Oligonucleotide conjugates inhibiting URAT1 expression and uses thereof

Abstract

The double-stranded oligonucleotide, the double-stranded oligonucleotide conjugate and the pharmaceutical composition provided by the invention can inhibit the expression of urate transporter 1, reduce the content of URAT1 protein and obviously reduce blood uric acid. The oligonucleotide conjugate and the pharmaceutical composition thereof provided by the disclosure are helpful for treating and / or preventing pathological conditions or diseases related to URAT1 expression, including hyperuricemia or gout.

Description

Technical Field

[0001] This disclosure belongs to the field of oligonucleotide drug technology, specifically relating to an oligonucleotide molecule with URAT1 inhibitory activity, its preparation method, and its application. Background Technology

[0002] Urate transporter 1 (URAT1) is the main transporter protein controlling the reabsorption of urate in the kidneys and a key target in the pathogenesis of hyperuricemia. Existing drugs that treat hyperuricemia by inhibiting URAT1 typically reduce blood uric acid levels by decreasing reabsorption, thus treating hyperuricemia and gout. However, these inhibitors usually have strong hepatotoxicity and kidney damage.

[0003] RNAi is one of the more mature treatment methods currently available. Reducing the expression of the gene encoding the URAT1 protein can decrease uric acid reabsorption and promote increased uric acid excretion, thereby lowering blood uric acid levels. However, there are currently no RNAi drugs specifically targeting URAT1 on the market. Summary of the Invention

[0004] The double-stranded oligonucleotides disclosed herein can inhibit the expression of urate transporter 1, reduce URAT1 protein content, and significantly reduce blood uric acid.

[0005] In a first aspect of this disclosure, a double-stranded oligonucleotide for inhibiting URAT1 gene expression is provided, the double-stranded oligonucleotide comprising a sense strand and an antisense strand, the sense strand and the antisense strand being 17 to 30 nucleotides in length, and the sense strand and the antisense strand being at least partially complementary.

[0006] In some embodiments of this disclosure, the antisense strand is 21 to 23 nucleotides in length; the sense strand is 19 to 21 nucleotides in length.

[0007] The antisense strand comprises at least 17 consecutive nucleotides whose first to 19th nucleotides differ by no more than 3 nucleotides from any of the nucleotide sequences shown in SEQ ID NO:70 to SEQ ID NO:138 in Table 3.

[0008] The positive chain comprises at least 17 consecutive nucleotides that differ by no more than 3 nucleotides from any of the nucleotide sequences shown in SEQ ID NO:1 to SEQ ID NO:69 in Table 3.

[0009] In some embodiments of this disclosure, the double-stranded oligonucleotide contains at least one modified nucleotide.

[0010] In some embodiments of this disclosure, each nucleotide in the double-stranded oligonucleotide is a modified nucleotide.

[0011] In some embodiments of this disclosure, the modified nucleotide is selected from: 2'-halogenated nucleotides, 2'-deoxygenated nucleotides, 2'-O-alkyl-modified nucleotides, and 2'-O-(CH2). n -OMe modified nucleotides (n selected from 1 or 2), 2'-amino modified nucleotides, nucleotides modified with 2',2'-[halogen, halogen] disubstituted or 2',2'-[halogen, alkyl] disubstituted groups, debased nucleotides, or nucleotide-like nucleotides.

[0012] In some embodiments of this disclosure, the modified double-stranded oligonucleotides are shown in Table 4.

[0013] In a second aspect of this disclosure, an oligonucleotide conjugate is provided, comprising the double-stranded oligonucleotide described in the first aspect of this disclosure and a targeting ligand group.

[0014] In some embodiments of this disclosure, the targeting ligand group is a polypeptide.

[0015] In some embodiments of this disclosure, the targeting ligand group includes any of the polypeptide fragments shown in 1)-6) below:

[0016] 1)-X1(KKEEE) n -K m - where n = 1-5, m = 0 or 1 or 2, and X1 is selected from any L-α amino acid or L-β amino acid other than K and E;

[0017] 2)-CLPVASC-, preferably in cyclic peptide form

[0018] 3)-CYFQNC-; Preferred cyclic peptide form

[0019] 4)-KIDRI-;

[0020] 5)-IDRI-(-Ile-Asp-Arg-Ile-);

[0021] 6)-dXa-Ser-dXb-X2-dXc-Gly-Xd-Ile-Asp-Arg(Ak)-Ile-; where X2 is selected from any non-natural amino acid; dXa, dXb, dXc are selected from any D-type amino acid; Arg(Ak) is selected from arginine or alkylated arginine; and Xd is selected from Hyp or Pro.

[0022] In a third aspect of this disclosure, a pharmaceutical composition is provided comprising the double-stranded oligonucleotides described in the first aspect of this disclosure and / or the double-stranded oligonucleotide conjugates described in the second aspect of this disclosure.

[0023] In some embodiments of this disclosure, the pharmaceutical composition further includes a pharmaceutically acceptable carrier or excipient.

[0024] In a fourth aspect of this disclosure, the present disclosure provides the use of the double-stranded oligonucleotides described in the first aspect of this disclosure and / or the double-stranded oligonucleotide conjugates described in the second aspect of this disclosure and / or the pharmaceutical compositions described in the third aspect of this disclosure in the preparation of a medicament for inhibiting URAT1 gene expression.

