Dsrna molecule for regulating expression of inhbe

Abstract

Provided are a double-stranded RNA for inhibiting the expression of an INHBE gene, a cell containing same, and a method for treating diseases or symptoms mediated by or associated with INHBE in a subject using the dsRNA or cell.

Description

dsRNA molecules that regulate INHBE expression

[0001] This application claims priority to Chinese Patent Application No. 202410667596.4, filed May 27, 2024, entitled "dsRNA molecule regulating INHBC / INHBE expression"; and Chinese Patent Application No. 202510376459.X, filed March 27, 2025, entitled "dsRNA molecule regulating INHBE expression", the entire contents of which are incorporated herein by reference. Technical Field

[0002] This invention relates to the field of RNA interference. Background Technology

[0003] Inhibin subunit beta E (INHBE) is a protein encoded by the INHBE gene. Its aberrant expression is associated with a range of diseases, including metabolic syndrome, type 2 diabetes, obesity, elevated triglyceride levels, lipid metabolism disorders, hepatitis, fatty liver, hypercholesterolemia, elevated liver enzymes, non-alcoholic steatohepatitis, cardiovascular disease, heart failure, and kidney disease.

[0004] There is a need in the art for compositions and methods for treating diseases related to INHBE.

[0005] One treatment approach is to reduce INHBE expression using small interfering RNA (siRNA) based on RNA interference mechanisms to treat neurodegenerative diseases associated with INHBE. Summary of the Invention

[0006] This invention provides novel double-stranded RNA (dsRNA) for inhibiting INHBE gene expression, cells, and pharmaceutical compositions and kits containing said dsRNA or cells, as well as methods for inhibiting or reducing INHBE gene expression or treating diseases or conditions that benefit from reduced INHBE gene expression using said dsRNA, cells, and pharmaceutical compositions and kits.

[0007] In a first aspect, the present invention provides a double-stranded RNA (dsRNA) for inhibiting INHBE gene expression, said dsRNA comprising a sense strand and an antisense strand forming a double-stranded region, wherein the length of each of the sense strand and the antisense strand is independently 15-30 nucleotides, and said antisense strand comprises a nucleotide sequence of at least 15 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO:243-484. In some embodiments, said sense strand comprises a nucleotide sequence of at least 15 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO:1-242.

[0008] In some embodiments, a hairpin loop is formed between the sense and antisense strands of the dsRNA. In other embodiments, the dsRNA is siRNA.

[0009] In some embodiments, the length of the positive and negative strands is independently 15-27 nucleotides, preferably 18-25 nucleotides, more preferably 19-21 nucleotides. In some embodiments, the length of the positive strand is 15-27 nucleotides, preferably 17-25 nucleotides, more preferably 18-23 nucleotides, more preferably 19-21 nucleotides, and most preferably 19 amino acids. In some embodiments, the length of the negative strand is 15-27 nucleotides, preferably 17-25 nucleotides, more preferably 18-23 nucleotides, more preferably 19-22 nucleotides, and most preferably 21 amino acids.

[0010] In some embodiments, the length of the double-stranded region is 15-25 nucleotide pairs, preferably 16-23 nucleotide pairs, more preferably 18-20 nucleotide pairs, and most preferably 19 nucleotide pairs.

[0011] In some embodiments, one or both of the sense strand and the antisense strand include a 3' overhang and / or a 5' overhang having at least one nucleotide, for example, one or both of the sense strand and the antisense strand include a 3' overhang and / or a 5' overhang having at least two nucleotides. In some embodiments, the antisense strand has a 3' overhang and / or a 5' overhang having at least one nucleotide, preferably the antisense strand includes a 3' overhang and / or a 5' overhang having two nucleotides. In some specific embodiments, the dsRNA has two nucleotide overhangs at the 3' end of the antisense strand and a blunt end at the 5' end of the antisense strand.

[0012] In some embodiments, the antisense strand comprises a nucleotide sequence of at least 16 consecutive nucleotides, at least 17 consecutive nucleotides, at least 18 consecutive nucleotides, at least 19 consecutive nucleotides, or at least 20 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO:243-484. In a preferred embodiment, the antisense strand comprises a nucleotide sequence shown in any one of SEQ ID NO:243-484.

[0013] In some embodiments, the positive strand comprises a nucleotide sequence of at least 16 consecutive nucleotides, at least 17 consecutive nucleotides, or at least 18 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO:1-242. In a preferred embodiment, the positive strand comprises a nucleotide sequence shown in any one of SEQ ID NO:1-242.

[0014] In some embodiments, the antisense strand comprises a nucleotide sequence of at least 15 consecutive nucleotides, at least 16 consecutive nucleotides, at least 17 consecutive nucleotides, at least 18 consecutive nucleotides, at least 19 consecutive nucleotides, or at least 20 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO: 273, 288, 310, 412, 479, 480, 481, 482, 483, and 484.

[0015] In some embodiments, the positive chain comprises a nucleotide sequence of at least 15 consecutive nucleotides, at least 16 consecutive nucleotides, at least 17 consecutive nucleotides, or at least 18 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO: 37, 52, 74, 176, 237, 238, 239, 240, 241, and 242.

[0016] In some embodiments, the dsRNA comprises any pair of paired sense and antisense sequences as shown in Table 3 of the specification.

[0017] In some embodiments, in the dsRNA of the present invention for inhibiting INHBE expression in cells,

[0018] (1) The sense strand contains the nucleotide sequence shown in SEQ ID NO:37, and the antisense strand contains the nucleotide sequence shown in SEQ ID NO:273;

[0019] (2) The sense strand contains the nucleotide sequence shown in SEQ ID NO:52, and the antisense strand contains the nucleotide sequence shown in SEQ ID NO:288;

[0020] (3) The sense strand contains the nucleotide sequence shown in SEQ ID NO:74, and the antisense strand contains the nucleotide sequence shown in SEQ ID NO:310;

[0021] (4) The sense strand contains the nucleotide sequence shown in SEQ ID NO:176, and the antisense strand contains the nucleotide sequence shown in SEQ ID NO:412;

[0022] (5) The sense strand contains the nucleotide sequence shown in SEQ ID NO:237, and the antisense strand contains the nucleotide sequence shown in SEQ ID NO:479;

[0023] (6) The sense strand contains the nucleotide sequence shown in SEQ ID NO:238, and the antisense strand contains the nucleotide sequence shown in SEQ ID NO:480;

[0024] (7) The sense strand contains the nucleotide sequence shown in SEQ ID NO:239, and the antisense strand contains the nucleotide sequence shown in SEQ ID NO:481;

[0025] (8) The sense strand contains the nucleotide sequence shown in SEQ ID NO:240, and the antisense strand contains the nucleotide sequence shown in SEQ ID NO:482;

[0026] (9) The sense strand contains the nucleotide sequence shown in SEQ ID NO:241, and the antisense strand contains the nucleotide sequence shown in SEQ ID NO:483; or

[0027] (10) The sense strand contains the nucleotide sequence shown in SEQ ID NO:242, and the antisense strand contains the nucleotide sequence shown in SEQ ID NO:484.

[0028] In some embodiments, substantially all nucleotides of the sense strand and / or substantially all nucleotides of the antisense strand are modified nucleotides.

[0029] In some embodiments, the sense strand and the antisense strand each independently comprise one or more modified nucleotides selected from the group consisting of: 2'-O-alkyl-modified nucleotides (e.g., 2'-O-methyl-modified nucleotides), 2'-methoxyethyl-modified nucleotides, 2'-fluoro-modified nucleotides, 2'-deoxy-modified nucleotides, inosine ribonucleotides, baseless nucleotides, reverse baseless deoxyribonucleotides, nucleotides containing a thiophosphate group, vinylphosphonate-modified nucleotides, locked nucleotides, unlocked nucleotides, 2'-amino-modified nucleotides, 2'-C-alkyl-modified nucleotides, 2'-O-allyl-modified nucleotides, morpholino-modified nucleotides, etc. nucleotides, aminophosphates, nucleotides containing non-natural bases, terminal nucleotides linked to cholesterol derivatives or dodecanoic acid didecylamide groups, deoxyribonucleotides, 3'-terminal deoxythymidine (dT) nucleotides, conformation-restricted nucleotides, restricted ethyl nucleotides, 2'-hydroxy modified nucleotides, nucleotides containing methylphosphonic acid groups, nucleotides containing 5'-phosphate, nucleotides containing 5'-phosphate mimics, diol modified nucleotides (GNA), 2-O-(N-methylacetamide) modified nucleotides, 2'-methoxyethoxyethoxy modified nucleotides, 2'-methoxyethoxy modified nucleotides, and SCP modified nucleotides.

[0030] In some embodiments, the sense strand and the antisense strand each independently comprise one or more nucleotide modifications selected from the group consisting of: 2'-O-methyl modified nucleotides, 2'-fluoro modified nucleotides and nucleotides containing a thiophosphate group, 2'-deoxy modified nucleotides, 2'-methoxyethoxyethoxy modified nucleotides, 2'-methoxyethoxy modified nucleotides, and SCP modified nucleotides.

[0031] In some embodiments, the sense strand and / or the antisense strand comprises at least two 2'-fluorinated nucleotides. In some embodiments, the sense strand and / or the antisense strand comprises at least eight 2'-O-methylated nucleotides. In some embodiments, the 3' and / or 5' ends of the sense strand and / or the antisense strand comprise 1-5 phosphate thioester nucleotide bonds, preferably 2-4 phosphate thioester nucleotide bonds.

[0032] In some embodiments, the sense strand of the dsRNA has a length of 19 nucleotides and has:

[0033] (i) (counting from the 5′ end) nucleotides with 2′-O-methyl modifications at positions 1 to 6 and 10 to 19, and nucleotides with 2′-fluoro modifications at positions 7 to 9; and / or

[0034] (ii) (Counting from the 5' end) Phosphophosphate nucleotide bonds located between nucleotide positions 1 and 2, between nucleotide positions 2 and 3, and between nucleotide positions 18 and 19.

[0035] In some embodiments, the antisense strand of the dsRNA has a length of 21 nucleotides and has

[0036] (i) (counting from the 5′ end) nucleotides with 2'-O-methyl modifications at nucleotide positions 1, 3 to 5, 7 to 13, 15, 17 to 21, and nucleotides with 2'-fluoro modifications at nucleotide positions 2, 6, 14, and 16; and / or

[0037] (ii) (Counting from the 5' end) Phosphophosphate nucleotide bonds located between nucleotide positions 1 and 2, nucleotide positions 2 and 3, nucleotide positions 19 and 20, and nucleotide positions 20 and 21.

[0038] In some embodiments, the antisense strand has a length of 21 nucleotides and has

[0039] (i) (counting from the 5' end) SCP-modified nucleotides located at position 1;

[0040] (ii) (counting from the 5' end) nucleotides with 2'-deoxy modification at positions 2, 5, 7 and 12;

[0041] (iii) (Counting from the 5' end) nucleotides with 2'-fluorine modification at position 14;

[0042] (iv) (counting from the 5' end) nucleotides with 2'-O-methyl modifications at positions 3, 4, 6, 8 to 11, 13 and 15 to 21;

[0043] (v) (counting from the 5' end) Phosphophosphate nucleotide bonds located between nucleotide positions 1 and 2, nucleotide positions 2 and 3, nucleotide positions 19 and 20, and nucleotide positions 20 and 21.

[0044] In some embodiments, the antisense strand has a length of 21 nucleotides and has

[0045] (i) (counting from the 5' end) SCP-modified nucleotides located at position 1;

[0046] (ii) (counting from the 5' end) nucleotides with 2'-fluorinated modification at positions 2, 5, 7, 12 and 14;

[0047] (iii) (Counting from the 5' end) nucleotides with 2'-O-methyl modifications at positions 3, 4, 6, 8 to 11, 13 and 15 to 21;

[0048] (iv) (Counting from the 5' end) Phosphophosphate nucleotide bonds located between nucleotide positions 1 and 2, nucleotide positions 2 and 3, nucleotide positions 19 and 20, and nucleotide positions 20 and 21.

[0049] In some embodiments, the antisense strand has a length of 21 nucleotides and has

[0050] (i) (counting from the 5' end) SCP-modified nucleotides located at position 1;

[0051] (ii) (counting from the 5' end) nucleotides with 2'-fluorinated modification at positions 2, 7, 12, 14 and 16;

[0052] (iii) (counting from the 5' end) nucleotides with 2'-O-methyl modification at positions 3 to 6, 8 to 11, 13, 15 and 17 to 21;

[0053] (iv) (Counting from the 5' end) Phosphophosphate nucleotide bonds located between nucleotide positions 1 and 2, nucleotide positions 2 and 3, nucleotide positions 19 and 20, and nucleotide positions 20 and 21.

[0054] In some embodiments, the antisense strand has a length of 21 nucleotides and has

[0055] (i) (counting from the 5' end) SCP-modified nucleotides located at position 1;

[0056] (ii) (counting from the 5' end) nucleotides with 2'-fluorinated modification at positions 2, 5, 12, 14 and 16;

[0057] (iii) (counting from the 5' end) nucleotides with 2'-O-methyl modification at positions 3, 4, 6 to 11, 13, 15 and 17 to 21;

[0058] (iv) (Counting from the 5' end) Phosphophosphate nucleotide bonds located between nucleotide positions 1 and 2, nucleotide positions 2 and 3, nucleotide positions 19 and 20, and nucleotide positions 20 and 21.

[0059] In some embodiments, the antisense strand has a length of 21 nucleotides and has

[0060] (i) (counting from the 5' end) SCP-modified nucleotides located at position 1;

[0061] (ii) (counting from the 5' end) nucleotides with 2'-fluorinated modification at positions 2, 6, 14 and 16;

[0062] (iii) (counting from the 5' end) nucleotides with 2'-O-methyl modifications at positions 3 to 5, 7 to 13, 15 and 17 to 21;

[0063] (iv) (Counting from the 5' end) Phosphophosphate nucleotide bonds located between nucleotide positions 1 and 2, nucleotide positions 2 and 3, nucleotide positions 19 and 20, and nucleotide positions 20 and 21.

[0064] In some implementations, the antisense chain further comprises:

[0065] (i) (counting from the 5' end) GNA located at position 7;

[0066] (ii) (counting from the 5' end) a nucleotide modified with 2'-methoxyethoxyethoxy at position 3; or

[0067] (iii) (Counting from the 5' end) nucleotides modified with 2'-methoxyethoxy at position 3.

[0068] In some embodiments, the antisense strand of the dsRNA comprises a modified nucleotide sequence of any one of the sequences shown in Tables 5A and 5B, and / or the sense strand comprises a modified nucleotide sequence of any one of the sequences shown in Table 4A. In some embodiments, the dsRNA is a pair of modified sense strand sequences and modified antisense strand sequences.

[0069] In some implementations, the dsRNA is further conjugated to a ligand, such as a vector that can deliver the dsRNA to the liver; for example, a ligand that targets ASGPR.

