected 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.

[0458] Ester-Based Cleavable Groups:Ester-based cleavable groups are cleaved by enzymes such as esterases and amidases within cells. Examples of ester-based cleavable groups include, but are not limited to, esters of alkylene, alkenylene, and alkynylene groups. Ester cleavable linking groups have the general formula -C(O)O-, or -OC(O)-. These candidates can be evaluated using methods similar to those described above.

[0459] Peptide-Based Cleavable Groups: Peptide-based cleavable groups are cleaved intracellularly by enzymes such as peptidases and proteases. 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 either alkylenes, alkenylenes, or alkynylenes. A peptide bond is a special 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 linking groups 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 a compound that is either a carbohydrate itself, made of one or more monosaccharide units (which may be linear, branched or cyclic) having at least six carbon atoms, with an oxygen, nitrogen or sulfur atom bonded to each carbon atom; or a compound that has a portion of a carbohydrate moiety, made of one or more monosaccharide units (which may be linear, branched or cyclic), each having at least six carbon atoms, 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), as well as polysaccharides such as starch, glycogen, cellulose and polysaccharide gums. Particular monosaccharides include sugars of C5 and above (preferably C5-C8); disaccharides and trisaccharides include sugars having two or three monosaccharide units (preferably C5-C8).

[0460] Previously, it has been reported that certain 1'-amino 2'-OTBS carbocyclic phosphoramidites have previously been prepared and incorporated into natural oligonucleotides to handle the conjugation of fluorophores for labeling of oligonucleotides (Org. Lett. 2021, 23, 6735-6739, incorporated herein by reference). Without wishing to be bound by theory, the linkers, scaffolds, and conjugates of the present disclosure may be distinguished from the previously reported 1'-amino 2'-OTBS carbocyclic phosphoramidites in various aspects, including chemical structure, conjugated oligonucleotides, use of the conjugates, and / or synthetic approaches.

[0461] Synthesis method In some aspects, the disclosure provides methods of preparing the compounds of the disclosure.

[0462] In some aspects, the disclosure provides compounds obtainable or obtained by a method of preparing a compound as described herein.

[0463] In some aspects, the disclosure provides intermediates as described herein that are suitable for use in the methods of preparing compounds as described herein.

[0464] The compounds of the present disclosure can be prepared by any suitable method known in the art. Specific processes for the preparation of these compounds are further illustrated in the accompanying examples.

[0465] In the description of the synthetic methods described herein, and any referenced synthetic methods used to prepare starting materials, it is to be understood that all proposed reaction conditions, including choice of solvent, reaction atmosphere, reaction temperature, duration of experiment, and work-up procedures, can be selected by one of ordinary skill in the art.

[0466] 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.

[0467] It will be understood that during the synthesis of the compounds of the present disclosure in the processes set forth herein, or during the synthesis of certain starting materials, it may be desirable to protect certain substituents to prevent their undesired reactions. A skilled chemist will be able to recognize when such protection is necessary and how such protecting groups are introduced and subsequently removed. For examples of protecting groups, see one of the many general textbooks on this subject, such as, for example, "Protective Groups in Organic Synthesis" by Theodora Green (publisher: John Wiley & Sons). Protecting groups may be removed by any convenient method described in the literature or known to a skilled chemist for the removal of the protecting group in question, and such methods are selected so that the removal of the protecting group can be carried out with minimal disruption of other groups of 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.

[0468] For example, suitable protecting groups for an amino or alkylamino group are, 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 can 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 can be, for example, acetals. A suitable protecting group for 1,3-diols can be, for example, tetraisopropyldisiloxanylidene (TIPDS).

[0469] The deprotection conditions for the protecting group will 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, for example, 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, for example, over a catalyst, such as palladium on carbon, or by treatment with a Lewis acid, for example, boron tris(trifluoroacetate). A suitable alternative protecting group for primary amino groups is the phthaloyl group, which can be removed by treatment with an alkylamine, for example, dimethylaminopropylamine, or with hydrazine.

[0470] 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, acyl groups such as alkanoyl or aroyl groups can be removed by hydrolysis, e.g. with a suitable base such as 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.

[0471] Suitable protecting groups for a carboxy group are, for example, an esterifying group, e.g. a methyl or ethyl group which may be removed, for example, by hydrolysis with a base such as sodium hydroxide, or, for example, a tert-butyl group which may be removed, for example, by treatment with an acid, for example an organic acid, such as trifluoroacetic acid, or, for example, a benzyl group which may be removed, for example, by hydrogenation over a catalyst, for example palladium on carbon.

[0472] Advantageously, the reaction of these compounds is carried out in the presence of a suitable solvent, preferably inert under the respective reaction conditions. Examples of suitable solvents are 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; glycol ethers, such as Examples of suitable solvents include, but are not limited to, ethylene glycol monomethyl or monoethyl ether or ethylene glycol dimethyl ether (diglyme); ketones, such as acetone, methyl isobutyl ketone (MIBK) or butanone; amides, such as acetamide, dimethylacetamide, dimethylformamide (DMF) or 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.

[0473] The reaction temperature is preferably about -100°C to 300°C depending on the reaction step and conditions used.

[0474] The reaction time generally ranges from a few minutes to several days, depending on the reactivity of each compound and each reaction condition. Suitable reaction times can be easily determined by methods known in the art, such as reaction monitoring. Based on the reaction temperatures shown above, suitable reaction times generally range from 10 minutes to 48 hours.

[0475] Moreover, by utilizing the procedures described herein in conjunction with ordinary skill 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 representative procedures can be used to prepare these compounds.

[0476] As will be appreciated by those skilled in the art of organic synthesis, the compounds of the present disclosure can be easily accessed by various synthetic routes, some of which are illustrated in the accompanying examples. Those skilled in the art will easily recognize what kind of reagents and reaction conditions are used to obtain the compounds of the present disclosure, and how they are applied and adapted in any particular case - whenever necessary or useful. Furthermore, some of the compounds of the present disclosure can be easily synthesized by converting one specific functional group present in the compounds of the present disclosure, or in a suitable precursor molecule thereof, to another 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; these methods are well known to those skilled in the art. Likewise, those skilled in the art will apply synthetic protecting groups (or protective groups) - whenever necessary or useful; suitable protecting groups and methods for their introduction and removal are well known to those skilled in the art of chemical synthesis and are described in more detail in PGM Wuts, TW Greene, "Greene's Protective Groups in Organic Synthesis", 4th edition (2006) (John Wiley & Sons).

[0477] A general route for the preparation of the compounds of the present application is illustrated herein in Scheme 1. [ka]

[0478] Biological assays The compounds, scaffolds, or conjugates designated, selected, prepared, and / or optimized by the methods described above 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 those assays described below, to determine whether they have the desired activity, such as target binding activity and / or specificity and / or stability.

[0479] Furthermore, high throughput screening can be used to speed up the analysis using such assays. As a result, it can 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.

[0480] A variety of in vitro or in vivo biological assays are suitable for detecting the effect of the disclosed compounds, scaffolds, or conjugates, including, but not limited to, enzyme activity assays, electrophoretic mobility shift assays, reporter gene assays, in vitro cell viability assays, and assays described herein.

[0481] In some embodiments, these biological assays are described in the Examples section herein.

[0482] Pharmaceutical Compositions In some aspects, the present disclosure provides pharmaceutical compositions containing, as an active ingredient, a compound, scaffold, or conjugate of the present disclosure.

[0483] The term "composition," as used herein, is intended to encompass a product containing the specified ingredients in the specified amounts, as well as any product that results directly or indirectly from the combination of the specified ingredients in the specified amounts.

[0484] 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, for example, a solvent or dispersion medium containing water, ethanol, polyol (e.g., glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof. 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. Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In many cases, it will be preferable to include an isotonic agent in the composition, such as sugars, polyalcohols such as mannitol and sorbitol, and sodium chloride. Prolonged absorption of the injectable composition can be brought about by including in the composition a substance that delays absorption, such as aluminum monostearate and gelatin.

[0485] Sterile injectable solutions can be prepared by mixing the required amount of the active compound in a suitable solvent with one or a combination of the above-listed ingredients, followed by sterilization by filtration as required.In general, dispersions are prepared by mixing the active compound into a sterile vehicle that contains a basic dispersion medium and other required ingredients from those listed above.In the case of sterile powders for preparing sterile injectable solutions, the preparation method is vacuum drying or freeze-drying, which produces a powder of the active ingredient plus any additional desired ingredients from their previously sterile-filtered solutions.

[0486] The formulation of the present disclosure may be in the form of an aqueous solution containing an aqueous vehicle. The aqueous vehicle component may contain water and at least one pharma- ceutically acceptable excipient. Suitable acceptable excipients include those selected from the group consisting of solubility enhancers, chelating agents, preservatives, tonicity agents, thickening / suspending agents, buffers, and pH modifiers, and mixtures thereof.