[0025] In a fifth aspect of this disclosure, the present disclosure provides the use of the double-stranded oligonucleotides described in the first aspect of this disclosure and / or the double-stranded oligonucleotide conjugates described in the second aspect of this disclosure and / or the pharmaceutical compositions described in the third aspect of this disclosure in the preparation of a medicament for treating and / or preventing pathological conditions or diseases associated with URAT1 expression.

[0026] In some embodiments of this disclosure, the pathological condition or disease is hyperuricemia or gout.

[0027] In a sixth aspect of this disclosure, this disclosure provides a method for inhibiting intracellular URAT1 expression, comprising administering to a subject the double-stranded oligonucleotides described in the first aspect of this disclosure and / or the double-stranded oligonucleotide conjugates described in the second aspect of this disclosure and / or the pharmaceutical compositions described in the third aspect of this disclosure.

[0028] In a seventh aspect of this disclosure, this disclosure provides a method for preventing and / or treating hyperuricemia or gout, the method comprising administering to a subject an effective amount of the double-stranded oligonucleotides described in the first aspect of this disclosure and / or the double-stranded oligonucleotide conjugates described in the second aspect of this disclosure and / or the pharmaceutical compositions described in the third aspect of this disclosure.

[0029] The double-stranded oligonucleotides, double-stranded oligonucleotide conjugates, and pharmaceutical compositions disclosed herein can inhibit the expression of urate transporter 1, reduce URAT1 protein levels, and significantly lower serum uric acid. The oligonucleotide conjugates and pharmaceutical compositions disclosed herein may help treat and / or prevent pathological conditions or diseases associated with URAT1 expression, including hyperuricemia or gout. Detailed Implementation

[0030] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Those skilled in the art can refer to the content of this article and appropriately improve the process parameters to achieve the desired results.

[0031] Terminology Explanation

[0032] Unless otherwise defined, all technical and scientific terms used in this disclosure have the same meaning as commonly understood by one of ordinary skill in the art. While similar or equivalent methods and materials to those described herein may be used in the practice or testing of this invention, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are incorporated herein by reference in their entirety. In case of conflict, this specification (including definitions) shall prevail. Furthermore, materials, methods, and examples are illustrative only and not intended to be limiting.

[0033] In this disclosure, the terms “comprising” or “including” are open-ended expressions used to refer to the phrase “including but not limited to” and are used interchangeably with it, meaning that they include the contents specified in this disclosure but do not exclude other contents.

[0034] In this disclosure, the terms “optionally,” “optionally,” or “optionally” generally refer to an event or condition that may, but may not, occur, and the description includes both cases in which the event or condition occurs and cases in which the event or condition does not occur.

[0035] In this disclosure, the term "small interfering RNA (siRNA)" refers to a double-stranded RNA of 17 to 25 nucleotides in length, comprising a sense strand and an antisense strand. siRNA mediates targeted cleavage of RNA transcripts via the RISC pathway by forming an RNA-induced silencing complex (RISC). Specifically, siRNA directs the specific degradation of mRNA sequences through a known RNA interference (RNAi) process, inhibiting the translation of mRNA into amino acids and its conversion into proteins.

[0036] In this disclosure, the terms "sequence" and "nucleotide sequence" refer to a sequence of nucleobases or nucleotides. As used herein, "base," "nucleotide base," or "nucleobase" is a pyrimidine or purine compound that is a component of a nucleotide and includes purine bases adenine and guanine, and pyrimidine bases cytosine, thymine, and uracil. Nucleobases may be further modified. The synthesis of modified nucleobases (including phosphorous amide compounds comprising modified nucleobases) is known in the art.

[0037] In this disclosure, the term "double-stranded oligonucleotide" refers to a double-stranded structure formed by two oligonucleotides through partial or complete base pairing. The two oligonucleotides include a sense strand and an antisense strand, which may or may not be of the same length. As long as at least some base-pairing regions exist to form a double-stranded region, the oligonucleotide having a double-stranded structure is considered a double-stranded oligonucleotide as described in this disclosure. The nucleotides constituting the double-stranded oligonucleotide in this disclosure can be modified or unmodified nucleotides. When referring to modified nucleotides, unless otherwise specified, the modification does not specifically refer to the modified site. In addition to the modification of the nucleotides, the linking bonds between the nucleotides in the double-stranded oligonucleotide in this disclosure may also be modified. Double-stranded oligonucleotides containing modified linking bonds between nucleotides are also considered double-stranded oligonucleotides as described in this invention. Besides the nucleotide portion, the double-stranded oligonucleotide in this disclosure may also contain compounds or modifiers acceptable in the art to improve the properties of the double-stranded oligonucleotide, such as linking ligands to form conjugates.

[0038] In this disclosure, the term "antisense strand (or guide strand)" includes a region substantially complementary to a target sequence. The term "sense strand (or follower strand)" refers to an iRNA strand containing an iRNA strand substantially complementary to the antisense strand. The term "substantially complementary" means completely complementary or at least partially complementary, for example, the antisense strand being completely complementary or at least partially complementary to the target sequence. In the case of partial complementarity, mismatches may be present in the internal or terminal regions of the molecule, wherein the most tolerant mismatches are present in the terminal regions, for example, within 5, 4, 3, or 2 nucleotides at the 5'- and / or 3' ends of the iRNA. It should be noted that "at least partially substantially complementary" of the antisense strand to the mRNA means that the antisense strand has a polynucleotide substantially complementary to a continuous portion of the mRNA of interest.