[0070] In some embodiments, the dsRNA is conjugated to a ligand moiety containing N-acetylgalactosamine, preferably via a phosphate ester group or a thiophosphate ester group.

[0071] In some embodiments, preferably the sense strand of the dsRNA is conjugated to the ligand portion, and preferably the 3' end of the sense strand is conjugated to the ligand portion.

[0072] In some implementations, the ligand portion has the following structure:

[0073] in This indicates the position where the dsRNA is linked via a phosphate group or a thiophosphate group.

[0074] In some embodiments, the sense strand comprises a modified nucleotide sequence of any one of the sequences shown in Table 4B of the specification, and preferably the antisense strand comprises a modified nucleotide sequence of any one of the sequences shown in Table 5B of the specification.

[0075] In some preferred embodiments, the dsRNA of the present invention comprises a sense strand and an antisense strand, wherein the dsRNA comprises a sense strand and an antisense strand selected from the following dsRNAs: DR011676, DR012642, DR012668, DR013589, DR013593, DR012657, DR013550, DR014064, DR012645, DR012671, DR011680, DR011614, DR013994, DR014012, DR014 047, DR012648, DR012674, DR013597, DR013598, DR013993, DR014011, DR011839, DR012651, DR014016, DR013980, DR013998, DR012664, DR012680, DR013590, DR013594, DR014294, DR014295, DR014297, DR014272, DR014274 and DR014275.

[0076] In a second aspect, the present invention provides a cell containing the dsRNA described in the first aspect of the present invention.

[0077] In a third aspect, the present invention provides a pharmaceutical composition comprising the dsRNA described in the first aspect of the invention or the cells described in the second aspect of the invention, and optionally a pharmaceutically acceptable carrier or excipient.

[0078] In a fourth aspect, the present invention provides a kit comprising the dsRNA described in the first aspect of the invention, the cells described in the second aspect of the invention, or the pharmaceutical composition described in the third aspect of the invention.

[0079] In a fifth aspect, the present invention provides a method for inhibiting INHBE gene expression in cells, the method comprising contacting the cells with dsRNA as described in the first aspect of the invention or a pharmaceutical composition as described in the third aspect of the invention. In some embodiments, the method is performed in vitro.

[0080] In a sixth aspect, the present invention provides a method for inhibiting the expression of the INHBE gene in cells of a subject, the method comprising administering to the subject dsRNA as described in the first aspect of the present invention, cells as described in the second aspect of the present invention, or a pharmaceutical composition as described in the third aspect of the present invention.

[0081] The present invention also provides a method for treating a disease or condition in a subject that benefits from reduced INHBE gene expression, the method comprising administering to the subject dsRNA as described in the first aspect of the invention, cells as described in the second aspect of the invention, or a pharmaceutical composition as described in the third aspect of the invention.

[0082] The present invention also provides a method for preventing at least one symptom in a subject suffering from a disease or condition that benefits from reduced INHBE gene expression, the method comprising administering to the subject dsRNA as described in the first aspect of the invention, cells as described in the second aspect of the invention, or a pharmaceutical composition as described in the third aspect of the invention.

[0083] The present invention also provides a method for preventing the progression of a disease or condition in a subject that benefits from reduced INHBE gene expression, the method comprising administering to the subject dsRNA as described in the first aspect of the invention, cells as described in the second aspect of the invention, or a pharmaceutical composition as described in the third aspect of the invention.

[0084] In some embodiments, the disease or condition that benefits from reduced INHBE gene expression is an INHBE-related disease. In some preferred embodiments, the INHBE-related disease is selected from the group consisting of: metabolic syndrome, type 2 diabetes, obesity, elevated triglyceride levels, lipid metabolism disorders, hepatitis, fatty liver, hypercholesterolemia, elevated liver enzymes, non-alcoholic steatohepatitis, cardiovascular disease, heart failure, and kidney disease.

[0085] In some embodiments, the dsRNA, cell, or pharmaceutical composition is administered subcutaneously.

[0086] In some implementations, the subject is a human being.

[0087] Invention Details

[0088] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention.

[0089] It should be understood that the scope of protection of the present invention is not limited to the specific embodiments described below; it should also be understood that the terminology used in the embodiments of the present invention is for describing specific embodiments and not for limiting the scope of protection of the present invention.

[0090] In this specification and claims, unless otherwise expressly stated herein, the singular forms “a,” “one,” and “this” include the plural forms.

[0091] When numerical ranges are given in the implementation scheme, it should be understood that, unless otherwise stated in this invention, both endpoints of each numerical range and any value between the two endpoints may be selected. Unless otherwise defined, all technical and scientific terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art. In addition to the specific methods, devices, and materials used in the embodiments, based on the prior art mastery of one of ordinary skill in the art and the description of this invention, any prior art methods, devices, and materials similar to or equivalent to those described, used, and materials in the embodiments of this invention may be used to implement this invention, and all such methods, devices, and materials fall within the protection scope of this invention. The embodiments of this invention are described in more detail below.

[0092] definition

[0093] In this paper, a “double-stranded region” refers to a region containing two antiparallel and complementary or substantially complementary nucleic acid strands.

[0094] As used herein, the term "double-stranded RNA" or "dsRNA" refers to a ribonucleic acid molecule or ribonucleic acid molecule complex containing the double-stranded region as defined above. The two parts forming the double-stranded region can be two distinct parts of a larger RNA molecule, or they can be separate RNA molecules.

[0095] When the two parts are separate RNA molecules, the dsRNA in this article is referred to as small interfering RNA or short interfering RNA, or simply siRNA.

[0096] When two parts are two distinct portions of a larger molecule, i.e., when the 3' end of one part is linked to the 5' end of the other part to form a double-stranded region by one or more uninterrupted nucleotides, the uninterrupted nucleotide used for the linkage is called a "hairpin loop." When the two parts are covalently linked to form a double-stranded region by a method other than a hairpin loop, the linkage structure is called a "linkosome." After such dsRNA is introduced into the cell, it is cleaved into siRNA by an intracellular endonuclease called Dicer.

[0097] The term "siRNA" in this article refers to a class of double-stranded RNA molecules comprising a sense strand and an antisense strand, which can mediate the silencing of target RNAs (e.g., mRNAs, such as transcripts of genes encoding proteins) that are complementary or substantially complementary to the antisense strand. siRNAs are typically double-stranded, consisting of an antisense strand complementary to the target RNA and a sense strand complementary or substantially complementary to that antisense strand. For convenience, such mRNAs are also referred to herein as mRNAs to be silenced. Such genes are also referred to as target genes. Typically, the RNA to be silenced is an endogenous gene or a pathogen gene. Additionally, RNAs other than mRNAs (e.g., tRNAs) and viral RNAs can also be targeted.

[0098] As used herein, the term "antisense strand" refers to a strand in dsRNA (especially siRNA) that contains regions that are fully or substantially complementary to the target sequence.

[0099] As used herein, the term "complementary region" refers to a region on the antisense strand that is perfectly or substantially complementary to the target mRNA sequence. In cases where the complementary region is not perfectly complementary to the target sequence, mismatches can occur within the molecule or at the ends. Typically, the most tolerant mismatches are located in the end regions, for example, within 5, 4, 3, 2, or 1 nucleotides at the 5' and / or 3' ends. The portion of the antisense strand most sensitive to mismatches is called the "seed region." For example, in a siRNA containing a 19-nt strand, the 19th position (from 5' to 3') can tolerate some mismatches.

[0100] When used in this context, the term "complementarity" refers to the ability of a first polynucleotide to hybridize with a second polynucleotide under certain conditions, such as stringent conditions. For example, stringent conditions may include 400 mM NaCl, 40 mM PIPES at pH 6.4, and 1 mM EDTA at 50 or 70°C for 12–16 hours.

[0101] As used herein, in order to satisfy the above requirements regarding their hybridization ability, "complementary" sequences may also include base pairs formed entirely from non-Watson-Crick base pairs and / or from non-natural and modified nucleotides. Such non-Watson-Crick base pairs include, but are not limited to, G:U swing base pairs or Hoogstein base pairs.

[0102] As used herein, a polynucleotide that is “at least partially complementary” or “substantially complementary” to messenger RNA (mRNA) refers to a polynucleotide that is substantially complementary to a continuous portion of the mRNA of interest (e.g., the mRNA encoding apolipoprotein(a)). For example, if the sequence is substantially complementary to an uninterrupted portion of the mRNA encoding apolipoprotein(a), then the polynucleotide is at least partially complementary to the mRNA encoding apolipoprotein(a).

[0103] The terms “complementary,” “fully complementary,” and “substantially complementary” as used herein can be used relative to the base pairing between the sense and antisense strands of dsRNA, particularly siRNA, or between the antisense strand of dsRNA, particularly siRNA, and the target sequence.

[0104] As used herein, the term "sense chain" refers to a chain of siRNA that includes regions substantially complementary to the regions defined herein as antisense chains.

[0105] "Nucleoside" is a compound composed of two substances: a purine or pyrimidine base and ribose or deoxyribose. "Nucleotide" is a compound composed of three substances: a purine or pyrimidine base, ribose or deoxyribose, and phosphate. "Oligonucleotide" refers to nucleic acid molecules (RNA or DNA) with a length of less than 100, 200, 300, or 400 nucleotides.

[0106] A "base" is the basic building block for the synthesis of nucleosides, nucleotides, and nucleic acids. Its constituent elements include nitrogen, hence it is also called a "nitrogenous base." In this article, unless otherwise specified, the capital letters A, U, T, G, and C represent the base composition of nucleotides, namely adenine, uracil, thymine, guanine, and cytosine, respectively.

[0107] As used herein, the term "nucleotide overhang" refers to at least one unpaired nucleotide that protrudes from the double-stranded region of siRNA. A nucleotide overhang exists, for example, when the 3' end of one strand of siRNA extends beyond the 5' end of the other strand, or vice versa. siRNA may contain overhangs having at least one nucleotide, at least two nucleotides, at least three nucleotides, at least four nucleotides, or at least five or more nucleotides. A nucleotide overhang may contain or consist of nucleotides / modified nucleotides (including deoxynucleotides / nucleosides). One or more overhangs may be located on the sense or antisense strand, or any combination thereof. An overhang having one or more nucleotides may be present at the 5' end, 3' end, or both ends of the antisense or sense strand of siRNA.

[0108] "Flat-ended" or "blunt-ended" means that there are no unpaired nucleotides at that end of the double-stranded siRNA, i.e., no nucleotide overhangs. "Flat-ended siRNA" is a double-stranded siRNA that is double-stranded along its entire length, meaning that there are no nucleotide overhangs at either end of the molecule.

[0109] The dsRNA (especially siRNA) of the present invention comprises substantially all modified nucleotides. For example, substantially all nucleotides of the sense strand are modified nucleotides, or substantially all nucleotides of the antisense strand are modified nucleotides, or substantially all nucleotides of both the sense and antisense strands are modified nucleotides. In other embodiments of the invention, all nucleotides of the dsRNA (especially siRNA) of the present invention are modified nucleotides. For example, substantially all nucleotides of the sense strand are modified nucleotides, or substantially all nucleotides of the antisense strand are modified nucleotides, or substantially all nucleotides of both the sense and antisense strands are modified nucleotides. As used herein, “substantially all nucleotides are modified” means that the majority, but not all, nucleotides of the dsRNA (especially siRNA) of the present invention are modified, and may include no more than 5, 4, 3, 2, or 1 unmodified nucleotides.

[0110] In this document, "modified nucleotides" include, but are not limited to, 2'-O-alkyl-modified nucleotides (e.g., 2'-O-methyl-modified nucleotides, 2'-methoxyethyl-modified nucleotides), 2'-fluoro-modified nucleotides, 2'-deoxy-modified nucleotides, inosine ribonucleotides, non-base nucleotides, reverse non-base deoxyribonucleotides, nucleotides containing a thiophosphate group, thiophosphate ester nucleotide bonds, vinylphosphonate-modified nucleotides, locked nucleotides, unlocked nucleotides, 2'-amino-modified nucleotides, 2'-C-alkyl-modified nucleotides, and 2'-O - Allyl-modified nucleotides, morpholinonucleotides, aminophosphates, nucleotides containing non-natural bases, terminal nucleotides linked to cholesterol derivatives or dodecanoic acid di ...

[0111] For example, "2'-fluorinated nucleotides" refer to nucleotides in which the 2'-hydroxyl group of the ribosome is replaced by fluorine. "2'-O-methyl nucleotides" refer to nucleotides in which the 2'-hydroxyl group of the ribosome is replaced by a methoxy group.

[0112] "A nucleotide containing a thiophosphate group" refers to a nucleotide in which one or more oxygen atoms on the phosphate group are replaced by sulfur atoms. "Modification of internucleotide thiophosphate bonding" refers to a modification in which two adjacent nucleotides are linked by a thiophosphate group.

[0113] In some embodiments, the sense strand of the dsRNA (especially siRNA) of this disclosure has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 5 (counting from the 5' end) and / or has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 5 (counting from the 3' end), and / or the antisense strand of the dsRNA (especially siRNA) of this disclosure has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 5 (counting from the 5' end) and / or has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 5 (counting from the 3' end).

[0114] In some embodiments, the sense strand of the dsRNA (especially siRNA) of this disclosure has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 4 (counting from the 5' end) and / or has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 4 (counting from the 3' end), and / or the antisense strand of the dsRNA (especially siRNA) of this disclosure has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 4 (counting from the 5' end) and / or has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 4 (counting from the 3' end).

[0115] In some embodiments, the sense strand of the dsRNA (especially siRNA) of this disclosure has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 3 (counting from the 5' end) and / or has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 3 (counting from the 3' end), and / or the antisense strand of the dsRNA (especially siRNA) of this disclosure has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 3 (counting from the 5' end) and / or has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 3 (counting from the 3' end).

[0116] In some embodiments, the sense strand of the dsRNA (especially siRNA) of this disclosure has a modification of one or two phosphate thioester nucleotides bonded at positions 1 and 2 (counting from the 5' end) and / or a modification of one or two phosphate thioester nucleotides bonded at positions 1 and 2 (counting from the 3' end), and / or the antisense strand of the dsRNA (especially siRNA) of this disclosure has a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 2 (counting from the 5' end) and / or a modification of one or two phosphate thioester nucleotides bonded at positions 1 to 2 (counting from the 3' end).

[0117] As used herein, the term “inhibition” is used interchangeably with “reduction,” “silence,” “downregulation,” and other similar terms, and includes any level of inhibition.

[0118] The phrase "inhibit the expression of the INHBE gene" refers to the inhibition of the expression of any INHBE gene, as well as variants or mutants of the INHBE gene. Therefore, the INHBE gene can be a wild-type INHBE gene, a mutant INHBE gene, or a transgenic INHBE gene in the case of genetically manipulated cells, cell groups, or organisms.