[0487] Any suitable solubility enhancer can be used. Examples of solubility 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, peracetylated-β-cyclodextrin, carboxymethyl-β-cyclodextrin, hydroxyethyl-β-cyclodextrin, 2-hydroxy-3-(trimethylammonio)propyl-β-cyclodextrin, glucosyl-β-cyclodextrin, β-cyclodextrin sulfate (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.

[0488] Any suitable chelating agent can 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, tetrasodium edetate, and the like, and mixtures thereof.

[0489] Any suitable preservative can be used. Examples of preservatives include those selected from the group consisting of 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.

[0490] The aqueous vehicle may also include an isotonicity agent to adjust tonicity (osmotic pressure), which may be selected from the group consisting of glycols (e.g., propylene glycol, diethylene glycol, triethylene glycol, etc.), glycerol, dextrose, glycerin, mannitol, potassium chloride, and sodium chloride, and mixtures thereof.

[0491] To adjust the formulation to an acceptable pH (typically a pH in the range of about 5.0 to about 9.0, more preferably about 5.5 to about 8.5, in particular 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 formulation may contain a pH modifier. The pH modifier is typically a mineral acid or metal hydroxide base selected from the group of potassium hydroxide, sodium hydroxide, and hydrochloric acid, and mixtures thereof, and preferably sodium hydroxide and / or hydrochloric acid. These acidic and / or basic pH modifiers are added to adjust the formulation to a target acceptable pH range. Thus, depending on the formulation - it may not be necessary to use both an acid and a base, and the addition of either an acid or a base may be sufficient to bring the mixture into the desired pH range.

[0492] The aqueous vehicle may also contain a buffering agent 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 salts thereof, including disodium tetraborate), citrate buffers (such as citric acid or salts thereof, including sodium citrate), and ε-aminocaproic acid, and mixtures thereof.

[0493] According to a further aspect of the present disclosure there is provided a pharmaceutical composition comprising a compound of the present disclosure as hereinbefore defined, or a pharma- ceutically acceptable salt, hydrate or solvate thereof, in association with a pharma- ceutically acceptable diluent or carrier.

[0494] The compositions of the present disclosure may be in a form suitable for oral use (e.g., tablets, lozenges, hard or soft capsules, aqueous or oily suspensions, emulsions, dispersible powders or granules, syrups or elixirs), topical use (e.g., creams, ointments, gels, or aqueous or oily solutions or suspensions), administration by inhalation (e.g., finely divided powders or liquid aerosols), administration by insufflation (e.g., finely divided powders), or parenteral administration (e.g., sterile aqueous or oily solutions for intravenous, subcutaneous, intramuscular, intraperitoneal or intramuscular administration, or suppositories for rectal administration).

[0495] The compositions of the present disclosure can be obtained by the usual procedures using the usual 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.

[0496] 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 alleviate the symptoms associated with an inflammasome-associated condition referred to herein.

[0497] 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 alleviate the symptoms associated with an inflammasome-associated condition referred to herein.

[0498] The magnitude of a therapeutic or prophylactic dose of a compound of formula (I) or (II) will naturally vary depending on 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.

[0499] 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 comprising administering to the subject a conjugate of the present disclosure.

[0500] 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 comprising administering to the subject a conjugate of the present disclosure.

[0501] In some aspects, the present disclosure provides a method of delivering a nucleic acid agent to a subject comprising administering to the subject a conjugate of the present disclosure.

[0502] 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.

[0503] 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.

[0504] In some aspects, the present disclosure provides a conjugate of the present disclosure for modulating (eg, reducing or eliminating) expression of a target gene in a cell or tissue of a subject.

[0505] In some aspects, the present disclosure provides a conjugate of the present disclosure for delivery of a nucleic acid agent to a subject.

[0506] In some aspects, the present disclosure provides a conjugate of the present disclosure for treating or preventing a disease in a subject in need thereof.

[0507] 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.

[0508] 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.

[0509] In some aspects, the present disclosure provides for the use of a conjugate of the present disclosure in the manufacture of a medicament for delivering a nucleic acid agent to a subject.

[0510] In some aspects, the present disclosure provides for the use of a conjugate of the present disclosure in the manufacture of a medicament for treating or preventing a disease in a subject in need thereof.

[0511] In some embodiments, the subject is a cell.

[0512] In some embodiments, the subject is a tissue.

[0513] In some embodiments, the subject is a human.

[0514] 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 ... The mutations are selected from the group consisting of GF gene, EGFR gene, cyclin A gene, cyclin E gene, WNT-1 gene, β-catenin gene, c-MET gene, PKC gene, NFKB gene, STAT3 gene, survivin gene, Her2 / Neu gene, topoisomerase I gene, topoisomerase IIα 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, and any combination thereof.

[0515] In some embodiments, the disease is characterized by unwanted expression of the target gene.

[0516] In some embodiments, the administration results in a reduction or elimination of expression of the target gene in the subject.

[0517] In some embodiments, the disease is a viral infection, such as an HCV, HBV, HPV, HSV, or HIV infection.

[0518] In some embodiments, the disease is cancer.

[0519] In some embodiments, the cancer is selected from the group consisting of cholangiocarcinoma, bladder cancer, transitional cell carcinoma, urothelial carcinoma, brain tumor, glioma, astrocytoma, breast cancer, metaplastic carcinoma, cervical cancer, squamous cell carcinoma of the cervix, 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 (CMML ... ), liver cancer, hepatocarcinoma, hepatoma, 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, gastric carcinoma, gastrointestinal stromal tumor (GIST), uterine cancer, or uterine sarcoma.

[0520] In some embodiments, the cancer is liver cancer, hepatocarcinoma, hepatoma, hepatocellular carcinoma, cholangiocarcinoma, or hepatoblastoma.

[0521] 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.

[0522] definition Unless otherwise stated, the following terms used in the specification and claims have the following meanings set forth below.

[0523] Without wishing to be limited by this reference, it is understood that while various options for the variables are described herein, the present disclosure is intended to encompass operative embodiments having combinations of the options, and the present disclosure may be construed to exclude inoperable embodiments caused by specific combinations of the options.

[0524] 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 (linear) saturated aliphatic hydrocarbon groups, as well as 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 moieties having 1 to 6 carbon atoms, such as, but not limited to, 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.

[0525] 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 can include, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylato, 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, thiocarboxylato, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or aromatic or heteroaromatic moieties.

[0526] The term "alkenyl" as used herein 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.

[0527] 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 carbons of the hydrocarbon backbone. Such substituents can include, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylato, 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, thiocarboxylato, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, heterocyclyl, alkylaryl, or aromatic or heteroaromatic moieties.

[0528] The term "alkynyl" as used herein includes unsaturated aliphatic groups similar 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, nonyl, 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.

[0529] 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 carbons of the hydrocarbon backbone. Such substituents can include, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylato, 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, thiocarboxylato, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or aromatic or heteroaromatic moieties.

[0530] Other optionally substituted moieties (such as optionally substituted cycloalkyl, heterocycloalkyl, aryl, or heteroaryl) refer to unsubstituted moieties and moieties having one or more 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.

[0531] 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 polycyclic cycloalkyl, only one ring in the cycloalkyl must be non-aromatic.

[0532] The term "heterocycloalkyl," as used herein, refers to a saturated or partially unsaturated 3-8 membered monocyclic, 7-12 membered bicyclic (fused, bridged, or spiro) or 11-14 membered tricyclic ring system (fused, bridged, or spiro) having one or more heteroatoms (e.g., O, N, S, P, or Se), for example, 1 or 1-2 or 1-3 or 1-4 or 1-5 or 1-6 heteroatoms, or for example, 1, 2, 3, 4, 5, or 6 heteroatoms independently selected from the group consisting of nitrogen, oxygen, and sulfur, unless otherwise specified.Examples of heterocycloalkyl groups are 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-oxazepanyl, 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[cyclohexane-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]pyridinyl midinyl, 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 ring in the heterocycloalkyl needs to be non-aromatic (eg, 4,5,6,7-tetrahydrobenzo[c]isoxazolyl).

[0533] The term "aryl" as used herein includes groups having aromatic character, including "conjugated" or polycyclic ring systems with one or more aromatic rings, but without heteroatoms within 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.

[0534] The term "heteroaryl" as used herein 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 substituents as defined). The nitrogen and sulfur heteroatoms may be optionally oxidized (i.e., N→O and S(O)). p, where 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, which are not aromatic to form a polycyclic ring system (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.

[0535] Furthermore, the terms "aryl" and "heteroaryl" include polycyclic aryl and heteroaryl groups, e.g., tricyclic, bicyclic, such as, for example, naphthalene, benzoxazole, benzodioxazole, benzothiazole, benzimidazole, benzothiophene, quinoline, isoquinoline, naphthyridine, indole, benzofuran, purine, benzofuran, deazapurine, indolizine, and the like.