[0039] In this disclosure, and unless otherwise stated, the term “complementary” when used to describe a first nucleobase or nucleotide sequence (e.g., the positive / antisense strand of an RNAi agent or a targeting mRNA) relative to a second nucleobase or nucleotide sequence means the ability of an oligonucleotide or polynucleotide comprising the first nucleotide sequence to hybridize with an oligonucleotide or polynucleotide comprising the second nucleotide sequence (forming base-pair hydrogen bonds under mammalian physiological conditions (or similar in vitro conditions)) and form a double-stranded or double-helical structure under certain standard conditions.

[0040] In this disclosure, "complete complementarity" means that in a hybridized nucleobase or nucleotide sequence pair, all (100%) bases of the sequential sequence of the first oligonucleotide will hybridize with the same number of bases of the sequential sequence of the second oligonucleotide. The sequential sequence may comprise all or part of the first or second nucleotide sequence. "Partial complementarity" means that in a hybridized nucleobase or nucleotide sequence pair, a majority, such as at least 70% (but not all), of the bases of the sequential sequence of the first oligonucleotide will hybridize with the same number of bases of the sequential sequence of the second oligonucleotide. The sequential sequence may comprise all or part of the first or second nucleotide sequence.

[0041] In this disclosure, "substantially complementary" means that in the hybridized nucleobase or nucleotide sequence pair, the vast majority, such as at least 85% or up to three nucleotide differences (but not all), of the sequential sequence of the first oligonucleotide will hybridize with the same number of bases in the sequential sequence of the second oligonucleotide. The sequential sequence may comprise all or part of the first or second nucleotide sequence.

[0042] In this disclosure, the terms “complementary,” “fully complementary,” “partially complementary,” and “substantially complementary” are used to refer to nucleobase or nucleotide matching between the sense and antisense strands of an RNAi agent, or between the antisense strand of an RNAi agent and the sequence of the target mRNA.

[0043] Unless otherwise specified, the term "complementarity" refers to the ability of an oligonucleotide in the first sequence to hybridize with an oligonucleotide in the second sequence under certain conditions and form a double-stranded structure. "At least partially complementary" means that the two sequences can be completely complementary, or have no more than 5, 4, 3, or 2 mismatched base pairs in total, while retaining the ability to hybridize under the relevant conditions. Furthermore, in cases where two oligonucleotides are designed to form one or more single-stranded overhangs upon hybridization, such overhangs should not be considered mismatches for determining complementarity. Correspondingly, in this invention, unless otherwise specified, "mismatch" means that in the siRNA double-stranded molecule, the bases at corresponding positions are not paired in a complementary manner.

[0044] In this disclosure, the terms "nucleotide difference," "nucleotide base difference," and "nucleotide sequence difference" are used interchangeably. A nucleotide difference refers to a change in the type of bases of nucleotides at the same or corresponding positions compared to the original nucleotide sequence. For example, if a nucleotide base in the original nucleotide sequence is A, and the nucleotide base at the same or corresponding position is changed to U, C, G, or dT, dC, dG, etc., then a nucleotide sequence difference is considered to exist at that position. It should be noted that if, compared to the original nucleotide sequence, the nucleotides at the same or corresponding positions differ only in the presence or type of modification, then a nucleotide sequence difference is not considered to exist at that position.

[0045] In this disclosure, the terms "ligand" or "conjugation group" refer to an atom or group of atoms that binds to an oligonucleotide or other oligomer. Generally, a conjugation group modifies one or more properties of the compound to which it is linked, including but not limited to pharmacodynamic, pharmacokinetic, binding, absorption, cellular distribution, cellular uptake, charge, and / or clearance properties. When referring to a link between two molecules, the term "link" as used herein means that the two molecules are directly or indirectly connected by a covalent bond, or that the two molecules are associated by a non-covalent bond (e.g., a hydrogen bond or an ionic bond).

[0046] In this disclosure, the terms “link” or “association” refer to the connection between two compounds or molecules, meaning that the two molecules are linked by a covalent bond or associated via a non-covalent bond (e.g., a hydrogen bond or an ionic bond). Unless otherwise stated, the terms “link” and “association” as used in this disclosure may refer to a link between a first compound and a second compound, with or without any inserted atoms or groups of atoms.

[0047] In this disclosure, a linking group is one or more atoms that connect one molecule or a portion of a molecule to another second molecule or a second portion of a molecule. A linking group can contain any number of atoms or functional groups. In some embodiments, the linking group is used solely to link two bioactive molecules.

[0048] Unless otherwise stated, any symbol used in this disclosure refers to any group or groups that may be attached thereto, which is consistent with the scope of the invention as described in this disclosure.

[0049] According to common knowledge in the art, a peptide is a compound formed by the linkage of two or more amino acids via amide bonds. Here, individual amino acids are linked in a specific sequence to form a chain. An amino acid is a compound carrying at least one amino group and at least one carboxyl group. This includes naturally occurring amino acids (amino acids that produce proteins in vivo), non-natural amino acids, or prepared amino acids that can be found in organisms.

[0050] In the peptides disclosed herein, unless otherwise specified, the amino acid units may be in the D- or L- form.