[0119] "Inhibition of INHBE gene expression" includes inhibition of the INHBE gene at any level, such as at least partial inhibition of INHBE gene expression. INHBE gene expression can be assessed based on the level or changes in the level of any variable associated with INHBE gene expression, such as the mRNA level encoding apolipoprotein(a) or the protein level of apolipoprotein(a). This level can be assessed in a single cell or in a group of cells, including, for example, samples derived from a subject.

[0120] Inhibition can be assessed by a decrease in the absolute or relative level of one or more variables associated with INHBE gene expression compared to a control level. The control level can be any type of control level utilized in the art, such as baseline levels before administration or levels determined from similar untreated or controlled (e.g., buffer-only control or inert agent control) subjects, cells, or samples.

[0121] As used herein, the terms “treatment,” “management,” etc., refer to the administration of a drug or the performance of a procedure to achieve an effect. These effects may be preventative in terms of complete or partial prevention of a disease or its symptoms, and / or therapeutic in terms of partial or complete cure of the disease and / or its symptoms. As used herein, “treatment” may include treating a disease or condition in mammals, particularly humans, and includes: (a) preventing the occurrence of the disease or its symptoms in subjects susceptible to the disease but not yet diagnosed with it (e.g., including diseases that may be related to or caused by the primary disease); (b) suppressing the disease, i.e., halting its development; and (c) alleviating the disease, i.e., causing its remission. Treatment may refer to any indication of success in treating, improving, or preventing cancer, including any objective or subjective parameter, such as elimination; relief; reduction of symptoms or making the disease condition more tolerable for the patient; slowing the rate of deterioration or decline; or weakening the endpoint of deterioration. Treatment or improvement of symptoms is based on one or more objective or subjective parameters; including the results of a physician’s examination. Therefore, the term "treatment" includes the application of the dsRNA, cellular, or pharmaceutical compositions disclosed in this invention to prevent or delay, alleviate, stop, or inhibit the development of disease-related symptoms or conditions. The term "therapeutic effect" refers to the reduction, elimination, or prevention of disease, disease symptoms, or disease side effects in a subject.

[0122] The term "effective amount" as used in this invention refers to an amount sufficient to treat a disease when administered to a subject for the purpose of treating that disease.

[0123] As used herein, the term “subject” refers to any mammalian subject for diagnosis, treatment, or therapy. For therapeutic purposes, “mammal” means any animal classified as a mammal, including humans, livestock, and laboratory animals, zoo animals, sporting animals, or pet animals, such as dogs, horses, cats, cattle, sheep, goats, pigs, mice, rats, rabbits, guinea pigs, monkeys, etc.

[0124] I.dsRNA

[0125] The present invention provides a double-stranded RNA (dsRNA) for inhibiting INHBE gene expression, the dsRNA comprising a sense strand and an antisense strand forming a double-stranded region, wherein the length of the sense strand and the antisense strand are each independently 15-30 nucleotides, and the antisense strand comprises a nucleotide sequence of at least 15 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO:243-484.

[0126] In some implementations, the bistranded regions formed by the sense and antisense strands are completely complementary. In other implementations, the bistranded regions formed by the sense and antisense strands are substantially complementary, and may contain one, two, three, four, or five non-complementary sites.

[0127] In some specific embodiments, the positive chain comprises a nucleotide sequence of at least 15 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO:1-242.

[0128] In some implementations, the lengths of the sense strand and the antisense strand are each independently 15-27 nucleotides, preferably 18-25 nucleotides, and more preferably 19-21 nucleotides.

[0129] In some embodiments, the length of the double-stranded region is 15-25 nucleotide pairs, preferably 16-23 nucleotide pairs, and more preferably 18-20 nucleotide pairs.

[0130] In some embodiments, the dsRNA of the present invention is siRNA. In other embodiments, a hairpin loop is formed between the sense and antisense strands of the dsRNA of the present invention.

[0131] One or both of the positive and negative strands comprise a 3' overhang and / or a 5' overhang having at least one nucleotide. In some embodiments, one or both of the positive and negative strands comprise a 3' overhang and / or a 5' overhang having at least one nucleotide. In some specific embodiments, one or both of the positive and negative strands comprise a 3' overhang and / or a 5' overhang having one nucleotide. In some specific embodiments, one or both of the positive and negative strands comprise a 3' overhang and / or a 5' overhang having two nucleotides. In some specific embodiments, one or both of the positive and negative strands comprise a 3' overhang and / or a 5' overhang having three nucleotides. In some specific embodiments, one or both of the positive and negative strands comprise a 3' overhang and / or a 5' overhang having four nucleotides.

[0132] In some specific embodiments, the antisense strand includes a 3' overhang and / or a 5' overhang of 1 nucleotide. In some embodiments, the antisense strand includes a 3' overhang and / or a 5' overhang of 2 nucleotides. In some embodiments, the antisense strand includes a 3' overhang and / or a 5' overhang of 3 nucleotides. In some embodiments, the antisense strand includes a 3' overhang and / or a 5' overhang of 4 nucleotides. In some preferred embodiments, the antisense strand has a 3' overhang and / or a 5' overhang of at least 2 nucleotides.

[0133] Preferably, the antisense strand comprises a 3' overhang and / or a 5' overhang with 2 nucleotides.

[0134] In some implementations, the justice chain and the antisense chain are of the same length.

[0135] In some implementations, the full length of the justice chain is complementary to the full length of the antisense chain, forming a double chain, i.e., having flat ends.

[0136] In other embodiments, the sense strand and the antisense strand are of the same length, and a portion of the sense strand is complementary to a portion of the antisense strand, i.e., both the sense strand and the antisense strand have a 5' overhang. In some embodiments, the sense strand and the antisense strand are of different lengths. In a preferred embodiment, the 5' end of the antisense strand has an overhang of at least one nucleotide, more preferably two or three nucleotides.

[0137] The dsRNA of the present invention includes dsRNA having a nucleotide overhang at one end (i.e., a reagent having one overhang and one blunt end) or having nucleotide overhangs at both ends. For example, the 5' end of the sense strand of the dsRNA includes an overhang of one or more nucleotides and the 3' end of the sense strand includes an overhang of one or more nucleotides. For example, the 5' end of the antisense strand of the dsRNA includes an overhang of one or more nucleotides and the 3' end of the antisense strand includes an overhang of one or more nucleotides. For example, the 5' end of the sense strand of the dsRNA includes an overhang of one or more nucleotides and the 5' end of the antisense strand includes an overhang of one or more nucleotides. For example, the 3' end of the sense strand of the dsRNA includes an overhang of one or more nucleotides and the 3' end of the antisense strand includes an overhang of one or more nucleotides. For example, the 5' end of the sense strand of the dsRNA includes an overhang of one or more nucleotides and the 3' end of the sense strand includes a blunt end. For example, the 3' end of the sense strand of dsRNA contains a protruding end with one or more nucleotides, and the 5' end of the sense strand contains a blunt end. Similarly, the 5' end of the antisense strand of dsRNA contains a protruding end with one or more nucleotides, and the 3' end of the antisense strand contains a blunt end.

[0138] In some preferred embodiments, the 3' end of the antisense strand of the dsRNA of the present invention includes a protrusion having one or more nucleotides, and the 5' end of the antisense strand includes a blunt end. In some more preferred embodiments, the 3' end of the antisense strand of the dsRNA of the present invention includes a protrusion having one, two, three, or four nucleotides, and the 5' end of the antisense strand includes a blunt end. In some more preferred embodiments, the 3' end of the antisense strand of the dsRNA of the present invention includes a protrusion having two nucleotides, and the 5' end of the antisense strand includes a blunt end.

[0139] In some embodiments, the antisense strand comprises a nucleotide sequence of at least 16 consecutive nucleotides, at least 17 consecutive nucleotides, at least 18 consecutive nucleotides, at least 19 consecutive nucleotides, or at least 20 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO:243-484, preferably the antisense strand comprises a nucleotide sequence shown in any one of SEQ ID NO:243-484.

[0140] In some embodiments, the positive chain comprises a nucleotide sequence of at least 16 consecutive nucleotides, at least 17 consecutive nucleotides, or at least 18 consecutive nucleotides of any of the nucleotide sequences shown in SEQ ID NO:1-242, preferably the positive chain comprises a nucleotide sequence shown in any of SEQ ID NO:1-242.

[0141] In some embodiments, the dsRNA comprises any pair of paired sense and antisense sequences as shown in Table 3.

[0142] II. Nucleotide Modification

[0143] In some embodiments, substantially all nucleotides of the sense strand and substantially all nucleotides of the antisense strand are modified nucleotides. In some embodiments, at least 80% of the nucleotides of the sense strand are modified nucleotides, and / or at least 80%, at least 85%, at least 90%, at least 92%, or at least 95% of the nucleotides of the antisense strand are modified nucleotides. In some embodiments, at least 80% of the nucleotides of the antisense strand are modified nucleotides, and / or at least 80%, at least 85%, at least 90%, at least 92%, or at least 95% of the nucleotides of the sense strand are modified nucleotides.

[0144] In some embodiments, all nucleotides of the sense strand are modified nucleotides and / or all nucleotides of the antisense strand are modified nucleotides.

[0145] The nucleotide modification described in this invention can be a modification on the phosphate group, ribose group and / or base group of the nucleotide.

[0146] In some specific embodiments, the sense strand and the antisense strand each independently comprise one or more nucleotide modifications selected from the group consisting of: 2'-O-alkyl-modified nucleotides (e.g., 2'-O-methyl-modified nucleotides), 2'-methoxyethyl-modified nucleotides, 2'-fluoro-modified nucleotides, 2'-deoxy-modified nucleotides, inosine ribonucleotides, baseless nucleotides, reverse baseless deoxyribonucleotides, nucleotides containing a thiophosphate group, vinylphosphonate-modified nucleotides, locked nucleotides, unlocked nucleotides, 2'-amino-modified nucleotides, 2'-C-alkyl-modified nucleotides, 2'-O-allyl-modified nucleotides, morpholino-modified nucleotides, etc. Nucleotides, aminophosphates, nucleotides containing non-natural bases, terminal nucleotides linked to cholesterol derivatives or dodecanoic acid didecylamide groups, deoxyribonucleotides, 3'-terminal deoxythymidine (dT) nucleotides, conformation-restricted nucleotides, restricted ethyl nucleotides, 2'-hydroxy-modified nucleotides, nucleotides containing methylphosphonic acid groups, nucleotides containing 5'-phosphate, nucleotides containing 5'-phosphate mimics, diol-modified nucleotides (GNA), 2-O-(N-methylacetamide)-modified nucleotides, 2'-methoxyethoxyethoxy-modified nucleotides, 2'-methoxyethoxy-modified nucleotides, and SCP-modified nucleotides.

[0147] In some preferred embodiments, the sense strand and the antisense strand each independently comprise one or more nucleotide modifications selected from the group consisting of: 2'-O-methyl modified nucleotides, 2'-fluoro modified nucleotides, and nucleotides containing a thiophosphate group, 2'-deoxy modified nucleotides, 2'-methoxyethoxyethoxy modified nucleotides, 2'-methoxyethoxy modified nucleotides, and SCP modified nucleotides.

[0148] In some preferred embodiments, the sense strand and / or the antisense strand comprises at least two 2'-fluoromodified nucleotides. In some preferred embodiments, the sense strand and / or the antisense strand comprises at least eight 2'-O-methylmodified nucleotides. In some preferred embodiments, the 3' and / or 5' ends of the sense strand and / or the antisense strand comprise 1-5 phosphate thioester nucleotide bonds, preferably 2-3 phosphate thioester nucleotide bonds.

[0149] In some preferred embodiments, the antisense strand comprises any of the modified nucleotide sequences shown in Tables 5A and 5B, and / or the sense strand comprises any of the modified nucleotide sequences shown in Table 4A. In some embodiments, the sense strand comprising the modified nucleotide sequences shown in Table 4A is paired with the antisense strand comprising the modified nucleotide sequences shown in Tables 5A and 5B.

[0150] In some implementations, the dsRNA contains:

[0151] (i) The justice chain contains CmsAmsGmUmCmGmUfCfCfCmAmGmAmAmUmAmAmCmsAm (SEQ ID No. 1022), and the antisense chain contains (SCP-U)sdGsUmUmdAUmdTCmUmGmGmdGAmCfGmAmCmUmGmsGmsUm (SEQ ID No. 987);

[0152] (ii) The justice chain contains CmsAmsGmUmCmGmUfCfCfCmAmGmAmAmUmAmCmsAm (SEQ ID No. 1023), and the antisense chain contains (SCP-U)sGfsUmUmAfUmUfCmUmGmGmGfAmCfGmAmCmUmGmsGmsUm (SEQ ID No. 988);

[0153] (iii) The justice chain contains CmsAmsGmUmCmGmUfCfCfCmAmGmAmAmUmAmAmCmsAm (SEQ ID No. 1024), and the antisense chain contains (SCP-U)sGfsUmUmAmUmUfCmUmGmGmGfAmCfGmAfCmUmGmsGmsUm (SEQ ID No. 989);

[0154] (iv) The justice chain contains CmsAmsGmUmCmGmUfCfCfCmAmGmAmAmUmAmAmCmsAm (SEQ ID No. 1025), and the antisense chain contains (SCP-U)sGfsUmUmAfUmUmCmUmGmGmGfAmCfGmAfCmUmGmsGmsUm (SEQ ID No. 990);

[0155] (v) The justice chain contains CmsAmsGmUmCmGmUfCfCfCmAmGmAmAmUmAmAmCmsAm (SEQ ID No. 1026), and the antisense chain contains (SCP-U)sGfsUmUmAmUfUmCmUmGmGmGmAmCfGmAfCmUmGmsGmsUm (SEQ ID No. 991);

[0156] (vi) The justice chain contains CmsAmsGmCmUmUmUfGfCfUmAmCmUmGmUmCmAmCmsAm (SEQ ID No. 1027), and the antisense chain contains (SCP-U)sGfsUmGmAfCmAfGmUmAmGmCfAmAfAmGmCmUmGmsAmsUm (SEQ ID No. 992);

[0157] (vii) The justice chain contains CmsAmsGmCmUmUmUfGfCfUmAmCmUmGmUmCmAmCmsAm (SEQ ID No. 1028), and the antisense chain contains (SCP-U)sdGsUmGmdACm(Agn)GmUmAmGmdCAmAfAmGmCmUmGmsAmsUm (SEQ ID No. 993);

[0158] (viii) The justice chain contains CmsAmsGmCmUmUmUfGfCfUmAmCmUmGmUmCmAmCmsAm (SEQ ID No. 1029), and the antisense chain contains (SCP-U)sGfs(Umoeoe)GmAfCmAfGmUmAmGmCfAmAfAmGmCmUmGmsAmsUm (SEQ ID No. 994);