[0536] The cycloalkyl ring, heterocycloalkyl ring, aryl ring, or heteroaryl ring may contain at one or more ring positions (e.g., ring-forming carbon or heteroatom such as N) a substituent as described above, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkoxy, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylato, alkylcarbonyl, alkylaminocarbonyl, aralkylaminocarbonyl, alkenylaminocarbonyl, alkylcarbonyl, arylcarbonyl, aralkylcarbonyl, alkenylcarbonyl, alkoxycarbonyl, a The aryl and heteroaryl groups may be substituted with aminocarbonyl, alkylthiocarbonyl, phosphate, phosphonato, phosphinato, amino (including alkylamino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylato, 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 to form polycyclic ring systems (e.g., tetralin, methylenedioxyphenyl, such as benzo[d][1,3]dioxol-5-yl, etc.).

[0537] The term "substituted" as used herein means that any one or more hydrogen atoms on the specified atom are replaced by a selection from the specified group, provided that the normal valence of the specified atom is not exceeded and that 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 do not occur in aromatic moieties. As used herein, a ring double bond 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, as well as formulation into an effective therapeutic agent.

[0538] When a bridge is shown where a bond to a substituent connects two atoms of a ring, such substituent may be bonded to any atom of that ring. When a substituent is recited without specifying the atom through which such substituent is bonded to the remainder of the compound of a given formula, such substituent may be bonded through any atom of such formula. Combinations of substituents and / or variables are permissible only if such combinations result in stable compounds.

[0539] When any variable (e.g., R) occurs more than one time in a 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 to 2 R moieties, that group is optionally substituted with up to 2 R moieties, and R at each occurrence may be independently selected from the definitions of R. Also, combinations of substituents and / or variables are permissible only if such combinations result in stable compounds.

[0540] As used herein, the term "hydroxy" or "hydroxyl" includes groups with -OH or -O-.

[0541] The term "halo" or "halogen" as used herein refers to fluoro, chloro, bromo and iodo.

[0542] The terms "haloalkyl" or "haloalkoxyl" refer to an alkyl or alkoxyl substituted with one or more halogen atoms.

[0543] 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 can include, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylato, 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, thiocarboxylato, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or aromatic or heteroaromatic moieties.

[0544] The term "alkoxy" or "alkoxyl" as used herein includes substituted and unsubstituted alkyl, alkenyl, and alkynyl groups covalently linked 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. These alkoxy groups can be substituted with groups such as alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylato, 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, thiocarboxylato, 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.

[0545] 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," and the like 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.

[0546] It should be understood that the present disclosure provides methods of synthesis of the compounds, scaffolds, and conjugates described herein. The present disclosure also provides detailed methods of synthesis of the various disclosed compounds, scaffolds, and conjugates according to the schemes herein and those shown in the Examples.

[0547] It should be understood that throughout this description, when a composition is described as having, including, or containing specific components, the composition is also intended to consist essentially of, or consist of, the recited components. Similarly, when a method or process is described as having, including, or containing specific processing steps, the process also consists essentially of, or consists of, the recited processing steps. Furthermore, it should be understood that the order of steps in the sequence for performing certain operations is not critical so long as the invention remains operable. Moreover, two or more steps or operations can be performed simultaneously.

[0548] It should be understood that the synthetic processes of the present disclosure can tolerate a wide variety of functional groups and thus can employ a variety of substituted starting materials. These processes generally provide the desired final compounds at or near the end of the overall process, however, in certain cases, further conversion of the compounds to their pharma- ceutical acceptable salts may be desirable.

[0549] It should be 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 utilizing standard synthetic methods and procedures that are either known to those skilled in the art or will be apparent to those skilled in the art in light of the present disclosure. 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 standard textbooks in the field. Examples of such methods and procedures include, but are not limited to, any one or several sources, including classic textbooks, such as Smith, MB, March, J., March's Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 5, pp. 111-115, 2002, which are incorporated herein by reference. th edition, John Wiley & Sons: New York, 2001;Greene, TW, Wuts, PGM, Protective Groups in Organic Synthesis, 3 rd edition, John Wiley & Sons: New York, 1999; R. Larock, Comprehensive Organic Transformations, VCH Publishers (1989); L. Fieser and M. Fieser, Fieser and Fieser's Reagents forganic Synthesis, John Wiley and Sons (1994); and L. Paquette, ed., Encyclopedia of Reagents forganic Synthesis, John Wiley and Sons (1995), are useful and recognized reference texts in organic synthesis known to those skilled in the art.

[0550] Those skilled in the art will note that the order of certain steps, such as the introduction and removal of protecting groups, may be varied during the reaction sequences and synthetic schemes described herein. Those skilled in the art will recognize that certain groups may require protection from the reaction conditions through the use of protecting groups. Protecting groups may also be used to distinguish similar functional groups within a molecule. For a list of protecting groups and how to introduce and remove these groups, see Greene, TW, Wuts, PGM, Protective Groups in Organic Synthesis, 3rd Edition, 1999. rd edition, John Wiley & Sons: New York, 1999.

[0551] Unless otherwise specified, any description of a method of treatment or prevention should be understood to include the use of the present compounds, scaffolds, and conjugates to provide treatment or prevention as described herein. Unless otherwise stated, any description of a method of treatment and prevention should be further understood to include the use of the present compounds, scaffolds, and conjugates for the preparation of 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.

[0552] Unless otherwise stated, any description of a method of treatment should be understood to include the use of the present compounds, scaffolds, and conjugates to provide treatment as described herein. Unless otherwise stated, any description of a method of treatment should be further understood to include the use of the present compounds, scaffolds, and conjugates for the preparation of a medicament for treating or preventing such a condition. Treatment includes the treatment of humans or non-human animals, including rodents and other disease models.

[0553] The term "subject" as used herein is interchangeable with the term "subject in need thereof," both of which refer to a subject having a disease or at increased risk of developing a disease. "Subject" includes mammals. The mammal can 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 can also be a bird or poultry. In some embodiments, the mammal is a human. The subject in need thereof can be a subject who has previously been diagnosed or determined to have a disease or disorder disclosed herein. The subject in need thereof can also be a subject suffering from a disease or disorder disclosed herein. Alternatively, the subject in need thereof can be a subject who has an increased risk of developing such a disease or disorder compared to the population as a whole (i.e., a subject who is predisposed to developing such a disorder compared to the population as a whole). The subject in need thereof may have a refractory or resistant 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 thereof has received and failed all known effective therapies for the disease or disorder disclosed herein. In some embodiments, the subject in need thereof has received at least one prior therapy.

[0554] The term "treating" or "treat" as used herein describes the management and care of a patient with the intent to eradicate the 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" also includes the treatment of cells in vitro or in animal models. It should be understood that reference to "treating" or "treatment" includes the alleviation of established symptoms of the condition. Thus, "treating" or "treatment" of a state, disorder, or condition includes: (1) preventing or delaying the appearance of clinical symptoms of a disease state, disorder, or condition occurring in a human suffering from or predisposed to the disease state, disorder, or condition, but who has not yet experienced or exhibited clinical or preclinical symptoms of the disease state, disorder, or condition; (2) inhibiting the disease state, disorder, or condition, i.e., arresting, relieving, or delaying the onset of the disease or its recurrence (in the case of maintenance therapy), or at least one clinical or preclinical symptom thereof; or (3) ameliorating or attenuating the disease, i.e., inducing regression of the disease state, disorder, or condition, or at least one clinical or preclinical symptom thereof.

[0555] It is to be understood that the compounds, scaffolds, and conjugates of the present disclosure, or their pharma- ceutically acceptable salts, polymorphs, or solvates, can also be or may be used to prevent the associated disease, condition, or disorder, or to identify suitable candidates for such purposes.

[0556] As used herein, the terms "preventing," "prevent" or "preventing against" describe the reduction or elimination of the onset of symptoms or complications of such a disease, condition or disorder.

[0557] It should be understood that the present disclosure also provides pharmaceutical compositions containing any of the compounds, scaffolds, or conjugates described herein in combination with at least one pharma- ceutically acceptable excipient or carrier.

[0558] The term "pharmaceutical composition" as used herein is a formulation that contains the disclosed compound, scaffold, or conjugate in a form suitable for administration to a subject. In some embodiments, the pharmaceutical composition is in bulk dosage form or unit dosage form. The unit dosage form is any of a variety of forms, including, for example, capsules, IV bags, tablets, aerosol inhalers, or single pumps on vials. The amount of active ingredient (e.g., formulations of the disclosed compounds or their salts, hydrates, solvates, or isomers) 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 is sometimes necessary to make routine variations in dosage depending on the age and condition of the patient. The dosage will also vary according to the route of administration. Various routes are contemplated, including oral, pulmonary, rectal, parenteral, transdermal, subcutaneous, intravenous, intramuscular, intraperitoneal, inhalation, buccal, sublingual, intrapleural, intrathecal, intranasal, and the like. 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- ceutical acceptable carrier, and with any preservatives, buffers, or propellants that are required.