[0051] As used in this disclosure, the term "standard amino acid" refers to the following twenty amino acids: alanine, arginine, asparagine, aspartic acid (aspartate), cysteine, glutamine, glutamic acid (glutamate), glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine.

[0052] As used in this disclosure, the term "non-standard amino acid" refers to an amino acid other than a "standard amino acid" as defined in this disclosure. "Non-standard amino acids" include, but are not limited to, N-formylmethionine, hydroxyproline, selenomethionine, isovaleine, citrulline (Cit), ornithine, α-methyl-aspartate (αMeD), α-methyl-leucine (αMeL), N-methylalanine, N-methyl-glycine (NMeG), N-methylleucine (NMeL), O-cyclohexyl-alanine (Cha), N-ethylalanine, N,N-ε-dimethyllysine (K(Me)2), methylarginine (Arg(Me)), dimethylarginine (R(Me)2), n-alkylated L-α amino acids, and other amino acid analogs or amino acid mimics that function in a manner similar to naturally occurring amino acids.

[0053] As used in this disclosure and as understood by those skilled in the art, a polyethylene glycol (PEG) unit refers to a repeating unit of the formula (CH2CH2O). It should be understood that in the chemical structures disclosed in this disclosure, a PEG unit can be described as (CH2CH2O), (OCH2CH2), or (CH2OCH2). It should also be understood that the number representing the number of repeating PEG units can be placed on either side of the parentheses representing the PEG unit. It should be further understood that terminal PEG units can be capped by atoms (e.g., hydrogen atoms) or some other part.

[0054] In this disclosure, the term "cyclic peptide" refers to a polypeptide chain in which two cysteine ​​residues are linked by a disulfide bond to form a cyclic polypeptide chain. In this disclosure, the disulfide bond between the two cysteine ​​residues in a "cyclic peptide" is represented by "︵".

[0055] In this disclosure, the term "pharmaceutical composition" or "composition" can refer to something used for the treatment of a disease or for use in in vitro cell culture experiments. When used for the treatment of a disease, the term "pharmaceutical composition" generally refers to a unit dose form and can be prepared by any method well known in the pharmaceutical industry. All methods involve the step of combining the active ingredient with excipients constituting one or more adjunct components. Typically, compositions are prepared by uniformly and adequately combining active siRNA with liquid excipients, finely pulverized solid excipients, or both.

[0056] In this disclosure, the term "pharmaceutical acceptable" means that a substance or composition must be chemically and / or toxicologically compatible with other components of the formulation and / or the mammals to which it is treated. Preferably, "pharmaceutical acceptable" as used in this disclosure means approved by a federal regulatory agency or national government, or listed in the United States Pharmacopeia or other generally recognized pharmacopoeia for use in animals, particularly in humans.

[0057] In this disclosure, the term "pharmaceutically acceptable carrier or excipient" may include any solvent, solid excipient, diluent, or other liquid excipient, etc., suitable for a specific target dosage form. The use of any conventional excipients that are incompatible with the siRNA of this disclosure, such as those that produce any adverse biological effects or interactions with any other component of the pharmaceutically acceptable composition in a harmful manner, is also within the scope of this disclosure.

[0058] In this disclosure, the terms “treatment,” “relief,” or “improvement” are used interchangeably. These terms refer to methods of achieving beneficial or desired outcomes, including, but not limited to, treatment benefits. A “treatment benefit” means the eradication or improvement of the underlying disorder being treated. Here, a treatment benefit is achieved by eradicating or improving one or more physical symptoms associated with the underlying disorder, thereby observing improvement in the subject, although the subject may still be suffering from the underlying disorder.

[0059] In this disclosure, the terms “prevention” and “avoidance” are used interchangeably to refer to methods for obtaining beneficial or desired results, including but not limited to preventive benefits. To obtain a “preventive benefit,” the conjugate, RNAi reagent, or composition may be given to a subject at risk of developing a specific disease, or to a subject who reports one or more physiological symptoms of a disease, even if a diagnosis of the disease may not have been made.

[0060] In this disclosure, the term "administration" generally refers to the introduction of a pharmaceutical preparation of this disclosure into the body of a subject by any route of introduction or delivery. Any method known to those skilled in the art for contacting cells, organs, or tissues with the drug may be employed. Administration may include, but is not limited to, intravenous, intra-arterial, intranasal, intraperitoneal, intramuscular, subcutaneous, or oral administration. A daily dose may be divided into one, two, or more doses in suitable forms to be administered at one, two, or more times during a period of time.

[0061] As in this disclosure, the term "regulation of gene expression" means that the expression of a gene, or the level of an RNA molecule or equivalent RNA molecule encoding one or more proteins or protein subunits, is upregulated or downregulated such that the expression, level, or activity is greater or less than that observed in the absence of a regulator. For example, the term "regulation" may mean "inhibition," but the use of the word "regulation" is not limited to this definition.

[0062] In this invention, unless otherwise specified, the term "inhibition" refers to the downregulation of target gene expression due to siRNA-mediated mRNA degradation. "Downregulation" means a decrease in target gene expression level of 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% or more compared to the absence of siRNA treatment.

[0063] In addition to any conventional excipients, the use of any range of siRNAs incompatible with the present disclosure, such as any adverse biological effects produced or interactions with any other component of a pharmaceutically acceptable composition in a harmful manner, is also within the scope of this disclosure.