[0159] (ix) The justice chain contains CmsAmsCmCmCmUmUfCfCfUmGmGmCmAmCmUmCmUmsAm (SEQ ID No. 1030), and the antisense chain contains (SCP-U)sAfsGmAmGfUmGfCmCmAmGmGfAmAfGmGmGmUmGmsGmsGm (SEQ ID No. 995);

[0160] (x) The justice chain contains CmsAmsCmCmCmUmUfCfCfUmGmGmCmAmCmUmCmUmsAm (SEQ ID No. 1031), and the antisense chain contains (SCP-U)sAfsGmAmGmUmGfCmCmAmGmGfAmAfGmGfGmUmGmsGmsGm (SEQ ID No. 996);

[0161] (xi) The justice chain contains CmsAmsCmCmCmUmUfCfCfUmGmGmCmAmCmUmCmUmsAm (SEQ ID No. 1032), and the antisense chain contains (SCP-U)sdAsGmAmdGUmdGCmCmAmGmdGAmAfGmGmGmUmGmsGmsGm (SEQ ID No. 997);

[0162] (xii) The justice chain contains UmsCmsCmUmGmGmCfAfCfUmCmUmUmUmGmCmUmUmsAm (SEQ ID No. 1033), and the antisense chain contains UmsdAsAmGmdCAmdAAmGmAmGmdTGmCfCmAmGmGmAmsAmsGm (SEQ ID No. 998);

[0163] (xiii) The justice chain contains UmsCmsCmUmGmGmCfAfCfUmCmUmUmUmGmCmUmsUmsAm (SEQ ID No. 1034), and the antisense chain contains (SCP-U)sAfsAmGmCfAmAfAmGmAmGmUfGmCfCmAmGmGmAmsAmsGm (SEQ ID No. 999);

[0164] (xiv) The justice chain contains UmsCmsCmUmGmGmCfAfCfUmCmUmUmUmGmCmUmsUmsAm (SEQ ID No. 1035), and the antisense chain contains (SCP-U)sAfsAmGmCmAmAfAmGmAmGmUfGmCfCmAfGmGmAmsAmsGm (SEQ ID No. 1000);

[0165] (xv) The justice chain contains UmsCmsCmUmGmGmCfAfCfUmCmUmUmUmGmCmUmUmsAm (SEQ ID No. 1036), and the antisense chain contains (SCP-U)sdAsAmGmdCAmdAAmGmAmGmdTGmdCCmAmGmGmAmsAmsGm (SEQ ID No. 1001);

[0166] (xvi) The justice chain contains UmsGmsAmGmAmAmGfUfCfUmGmGmUmGmUmCmCmUmsAm (SEQ ID No. 1037), and the antisense chain contains (SCP-U)sAfsGmGmAfCmAfCmCmAmGmAfCmUfUmCmUmCmAmsCmsCm (SEQ ID No. 1002);

[0167] (xvii) The justice chain contains UmsGmsAmGmAmAmGfUfCfUmGmGmUmGmUmCmCmUmsAm (SEQ ID No. 1038), and the antisense chain contains (SCP-U)sAfsGmGmAmCmAfCmCmAmGmAfCmUfUmCfUmCmAmsCmsCm (SEQ ID No. 1003);

[0168] (xviii) The justice chain contains UmsGmsAmGmAmAmGfUfCfUmGmGmUmGmUmCmCmUmsAm (SEQ ID No. 1039), and the antisense chain contains (SCP-U)sAfsGmGmAfCmAmCmCmAmGmAfCmUfUmCfUmCmAmsCmsCm (SEQ ID No. 1004);

[0169] (xix) The justice chain contains UmsGmsAmGmAmAmGfUfCfUmGmGmUmGmUmCmCmUmsAm (SEQ ID No. 1040), and the antisense chain contains (SCP-U)sAfsGmGmAmCfAmCmCmAmGmAmCmUfUmCfUmCmAmsCmsCm (SEQ ID No. 1005);

[0170] (xx) The justice chain contains GmsAmsAmGmUmCmUfGfGfUmGmUmCmCmUmGmAmAmsAm (SEQ ID No. 1041), and the antisense chain contains (SCP-U)sUfsUmCmAfGmGfAmCmAmCmCfAmGfAmCmUmUmCmsUmsCm (SEQ ID No. 1006);

[0171] (xxi) The justice chain contains GmsAmsAmGmUmCmUfGfGfUmGmUmCmCmUmGmAmAmsAm (SEQ ID No. 1042), and the antisense chain contains (SCP-U)sUfsUmCmAmGmGfAmCmAmCmCfAmGfAmCfUmUmCmsUmsCm (SEQ ID No. 1007);

[0172] (xxii) The justice chain contains CmsCmsUmAmGmAmGfCfUfUmAmAmGmAmUmCmCmGmsAm (SEQ ID No. 1043), and the antisense chain contains (SCP-U)sdCsGmGmdAUmdCUmUmAmAmdGCmUfCmUmAmGmGmsAmsAm (SEQ ID No. 1008);

[0173] (xxiii) The justice chain contains CmsCmsUmAmGmAmGfCfUfUmAmAmGmAmUmCmCmGmsAm (SEQ ID No. 1044), and the antisense chain contains (SCP-U)sCfsGmGmAfUmCfUmUmAmAmGfCmUfCmUmAmGmGmsAmsAm (SEQ ID No. 1009);

[0174] (xxiv) The justice chain contains CmsCmsUmAmGmAmGfCfUfUmAmAmGmAmUmCmCmGmsAm (SEQ ID No. 1045), and the antisense chain contains (SCP-U)sCfs(Gmoe)GmAfUmCfUmUmAmAmGfCmUfCmUmAmGmGmsAmsAm (SEQ ID No. 1010);

[0175] (xxv) The justice chain contains UmsGmsGmGmCmAmCfUfUfUmCmUmUmGmUmCmUmGmsAm (SEQ ID No. 1046), and the antisense chain contains (SCP-U)sCfsAmGmAfCmAfAmGmAmAmAfGmUfGmCmCmCmAmsUmsUm (SEQ ID No. 1011);

[0176] (xxvi) The justice chain contains UmsGmsGmGmCmAmCfUfUfUmCmUmUmGmUmCmUmGmsAm (SEQ ID No. 1047), and the antisense chain contains (SCP-U)sCfsAmGmAmCmAfAmGmAmAmAfGmUfGmCfCmCmAmsUmsUm (SEQ ID No. 1012);

[0177] (xxvii) The justice chain contains GmsGmsCmAmCmUmUfUfCfUmUmGmUmCmUmGmAmGmsAm (SEQ ID No. 1048), and the antisense chain contains (SCP-U)sCfsUmCmAfGmAfCmAmAmGmAfAmAfGmUmGmCmCmsCmsAm (SEQ ID No. 1013);

[0178] (xxviii) The justice chain contains GmsGmsCmAmCmUmUfUfCfUmUmGmUmCmUmGmAmGmsAm (SEQ ID No. 1049), and the antisense chain contains (SCP-U)sCfsUmCmAmGmAfCmAmAmGmAfAmAfGmUfGmCmCmsCmsAm (SEQ ID No. 1014);

[0179] The justice chain (xxix) contains GmsGmsCmAmCmUmUfUfCfUmUmGmUmCmUmGmAmGmsAm (SEQ ID No. 1050), and the antisense chain contains (SCP-U)sCfsUmCmAfGmAmCmAmAmGmAfAmAfGmUfGmCmCmsCmsAm (SEQ ID No. 1015);

[0180] The justice chain (xxx) contains GmsGmsCmAmCmUmUfUfCfUmUmGmUmCmUmGmAmGmsAm (SEQ ID No. 1051), and the antisense chain contains (SCP-U)sCfsUmCmAmGfAmCmAmAmGmAmAmAfGmUfGmCmCmsCmsAm (SEQ ID No. 1016);

[0181] (xxxi) The justice chain contains CmsAmsGmCmUmUmUfGfCfUmAmCmUmGmUmCmAmsCmsAm (SEQ ID No. 536), and the antisense chain contains UmsGfsUmGfAmCfAmGfUmAfGmCfAmAfAmGfCmUfGmsAmsUm (SEQ ID No. 802);

[0182] (xxxii) The justice chain contains GmsAmsAmGmUmCmUfGfGfUmGmUmCmCmUmGmAmsAmsAm (SEQ ID No. 558), and the antisense chain contains UmsUfsUmCfAmGfGmAfCmAfCmCfAmGfAmCfUmUfCmsUmsCm (SEQ ID No. 824); or

[0183] (xxxii) The justice chain contains GmsGmsCmAmCmUmUfUfCfUmUmGmUmCmUmGmAmsGmsAm (SEQ ID No. 660), and the antisense chain contains UmsCfsUmCfAmGfAmCfAmAfGmAfAmAfGmUfGmCfCmsCmsAm (SEQ ID No. 926).

[0184] III. Ligand Part

[0185] In some embodiments, the dsRNA is further conjugated to a ligand, such as a vector that can deliver the dsRNA to the liver; for example, a ligand targeting ASGPR. In some embodiments, the dsRNA is conjugated to a ligand containing N-acetylgalactosamine, preferably via a phosphate ester group or a thiophosphate ester group. In some embodiments, preferably the sense strand of the dsRNA is conjugated to the ligand, and preferably the 3' end of the sense strand is conjugated to the ligand.

[0186] In some embodiments, the ligand moiety comprises a conjugating group represented by formula (X'):

[0187] in,

[0188] Indicates the location where it is linked to dsRNA;

[0189] Q is independent of H,

[0190] Where L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O))a-;

[0191] L2 is a chemical bond or -CH2CH2C(O)-;

[0192] L3 is a chemical bond, -(NHCH2CH2)b-, -(NHCH2CH2CH2)b-, or -C(O)CH2-;

[0193] L4 is -(OCH2CH2)c-, -(OCH2CH2CH2)c-, -(OCH2CH2CH2CH2)c-, -(OCH2CH2CH2CH2CH2)c- or -NHC(O)-(CH2)d-;

[0194] Where a = 0, 1, 2 or 3;

[0195] b = 1, 2, 3, 4 or 5;

[0196] c = 1, 2, 3, 4 or 5;

[0197] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0198] L represents a chemical bond, -CH2O- or -NHC(O)-;

[0199] L' is a chemical bond, -C(O)NH-, -NHC(O)- or -O(CH2CH2O)e-;

[0200] Where e is 1, 2, 3, 4 or 5;

[0201] T represents a chemical bond, -CH2-, -C(O)-, -M-, -CH2-M-, or -C(O)-M-;

[0202] Where M is

[0203] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0204] Alternatively, R1 and R3 together form a -C1-2 alkylene group, and R2 is H;

[0205] Wherein R is -OR', -CH2OR' or -CH2CH2OR', where R' is H, a hydroxyl protecting group or a solid support, and the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0206] m = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0207] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0208] In some embodiments, the conjugating group is as shown in formula (I'):

[0209] in,

[0210] Indicates the location where it is linked to dsRNA;

[0211] Q is independent of H,

[0212] Where L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O))a-;

[0213] L2 is a chemical bond or -CH2CH2C(O)-;

[0214] L3 is a chemical bond, -(NHCH2CH2)b-, -(NHCH2CH2CH2)b-, or -C(O)CH2-;

[0215] L4 is -(OCH2CH2)c-, -(OCH2CH2CH2)c-, -(OCH2CH2CH2CH2)c-, -(OCH2CH2CH2CH2CH2)c- or -NHC(O)-(CH2)d-;

[0216] Where a = 0, 1, 2 or 3;

[0217] b = 1, 2, 3, 4 or 5;

[0218] c = 1, 2, 3, 4 or 5;

[0219] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0220] L is either -CH2O- or -NHC(O)-;

[0221] L' is a chemical bond, -C(O)NH- or -NHC(O)-;

[0222] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0223] Alternatively, R1 and R3 together form a -C1-2 alkylene group, and R2 is H;

[0224] Wherein R is -OR', -CH2OR' or -CH2CH2OR', where R' is H, a hydroxyl protecting group or a solid support, and the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0225] m = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0226] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0227] In some specific implementation plans, where,

[0228] Q is independently of H or

[0229] Where L1 is -CH2O- or -NHC(O)-(CH2NHC(O))a-;

[0230] L2 is -CH2CH2C(O)-;

[0231] L3 is -(NHCH2CH2)b- or -(NHCH2CH2CH2)b-;

[0232] L4 is -(OCH2CH2)c- or -NHC(O)-(CH2)d-;

[0233] Where a = 0, 1, 2 or 3;

[0234] b = 1, 2, 3, 4 or 5;

[0235] c = 1, 2, 3, 4 or 5;

[0236] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0237] L stands for -CH2O-;

[0238] L' is a chemical bond;

[0239] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0240] Alternatively, R1 and R3 together form a -C1-2 alkylene group, and R2 is H;

[0241] Wherein R is -OR', -CH2OR' or -CH2CH2OR', where R' is H, a hydroxyl protecting group or a solid support, and the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0242] m = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0243] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0244] In some embodiments, the conjugating group is shown as in formula (I'-1), formula (I'-2), or formula (I'-3):

[0245] in,

[0246] Indicates the location where it is linked to dsRNA;

[0247] Q is

[0248] Where L1 is -CH2O- or -NHC(O)-;

[0249] L2 is -CH2CH2C(O)-;

[0250] L3 is -(NHCH2CH2)b- or -(NHCH2CH2CH2)b-;

[0251] L4 is -(OCH2CH2)c- or -NHC(O)-(CH2)d-;

[0252] Where b = 1, 2, 3, 4 or 5;

[0253] c = 1, 2, 3, 4 or 5;

[0254] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0255] L stands for -CH2O-;

[0256] R' is H, a hydroxyl protecting group, or a solid support, wherein the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0257] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0258] In some specific implementation plans, where,

[0259] Q is independent of H,

[0260] Where L1 is -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O))a-;

[0261] L3 is -(NHCH2CH2)b-, -(NHCH2CH2CH2)b-, or -C(O)CH2-;

[0262] L4 is -(OCH2CH2)c- or -NHC(O)-(CH2)d-;

[0263] Where a = 0, 1, 2 or 3;

[0264] b = 1, 2, 3, 4 or 5;

[0265] c = 1, 2, 3, 4 or 5;

[0266] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0267] L is either -CH2O- or -NHC(O)-;

[0268] L' represents a chemical bond or -C(O)NH-;

[0269] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0270] Alternatively, R1 and R3 together form a -C1-2 alkylene group, and R2 is H;

[0271] Wherein R is -OR', -CH2OR' or -CH2CH2OR', where R' is H, a hydroxyl protecting group or a solid support, and the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0272] m = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0273] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0274] In some embodiments, the conjugating group is as shown in formula (II'-1) or formula (II'-2):

[0275] in,

[0276] Indicates the location where it is linked to dsRNA;

[0277] Q independently for

[0278] Where L1 is -CH2O- or -CH2O-CH2CH2O-;

[0279] L3 is -(NHCH2CH2)b-, -(NHCH2CH2CH2)b-, or -C(O)CH2-;