[0559] As used herein, the term "pharmaceutical acceptable" refers to those compounds, scaffolds, conjugates, anions, cations, materials, compositions, carriers, and / or dosage forms that are suitable for use in contact with the tissues of human beings and animals without undue toxicity, irritation, allergic response, or other problem or complication, within the scope of sound medical judgment, commensurate with a reasonable benefit / risk ratio.

[0560] The term "pharmaceutical acceptable excipient" as used herein means an excipient that is generally safe, non-toxic, and not biologically or otherwise undesirable, which is useful in the preparation of pharmaceutical compositions, and includes excipients acceptable for veterinary use as well as human pharmaceutical use. As used in the specification and claims, "pharmaceutical acceptable excipient" includes both one or more of such excipients.

[0561] It should be understood that the pharmaceutical composition of the present disclosure is formulated to be compatible with its intended route of administration. Examples of routes of administration include parenteral, e.g., intravenous, intradermal, subcutaneous, oral (e.g., ingested), inhalation, transdermal (topical), and transmucosal administration. Solutions or suspensions used for parenteral, intradermal, or subcutaneous application can contain the following components: a sterile diluent, such as water for injection, saline, fixed oils, polyethylene glycols, glycerin, propylene glycol, or other synthetic solvents; antibacterial agents, such as benzyl alcohol or methylparaben; antioxidants, such as ascorbic acid or sodium bisulfite; chelating agents, such as ethylenediaminetetraacetic acid; buffers, such as acetates, citrates, or phosphates, and substances for adjusting tonicity, such as 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.

[0562] It should be understood that the compounds or pharmaceutical compositions of the present disclosure can be administered to a subject by many well-known methods currently used for chemotherapy. For example, the compounds of the present disclosure can be injected into the bloodstream or body cavity, or taken orally, or applied through the skin by a patch. The dosage selected must be sufficient to constitute an effective treatment, but not so high as to cause unacceptable side effects. The pathology of the disease state (e.g., a disease or disorder disclosed herein) and the health status of the patient must be closely monitored, preferably during and for a reasonable period after treatment.

[0563] The term "therapeutically effective amount" as used herein refers to an amount of a pharmaceutical agent to treat, ameliorate, or prevent a defined disease or condition, or to exhibit a detectable therapeutic or inhibitory effect. This effect can be detected by any assay method known in the art. The exact effective amount for a subject will depend 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.

[0564] The term "therapeutically effective amount" as used herein refers to an amount of a pharmaceutical agent to treat or ameliorate a defined disease or condition, or to exhibit a detectable therapeutic or inhibitory effect. This effect can be detected by any assay method known in the art. The exact effective amount for a subject will depend 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.

[0565] It should be understood that for any compound, the therapeutically effective amount can be estimated initially either in cell culture assays, e.g., 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 for administration in humans. Therapeutic / prophylactic efficacy and toxicity can be assessed, for example, by measuring the ED 50 (the dose therapeutically effective in 50% of the population) and LD 50The LD (the dose lethal to 50% of the population) may be determined by standard pharmacological procedures in cell cultures or experimental animals. The dose ratio between toxic and therapeutic effects is the therapeutic index, and this is known as the LD 50 / ED 50 It can be expressed as a ratio. Pharmaceutical compositions that exhibit large therapeutic indices are preferred. The dosage may vary within this range depending on the dosage form utilized, the sensitivity of the patient, and the route of administration.

[0566] Dosage and administration are adjusted to provide sufficient levels of the active agent(s) or to maintain the desired effect. Factors that may be considered include the severity of the disease condition, the subject's general health, the subject's age, weight, and sex, diet, time and frequency of administration, concomitant medication(s), reaction sensitivities, and tolerance / response to therapy. Long-acting pharmaceutical compositions may 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.

[0567] The pharmaceutical composition containing the active compound of the present disclosure may be prepared in a generally known manner, for example, by the usual mixing, dissolving, granulating, dragee-making, wet grinding, emulsifying, encapsulating, encapsulating or lyophilizing process.The pharmaceutical composition may be formulated in a conventional manner using one or more pharma-ceutical acceptable carriers, including excipients and / or auxiliaries that facilitate the processing of the active compound into a preparation that can be used as a medicament.Of course, the suitable formulation depends on the selected route of administration.

[0568] 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, for example, a solvent or dispersion medium containing water, ethanol, polyol (e.g., glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof. 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. Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In many cases, it will be preferable to include an isotonic agent in the composition, such as sugars, polyalcohols such as mannitol and sorbitol, and sodium chloride. Prolonged absorption of the injectable composition can be brought about by including in the composition a substance that delays absorption, such as aluminum monostearate and gelatin.

[0569] Sterile injectable solutions can be prepared by mixing the required amount of the active compound in a suitable solvent with one or a combination of the above-listed ingredients, followed by sterilization by filtration as required.Generally, dispersions are prepared by mixing the active compound into a sterile vehicle that contains a basic dispersion medium and other required ingredients from those listed above.In the case of sterile powders for preparing sterile injectable solutions, the preparation method is vacuum drying or freeze-drying, which produces a powder of the active ingredient plus any additional desired ingredients from their previously sterilized-filtered solutions.

[0570] Oral compositions generally contain an inert diluent or an edible medicament acceptable carrier. These can be enclosed in gelatin capsules or compressed into tablets. For the purpose of oral therapeutic administration, the active compound can be mixed with an excipient and used in the form of tablets, troches, or capsules. Oral compositions can also be prepared using a liquid carrier for use as a mouthwash, where the compound in the liquid carrier is applied orally, expectorated and expectorated or swallowed. Pharmaceutically compatible binding agents, and / or adjuvant materials can be included as part of the composition. Tablets, pills, capsules, troches and the like may 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 disintegrant 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, orange flavor, and the like.

[0571] 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.

[0572] 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, a drop, or a swab. In some embodiments, the compound is delivered as a powder. In some embodiments, the compound is included in a kit that further comprises a nasal applicator.

[0573] Systemic administration can also 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 intranasal sprays or suppositories. For transdermal administration, the active compound is formulated into ointments, salves, gels, or creams, as generally known in the art.

[0574] The active compound can be prepared with a pharma- ceutically acceptable carrier that will protect the compound against rapid elimination from the body, such as a controlled release formulation, including implants and microencapsulated delivery systems. Biodegradable polymers, 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. These materials are also commercially available from Alza Corporation and Nova Pharmaceuticals, Inc. Liposomal suspensions (containing liposomes targeted to infected cells with monoclonal antibodies against viral antigens) can also be used as pharma-ceutically acceptable carriers. These can be prepared by methods known to those skilled in the art, such as those disclosed in U.S. Pat. No. 4,522,811.

[0575] For ease of administration and uniform dose, it is particularly advantageous to formulate oral or parenteral compositions in unit dosage form.The unit dosage form used herein refers to a physically separate unit suitable as a uniform dose for the subject to be treated; each unit contains a predetermined amount of active compound calculated to produce desired therapeutic effect in association with necessary pharmaceutical carrier.The details of the unit dosage form of the present disclosure are specified and directly depend on the unique characteristics of active compound and the specific therapeutic effect to be achieved.

[0576] In therapeutic applications, the dosage of pharmaceutical compositions used in accordance with the present disclosure will vary depending on the substance, the age, weight, and clinical condition of the recipient patient, and the experience and judgment of the clinician or practitioner administering the therapy, among other factors that affect the selected dosage. In general, the dosage should be sufficient to cause a delay, and preferably regression, of symptoms of the disease or disorder disclosed herein, and also preferably complete regression of the disease or disorder. Doses can range from about 0.01 mg / kg / day to about 5000 mg / kg / day. An effective amount of a pharmaceutical agent is one that provides an objectively determinable improvement noted by a clinician or other qualified observer. Improved survival and growth refers to 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.

[0577] It should be understood that the pharmaceutical compositions can be included in a container, pack, or dispenser together with instructions for administration.

[0578] With respect to compounds, scaffolds, or conjugates of the present disclosure that are capable of further forming salts, it should be understood that all of these forms are also intended to be within the scope of the claimed disclosure.

[0579] The term "pharmaceutical acceptable salts" as used herein refers to derivatives of the compounds of the present disclosure, which are modified by making their acid salts or base salts. Examples of pharmaceutical acceptable salts include, but are not limited to, mineral organic acid salts of basic residues such as amines, alkali organic salts of acidic residues such as carboxylic acids, and the like. These pharmaceutical acceptable salts include the usual non-toxic salts or quaternary ammonium salts formed from the parent compound, for example, from non-toxic inorganic acids, 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, hexylresorcylic acid, hydrabamic acid, hydrobromic acid, hydrochloric acid, hydroiodic acid, hydroxymaleic acid, hydroxynaphthoic acid, isethionic acid, lactic acid, lactobionic acid, These include, but are not limited to, those derived from inorganic and organic acids selected from laurylsulfonic acid, maleic acid, malic acid, mandelic acid, methanesulfonic acid, naphsylic acid, nitric acid, oxalic acid, pamoic acid, pantothenic acid, phenylacetic acid, phosphoric acid, polygalacturonic acid, propionic acid, salicylic acid, stearic acid, subacetic acid, succinic acid, sulfamic acid, sulfanilic acid, sulfuric acid, tannic acid, tartaric acid, toluenesulfonic acid, and commonly occurring amino acids such as glycine, alanine, phenylalanine, arginine, and the like.