[0064] To make the objectives, technical solutions, and advantages of this application clearer, the implementation schemes of this application will be further described in detail below with reference to the embodiments.

[0065] Double-stranded oligonucleotides

[0066] In a first aspect of this disclosure, a double-stranded oligonucleotide for inhibiting URAT1 gene expression is provided, the double-stranded oligonucleotide comprising a sense strand and an antisense strand, the sense strand and the antisense strand being 17 to 30 nucleotides in length, and the sense strand and the antisense strand being at least partially complementary.

[0067] In some embodiments of this disclosure, the antisense strand is 21 to 23 nucleotides in length; the sense strand is 19 to 21 nucleotides in length.

[0068] The antisense strand comprises at least 17 consecutive nucleotides whose first to 19th nucleotides differ by no more than 3 nucleotides from any of the nucleotide sequences shown in SEQ ID NO:70 to SEQ ID NO:138 in Table 3.

[0069] The positive chain comprises at least 17 consecutive nucleotides that differ by no more than 3 nucleotides from any of the nucleotide sequences shown in SEQ ID NO:1 to SEQ ID NO:69 in Table 3.

[0070] In some embodiments of this disclosure, the antisense strand differs from the nucleotides 1-19 of any of the nucleotide sequences shown in SEQ ID NO:70 to SEQ ID NO:138 by no more than 3 nucleotides;

[0071] In some embodiments of this disclosure, the antisense strand contains no more than two nucleotides differing from the nucleotide sequences shown in any of SEQ ID NO:70 to SEQ ID NO:138 at positions 1-19;

[0072] In some embodiments of this disclosure, the antisense strand contains no more than one nucleotide differing from the nucleotide sequences shown in any of SEQ ID NO:70 to SEQ ID NO:138 in positions 1-19;

[0073] In some embodiments of this disclosure, the antisense strand comprises nucleotides 1-19 selected from any of the nucleotide sequences shown in SEQ ID NO:70 to SEQ ID NO:138;

[0074] In some embodiments of this disclosure, the antisense strand is selected from any of the nucleotide sequences shown in SEQ ID NO:70 to SEQ ID NO:138.

[0075] In some embodiments of this disclosure, the sense strand and the antisense strand have a mismatch of no more than 3 nucleotides.

[0076] In some embodiments of this disclosure, the sense strand and the antisense strand have a mismatch of no more than 2 nucleotides.

[0077] In some embodiments of this disclosure, the sense strand and the antisense strand have a mismatch of no more than one nucleotide.

[0078] In some embodiments of this disclosure, the justice chain and the antisense chain are completely complementary.

[0079] In some specific embodiments of this disclosure, the double-stranded oligonucleotide is selected from the sense strand sequence of any of the double strands 1 to 69 shown in Table 3, and the 1st to 19th nucleotide sequences of the antisense strand sequence.

[0080] In some embodiments of this disclosure, the double-stranded oligonucleotide is selected from:

[0081] The double-stranded body 1, wherein the antisense strand sequence of the double-stranded body 1 is shown in SEQ ID NO:70 and the sense strand sequence is shown in SEQ ID NO:1;

[0082] Double strand 2, wherein the antisense strand sequence of the double strand 2 is shown in SEQ ID NO:71 and the sense strand sequence is shown in SEQ ID NO:2;

[0083] The double-stranded body 3, wherein the antisense strand sequence of the double-stranded body 3 is shown in SEQ ID NO:72 and the sense strand sequence is shown in SEQ ID NO:3;

[0084] The double-stranded body 4, wherein the antisense strand sequence of the double-stranded body 4 is shown in SEQ ID NO:73 and the sense strand sequence is shown in SEQ ID NO:4;

[0085] The double-stranded body 5, wherein the antisense strand sequence of the double-stranded body 5 is shown in SEQ ID NO:74 and the sense strand sequence is shown in SEQ ID NO:5;

[0086] The double-stranded body 6, wherein the antisense strand sequence of the double-stranded body 6 is shown in SEQ ID NO:75 and the sense strand sequence is shown in SEQ ID NO:6;

[0087] The double-stranded body 7, wherein the antisense strand sequence of the double-stranded body 7 is shown in SEQ ID NO:76 and the sense strand sequence is shown in SEQ ID NO:7;

[0088] The double-stranded body 8, wherein the antisense strand sequence of the double-stranded body 8 is shown in SEQ ID NO:77 and the sense strand sequence is shown in SEQ ID NO:8;

[0089] The double-stranded body 9, wherein the antisense strand sequence of the double-stranded body 9 is shown in SEQ ID NO:78 and the sense strand sequence is shown in SEQ ID NO:9;

[0090] The double-stranded body 10, wherein the antisense strand sequence of the double-stranded body 10 is shown in SEQ ID NO:79 and the sense strand sequence is shown in SEQ ID NO:10;

[0091] The double-stranded body 11, wherein the antisense chain sequence of the double-stranded body 11 is shown in SEQ ID NO:80 and the sense chain sequence is shown in SEQ ID NO:11;

[0092] The double-stranded body 12, wherein the antisense strand sequence of the double-stranded body 12 is shown in SEQ ID NO:81 and the sense strand sequence is shown in SEQ ID NO:12;