[0280] L4 is -(OCH2CH2)c- or -NHC(O)-(CH2)d-;

[0281] Where b = 1, 2, 3, 4 or 5;

[0282] c = 1, 2, 3, 4 or 5;

[0283] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0284] L is -NHC(O)-;

[0285] L' represents a chemical bond or -C(O)NH-;

[0286] R' is H, a hydroxyl protecting group, or a solid support, wherein the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0287] m = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0288] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0289] In some specific implementation plans, where,

[0290] Q is independent of H,

[0291] Where L1 is -CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O- or -NHC(O)-(CH2NHC(O))a-;

[0292] L2 is a chemical bond;

[0293] L3 is -(NHCH2CH2)b-, -(NHCH2CH2CH2)b-, or -C(O)CH2-;

[0294] L4 is -(OCH2CH2)c- or -NHC(O)-(CH2)d-;

[0295] Where a = 0, 1, 2 or 3;

[0296] b = 1, 2, 3, 4 or 5;

[0297] c = 1, 2, 3, 4 or 5;

[0298] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0299] L is either -CH2O- or -NHC(O)-;

[0300] L' represents a chemical bond or -C(O)NH-;

[0301] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0302] Alternatively, R1 and R3 together form a -C1-2 alkylene group, and R2 is H;

[0303] Wherein R is -OR', -CH2OR' or -CH2CH2OR', where R' is H, a hydroxyl protecting group or a solid support, and the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0304] m = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0305] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0306] In some embodiments, the conjugated group is as shown in formula (II'-2):

[0307] in,

[0308] Indicates the location where it is linked to dsRNA;

[0309] Q independently for

[0310] Where L1 is -CH2- or -C(O)-;

[0311] L3 is -(NHCH2CH2)b-;

[0312] L4 is -(OCH2CH2)c-;

[0313] Where b = 1, 2, 3, 4 or 5;

[0314] c = 1, 2, 3, 4 or 5;

[0315] L is either -CH2O- or -NHC(O)-;

[0316] R' is H, a hydroxyl protecting group, or a solid support, wherein the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0317] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0318] In some specific implementation schemes, where:

[0319] Q is independent of H,

[0320] Where L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O))a-;

[0321] L2 is a chemical bond or -CH2CH2C(O)-;

[0322] L3 is a chemical bond, -(NHCH2CH2)b-, -(NHCH2CH2CH2)b-, or -C(O)CH2-;

[0323] L4 is -(OCH2CH2)c-, -(OCH2CH2CH2)c-, -(OCH2CH2CH2CH2)c-, -(OCH2CH2CH2CH2CH2)c- or -NHC(O)-(CH2)d-;

[0324] Where a = 0, 1, 2 or 3;

[0325] b = 1, 2, 3, 4 or 5;

[0326] c = 1, 2, 3, 4 or 5;

[0327] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0328] L represents a chemical bond, -CH2O- or -NHC(O)-;

[0329] L' is a chemical bond, -C(O)NH-, -NHC(O)- or -O(CH2CH2O)e-;

[0330] Where e is 1, 2, 3, 4 or 5;

[0331] T represents a chemical bond, -CH2-, -M-, -CH2-M-, or -C(O)-M-;

[0332] Where M is

[0333] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0334] Alternatively, R1 and R3 together form a -C1-2 alkylene group, and R2 is H;

[0335] Wherein R is -OR', -CH2OR' or -CH2CH2OR', where R' is H, a hydroxyl protecting group or a solid support, and the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0336] m = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0337] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0338] In some specific implementation plans, where,

[0339] T is -M-, -CH2-M-, or -C(O)-M-, where M is

[0340] In some specific implementation plans, where,

[0341] Q is independently of H or

[0342] Where L1 is -CH2O- or -NHC(O)-(CH2NHC(O))a-;

[0343] L2 is -CH2CH2C(O)-;

[0344] L3 is -(NHCH2CH2)b- or -(NHCH2CH2CH2)b-;

[0345] L4 is -(OCH2CH2)c- or -NHC(O)-(CH2)d-;

[0346] Where a = 0, 1, 2 or 3;

[0347] b = 1, 2, 3, 4 or 5;

[0348] c = 1, 2, 3, 4 or 5;

[0349] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0350] L represents a chemical bond or -CH2O-;

[0351] L' represents a chemical bond or -O(CH2CH2O)e-;

[0352] Where e is 1, 2, 3, 4 or 5;

[0353] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0354] Alternatively, R1 and R3 together form a -C1-2 alkylene group, and R2 is H;

[0355] Wherein R is -OR', -CH2OR' or -CH2CH2OR', where R' is H, a hydroxyl protecting group or a solid support, and the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0356] m = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0357] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0358] Where T is as defined in the above implementation scheme.

[0359] In some embodiments, the conjugated group is as shown in formula (III'-1), formula (III'-2), or formula (III'-3):

[0360] in,

[0361] Q is

[0362] Where L1 is -CH2O- or -NHC(O)-;

[0363] L2 is -CH2CH2C(O)-;

[0364] L3 is -(NHCH2CH2)b- or -(NHCH2CH2CH2)b-;

[0365] L4 is -(OCH2CH2)c- or -NHC(O)-(CH2)d-;

[0366] Where b = 1, 2, 3, 4 or 5;

[0367] c = 1, 2, 3, 4 or 5;

[0368] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0369] L represents a chemical bond or -CH2O-;

[0370] Wherein R' is H, a hydroxyl protecting group, or a solid support, and the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0371] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0372] Where T is as defined in the above implementation scheme.

[0373] In some specific implementation plans, where,

[0374] Q is independent of H,

[0375] Where L1 is -CH2-, -CH2O-, or -C(O)-;

[0376] L2 is a chemical bond;

[0377] L3 is -(NHCH2CH2)b-, -(NHCH2CH2CH2)b-, or -C(O)CH2-;

[0378] L4 is -(OCH2CH2)c- or -NHC(O)-(CH2)d-;

[0379] Where b = 1, 2, 3, 4 or 5;

[0380] c = 1, 2, 3, 4 or 5;

[0381] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0382] L represents a chemical bond or -NHC(O)-;

[0383] L' is a chemical bond;

[0384] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0385] Alternatively, R1 and R3 together form a -C1-2 alkylene group, and R2 is H;

[0386] Wherein R is -OR', -CH2OR' or -CH2CH2OR', where R' is H, a hydroxyl protecting group or a solid support, and the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0387] m = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0388] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0389] Where T is as defined in the above implementation scheme.

[0390] In some embodiments, the conjugating group is as shown in formula (IV-1) or formula (IV-2):

[0391] in,

[0392] Q independently for

[0393] Where L1 is -CH2-, -CH2O-, or -C(O)-;

[0394] L3 is -(NHCH2CH2)b-, -(NHCH2CH2CH2)b-, or -C(O)CH2-;

[0395] L4 is -(OCH2CH2)c- or -NHC(O)-(CH2)d-;

[0396] Where b = 1, 2, 3, 4 or 5;

[0397] c = 1, 2, 3, 4 or 5;

[0398] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0399] L represents a chemical bond or -NHC(O)-;

[0400] L' is a chemical bond;

[0401] Wherein R' is H, a hydroxyl protecting group, or a solid support, and the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0402] m = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0403] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0404] Where T is as defined in the above implementation scheme.

[0405] In some specific implementation schemes, where:

[0406] Q is independent of H,

[0407] Where L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O))a-;

[0408] L2 is a chemical bond or -CH2CH2C(O)-;

[0409] L3 is a chemical bond, -(NHCH2CH2)b-, -(NHCH2CH2CH2)b-, or -C(O)CH2-;

[0410] L4 is -(OCH2CH2)c-, -(OCH2CH2CH2)c-, -(OCH2CH2CH2CH2)c-, -(OCH2CH2CH2CH2CH2)c- or -NHC(O)-(CH2)d-;

[0411] Where a = 0, 1, 2 or 3;

[0412] b = 1, 2, 3, 4 or 5;

[0413] c = 1, 2, 3, 4 or 5;

[0414] d = 1, 2, 3, 4, 5, 6, 7 or 8;

[0415] L represents a chemical bond, -CH2O- or -NHC(O)-;

[0416] L' is -O(CH2CH2O)e-;

[0417] Where e is 1, 2, 3, 4 or 5;

[0418] T represents a chemical bond, -CH2-, -C(O)-, -M-, -CH2-M-, or -C(O)-M-;

[0419] Where M is

[0420] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0421] Alternatively, R1 and R3 together form a -C1-2 alkylene group, and R2 is H;

[0422] Wherein R is -OR', -CH2OR' or -CH2CH2OR', where R' is H, a hydroxyl protecting group or a solid support, and the hydroxyl protecting group is preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytriphenylmethyl;

[0423] m = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0424] n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0425] In some preferred embodiments, the conjugating group is selected from the following Table A:

[0426] Table 1

[0427] In some preferred embodiments, the conjugating group is selected from Table B below:

[0428] Table 2

[0429] In some embodiments, the ligand targets the desialyl glycoprotein receptor (ASGPR). In some embodiments, the ligand targets the desialyl glycoprotein receptor (ASGPR) on hepatocytes.

[0430] In a preferred embodiment, the ligand has the following structure:

[0431] in This indicates the location where the dsRNA is linked via a phosphate group or a thiophosphate group.

[0432] In a preferred embodiment, the ligand has the following structure:

[0433] in This indicates the location where the dsRNA is linked via a phosphate group or a thiophosphate group.

[0434] In a preferred embodiment, the ligand has the following structure:

[0435] in This indicates the position where the ligand is attached to the positive strand of the dsRNA via a phosphate ester group or a thiophosphate ester group. In a preferred embodiment, the ligand has the following structure:

[0436] in This indicates the position where the dsRNA is attached to the positive strand via a phosphate group or a thiophosphate group.

[0437] In some embodiments, the sense strand comprises a modified nucleotide sequence of any one of the sequences shown in Table 4B of the specification, and preferably the antisense strand comprises a modified nucleotide sequence of any one of the sequences shown in Table 5B of the specification.

[0438] In some preferred embodiments, the dsRNA of the present invention comprises a sense strand and an antisense strand, wherein the dsRNA comprises a sense strand and an antisense strand selected from the following dsRNAs: DR011676, DR012642, DR012668, DR013589, DR013593, DR012657, DR013550, DR014064, DR012645, DR012671, DR011680, DR011614, DR013994, DR014012, DR014 047, DR012648, DR012674, DR013597, DR013598, DR013993, DR014011, DR011839, DR012651, DR014016, DR013980, DR013998, DR012664, DR012680, DR013590, DR013594, DR014294, DR014295, DR014297, DR014272, DR014274 and DR014275.

[0439] IV. Inhibition of INHBE gene expression

[0440] The dsRNA of the present invention is capable of inhibiting INHBE gene expression by at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99%.

[0441] Inhibition of INHBE gene expression can be manifested by a reduction in the amount of mRNA expressed by a first cell or cell group (such cells may be present, for example, in a sample derived from a subject), wherein the INHBE gene is transcribed and the cell or these cells have been treated (e.g., by contacting the cell or these cells with the dsRNA of the present invention, or by administering the dsRNA of the present invention to a subject who currently or previously has these cells), such that INHBE gene expression is inhibited compared to a second cell or cell group (one or more control cells) that is substantially the same as the first cell or cell group but has not been so treated.

[0442] In a preferred embodiment, the inhibition is assessed by expressing the level of mRNA in the treated cells as a percentage of the level of mRNA in the control cells using the following formula. In some specific embodiments, 2 is calculated. -△△Ct The value is used to compare the difference between the experimental group and the control group, where △Ct=[(Ct target gene of experimental group - Ct internal reference of experimental group) - (Ct target gene of control group - Ct internal reference of control group)].

[0443] Control cells or cell populations that can be used to assess the inhibition of INHBE gene expression include cells or cell populations that have not been contacted with the dsRNA of the present invention. For example, such control cells or cell populations may be derived from individual subjects (e.g., human or animal subjects) prior to treatment with the dsRNA.

[0444] V. Cells

[0445] The present invention provides a cell containing the dsRNA described herein.

[0446] VI. Pharmaceutical Composition

[0447] The present invention provides pharmaceutical compositions comprising the dsRNA or cells described herein, and optionally pharmaceutically acceptable carriers or excipients.

[0448] As used in this article, "pharmaceutically acceptable" means compounds, materials, compositions, and / or dosage forms that, to the extent of proper medical judgment, are suitable for contact with the tissues of human and animal subjects without excessive toxicity, irritation, allergic reactions, or other problems or complications, and are commensurate with a reasonable benefit / risk ratio.

[0449] In this article, pharmaceutically acceptable carriers refer to drug carriers that facilitate the delivery of dsRNA or its containing cells to the human body and / or promote its absorption or efficacy. Examples include: diluents, excipients such as water; fillers such as starch and sucrose; binders such as cellulose derivatives, alginate, gelatin, and polyvinylpyrrolidone; humectants such as glycerin; disintegrants such as agar, calcium carbonate, and sodium bicarbonate; absorption enhancers such as quaternary ammonium compounds; surfactants such as hexadecyl alcohol; adsorbents such as kaolin and soap clay; and lubricants such as talc, calcium / magnesium stearate, and polyethylene glycol. Other excipients such as flavoring agents and sweeteners may also be added to the composition.

[0450] The pharmaceutical compositions of the present invention may contain a pharmaceutically acceptable diluent or a sustained-release matrix in which the dsRNA or cells of the present invention are embedded.

[0451] The pharmaceutical compositions of the present invention may comprise a drug delivery system for delivering dsRNA. The drug delivery systems of the present invention include, but are not limited to, nanoparticles (e.g., lipid nanoparticles, polymer-based nanoparticles), polymers, PEG, or cationic delivery systems, polylactic acid (PLA) microspheres, polylactic acid-glycolic acid copolymer (PLGA) microspheres, liposomes, micelles, inverse micelles, lipid cochleates, lipid microtubules, cholesterol, PEG lipids PEG-2000-C-DMG, PEG-2000-DMG (Moderna), ALC-0159, or DSPC.

[0452] In some embodiments, the dsRNA in the pharmaceutical compositions of the present invention may be contained in polymers and polymer-based nanoparticles.

[0453] In some specific embodiments, the polymer is based on poly(lactic-co-glycolic acid) copolymer (PLGA). In some specific embodiments, the PLGA-based polymer is modified to contain individual cationic groups.

[0454] In some specific embodiments, the polymer contains amine groups capable of becoming cationic, such as polyethyleneimine (PEI) and poly(L-lysine) (PLL), which can form a complex with dsRNA via electrostatic interactions and deliver dsRNA into cells. In some embodiments, PEG and PLL are chemically modified to improve in vivo potency and tolerability.

[0455] In some embodiments, the siRNA or cells in the pharmaceutical composition of the present invention can be delivered via the cationic polymer poly(β-amino ester) (PBAE).