[0580] 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.

[0581] Other illustrative 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, tertiary butylacetic acid, muconic acid, and the like. The present disclosure also encompasses salts formed when an acidic proton present in the parent compound is either replaced by a metal ion, e.g., an alkali metal ion, an alkaline earth ion, or an aluminum ion; or is coordinated with an organic base, 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.

[0582] It should be understood that all references to pharma- ceutically acceptable salts include the solvent addition forms (solvates) or crystal forms (polymorphs) of such salts, as defined herein.

[0583] The compounds, or their pharma- ceutically acceptable salts, can be administered orally, nasally, transdermally, pulmonary, inhalation, buccal, sublingual, intraperitoneally, subcutaneously, intramuscularly, intravenously, rectally, intrapleurally, intrathecally and parenterally.In some embodiments, the compounds are administered orally.Those skilled in the art will recognize the advantages of certain administration routes.

[0584] The dosing regimen utilizing the present compounds 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 being treated; the route of administration; the renal and hepatic function of the patient; and the particular compound or salt thereof being utilized. A physician or veterinarian of ordinary skill can readily determine and prescribe the effective amount of drug required to prevent, reverse, or stop the progression of the condition. A physician or veterinarian of ordinary skill can readily determine and prescribe the effective amount of drug required to reverse or stop the progression of the condition.

[0585] Techniques for formulation and administration of the disclosed compounds of this disclosure are described in Remington: The Science and Practice of Pharmacy, 1999. th edition, Mack Publishing Co., Easton, PA (1995). In some embodiments, 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 aqueous organic solutions. The compounds will be present in such pharmaceutical compositions in an amount sufficient to provide the desired dosage within the range described herein.

[0586] 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 various components and methodologies useful in practicing the present disclosure. These examples do not limit the disclosure as claimed. Based on the present disclosure, one skilled in the art can determine and utilize other components and methodologies useful for practicing the present disclosure.

[0587] In the synthetic schemes described herein, the compounds are depicted in one particular configuration for the sake of simplicity. Such a particular configuration should not be construed as limiting the disclosure to one or another isomer, tautomer, positional isomer, or stereoisomer, nor does it 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.

[0588] All publications and patent documents cited in this specification are incorporated herein by reference as if each such publication or document was specifically and individually indicated to be incorporated herein by reference. The citation of any publication or patent document is not intended as an admission that it is prior art, nor does it constitute any admission of the contents or date of the publication or patent document. The invention has now been described by written description, and those skilled in the art will recognize that the 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 following claims.

[0589] Illustrative Embodiments Illustrative Embodiment 1 Compounds of formula (I) or (II): [ka] or a pharma- ceutically acceptable salt thereof, wherein: W is H, C1-C6 alkyl optionally substituted with one or more halogens, or an amino substituent; X is H, halogen, or -OR X and; R X is H, C1-C6 alkyl, or -(C1-C6 alkyl)-(C6-C 10 aryl), where C1-C6 alkyl or -(C1-C6 alkyl)-(C6-C10 aryl) may be one or more R Xa optionally replaced by; Each R Xa is independently halogen, C1-C6 alkyl, or -O-(C1-C6 alkyl), where C1-C6 alkyl or -O-(C1-C6 alkyl) is optionally substituted with one or more halogens; 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; Alternatively, Y and Z together in formula (I) represent -Si(R L )2-O-Si(R L )2-, where each R L is independently H or C1-C6 alkyl; 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; and Each R 5 is independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens; and Each R 6 is independently H, halogen, or C1-C6 alkyl optionally substituted with one or more halogens.

[0590] Illustrative Embodiment 2 or a pharma- ceutically acceptable salt thereof, wherein the scaffold is: (i) a ligand; and (ii) comprising a linker unit, wherein the linker unit is: [ka] where the variable R 1 , R 2 , R 3 , R 4 , R 5 , R6 , X, Y, and Z are as described in illustrative embodiment 1, and # indicates connection to a ligand.

[0591] Illustrative Embodiment 3 or a pharma- ceutically acceptable salt thereof, wherein the scaffold is: (i) one or more nucleic acid agents; and (ii) comprises one or more linker units, where each linker unit is independently: [ka] where the variable R 1 , R 2 , R 3 , R 4 , R 5 , R 6 A scaffold, wherein, W, X, Y, and Z are as described in illustrative embodiment 1, and ## indicates connection to a nucleic acid material.

[0592] Illustrative embodiment 4. or a pharma- ceutically acceptable salt thereof, wherein the conjugate is: (i) one or more nucleic acid agents; (ii) one or more ligands; and (iii) comprising one or more linker units, wherein each linker unit is independently: [ka] where the variable R 1 , R 2 , R 3 , R 4 , R 5 , R 6 A conjugate, wherein X, Y, and Z are as described in exemplary embodiment 1, # indicates connection to the ligand, and ## indicates connection to the nucleic acid agent.

[0593] Illustrative Embodiment 5 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein W is H.

[0594] Illustrative embodiment 6. The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein W is C1-C6 alkyl optionally substituted with one or more halogens.

[0595] Illustrative Embodiment 7 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein W is an amino substituent.

[0596] Illustrative embodiment 8. The compound, scaffold or conjugate of any one of the preceding illustrative 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).

[0597] Illustrative embodiment 9. The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein W is optionally substituted acyl.

[0598] Illustrative Embodiment 10 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein W is trifluoroacetyl (TFA).

[0599] Illustrative embodiment 11. The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein X is H.

[0600] Illustrative embodiment 12. The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein X is a halogen.

[0601] Illustrative Embodiment 13 . X is -OR XThe compound, scaffold or conjugate according to any one of the preceding illustrative embodiments,

[0602] Illustrative Embodiment 14 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein X is -OH.

[0603] Illustrative Embodiment 15 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein X is -O-(C1-C6 alkyl).

[0604] Illustrative Embodiment 16 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein X is -O-(C1-C6 alkyl)-O-(C1-C6 alkyl).

[0605] Illustrative Embodiment 17 X is one or more R Xa -O-(C1-C6 alkyl)-(C6-C 10 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0606] Illustrative Embodiment 18 X is -O-(C1-C6 alkyl)-(C6-C 10 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0607] Illustrative Embodiment 19 .R X The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0608] Illustrative Embodiment 20 .R X The compound, scaffold or conjugate of any one of the preceding illustrative embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens, or -O-(C1-C6 alkyl) optionally substituted with one or more halogens.

[0609] Illustrative Embodiment 21 .R X is optionally substituted with one or more halogens, -(C1-C6 alkyl)-(C6-C 10 The compound, scaffold or conjugate of any one of the preceding illustrative embodiments, wherein -C1-C6 alkyl or -O-(C1-C6 alkyl) is optionally substituted with one or more halogens.

[0610] Illustrative Embodiment 22 .R X -(C1-C6 alkyl)-(C6-C 10 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Illustrative Embodiment 23 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Y is H.

[0611] Illustrative Embodiment 24 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Y is C1-C6 alkyl optionally substituted with one or more halogens.

[0612] Illustrative Embodiment 25 . 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 illustrative embodiments.

[0613] Illustrative Embodiment 26 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Y is a hydroxy protecting group.

[0614] Illustrative Embodiment 27 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Y is silyl.

[0615] Illustrative Embodiment 28 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Y is triphenylmethyl (Tr) or 4,4'-dimethoxytrityl (DMTr).

[0616] Illustrative Embodiment 29 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Y is optionally substituted acyl or benzyl.

[0617] Illustrative embodiment 30 At least one R Y The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0618] Illustrative Embodiment 31 At least one R Y The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0619] Illustrative Embodiment 32 At least one R Y is H, and at least one R Y The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0620] Illustrative Embodiment 33 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein when X is -OH then Y is not H or a hydroxy protecting group.

[0621] Illustrative Embodiment 34 When X is -OH, Y is 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, or -P(=S)(SR Y )2. The compound, scaffold or conjugate of any one of the preceding illustrative embodiments.

[0622] Illustrative Embodiment 35 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein when Y is H or a hydroxy protecting group, then X is not -OH.

[0623] Illustrative Embodiment 36 When Y is H or a hydroxy protecting group, X is H, a halogen, or -OR X and R X is C1-C6 alkyl or -(C1-C6 alkyl)-(C6-C 10 aryl), wherein C1-C6 alkyl or -(C1-C6 alkyl)-(C6-C 10 aryl) may be one or more R Xa The compound, scaffold or conjugate of any one of the preceding illustrative embodiments, optionally substituted by:

[0624] Illustrative Embodiment 37 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Z is H.