[0093] The double-stranded body 13, wherein the antisense strand sequence of the double-stranded body 13 is shown in SEQ ID NO:82 and the sense strand sequence is shown in SEQ ID NO:13;

[0094] The double-stranded body 14, wherein the antisense strand sequence of the double-stranded body 14 is shown in SEQ ID NO:83 and the sense strand sequence is shown in SEQ ID NO:14;

[0095] The double-stranded body 15, wherein the antisense strand sequence of the double-stranded body 15 is shown in SEQ ID NO:84 and the sense strand sequence is shown in SEQ ID NO:15;

[0096] The double-stranded body 16, wherein the antisense strand sequence of the double-stranded body 16 is shown in SEQ ID NO:85 and the sense strand sequence is shown in SEQ ID NO:16;

[0097] The double-stranded body 17, wherein the antisense strand sequence of the double-stranded body 17 is shown in SEQ ID NO:86 and the sense strand sequence is shown in SEQ ID NO:17;

[0098] The double-stranded body 18, wherein the antisense strand sequence of the double-stranded body 18 is shown in SEQ ID NO:87 and the sense strand sequence is shown in SEQ ID NO:18;

[0099] The double-stranded body 19, wherein the antisense strand sequence of the double-stranded body 19 is shown in SEQ ID NO:88 and the sense strand sequence is shown in SEQ ID NO:19;

[0100] The double-stranded body 20, wherein the antisense strand sequence of the double-stranded body 20 is shown in SEQ ID NO:89 and the sense strand sequence is shown in SEQ ID NO:20;

[0101] The double-stranded body 21, wherein the antisense chain sequence of the double-stranded body 21 is shown in SEQ ID NO:90 and the sense chain sequence is shown in SEQ ID NO:21;

[0102] The double-stranded body 22, wherein the antisense strand sequence of the double-stranded body 22 is shown in SEQ ID NO:91 and the sense strand sequence is shown in SEQ ID NO:22;

[0103] The double-stranded body 23, wherein the antisense strand sequence of the double-stranded body 23 is shown in SEQ ID NO:92 and the sense strand sequence is shown in SEQ ID NO:23;

[0104] The double-stranded body 24, wherein the antisense strand sequence of the double-stranded body 24 is shown in SEQ ID NO:93 and the sense strand sequence is shown in SEQ ID NO:24;

[0105] The double-stranded body 25, wherein the antisense strand sequence of the double-stranded body 25 is shown in SEQ ID NO:94 and the sense strand sequence is shown in SEQ ID NO:25;

[0106] The double-stranded body 26, wherein the antisense strand sequence of the double-stranded body 26 is shown in SEQ ID NO:95 and the sense strand sequence is shown in SEQ ID NO:26;

[0107] The double-stranded body 27, wherein the antisense strand sequence of the double-stranded body 27 is shown in SEQ ID NO:96 and the sense strand sequence is shown in SEQ ID NO:27;

[0108] The double-stranded body 28, wherein the antisense strand sequence of the double-stranded body 28 is shown in SEQ ID NO:97 and the sense strand sequence is shown in SEQ ID NO:28;

[0109] The double-stranded body 29, wherein the antisense strand sequence of the double-stranded body 29 is shown in SEQ ID NO:98 and the sense strand sequence is shown in SEQ ID NO:29;

[0110] The double-stranded body 30, wherein the antisense strand sequence of the double-stranded body 30 is shown in SEQ ID NO:99 and the sense strand sequence is shown in SEQ ID NO:30;

[0111] The double-stranded body 31, wherein the antisense chain sequence of the double-stranded body 31 is shown in SEQ ID NO:100 and the sense chain sequence is shown in SEQ ID NO:31;

[0112] The double-stranded body 32, wherein the antisense strand sequence of the double-stranded body 32 is shown in SEQ ID NO:101 and the sense strand sequence is shown in SEQ ID NO:32;

[0113] The double-stranded body 33, wherein the antisense strand sequence of the double-stranded body 33 is shown in SEQ ID NO:102 and the sense strand sequence is shown in SEQ ID NO:33;

[0114] The double-stranded body 34, wherein the antisense strand sequence of the double-stranded body 34 is shown in SEQ ID NO:103 and the sense strand sequence is shown in SEQ ID NO:34;

[0115] The double-stranded body 35, wherein the antisense strand sequence of the double-stranded body 35 is shown in SEQ ID NO:104 and the sense strand sequence is shown in SEQ ID NO:35;

[0116] The double-stranded body 36, wherein the antisense strand sequence of the double-stranded body 36 is shown in SEQ ID NO:105 and the sense strand sequence is shown in SEQ ID NO:36;

[0117] The double-stranded body 37, wherein the antisense strand sequence of the double-stranded body 37 is shown in SEQ ID NO:106 and the sense strand sequence is shown in SEQ ID NO:37;

[0118] The double-stranded body 38, wherein the antisense strand sequence of the double-stranded body 38 is shown in SEQ ID NO:107 and the sense strand sequence is shown in SEQ ID NO:38;

[0119] The double-stranded body 39, wherein the antisense strand sequence of the double-stranded body 39 is shown in SEQ ID NO:108 and the sense strand sequence is shown in SEQ ID NO:39;

[0120] The double-stranded body 40, wherein the antisense strand sequence of the double-stranded body 40 is shown in SEQ ID NO:109 and the sense strand sequence is shown in SEQ ID NO:40;