[0456] VII. Reagent Kit

[0457] This invention provides a kit containing the dsRNA or cells described herein.

[0458] The present invention also provides a kit for using the dsRNA or cells described herein and / or performing the methods of the present invention. Such a kit comprises one or more of the dsRNA or cells described herein, and may further comprise instructions for use. These instructions may describe methods for inhibiting INHBE gene expression in cells by contacting the cells with the dsRNA described herein in an amount that effectively inhibits INHBE gene expression.

[0459] In the case of in vitro contact between the dsRNA described in this invention and cells, the kit of this invention may optionally include a tool for contacting cells with the dsRNA described in this invention (e.g., an injection device) or a tool for measuring the inhibitory effect on the INHBE gene (e.g., a device for measuring the inhibition of INHBE mRNA or protein). Such a device for measuring the inhibition of the INHBE gene may include a device for obtaining a sample from a subject.

[0460] When administering the dsRNA of the present invention or cells that have already been introduced with dsRNA in vitro into the body, the kit of the present invention may optionally include means for administering the dsRNA or cells of the present invention to a subject or means for determining a therapeutically effective amount or a preventatively effective amount.

[0461] VIII. Treatment methods and pharmaceutical uses

[0462] This invention provides a method for inhibiting INHBE gene expression in cells, the method comprising contacting the cells with a dsRNA or pharmaceutical composition as described herein. In some embodiments, the method is performed in vitro. This invention provides the dsRNA or pharmaceutical composition described herein for inhibiting INHBE gene expression in cells.

[0463] This invention provides a method for inhibiting INHBE gene expression in cells of a subject, the method comprising administering to the subject a dsRNA, cell, or pharmaceutical composition as described herein. This invention provides dsRNA, cell, or pharmaceutical compositions of this invention for inhibiting INHBE gene expression in cells of a subject. This invention provides the use of dsRNA, cell, or pharmaceutical compositions of this invention in the preparation of a medicament for inhibiting INHBE gene expression in cells of a subject.

[0464] The present invention also provides a method for treating a disease or condition in which a subject benefits from reduced INHBE gene expression, the method comprising administering to the subject a dsRNA, cell, or pharmaceutical composition as described herein. The present invention also provides a dsRNA, cell, or pharmaceutical composition of the present invention for treating a disease or condition in which a subject benefits from reduced INHBE gene expression. The present invention further provides the use of the dsRNA, cell, or pharmaceutical composition of the present invention in the preparation of a medicament for treating a disease or condition in which a subject benefits from reduced INHBE gene expression.

[0465] The present invention also provides a method for preventing at least one symptom in a subject suffering from a disease or condition that benefits from reduced INHBE gene expression, the method comprising administering to the subject a dsRNA, cell, or pharmaceutical composition as described in the present invention. The present invention also provides dsRNA, cell, or pharmaceutical compositions of the present invention for preventing a disease or condition that benefits from reduced INHBE gene expression. The present invention further provides the use of dsRNA, cell, or pharmaceutical compositions of the present invention in the preparation of a medicament for preventing a disease or condition that benefits from reduced INHBE gene expression.

[0466] The present invention also provides a method for preventing the progression of a disease or condition in which a subject benefits from reduced INHBE gene expression, the method comprising administering to the subject a dsRNA, cell, or pharmaceutical composition as described in the present invention. The present invention also provides dsRNA, cell, or pharmaceutical compositions of the present invention for preventing the progression of a disease or condition in which a subject benefits from reduced INHBE gene expression. The present invention further provides the use of dsRNA, cell, or pharmaceutical compositions of the present invention in the preparation of a medicament for preventing the progression of a disease or condition in which a subject benefits from reduced INHBE gene expression.

[0467] In some embodiments, the disease or condition that benefits from reduced INHBE gene expression is an INHBE-related disease. In some preferred embodiments, the INHBE-related disease is selected from the group consisting of: metabolic syndrome, type 2 diabetes, obesity, elevated triglyceride levels, lipid metabolism disorders, hepatitis, fatty liver, hypercholesterolemia, elevated liver enzymes, non-alcoholic steatohepatitis, cardiovascular disease, heart failure, and kidney disease.

[0468] In some embodiments, the dsRNA, cell, or pharmaceutical composition is administered subcutaneously.

[0469] In some embodiments, the subject is a mammal. In some embodiments, the subject is a primate mammal. In some embodiments, the subject is a human.

[0470] sequence

[0471] The RNA sequence provided by this invention targets the human INHBE gene (or target gene, target mRNA sequence, target sequence). The target INHBE mRNA sequence is, for example, the gene shown in GenBank accession number NM_031479.5.

[0472] Table 3. Nucleotide sequences of the sense and antisense strands of the INHBE-targeting mRNA

[0473] Tables 4A-B and 5A-B show the modified RNA sequences used in this invention.

[0474] The meanings of the abbreviations in this article are as follows:

[0475] The distributions A, U, G, and C represent naturally occurring adenine ribonucleotides, uracil ribonucleotides, guanine ribonucleotides, and cytosine ribonucleotides.

[0476] The 'd' indicates that the nucleotide adjacent to its right is a deoxyribonucleotide. For example, dA, dT, dG, and dC represent adenine deoxyribonucleotide, thymine deoxyribonucleotide, guanine deoxyribonucleotide, and cytosine deoxyribonucleotide, respectively.

[0477] i represents inosine ribonucleotide.

[0478] The 'm' indicates that the nucleotide adjacent to it on the left is a nucleotide modified with 2'-OCH3. For example, Am, Um, Gm, and Cm represent A, U, G, and C modified with 2'-OCH3, respectively.

[0479] The 'f' indicates that the nucleotide adjacent to it on the left is a 2'-fluoro modified nucleotide. For example, Af, Uf, Gf, and Cf represent 2'-fluoro modified A, U, G, and C, respectively.

[0480] “s” or s- indicates that the two adjacent nucleotides and / or delivery carriers are linked by a phosphate thioester.

[0481] Agn is the S-isomer of adenosine-glycol nucleic acid (GNA), which is well known in the art. See, for example, PCT publications WO2019222166A1 and WO2020132227A2. Its structure is as follows: B represents adenine.

[0482] GL6 represents the GalNAc delivery vector with the following structure, where Indicates the position where it is linked to dsRNA via a phosphate ester group or a thiophosphate ester group.

[0483] "SCP" refers to nucleotides with the following modified structures: The Base can be an unmodified base, including adenine, guanine, uracil, and cytosine bases. For example, SCP-U indicates that the Base is a uracil base.

[0484] Umoeoe represents U carrying a 2'-methoxyethoxyethoxy modification, with the following specific structure:

[0485] Gmoe represents G carrying a 2'-methoxyethoxy modification, with the specific structure as follows:

[0486] UNA represents a nucleotide with the following structure known in the art, where B is a base. For example, A(UNA) represents the following structure where B is an adenine base.

[0487] Table 4A. Sensitive strand sequences of modified siRNAs targeting INHBE mRNA

[0488] " / " indicates the sequence of the positive chain in Table 4B that is not connected to a ligand, and its order corresponds to SEQ ID NO.721-SEQ ID NO.750 in Table 4B.

[0489] Table 4B. Sensitive strand (conjugated ligand) of modified siRNAs targeting INHBE mRNA

[0490] Table 5A. Antisense strand sequences of modified siRNAs targeting INHBE mRNA

[0491] Table 5B. Antisense strands of modified siRNAs targeting INHBE mRNA

[0492] Paired siRNAs targeting INHBE mRNA, with positive and negative strands.

[0493] Unless otherwise specified, the siRNA compounds described below consist of the corresponding paired sense and antisense strands in the table above. For example, DR010514 consists of the sense strand DR010514-SS and the antisense strand DR010514-AS.

[0494] Those skilled in the art should understand that the number and position of the inter-nucleotide bonds of thiophosphate in the siRNA compounds of the present invention are not limited to the number and position shown in the sequences shown in Tables 4 and 5. Those skilled in the art can adjust the number and position of the inter-nucleotide bonds of thiophosphate without changing the sequence and other modifications such as methoxy / fluorination, and add or not add a delivery vector, such as a delivery vector containing GalNAc, based on the teachings of Tables 4 and 5. These adjusted sense strands, antisense strands and paired siRNAs are also included within the scope of the present invention.

[0495] The present invention will be further described below with reference to embodiments. It should be understood that the following embodiments are merely illustrative and should not be considered as limiting the scope of the present invention. Example

[0496] Unless otherwise specified, the materials used in the embodiments are from the following sources:

[0497] Hep3B cell line was purchased from Nanjing Kebai, catalog number CBP60197;

[0498] HepG2 cell line was purchased from Nanjing Kebai, catalog number CBP60199.

[0499] Example 1: Preparation of siRNA

[0500] The siRNA of the present invention was prepared using the solid-phase phosphoramide method well known in the art. Specific methods can be found, for example, in PCT publications WO2016081444 and WO2019105419, and are briefly described below.

[0501] 1.1 Synthesis of the Justice Chain (SS Chain)

[0502] The oligonucleotide was synthesized using a solid-phase phosphoramide method, starting with a blank CPG solid support. Nucleoside monomers were sequentially linked from the 3'-5' direction according to the nucleotide arrangement of the positive strand. Each linkage of a nucleoside monomer involved four steps: deprotection, coupling, capping, and oxidation or thiolation. The synthetic conditions for a scale of 5 μmol of oligonucleotides are as follows:

[0503] Commercially available phosphoramids with 2'-F, 2'-O-methyl, and other modifications were used. The nucleoside monomers were provided in a 0.05 mol / L acetonitrile solution. Each reaction step was performed under identical conditions: 25°C. Deprotection was performed three times using a 3% trichloroacetic acid-dichloromethane solution. Coupling was performed twice using a 0.25 mol / L ETT-acetonitrile solution as the activator. Capping was performed twice using a 10% acetic anhydride-acetonitrile and pyridine / N-methylimidazole / acetonitrile mixture (10:14:76, v / v / v). Oxidation was performed twice using a 0.05 mol / L iodine / tetrahydrofuran / pyridine / water mixture (70 / 20 / 10, v / v / v). Thiolysis was performed twice using 0.2 mol / L PADS in an acetonitrile / 3-methylpyridine mixture (1 / 1, v / v).

[0504] 1.2 Synthesis of the antisense chain (AS chain)

[0505] The solid-phase phosphoramide synthesis method utilizes a blank CPG solid-phase support as the starting cycle, and nucleotide monomers are sequentially linked from the 3'-5' direction according to the nucleotide arrangement sequence of the antisense strand. Each linkage of a nucleotide monomer involves four steps: deprotection, coupling, capping, and oxidation or thiolation. The synthesis conditions for 5 μmol oligonucleotides of the antisense strand are the same as those for the sense strand.

[0506] 1.3 Purification and Annealing of Oligonucleotides

[0507] 1.3.1 Ammonolysis

[0508] The synthesized solid support (sense or antisense chain) was added to a 5 mL centrifuge tube, and 3% diethylamine / ammonia (v / v) was added. The mixture was reacted in a 35°C water bath for 16 hours (or in a 55°C water bath for 8 hours). After filtration, the solid support was washed three times with 1 mL of ethanol / water each time. The filtrate was centrifuged and concentrated, and the crude product was purified.

[0509] 1.3.2 Purification

[0510] Purification and desalting methods are well known to those skilled in the art. For example, a column packed with strong anion exchange media can be used for elution purification with a sodium chloride-sodium hydroxide system, and the product can be collected and piped. Desalting can be performed using a gel-packed purification column with pure water as the elution system.

[0511] 1.3.3 Annealing

[0512] According to Table 6, the sense chain (SS chain) and the antisense chain (AS chain) are mixed in a molar ratio (SS chain / AS chain = 1 / 1.05), heated in a water bath to 70-95℃, held for 3-5 minutes, naturally cooled to room temperature, and then freeze-dried to obtain the product.

[0513] Example 2: Screening of Hep3B cell line viability

[0514] Cell transfection

[0515] On day 1, after digesting the cell lines, resuspend them and count the cells. Spread the cell resuspended solution onto 96-well plates at 100 μL / well, 1×10⁶ cells / well. 4 Cells per well, transfection performed 18 hours later.

[0516] The next day, the 20 μM siRNA stock solution was diluted with Opti-MEM. 198 μL of Opti-MEM was added to 2 μL of siRNA stock solution. The final siRNA concentration is shown below. Mix well by pipetting and aspiration, and set aside for use.

[0517] The next day, take 14.1 μL of Opti-MEM and dilute 0.9 μL of Lipofectamine. TM RNAiMAX (Thermo, 13778150) was gently mixed by pipetting and incubated at room temperature for 5 minutes. Then, 15 μL of the prepared RNAi-MAX mixture and 15 μL of diluted siRNA were gently mixed by pipetting, avoiding air bubbles. The mixture was incubated at room temperature for 10 minutes. The resulting solution was then added to a 96-well plate containing cell resuspension at 10 μL / well. The plates were incubated at 37°C in a 5% CO2 incubator for 24 hours (control group did not receive siRNA).

[0518] RNA extraction

[0519] Cell RNA was extracted using a nucleic acid extractor (Hangzhou Aosheng, Auto-pure96) following the instructions of the high-throughput cell RNA extraction kit (Shanghai Fushen Biotechnology, FSF0035-CS).

[0520] RNA reverse transcription

[0521] Refer to PrimeScript for the preparation of denaturing reaction mixtures. TM II 1st Strand cDNA Synthesis Kit (Takara, 6210B) Single well preparation volume: Oligo dT Primer 1μL, dNTP Mixture 1μL, template RNA 12.5μL. Incubate at 65℃ in a standard PCR instrument for 5 minutes, then rapidly cool on ice for 2 minutes.

[0522] The reverse transcription reaction solution was prepared according to PrimeScript. TMII 1st Strand cDNA Synthesis Kit (Takara, 6210B). Each well contains 4 μL of 5×Prime Script II Buffer, 0.5 μL of RNase Inhibitor, and 1 μL of Prime Script II RTase.

[0523] 14.5 μL of the denatured reaction solution was slowly mixed with the reverse transcription reaction solution, and the reverse transcription was performed by incubating at 42°C for 45 minutes in a conventional PCR instrument. The enzyme was then inactivated by incubating at 95°C for 5 minutes, and the reverse transcription product (cDNA) was cooled at 4°C.

[0524] After the reversal is complete, add 30 μL of distilled water (free of DNase and RNase) to each well of the cDNA sample.

[0525] Real-time PCR

[0526] ReferenceTaqMan TM The procedure for using Fast Advanced Master Mix (ABI, 4444557) was followed by quantitative real-time PCR (qPCR) in a 20 μL volume (ABI, QuantStudio3). The reaction program was: (50℃, 2 min) × 1 cycle; (95℃, 20 s) × 1 cycle; (95℃, 1 s; 60℃, 24 s) × 40 cycles.