[0625] Illustrative Embodiment 38 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Z is C1-C6 alkyl optionally substituted with one or more halogens.

[0626] Illustrative Embodiment 39 . 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 illustrative embodiments.

[0627] Illustrative Embodiment 40 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Z is a hydroxy protecting group.

[0628] Illustrative embodiment 41 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Z is silyl.

[0629] Illustrative Embodiment 42 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Z is triphenylmethyl (Tr) or 4,4'-dimethoxytrityl (DMTr).

[0630] Illustrative Embodiment 43 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein Z is substituted acyl or benzyl.

[0631] Illustrative Embodiment 44 At least one R Z The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0632] Illustrative Embodiment 45 At least one R Z The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0633] Illustrative embodiment 46. At least one R Z is H, and at least one R Z The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0634] Illustrative Embodiment 47 In formula (I), Y and Z together represent -Si(R L )2-O-Si(R L )2-,

[0635] Illustrative Embodiment 48 At least one R L The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0636] Illustrative Embodiment 49 .Each R L is independently C1-C6 alkyl.

[0637] Illustrative embodiment 50 .R1 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0638] Illustrative Embodiment 51 .R 1 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is a halogen.

[0639] Illustrative Embodiment 52 .R 1 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens.

[0640] Illustrative Embodiment 53 .R 2 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0641] Illustrative Embodiment 54 .R 2 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is a halogen.

[0642] Illustrative Embodiment 55 .R 2 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens.

[0643] Illustrative Embodiment 56 .R 3 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0644] Illustrative Embodiment 57 .R 3 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is a halogen.

[0645] Illustrative Embodiment 58 .R 3The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens.

[0646] Illustrative Embodiment 59 .R 4 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0647] Illustrative embodiment 60 .R 4 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is a halogen.

[0648] Illustrative embodiment 61 .R 4 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens.

[0649] Illustrative embodiment 62 .R 5 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0650] Illustrative embodiment 63 .R 5 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is a halogen.

[0651] Illustrative Embodiment 64 .R 5 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens.

[0652] Illustrative Embodiment 65 .R 6 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein

[0653] Illustrative Embodiment 66 .R 6The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is a halogen.

[0654] Illustrative Embodiment 67 .R 6 The compound, scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein is C1-C6 alkyl optionally substituted with one or more halogens.

[0655] Illustrative Embodiment 68 .R 1 , R 2 , R 3 , R 4 , R 5 , and R 6 The compound, scaffold or conjugate of any one of the preceding illustrative embodiments, wherein each of

[0656] Illustrative Embodiment 69 The compound has the formula (I'-1), (I'-2), (II'-1), or (II'-2): [ka] or a pharma- ceutically acceptable salt thereof.

[0657] Illustrative embodiment 70 The compound has the formula (IA) or (II-A): [ka] or a pharma- ceutically acceptable salt thereof.

[0658] Illustrative embodiment 71 The compound has the formula (I-A'-1), (I-A'-2), (II-A'-1), or (II-A'-2): [ka] or a pharma- ceutically acceptable salt thereof.

[0659] Illustrative embodiment 72 The compound has formula (IB) or (II-B): [ka] or a pharma- ceutically acceptable salt thereof.

[0660] Illustrative embodiment 73. The compound has formula (I-B'-1), (I-B'-2), (II-B'-1), or (II-B'-2): [ka] or a pharma- ceutically acceptable salt thereof.

[0661] Illustrative Embodiment 74 Y is a hydroxy protecting group and Z is a hydroxy protecting group; or In the formula (I), (I'-1), (I'-2), (IA), (I-A'-1), (I-A'-2), (IB), (I-B'-1), or (I-B'-2), Y and Z together represent -Si(R L )2-O-Si(R L )2-, where each R L is independently H or C1-C6 alkyl.

[0662] Illustrative embodiment 75 The compound is: [ka] or a pharma- ceutically acceptable salt thereof, wherein: 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 )RY , -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 -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; and Each R Z is independently H, or C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0663] Illustrative Embodiment 76 The compound of any one of the preceding illustrative embodiments, wherein the compound is selected from the compounds set forth in Table L and pharma- ceutically acceptable salts thereof.

[0664] Illustrative embodiment 77 A compound which is an isotopic derivative of any one of the compounds of the preceding illustrative embodiments.

[0665] Illustrative Embodiment 78 . The scaffold is (linker unit) p -((nucleic acid material)-(linker unit)s ) r -(Nucleic acid material) q where: each linker unit is independent of other linker units, and each nucleic acid entity is independent of other nucleic acid entities; 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; and The scaffold of any one of the preceding illustrative embodiments, wherein the scaffold comprises at least one linker unit and at least one nucleic acid agent.

[0666] Illustrative Embodiment 79 . The scaffold is: [ka] or a pharma- ceutically acceptable salt thereof, wherein: 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; 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; and Each R Z is independently H, or C1-C6 alkyl optionally substituted with one or more halogen or cyano.

[0667] Illustrative embodiment 80 . The scaffold of any one of the preceding illustrative embodiments, wherein the scaffold is selected from the scaffolds listed in Table S1.

[0668] Illustrative embodiment 81 . The scaffold: [ka] or a pharma- ceutically acceptable salt thereof, wherein: The scaffold of any one of the preceding illustrative embodiments, wherein W is an amino substituent.

[0669] Illustrative embodiment 82 . The scaffold of any one of the preceding illustrative embodiments, wherein the scaffold is selected from the scaffolds listed in Table S2.

[0670] Illustrative embodiment 83 The conjugate may be a (linker unit-(ligand) 0-1 ) p -((nucleic acid material)-(linker unit-(ligand) 0-1 ) s ) r -(Nucleic acid material) 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; and The conjugate of any one of the preceding illustrative embodiments, wherein the conjugate comprises at least one linker unit, at least one nucleic acid agent, and at least one ligand.

[0671] Illustrative Embodiment 84 The conjugate of any one of the preceding illustrative embodiments, wherein the conjugate is selected from the conjugates listed in Table C.

[0672] Illustrative Embodiment 85 . The scaffold or conjugate of any one of the preceding illustrative embodiments, wherein the linker unit is of formula (I) where W is replaced by a connection to a ligand.

[0673] Illustrative Embodiment 86 . The scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein the linker unit is of formula (I), wherein Y and / or Z is replaced by a connection to a nucleic acid agent.

[0674] Illustrative Embodiment 87 The scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein the ligand comprises a carbohydrate moiety.

[0675] Illustrative Embodiment 88 The scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein the carbohydrate moiety comprises a monosaccharide, a disaccharide, a trisaccharide, or a tetrasaccharide.

[0676] Illustrative Embodiment 89 The scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein the carbohydrate moiety comprises galactose or a derivative thereof.

[0677] Illustrative embodiment 90 The ligand is: [ka] The scaffold or conjugate according to any one of the preceding illustrative embodiments.

[0678] Illustrative embodiment 91 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0679] Illustrative embodiment 92 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0680] Illustrative embodiment 93 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0681] Illustrative embodiment 94 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0682] Illustrative embodiment 95 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0683] Illustrative embodiment 96. The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0684] Illustrative embodiment 97. The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0685] Illustrative embodiment 98 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0686] Illustrative embodiment 99 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0687] Illustrative embodiment 100 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0688] Illustrative embodiment 101. The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0689] Illustrative embodiment 102 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0690] Illustrative embodiment 103 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0691] Illustrative Embodiment 104 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0692] Illustrative Embodiment 105 The ligand is: [ka] The scaffold or conjugate of any one of the preceding illustrative embodiments, comprising:

[0693] Illustrative Embodiment 106 The scaffold or conjugate of any one of the preceding illustrative embodiments, wherein the ligand comprises a lipid.

[0694] Illustrative Embodiment 107 The scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein the ligand comprises a peptide moiety.

[0695] Illustrative Embodiment 108 The scaffold or conjugate of any one of the preceding illustrative embodiments, wherein the ligand comprises an antibody moiety.

[0696] Illustrative Embodiment 109. The scaffold or conjugate of any one of the preceding illustrative embodiments, wherein the nucleic acid material comprises an oligonucleotide.

[0697] Illustrative embodiment 110 . The scaffold or conjugate of any one of the preceding illustrative embodiments, wherein the nucleic acid material comprises one or more phosphate groups, or one or more phosphate group analogs.

[0698] Illustrative embodiment 111 . The scaffold or conjugate according to any one of the preceding illustrative embodiments, wherein the linker unit is connected to the nucleic acid material via a phosphate group, or a phosphate group analogue, in the nucleic acid material.

[0699] Illustrative embodiment 112 . The scaffold or conjugate of any one of the preceding illustrative embodiments, wherein the nucleic acid material comprises RNA.