[0121] The double-stranded body 41, wherein the antisense strand sequence of the double-stranded body 41 is shown in SEQ ID NO:110 and the sense strand sequence is shown in SEQ ID NO:41;

[0122] The double-stranded body 42, wherein the antisense strand sequence of the double-stranded body 42 is shown in SEQ ID NO:111 and the sense strand sequence is shown in SEQ ID NO:42;

[0123] The double-stranded body 43, wherein the antisense strand sequence of the double-stranded body 43 is shown in SEQ ID NO:112 and the sense strand sequence is shown in SEQ ID NO:43;

[0124] The double-stranded body 44, wherein the antisense strand sequence of the double-stranded body 44 is shown in SEQ ID NO:113 and the sense strand sequence is shown in SEQ ID NO:44;

[0125] The double-stranded body 45, wherein the antisense strand sequence of the double-stranded body 45 is shown in SEQ ID NO:114 and the sense strand sequence is shown in SEQ ID NO:45;

[0126] The double-stranded body 46, wherein the antisense strand sequence of the double-stranded body 46 is shown in SEQ ID NO:115 and the sense strand sequence is shown in SEQ ID NO:46;

[0127] The double-stranded body 47, wherein the antisense strand sequence of the double-stranded body 47 is shown in SEQ ID NO:116 and the sense strand sequence is shown in SEQ ID NO:47;

[0128] The double-stranded body 48, wherein the antisense strand sequence of the double-stranded body 48 is shown in SEQ ID NO:117 and the sense strand sequence is shown in SEQ ID NO:48;

[0129] The double-stranded body 49, wherein the antisense strand sequence of the double-stranded body 49 is shown in SEQ ID NO:118 and the sense strand sequence is shown in SEQ ID NO:49;

[0130] The double-stranded body 50, wherein the antisense strand sequence of the double-stranded body 50 is shown in SEQ ID NO:119 and the sense strand sequence is shown in SEQ ID NO:50;

[0131] The double-stranded body 51, wherein the antisense strand sequence of the double-stranded body 51 is shown in SEQ ID NO:120 and the sense strand sequence is shown in SEQ ID NO:51;

[0132] The double-stranded body 52, wherein the antisense strand sequence of the double-stranded body 52 is shown in SEQ ID NO:121 and the sense strand sequence is shown in SEQ ID NO:52;

[0133] The double-stranded body 53, wherein the antisense strand sequence of the double-stranded body 53 is shown in SEQ ID NO:122 and the sense strand sequence is shown in SEQ ID NO:53;

[0134] The double-stranded body 54, wherein the antisense strand sequence of the double-stranded body 54 is shown in SEQ ID NO:123 and the sense strand sequence is shown in SEQ ID NO:54;

[0135] The double-stranded body 55, wherein the antisense strand sequence of the double-stranded body 55 is shown in SEQ ID NO:124 and the sense strand sequence is shown in SEQ ID NO:55;

[0136] The double-stranded body 56, wherein the antisense strand sequence of the double-stranded body 56 is shown in SEQ ID NO:125 and the sense strand sequence is shown in SEQ ID NO:56;

[0137] The double-stranded body 57, wherein the antisense strand sequence of the double-stranded body 57 is shown in SEQ ID NO:126 and the sense strand sequence is shown in SEQ ID NO:57;

[0138] The double-stranded body 58, wherein the antisense strand sequence of the double-stranded body 58 is shown in SEQ ID NO:127 and the sense strand sequence is shown in SEQ ID NO:58;

[0139] The double-stranded body 59, wherein the antisense strand sequence of the double-stranded body 59 is shown in SEQ ID NO:128 and the sense strand sequence is shown in SEQ ID NO:59;

[0140] The double-stranded body 60, wherein the antisense strand sequence of the double-stranded body 60 is shown in SEQ ID NO:129 and the sense strand sequence is shown in SEQ ID NO:60;

[0141] The double-stranded body 61, wherein the antisense strand sequence of the double-stranded body 61 is shown in SEQ ID NO:130 and the sense strand sequence is shown in SEQ ID NO:61;

[0142] The double-stranded body 62, wherein the antisense strand sequence of the double-stranded body 62 is shown in SEQ ID NO:131 and the sense strand sequence is shown in SEQ ID NO:62;

[0143] The double-stranded body 63, wherein the antisense strand sequence of the double-stranded body 63 is shown in SEQ ID NO:132 and the sense strand sequence is shown in SEQ ID NO:63;

[0144] The double-stranded body 64, wherein the antisense strand sequence of the double-stranded body 64 is shown in SEQ ID NO:133 and the sense strand sequence is shown in SEQ ID NO:64;

[0145] The double-stranded body 65, wherein the antisense strand sequence of the double-stranded body 65 is shown in SEQ ID NO:134 and the sense strand sequence is shown in SEQ ID NO:65;

[0146] The double-stranded body 66, wherein the antisense strand sequence of the double-stranded body 66 is shown in SEQ ID NO:135 and the sense strand sequence is shown in SEQ ID NO:66;

[0147] The double-stranded body 67, wherein the antisense strand sequence of the double-stranded body 67 is shown in SEQ ID NO:136 and the sense strand sequence is shown in SEQ ID NO:67;

[0148] The double-stranded body 68, wherein the antisense strand sequence of the double-stranded body 68 is shown in SEQ ID NO:137 and the sense strand sequence is shown in SEQ ID NO:68;

[0149] The double strand 69, wherein the antisense strand sequence of the double strand 69 is shown in SEQ ID NO:138 and the sense strand sequence is shown in SEQ ID NO:69.