[0527] Table 6. Primer Information

[0528] Data statistics

[0529] Calculate 2 -△△Ct The values ​​are then converted to percentages to obtain the residual inhibition rate;

[0530] △△Ct=[(Ct experimental group target gene - Ct experimental group internal reference) - (Ct control group target gene - Ct control group internal reference)].

[0531] The target gene is hINHBE, and the internal control is hGAPDH.

[0532] Using the Hep3B cell line (Nanjing Kebai, CBP60197), high-throughput screening of siRNA compound cell line activity was performed with final concentrations of 10 nM and 0.1 nM. The experimental screening results are shown in Table 7.

[0533] Table 7. Results of Hep3B cell line viability screening

[0534] Example 3: Screening of Hep3B cell line viability

[0535] Following the method in Example 2, Hep3B cell line was used to perform high-throughput screening of siRNA compound cell line activity using a starting concentration of 10 nM and 7 concentration points (10 nM, 3.33 nM, 1.11 nM, 0.37 nM, 0.123 nM, 0.041 nM, 0.0137 nM) with 3-fold serial dilutions. The experimental screening results are shown in Table 10.

[0536] Table 8. Results of Hep3B cell line viability screening

[0537] Example 4: Activity Detection of AAV Humanized Mouse Model

[0538] AAV8 virus carrying the human INHBE gene fragment (purchased from Heyuan Biotechnology) was injected into mice via the tail vein at a dose of 1 x 10⁻⁶. 11 vg. Day -14 was defined as the day of viral injection. siRNA was administered subcutaneously on day 0 at a dose of 3 mg / kg, with 3 mice per group. Mice were euthanized on day 14 (D14). Subsequent tissue collection was performed: liver tissue was harvested; right lobe liver tissue was placed in an RNAlater and incubated overnight at 4°C for RNA extraction; the remaining tissue was stored at -80°C.

[0539] RNA extraction was performed using a nucleic acid extractor (Hangzhou Aosheng, Auto-pure96) following the instructions of the high-throughput cell RNA extraction kit (Shanghai Fushen Biotechnology, FSF0035-TS).

[0540] The methods for RNA reverse transcription and quantitative real-time PCR are as described above, and the primer sequences used are shown in the table below. The residual inhibition rate was calculated using the same method as described above, with hINHBE as the target gene and mGAPDH as the internal control.

[0541] Table 9. Primer Information

[0542] Table 10. Activity evaluation results in AAV mice

[0543] Example 5: Activity Detection of Transgenic Mouse Model

[0544] Experimental animals were selected from 6-8 week old hINHBE KI mice (purchased from Biocytogen), with a quantity of 3 mice per compound per test point. Two days prior to administration (D-2), mice were fasted for 4 hours and serum was collected. On the day of administration (D0), the mice were weighed and the test compound was administered subcutaneously at a volume of 5 ml / kg, as a single dose. Depending on the experimental objective, the mice were weighed on D6 / D13, and euthanized after a 16-hour fast on D7 / D14.

[0545] The final sampling was then performed: liver tissue was taken, liver weight was measured, the right lobe liver tissue was placed in an RNAlater, incubated overnight at 4°C for RNA extraction, and the remaining tissue was stored at -80°C.

[0546] RNA extraction and quantitative PCR methods were described above, and the primer sequences used are listed in Table 9. The residual inhibition rate was calculated using the same method as described above.

[0547] Table 11. Activity assays of the compounds of this invention in transgenic mouse models (Experiment 1)

[0548] Table 12. Activity assays of the compounds of this invention in transgenic mouse models (Experiment 2)

[0549] Table 13. Activity assays of the compounds of this invention in transgenic mouse models (Experiment 3)

[0550] ND: Not detected

[0551] Table 14. Activity assays of the compounds of this invention in transgenic mouse models (Experiment 4)

[0552] Table 15. Activity assays of the compounds of this invention in transgenic mouse models (Experiment 5)

[0553] Table 16. Activity assays of the compounds of this invention in transgenic mouse models (Experiment 6)

[0554] Table 17. Activity assays of the compounds of this invention in transgenic mouse models (Experiment 6)

Claims

1. A double-stranded RNA (dsRNA) for inhibiting INHBE gene expression, said dsRNA comprising a sense strand and an antisense strand forming a double-stranded region, wherein the length of the sense strand and the antisense strand is each independently 15-30 nucleotides, and said antisense strand comprises a nucleotide sequence of at least 15 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO:243-484.

2. The dsRNA of claim 1, wherein the sense strand comprises a nucleotide sequence of at least 15 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO:1-242.

3. The dsRNA of claim 1 or 2, wherein the dsRNA is siRNA.

4. The dsRNA of any one of claims 1-3, wherein the length of the sense strand and the antisense strand is each independently 15-27 nucleotides, preferably 18-25 nucleotides, more preferably 19-21 nucleotides.

5. The dsRNA of any one of claims 1-4, wherein the dsRNA has two nucleotide overhangs at the 3' end of the antisense strand and a blunt end at the 5' end of the antisense strand.

6. The dsRNA of any one of claims 1-5, wherein the antisense strand comprises a nucleotide sequence of at least 16 consecutive nucleotides, at least 17 consecutive nucleotides, at least 18 consecutive nucleotides, at least 19 consecutive nucleotides, or at least 20 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO: 243-484, preferably the antisense strand comprises a nucleotide sequence shown in any one of SEQ ID NO: 243-484.

7. The dsRNA of any one of claims 1-6, wherein the positive strand comprises a nucleotide sequence of at least 16 consecutive nucleotides, at least 17 consecutive nucleotides, or at least 18 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO: 1-242, preferably the positive strand comprises a nucleotide sequence shown in any one of SEQ ID NO: 1-242.

8. The dsRNA of any one of claims 1-7, wherein the antisense strand comprises a nucleotide sequence of at least 15 consecutive nucleotides, at least 16 consecutive nucleotides, at least 17 consecutive nucleotides, at least 18 consecutive nucleotides, at least 19 consecutive nucleotides, or at least 20 consecutive nucleotides of the nucleotide sequence shown in any one of SEQ ID NO: 273, 288, 310, 412, 479, 480, 481, 482, 483, and 484.

9. The dsRNA of any one of claims 1-8, wherein the sense strand comprises a nucleotide sequence of at least 15 contiguous nucleotides, a nucleotide sequence of at least 16 contiguous nucleotides, a nucleotide sequence of at least 17 contiguous nucleotides, or a nucleotide sequence of at least 18 contiguous nucleotides of the nucleotide sequence set forth in any one of SEQ ID NOs: 37, 52, 74, 176, 237, 238, 239, 240, 241, and 242.

10. The dsRNA of any one of claims 1-9, wherein the siRNA comprises any one of the paired sense strand sequence and antisense strand sequence as set forth in Table 3 of the specification.

11. The dsRNA of claim 10, wherein: (1) the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 37, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 273; (2) the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 52, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 288; (3) the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 74, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 310; (4) the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 176, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 412; (5) the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 237, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 479; (6) the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 238, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 480; (7) the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 239, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 481; (8) the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 240, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 482; (9) the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 241, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 483; or (10) the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 242, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO:

484.

12. The dsRNA of any one of claims 1-11, wherein substantially all of the nucleotides of the sense strand and / or substantially all of the nucleotides of the antisense strand are modified nucleotides, or all of the nucleotides of the sense strand and / or all of the nucleotides of the antisense strand are modified nucleotides.

13. The dsRNA of claim 12, wherein each of the sense strand and the antisense strand independently comprises one or more modified nucleotides selected from the group consisting of 2'-0-alkyl modified nucleotides (e.g., 2'-0-methyl modified nucleotides), 2'-methoxyethyl modified nucleotides, 2'-fluoro modified nucleotides, 2'-deoxy-modified nucleotides, inosine ribonucleotides, abasic nucleotides, inverted abasic deoxyribonucleotides, nucleotides comprising phosphorothioate groups, vinyl phosphonate modified nucleotides, locked nucleotides, unlocked nucleotides, 2'-amino-modified nucleotides, 2'-C-alkyl-modified nucleotides, 2'-0-allyl modified nucleotides, morpholino nucleotides, phosphoramidates, nucleotides comprising non-natural bases, terminal nucleotides linked to a cholesteryl derivative or dodecanoic acid didecanoyl amide group, deoxyribonucleotides, 3'-terminal deoxythymine (dT) nucleotides, conformationally restricted nucleotides, constrained ethyl nucleotides, 2'-hydroxyl modified nucleotides, nucleotides comprising methylphosphonate groups, nucleotides comprising 5'-phosphates, nucleotides comprising 5'-phosphate mimics, glycol modified nucleotides (GNAs), 2-O-(N-methylacetamide) modified nucleotides, 2'-methoxyethyloxyethyloxy modified nucleotides, 2'-methoxyethyloxy modified nucleotides, and SCP modified nucleotides.

14. The dsRNA of claim 13, wherein each of the sense strand and the antisense strand independently comprises one or more nucleotide modifications selected from the group consisting of 2'-0-methyl modified nucleotides, 2'-fluoro modified nucleotides, 2'-deoxy modified nucleotides, nucleotides comprising phosphorothioate groups, 2'-methoxyethyloxyethyloxy modified nucleotides, 2'-methoxyethyloxy modified nucleotides, and SCP modified nucleotides.

15. The dsRNA of any one of claims 1-14, wherein the sense strand has a length of 19 nucleotides and has: (i) (counting from the 5' end) 2'-0-methyl modified nucleotides at positions 1 to 6, 10 to 19, and 2'-fluoro modified nucleotides at positions 7-9; and / or (ii) (counting from the 5' end) phosphorothioate internucleotide linkages between nucleotide positions 1 and 2, between nucleotide positions 2 and 3, and between nucleotide positions 18 and 19.

16. The dsRNA of any one of claims 1-15, wherein the antisense strand has a length of 21 nucleotides and has (i) (counting from the 5' end) 2'-0-methyl modified nucleotides at positions 1, 3 to 5, 7 to 13, 15, 17 to 21, and 2'-fluoro modified nucleotides at positions 2, 6, 14, and 16; and / or (ii) (counting from the 5' end) phosphorothioate internucleotide linkages between nucleotide positions 1 and 2, between nucleotide positions 2 and 3, between nucleotide positions 19 and 20, and between nucleotide positions 20 and 21.

17. The dsRNA of any one of claims 1-16, wherein the antisense strand has a length of 21 nucleotides and has (i) an SCP-modified nucleotide at position 1 (counting from the 5' end); (ii) 2'-deoxy-modified nucleotides at positions 2, 5, 7, and 12 (counting from the 5' end); (iii) a 2'-fluoro-modified nucleotide at position 14 (counting from the 5' end); (iv) 2'-O-methyl-modified nucleotides at positions 3, 4, 6, 8 to 11, 13, and 15 to 21 (counting from the 5' end); (v) phosphorothioate internucleotide linkages between nucleotide positions 1 and 2, between nucleotide positions 2 and 3, between nucleotide positions 19 and 20, and between nucleotide positions 20 and 21 (counting from the 5' end).

18. The dsRNA of any one of claims 1-16, wherein the antisense strand has a length of 21 nucleotides and has (i) an SCP-modified nucleotide at position 1 (counting from the 5' end); (ii) 2'-fluoro-modified nucleotides at positions 2, 5, 7, 12, and 14 (counting from the 5' end); (iii) 2'-O-methyl-modified nucleotides at positions 3, 4, 6, 8 to 11, 13, and 15 to 21 (counting from the 5' end); (iv) phosphorothioate internucleotide linkages between nucleotide positions 1 and 2, between nucleotide positions 2 and 3, between nucleotide positions 19 and 20, and between nucleotide positions 20 and 21 (counting from the 5' end).

19. The dsRNA of any one of claims 1-16, wherein the antisense strand has a length of 21 nucleotides and has (i) an SCP-modified nucleotide at position 1 (counting from the 5' end); (ii) 2'-fluoro-modified nucleotides at positions 2, 7, 12, 14, and 16 (counting from the 5' end); (iii) 2'-O-methyl-modified nucleotides at positions 3 to 6, 8 to 11, 13, 15, and 17 to 21 (counting from the 5' end); (iv) phosphorothioate internucleotide linkages between nucleotide positions 1 and 2, between nucleotide positions 2 and 3, between nucleotide positions 19 and 20, and between nucleotide positions 20 and 21 (counting from the 5' end).

20. The dsRNA of any one of claims 1-16, wherein the antisense strand has a length of 21 nucleotides and has (i) an SCP-modified nucleotide at position 1 (counting from the 5' end); (ii) 2'-fluoro-modified nucleotides at positions 2, 5, 12, 14, and 16 (counting from the 5' end); (iii) 2'-O-methyl-modified nucleotides at positions 3, 4, 6 to 11, 13, 15, and 17 to 21 (counting from the 5' end); (iv) a phosphorothioate internucleotide linkage between nucleotide positions 1 and 2, 2 and 3, 19 and 20, and 20 and 21 (counting from the 5' end).

21. The dsRNA of any one of claims 1-16, wherein the antisense strand has a length of 21 nucleotides and has (i) an SCP-modified nucleotide at position 1 (counting from the 5' end); (ii) 2'-fluoro-modified nucleotides at positions 2, 6, 14, and 16 (counting from the 5' end); (iii) 2'-0-methyl-modified nucleotides at positions 3 to 5, 7 to 13, 15, and 17 to 21 (counting from the 5' end); (iv) a phosphorothioate internucleotide linkage between nucleotide positions 1 and 2, 2 and 3, 19 and 20, and 20 and 21 (counting from the 5' end).

22. The dsRNA of any one of claims 17-21, wherein the antisense strand further has: (i) a GNA at position 7 (counting from the 5' end); (ii) a 2'-methoxyethoxyethoxy-modified nucleotide at position 3 (counting from the 5' end); or (iii) a 2'-methoxyethoxy-modified nucleotide at position 3 (counting from the 5' end).

23. The dsRNA of any one of claims 1-22, wherein the sense strand of the dsRNA comprises the modified nucleotide sequence of any one set forth in Table 4A of the specification, and / or the antisense strand comprises the modified nucleotide sequence of any one set forth in Tables 5A and 5B of the specification.