[0700] Illustrative embodiment 113 . The scaffold or conjugate of any one of the preceding illustrative embodiments, wherein the oligonucleotide is an siRNA, microRNA, anti-microRNA, microRNA mimic, anti-microRNA, antagomir, dsRNA, ssRNA, aptamer, immunostimulatory oligonucleotide, decoy oligonucleotide, splicing modulation oligonucleotide, triplex forming oligonucleotide, G-quadruplex, or antisense oligonucleotide.

[0701] Illustrative embodiment 114 A pharmaceutical composition comprising a compound, scaffold or conjugate according to any one of the preceding illustrative embodiments.

[0702] Illustrative embodiment 115 A method for regulating expression of a target gene in a subject comprising administering to the subject a conjugate according to any one of the preceding illustrative embodiments.

[0703] Illustrative embodiment 116 A method of delivering a nucleic acid material to a subject comprising administering to the subject a conjugate according to any one of the preceding illustrative embodiments.

[0704] Illustrative Embodiment 117 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 according to any one of the preceding illustrative embodiments.

[0705] Illustrative Embodiment 118 . A conjugate according to any one of the preceding illustrative embodiments for modulating expression of a target gene in a subject.

[0706] Illustrative Embodiment 119 . A conjugate according to any one of the preceding illustrative embodiments for delivery of a nucleic acid material to a subject.

[0707] Illustrative embodiment 120 A conjugate according to any one of the preceding illustrative embodiments for treating or preventing a disease in a subject in need thereof.

[0708] Illustrative embodiment 121 . Use of a conjugate according to any one of the preceding illustrative embodiments in the manufacture of a medicament for modulating expression of a target gene in a subject.

[0709] Illustrative embodiment 122 . Use of a conjugate according to any one of the preceding illustrative embodiments in the manufacture of a medicament for delivering a nucleic acid substance to a subject.

[0710] Illustrative embodiment 123 Use of a conjugate according to any one of the preceding illustrative embodiments in the manufacture of a medicament for treating or preventing a disease in a subject in need thereof.

[0711] Illustrative Embodiment 124 The method, conjugate, or use of any one of the preceding illustrative embodiments, wherein the subject is a human. EXAMPLES

[0712] Working Example Example 1. Synthesis of GalNAc Compounds [ka] (6aR,8R,9S,9aR)-8-Amino-2,2,4,4-tetraisopropylhexahydrocyclopenta[f][1,3,5,2,4]-trioxadisiloxin-9-ol (1-2). To a solution of compound 1-1 (10.0 g, 54.5 mmol) in pyridine (100 mL) was added TIPDSiCl2 (18.9 g, 59.9 mmol) at 0 °C, and the mixture was stirred at 25 °C for 16 h. The reaction was quenched with MeOH and concentrated in vacuo. The remaining residue was dissolved in EtOAc (200 mL) and washed with aqueous citric acid (200 mL x 2) and brine (200 mL). The organic layer was dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to provide compound 1-2 (42.0 g, 99.0% yield) as a yellow oil. 1 H NMR: 400 MHz, DMSO-d6, δ 8.15 (s, 2H), 4.75 (d, J = 4.4 Hz, 1H), 4.00-3.96 (m, 1H), 3.85 (d, J = 3.2 Hz, 2H), 3.82-3.67 (m, 1H), 3.19-3.16 (m, 1H), 2.05-1.95 (m, 2H), 1.19-1.15 (m, 1H), 1.03-0.84 (m, 30H).

[0713] Tert-Butyl ((6aR,8R,9S,9aR)-9-hydroxy-2,2,4,4-tetraisopropylhexahydrocyclopenta[f][1,3,5,2,4]trioxadisiloxin-8-yl)carbamate (1-3) To a solution of compound 1-2 (21.0 g, 53.9 mol) in dioxane (210 mL) was added Boc2O (17.6 g, 80.8 mmol) and aqueous NaHCO3 (11.3 g, 135 mmol). The mixture was stirred at 20 °C for 16 h. The reaction mixture was quenched with aqueous NH4Cl (400 mL) and extracted with EtOAc (400 mL x 2). The organic layers were washed with brine (400 mL), dried over Na2SO4, filtered and concentrated under vacuum. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 20 / 1 to 1 / 1) to give compound 1-3 (37.3 g, 70.7% yield) as a colorless oil. 1 H NMR: 400 MHz, DMSO-d 6,δ 6.96 (d, J = 6.8 Hz, 1H), 4.22 (d, J = 4.8 Hz, 1H), 3.92-3.91 (m, 1H), 3.82-3.78 (m, 1H), 3.67-3.57 (m, 2H), 3.34(s, 1H), 1.99-1.96 (m, 1H), 1.38 (s, 9H), 1.17-0.89 (m, 29H).

[0714] Tert-Butyl ((6aR,8R,9S,9aR)-2,2,4,4-tetraisopropyl-9-methoxyhexahydro-cyclopenta[f][1,3,5,2,4]trioxadisilosin-8-yl)carbamate (1-4) To a solution of compound 1-3 (12.4 g, 25.3 mmol) in MeI (124 mL), Ag2O (29.3 g, 126 mmol) was added at 25° C., and the mixture was stirred at 50° C. for 72 h. The reaction mixture was then filtered and concentrated in vacuum. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate=20 / 1 to 1 / 1) to give compound 1-4 (15.2 g, yield 39.7%) as a yellow oil. 1 H NMR: 400 MHz, CDCl3, δ 4.36 (s, 1H), 3.98-3.94 (m, 2H), 3.92 (s, 1H), 3.72-3.68 (m, 2H), 3.54 (s, 3H), 3.41 (s, 1H), 2.22-2.13 (m, 2H), 1.46 (d, J = 5.2 Hz, 9H), 1.08-0.91 (m, 29H).

[0715] (1R,2S,3R,5R)-3-Amino-5-(hydroxymethyl)-2-methoxycyclopentan-1-ol hydrochloride (1-5) To a solution of compound 1-4 (13.2 g, 26.2 mmol) in MeOH (10 ml) was added HCl / MeOH (4 M, 264 mL, 1.06 mol). The mixture was stirred at 25° C. for 2 h and concentrated in vacuo to give compound 1-5 (13.6 g, crude) as a yellow oil, which was used in the next step without further purification.

[0716] (2S,3S,4S,5S,6S)-5-acetamido-2-(acetoxymethyl)-6-((5-(((1R,2S,3R,4R)-3-hydroxy-4-(hydroxymethyl)-2-methoxycyclopentyl)amino)-5-oxopentyl)oxy)tetrahydro-2H-pyran-3,4-diyl diacetate (1-6)To a solution of compound 1-5 (5.18 g, 32.1 mmol) and GalNAc-NHS ester (17.5 g, 32.1 mmol) in DMF (52 mL) was added DIPEA (12.5 g, 96.4 mmol) to reach pH 8. The mixture was stirred at 25 °C for 1 h, quenched with aqueous NaHCO3 (200 mL), and extracted with EtOAc (100 mL x 2). The organic layers were washed with brine (100 mL), dried over 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 1-6 (14.3 g, 75.4% yield) as a yellow oil. 1 H NMR: 400 MHz, DMSO-d 6, δ 9.11 (s, 1H), 7.85 (d, J = 9.2 Hz, 1H), 7.76 (d, J = 8.0 Hz, 1H), 5.21 (d, J = 3.2 Hz, 1H), 4.98-4.94 (m, 1H), 4.56 (t, J = 2.6 Hz, 1H), 4.48 (d, J = 8.8 Hz, 1H), 4.31 (d, J = 5.6 Hz, 1H), 4.02-4.01 (m, 4H), 3.75-3.74 (m, 3H), 3.30 (s, 3H), 3.28-3.17 (m, 3H),2.10 (s, 3H), 2.04-1.98 (m, 6H), 1.89 (s, 3H), 1.77 (s, 3H), 1.47-1.45 (m, 4H), 1.30-1.27 (m, 8H), 1.20-0.97(m, 1H).

[0717] (2S,3S,4S,5S,6S)-5-acetamido-2-(acetoxymethyl)-6-((5-(((1R,2S,3R,4R)-4-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-3-hydroxy-2-methoxycyclopentyl)amino)-5-oxopentyl)oxy)tetrahydro-2H-pyran-3,4-diyl diacetate (1-7)To a solution of compound 1-6 (8.00 g, 13.55 mmol) in pyridine (80 mL) was added DMTrCl (5.05 g, 14.9 mmol) and the mixture was stirred at 25 °C for 2 h. The reaction was quenched with aqueous citric acid (100 mL) and extracted with EtOAc (100 mL x 2). The organic layer was washed with brine (100 mL), dried over Na2SO4, filtered and concentrated under vacuum. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 2 / 1 to 0 / 1) to afford compound 1-7 (8.40 g, yield 40.0%) as a yellow oil. 1 H NMR: 400 MHz, DMSO-d 6, δ 7.82 (d, J = 9.2 Hz, 1H), 7.75 (d, J = 8.0 Hz, 1H), 7.38-7.21 (m, 9H), 6.88 (d, J = 8.4 Hz, 4H), 5.21 (d, J = 3.2 Hz, 1H), 4.98-4.94 (m, 1H), 4.47 (d, J = 8.8 Hz, 1H), 4.40 (d, J = 6.4 Hz, 1H), 4.05-4.00 (m, 5H), 3.88 (s, 1H), 3.73-3.71 (m, 8H), 3.33 (s, 3H), 3.31-2.85 (m, 3H), 2.12-1.99 (m, 11H), 1.89 (s, 3H), 1.76 (s, 3H), 1.46-1.45 (m, 4H), 1.03-1.00 (s, 1H).

[0718] (2S,3S,4S,5S,6S)-5-acetamido-2-(acetoxymethyl)-6-((5-(((1R,2S,3R,4R)-4-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-3-(((2-cyanoethoxy)(diisopropylamino)phosphanyl)oxy)-2-methoxycyclopentyl)amino)-5-oxopentyl)oxy)tetrahydro-2H-pyran-3,4-diyl diacetate (1-8)To a solution of compound 1-7 (7.40 g, 8.29 mmol) in DCM (74 mL), DCI (1.47 g, 12.4 mmol) and 2-cyanoethyl-N,N,N',N'-tetraisopropylphosphoro-diamidite (5.00 g, 16.6 mmol) were added at 25° C. The reaction was stirred at 25° C. for 1 h. The mixture was then poured into aqueous NaHCO3 (100 mL) and extracted with DCM (100 mL×3). The organic layer was washed with brine (100 mL), dried over sodium sulfate, filtered, and concentrated in vacuo. The mixture was purified by column chromatography (SiO2, petroleum ether:ethyl acetate=3 / 1 to 0 / 1, 0.1% TEA) to give GalNAc amidite compound 1-8 (6.90 g, 68.0% yield) as a white solid. 1 H NMR: 400 MHz, CD3CN, δ 7.44-7.22 (m, 9H), 6.88-6.85 (m, 4H), 6.48-6.46 (m, 2H), 5.28 (d, J = 3.6 Hz, 1H), 5.01-4.97 (m, 1H), 4.50 (d, J = 8.4 Hz, 1H), 4.15-4.05 (m, 6H), 3.92 (s, 9H), 3.65-3.45 (m, 4H), 3.36 (t, J = 4.8 Hz, 4H), 3.19-2.95 (m, 2H), 2.64-2.30 (m, 3H), 2.17 (s, 2H), 2.05-1.98 (m, 2H), 1.94 (s, 3H), 1.91 (s, 3H), 1.81 (s, 3H), 1.51-1.49 (m, 4H), 1.16-1.12 (m, 9H), 1.04 (d, J = 6.8 Hz, 4H).

[0719] Example 2. mRNA knockdown activity of illustrative siRNA duplexes conjugated with GalNAc G3 to target gene 1 Gene silencing activity was studied using the exemplary siRNA duplexes listed in Table 1. These siRNA duplexes were conjugated with either GalNAc L96 or GalNAc G3 for hepatic delivery to target gene 1. As shown in Figure 1, GalNAc G3 provided comparable delivery efficiency and KD activity as GalNAc L96.

[0720] CD-1 female mice were subcutaneously administered 0.5 mg / kg of GalNAc-conjugated siRNA duplex. The control group was dosed with phosphate-buffered saline (PBS). Then, on day 4 after treatment, the animals were hydrodynamically injected (HDI) with 10 μg of human gene 1 in pcDNA3.1(+) through the tail vein. The mice were sacrificed on day 1 after treatment. Liver tissue was collected and stored overnight at 4° C. in RNAlater®, and transferred to −80° C. for mRNA analysis after removal of RNAlater. The reduction of target mRNA was measured by qPCR using the 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 1] The lower case 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 ethyl phosphonate modification at the 5'-terminus; L96 and G3 indicate the GalNAc structure shown below: [ka]

[0721] equivalent The details of one or more embodiments of the present disclosure are described in the accompanying description 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 description and claims. In this specification and the appended claims, the singular forms include plural referents unless the context clearly dictates 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.

[0722] 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 to be limited by the claims appended hereto.

Claims

1. A scaffold or a pharmaceutically acceptable salt thereof, wherein the scaffold is: It includes the following (i) and (ii): (i) 【Chemical 1】 a ligand comprising: (ii) 【Chemistry 2】 a linker unit which is During the ceremony, X is H, halogen, or -OR Ｘ and R Ｘ is H, C １ -C ６ alkyl, or -(C １ -C ６ alkyl)-(C ６ -C １０ aryl), where C １ -C ６ Alkyl or -(C １ -C ６ alkyl)-(C ６ -C １０ aryl) is one or more R Ｘａ optionally substituted by; Each R Ｘａ are independently halogen, C １ -C ６ Alkyl, or —O—(C １ -C ６ alkyl), where C １ -C ６ Alkyl or —O—(C １ -C ６ alkyl) is optionally substituted with one or more halogens; 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, 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 alkyl; Alternatively, Y and Z together represent —Si(R Ｌ ) ２ —O—Si(R Ｌ ) ２ -, where each R Ｌ are independently H or C 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; Each R ６ are independently H, halogen, or C optionally substituted with one or more halogens. １ -C ６ is alkyl; and # indicates connection to a ligand, scaffold or a pharmaceutically acceptable salt thereof.

2. A conjugate or a pharmaceutically acceptable salt thereof, wherein the conjugate comprises: (i) one or more nucleic acid agents; (ii) one or more ligands, wherein each ligand is independently 【Chemistry 3】 Contains; and (iii) one or more linker units, wherein each linker unit is independently: 【Chemistry 4】 and During the ceremony, X is H, halogen, or -OR Ｘ and R Ｘ is H, C １ -C ６ alkyl, or -(C １ -C ６ alkyl)-(C ６ -C １０ aryl), where C １ -C ６ Alkyl or -(C １ -C ６ alkyl)-(C ６ -C １０ aryl) is one or more R Ｘａ optionally substituted by; Each R Ｘａ are independently halogen, C １ -C ６ Alkyl, or —O—(C １ -C ６ alkyl), where C １ -C ６ Alkyl or —O—(C １ -C ６ alkyl) is optionally substituted with one or more halogens; 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 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; Each R ６ are independently H, halogen, or C optionally substituted with one or more halogens. １ -C ６ is alkyl; # indicates a connection to a ligand; and ## indicates the connection to the nucleic acid material, conjugate or a pharmaceutically acceptable salt thereof.

3. The scaffold, 【Chemistry 5】 and During the ceremony, 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; Each R Ｙ are independently H or C optionally substituted with one or more halogens or cyanos. １ -C ６ is alkyl; 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; and Each R Ｚ are independently H or C optionally substituted with one or more halogens or cyanos. １ -C ６ is alkyl, 2. The scaffold of claim 1 or a pharmaceutically acceptable salt thereof. 【Request 4】 【Chemical 6-1】 【Chemistry 6-2】 A scaffold selected from: 【Request 5】 【Chemical 7】 The scaffold of claim 4, wherein 【Request 6】 【Chemical 8-1】 【Chemistry 8-2】 【Chemistry 8-3】 【Chemistry 8-4】 【Chemistry 8-5】 【Chemistry 8-6】 【Chemistry 8-7】 【Hua 8-8】 【Chemistry 8-9】 【Chemistry 8-10】 【Chemistry 8-11】 【Chemistry 8-12】 【Chemistry 8-13】 【Chemistry 8-14】 【Chemistry 8-15】 【Chemistry 8-16】 【Chemistry 8-17】 【Chemistry 8-18】 A conjugate comprising one or more of: The conjugate further comprises one or more nucleic acid entities bound to ##; During the ceremony, 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, 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; and Each R Ｚ are independently H or C optionally substituted with one or more halogens or cyanos. １ -C ６ is alkyl, Conjugates. 【Request 7】 【Chemical 9】 7. The conjugate of claim 6, comprising one or more of:

8. The ligand is 【Chemistry 10】 Including, In the formula, the linking moiety is C １ -C １５ an alkylene chain, wherein optionally one or more carbon atoms in the alkylene chain are independently replaced by one or more -C(O)-, -C(O)O-, -OC(O)-, -C(O)NH-, -NHC(O)-, -NHC(O)NH-, -C(S)-, -C(S)O-, -OC(S)-, -C(S)NH-, -NHC(S)-, or -NHC(S)NH-; and Each alkylene chain is １ -C ６ Alkyl, halogen, OH, NH ２ , C １ -C ６ optionally substituted with one or more groups independently selected from alkoxy, CN, and COOH; A scaffold according to claim 1 or a conjugate according to claim 2.

9. The conjugate of any one of claims 2 and 6 to 8, wherein each nucleic acid agent comprises an oligonucleotide.

10. A pharmaceutical composition comprising a compound, scaffold or conjugate according to any one of claims 1 to 7.

11. A conjugate according to any one of claims 2 and 6 to 9 for use in the treatment or prevention of a disease in a subject in need thereof.

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