24. The dsRNA of any one of claims 1-23, wherein: (i) the sense strand comprises CmsAmsGmUmCmGmUfCfCfCmAmGmAmAmUmAmAmCmsAm (SEQ ID No. 1023) and the antisense strand comprises (SCP-U)sdGsUmUmdAUmdTCmUmGmGmdGAmCfGmAmCmUmGmsGmsUm (SEQ ID No. 987); (ii) the sense strand comprises CmsAmsGmUmCmGmUfCfCfCmAmGmAmAmUmAmAmCmsAm (SEQ ID No. 1024) and the antisense strand comprises (SCP-U)sGfsUmUmAfUmUfCmUmGmGmGfAmCfGmAmCmUmGmsGmsUm (SEQ ID No. 988); (iii) the sense strand comprises CmsAmsGmUmCmGmUfCfCfCmAmGmAmAmUmAmAmCmsAm (SEQ ID No. 1025) and the antisense strand comprises (SCP-U) sGfsUmUmAmUmUfCmUmGmGmGfAmCfGmAfCmUmGmsGmsUm (SEQ ID No. 989); (iv) the sense strand comprises CmsAmsGmUmCmGmUfCfCfCmAmGmAmAmUmAmAmCmsAm (SEQ ID No. 1026) and the antisense strand comprises (SCP-U) sGfsUmUmAfUmUmCmUmGmGmGfAmCfGmAfCmUmGmsGmsUm (SEQ ID No. 990); (v) the sense strand comprises CmsAmsGmUmCmGmUfCfCfCmAmGmAmAmUmAmAmCmsAm (SEQ ID No. 1027) and the antisense strand comprises (SCP-U) sGfsUmUmAmUfUmCmUmGmGmGmAmCfGmAfCmUmGmsGmsUm (SEQ ID No. 991); (vi) the sense strand comprises CmsAmsGmCmUmUmUfGfCfUmAmCmUmGmUmCmAmCmsAm (SEQ ID No. 1028) and the antisense strand comprises (SCP-U) sGfsUmGmAfCmAfGmUmAmGmCfAmAfAmGmCmUmGmsAmsUm (SEQ ID No. 992); (vii) the sense strand comprises CmsAmsGmCmUmUmUfGfCfUmAmCmUmGmUmCmAmCmsAm (SEQ ID No. 1029) and the antisense strand comprises (SCP-U) sdGsUmGmdACm(Agn)GmUmAmGmdCAmAfAmGmCmUmGmsAmsUm (SEQ ID No. 993); (viii) the sense strand comprises CmsAmsGmCmUmUmUfGfCfUmAmCmUmGmUmCmAmCmsAm (SEQ ID No. 1030) and the antisense strand comprises (SCP-U) sGfs(Umoeoe)GmAfCmAfGmUmAmGmCfAmAfAmGmCmUmGmsAmsUm (SEQ ID No. 994); (ix) the sense strand comprises CmsAmsCmCmCmUmUfCfCfUmGmGmCmAmCmUmCmUmsAm (SEQ ID No. 1031) and the antisense strand comprises (SCP-U) sAfsGmAmGfUmGfCmCmAmGmGfAmAfGmGmGmUmGmsGmsGm (SEQ ID No. 995); (x) the sense strand comprises CmsAmsCmCmCmUmUfCfCfUmGmGmCmAmCmUmCmUmsAm (SEQ ID No. 1032) and the antisense strand comprises (SCP-U) sAfsGmAmGmUmGfCmCmAmGmGfAmAfGmGfGmUmGmsGmsGm (SEQ ID No. 996); (xi) the sense strand comprises CmsAmsCmCmCmUmUfCfCfUmGmGmCmAmCmUmCmUmsAm (SEQ ID No. 1033) and the antisense strand comprises (SCP-U) sdAsGmAmdGUmdGCmCmAmGmdGAmAfGmGmGmUmGmsGmsGm (SEQ ID No. 997); (xii) the sense strand comprises UmsCmsCmUmGmGmCfAfCfUmCmUmUmUmGmCmUmUmsAm (SEQ ID No. 1034) and the antisense strand comprises UmsdAsAmGmdCAmdAAmGmAmGmdTGmCfCmAmGmGmAmsAmsGm (SEQ ID No. 998); (xiii) the sense strand comprises UmsCmsCmUmGmGmCfAfCfUmCmUmUmUmGmCmUmsUmsAm (SEQ ID No. 1035) and the antisense strand comprises (SCP-U) sAfsAmGmCfAmAfAmGmAmGmUfGmCfCmAmGmGmAmsAmsGm (SEQ ID No. 999); (xiv) the sense strand comprises UmsCmsCmUmGmGmCfAfCfUmCmUmUmUmGmCmUmsUmsAm (SEQ ID No. 1036) and the antisense strand comprises (SCP-U) sAfsAmGmCmAmAfAmGmAmGmUfGmCfCmAfGmGmAmsAmsGm (SEQ ID No. 1000); (xvii) the sense strand comprises UmsGmsAmGmAmAmGfUfCfUmGmGmUmGmUmCmCmUmsAm (SEQ ID No. 1039), and the antisense strand comprises (SCP-U) sAfsGmGmAmCmAfCmCmAmGmAfCmUfUmCfUmCmAmsCmsCm (SEQ ID No. 1003); (xvii) the sense strand comprises UmsGmsAmGmAmAmGfUfCfUmGmGmUmGmUmCmCmUmsAm (SEQ ID No. 1039), and the antisense strand comprises (SCP-U) sAfsGmGmAmCmAfCmCmAmGmAfCmUfUmCfUmCmAmsCmsCm (SEQ ID No. 1003); (xvii) the sense strand comprises UmsGmsAmGmAmAmGfUfCfUmGmGmUmGmUmCmCmUmsAm (SEQ ID No. 1039), and the antisense strand comprises (SCP-U) sAfsGmGmAmCmAfCmCmAmGmAfCmUfUmCfUmCmAmsCmsCm (SEQ ID No. 1003); (xvii) the sense strand comprises UmsGmsAmGmAmAmGfUfCfUmGmGmUmGmUmCmCmUmsAm (SEQ ID No. 1039), and the antisense strand comprises (SCP-U) sAfsGmGmAmCmAfCmCmAmGmAfCmUfUmCfUmCmAmsCmsCm (SEQ ID No. 1003); (xvii) the sense strand comprises UmsGmsAmGmAmAmGfUfCfUmGmGmUmGmUmCmCmUmsAm (SEQ ID No. 1039), and the antisense strand comprises (SCP-U) sAfsGmGmAmCmAfCmCmAmGmAfCmUfUmCfUmCmAmsCmsCm (SEQ ID No. 1003); (xx) the sense strand comprises GmsAmsAmGmUmCmUfGfGfUmGmUmCmCmUmGmAmAmsAm (SEQ ID No. 1042), and the antisense strand comprises (SCP-U) sUfsUmCmAfGmGfAmCmAmCmCfAmGfAmCmUmUmCmsUmsCm (SEQ ID No. 1006); (xxxii) the sense strand comprises CmsCmsUmAmGmAmGfCfUfUmAmAmGmAmUmCmCmGmsAm (SEQ ID No. 1044) and the antisense strand comprises (SCP-U) sdCsGmGmdAUmdCUmUmAmAmdGCmUfCmUmAmGmGmsAmsAm (SEQ ID No. 1008); (xxxii) the sense strand comprises CmsCmsUmAmGmAmGfCfUfUmAmAmGmAmUmCmCmGmsAm (SEQ ID No. 1044) and the antisense strand comprises (SCP-U) sdCsGmGmdAUmdCUmUmAmAmdGCmUfCmUmAmGmGmsAmsAm (SEQ ID No. 1008); (xxxii) the sense strand comprises CmsCmsUmAmGmAmGfCfUfUmAmAmGmAmUmCmCmGmsAm (SEQ ID No. 1044) and the antisense strand comprises (SCP-U) sdCsGmGmdAUmdCUmUmAmAmdGCmUfCmUmAmGmGmsAmsAm (SEQ ID No. 1008); (xxxii) the sense strand comprises CmsCmsUmAmGmAmGfCfUfUmAmAmGmAmUmCmCmGmsAm (SEQ ID No. 1044) and the antisense strand comprises (SCP-U) sdCsGmGmdAUmdCUmUmAmAmdGCmUfCmUmAmGmGmsAmsAm (SEQ ID No. 1008); (xxxii) the sense strand comprises CmsCmsUmAmGmAmGfCfUfUmAmAmGmAmUmCmCmGmsAm (SEQ ID No. 1044) and the antisense strand comprises (SCP-U) sdCsGmGmdAUmdCUmUmAmAmdGCmUfCmUmAmGmGmsAmsAm (SEQ ID No. 1008); (xxxii) the sense strand comprises CmsCmsUmAmGmAmGfCfUfUmAmAmGmAmUmCmCmGmsAm (SEQ ID No. 1044) and the antisense strand comprises (SCP-U) sdCsGmGmdAUmdCUmUmAmAmdGCmUfCmUmAmGmGmsAmsAm (SEQ ID No. 1008); (xxvii) the sense strand comprises GmsGmsCmAmCmUmUfUfCfUmUmGmUmCmUmGmAmGmsAm (SEQ ID No. 1049), and the antisense strand comprises (SCP-U) sCfsUmCmAfGmAfCmAmAmGmAfAmAfGmUmGmCmCmsCmsAm (SEQ ID No. 1013); (xxviii) the sense strand comprises GmsGmsCmAmCmUmUfUfCfUmUmGmUmCmUmGmAmGmsAm (SEQ ID No. 1050), and the antisense strand comprises (SCP-U) sCfsUmCmAmGmAfCmAmAmGmAfAmAfGmUfGmCmCmsCmsAm (SEQ ID No. 1014); (xxix) the sense strand comprises GmsGmsCmAmCmUmUfUfCfUmUmGmUmCmUmGmAmGmsAm (SEQ ID No. 1051), and the antisense strand comprises (SCP-U) sCfsUmCmAfGmAmCmAmAmGmAfAmAfGmUfGmCmCmsCmsAm (SEQ ID No. 1015); (xxx) the sense strand comprises GmsGmsCmAmCmUmUfUfCfUmUmGmUmCmUmGmAmGmsAm (SEQ ID No. 1052), and the antisense strand comprises (SCP-U) sCfsUmCmAmGfAmCmAmAmGmAmAmAfGmUfGmCmCmsCmsAm (SEQ ID No. 1016); (xxxi) the sense strand comprises CmsAmsGmCmUmUmUfGfCfUmAmCmUmGmUmCmAmsCmsAm (SEQ ID No. 536), and the antisense strand comprises UmsGfsUmGfAmCfAmGfUmAfGmCfAmAfAmGfCmUfGmsAmsUm (SEQ ID No. 802); (xxxii) the sense strand comprises GmsAmsAmGmUmCmUfGfGfUmGmUmCmCmUmGmAmsAmsAm (SEQ ID No. 558), and the antisense strand comprises UmsUfsUmCfAmGfGmAfCmAfCmCfAmGfAmCfUmUfCmsUmsCm (SEQ ID No. 824); or (xxxiii) the sense strand comprises CmsAmsGmCmUmUmUfGfCfUmAmCmUmGmUmCmAmsCmsAm (SEQ ID No. 537), and the antisense strand comprises UmsGfsUmGfAmCfAmGfUmAfGmCfAmAfAmGfCmUfGmsAmsUm (SEQ ID No. 803); or (xxxiv) the sense strand comprises GmsAmsAmGmUmCmUfGfGfUmGmUmCmCmUmGmAmsAmsAm (SEQ ID No. 559), and the antisense strand comprises UmsUfsUmCfAmGfGmAfCmAfCmCfAmGfAmCfUmUfCmsUmsCm (SEQ ID No. 825). (xxxii) The justice chain contains GmsGmsCmAmCmUmUfUfCfUmUmGmUmCmUmGmAmsGmsAm (SEQ ID No. 660), and the antisense chain contains UmsCfsUmCfAmGfAmCfAmAfGmAfAmAfGmUfGmCfCmsCmsAm (SEQ ID No. 926).

25. The dsRNA of any one of claims 1-24, wherein the dsRNA is further conjugated to a ligand portion. Preferably, the dsRNA is conjugated to a ligand moiety containing N-acetylgalactosamine, more preferably conjugated to the ligand moiety containing N-acetylgalactosamine via a phosphate ester group or a thiophosphate ester group; Preferably, the sense strand of the dsRNA is conjugated to the ligand portion, and more preferably, the 3' end of the sense strand is conjugated to the ligand portion; Preferably, the ligand portion has the following structure: wherein This indicates the position where the dsRNA is linked via a phosphate group or a thiophosphate group.

26. The dsRNA of claim 25, wherein the sense strand comprises a modified nucleotide sequence of any one of those shown in Table 4B of the specification, and preferably the antisense strand comprises a modified nucleotide sequence of any one of those shown in Table 5B of the specification.

27. The dsRNA of claim 26, wherein the dsRNA comprises a sense strand and an antisense strand selected from the following dsRNAs: DR011676, DR012642, DR012668, DR013589, DR013593, DR012657, DR013550, DR014064, DR012645, DR012671, DR011680, DR011614, DR013994, DR014012, DR014047, DR0 12648, DR012674, DR013597, DR013598, DR013993, DR014011, DR011839, DR012651, DR014016, DR013980, DR013998, DR012664, DR012680, DR013590, DR013594, DR014294, DR014295, DR014297, DR014272, DR014274 and DR014275.

28. A cell containing dsRNA as described in any one of claims 1-27.

29. A pharmaceutical composition comprising dsRNA as described in any one of claims 1-27, or cells as described in claim 28, and optionally a pharmaceutically acceptable carrier or excipient.

30. A kit comprising dsRNA as described in any one of claims 1-27, cells as described in claim 28, or a pharmaceutical composition as described in claim 29.

31. A method of inhibiting expression of an INHBE gene in a cell, the method comprising contacting the cell with the dsRNA of any one of claims 1-27 or the pharmaceutical composition of claim 31.

32. The method of claim 31, performed in vitro.

33. A method of inhibiting expression of an INHBE gene in a cell in a subject, the method comprising administering to the subject the dsRNA of any one of claims 1-27, the cell of claim 28, or the pharmaceutical composition of claim 29.

34. A method of treating a disease or disorder in a subject that benefits from a reduction in INHBE gene expression, the method comprising the step of administering to the subject the dsRNA of any one of claims 1-27, the cell of claim 28, or the pharmaceutical composition of claim 29.

35. A method of preventing at least one symptom in a subject having a disease or disorder that benefits from a reduction in INHBE gene expression, the method comprising the step of administering to the subject the dsRNA of any one of claims 1-27, the cell of claim 28, or the pharmaceutical composition of claim 29.

36. A method of preventing progression of a disease or disorder in a subject that benefits from a reduction in INHBE gene expression, the method comprising the step of administering to the subject the dsRNA of any one of claims 1-27, the cell of claim 28, or the pharmaceutical composition of claim 29.

37. The method of any one of claims 34-36, wherein the disease or disorder that benefits from a reduction in INHBE gene expression is an INHBE-associated disease.

38. The method of claim 37, wherein the INHBE-associated disease is selected from the group consisting of metabolic syndrome, type 2 diabetes, obesity, elevated triglyceride levels, lipodystrophy, hepatitis, fatty liver, hypercholesterolemia, elevated liver enzymes, nonalcoholic steatohepatitis, cardiovascular disease, heart failure, and kidney disease.

39. The method of any one of claims 33-38, wherein the dsRNA, cell, or pharmaceutical composition is administered subcutaneously.

40. The method of any one of claims 33-39, wherein the subject is a human.

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