[0150] In some embodiments of this disclosure, the double-stranded oligonucleotide contains at least one modified nucleotide.

[0151] In some embodiments of this disclosure, each nucleotide in the double-stranded oligonucleotide is a modified nucleotide.

[0152] In some embodiments of this disclosure, the modified nucleotide is selected from: 2'-halogenated nucleotides, 2'-deoxygenated nucleotides, 2'-O-alkyl-modified nucleotides, and 2'-O-(CH2). n -OMe modified nucleotides (n selected from 1 or 2), 2'-amino modified nucleotides, nucleotides modified with 2',2'-[halogen, halogen] disubstituted or 2',2'-[halogen, alkyl] disubstituted groups, debased nucleotides, or nucleotide-like nucleotides.

[0153] In some embodiments of this disclosure, the modified nucleotide is selected from: 2'-fluoro-modified nucleotides, 2'-deoxy-modified nucleotides, 2'-O-methyl-modified nucleotides, 2'-O-methoxyethyl-modified nucleotides, and 2',2'-[fluoro,methyl]di-substituent-modified nucleotides.

[0154] In some embodiments of this disclosure, the nucleotide modified with 2',2'-[fluoro,methyl] disubstituents is located at the overhang (or 3'-terminus) of the antisense strand.

[0155] For example, when the antisense strand is 21 nucleotides long, at least one nucleotide modified with a 2',2'-[fluoro,methyl] disubstituent is located at position 20-21 of the antisense strand; preferably, positions 20-21 of the antisense strand are all nucleotides modified with a 2',2'-[fluoro,methyl] disubstituent.

[0156] In some embodiments of this disclosure, the antisense strand of the double-stranded oligonucleotide is 21 nucleotides long, the sense strand is 19 nucleotides long, and each nucleotide in the double-stranded oligonucleotide is a modified nucleotide; wherein:

[0157] In the direction from the 5' end to the 3' end, at least four nucleotides at positions 2, 6, 9, 14, and 16 of the antisense strand are selected from 2'-F modified nucleotides, and the nucleotides at the remaining positions are selected from 2'-methoxy modified nucleotides or 2'-O-methoxyethyl modified nucleotides; at least three nucleotides at positions 7-10 of the sense strand are selected from 2'-F modified nucleotides, and the remaining positions are 2'-methoxy modified nucleotides.

[0158] In some embodiments of this disclosure, the nucleotides at positions 2, 6, 9, 14, and 16 of the antisense strand, in the direction from the 5' end to the 3' end, are selected from 2'-F modified nucleotides, the nucleotide at position 15 is selected from 2'-O-methoxyethyl modified nucleotides or 2'-methoxy modified nucleotides, and the nucleotides at the remaining positions are 2'-methoxy modified nucleotides or other modified nucleotides besides the above modifications; the nucleotides at positions 7-10 of the sense strand are selected from 2'-F modified nucleotides, and the nucleotides at the remaining positions are selected from 2'-methoxy modified nucleotides or other modified nucleotides besides the above modifications.

[0159] In some embodiments of this disclosure, the nucleotides at positions 2, 6, 9, 14, and 16 of the antisense strand are selected from 2'-F modified nucleotides, the nucleotide at position 15 is selected from 2'-O-methoxyethyl modified nucleotides, and the nucleotides at the remaining positions are 2'-methoxy modified nucleotides; the nucleotides at positions 7-10 of the sense strand are selected from 2'-F modified nucleotides, and the nucleotides at the remaining positions are selected from 2'-methoxy modified nucleotides.

[0160] In some embodiments of this disclosure, the double-stranded oligonucleotide has a modified phosphate group selected from thiophosphate groups, and the thiophosphate group is present at at least one of the following positions: 1) between the first and second nucleotides at the 5' end of the sense strand, and / or between the second and third nucleotides; 2) between the first and second nucleotides at the 3' end of the sense strand, and / or between the second and third nucleotides; 3) between the first and second nucleotides at the 5' end of the antisense strand, and / or between the second and third nucleotides; 4) between the first and second nucleotides at the 3' end of the antisense strand, and / or between the second and third nucleotides, and / or between the third and fourth nucleotides; 5) between the tenth and eleventh nucleotides at the 5' end of the antisense strand.

[0161] In some embodiments of this disclosure, the antisense strand comprises at least 17 consecutive nucleotides whose first nucleotides differ by no more than 3 nucleotides from the nucleotide sequences shown in any of SEQ ID NO:208 to SEQ ID NO:276 in Table 4; and the sense strand comprises at least 17 consecutive nucleotides whose nucleotide sequences differ by no more than 3 nucleotides from the nucleotide sequences shown in any of SEQ ID NO:139 to SEQ ID NO:207 in Table 4.

[0162] In some embodiments of this disclosure, the antisense strand contains no more than three nucleotides different from the nucleotides at positions 1-19 of any of the nucleotide sequences shown in SEQ ID NO:208 to SEQ ID NO:276.

[0163] In some embodiments of this disclosure, the antisense strand contains no more than two nucleotides differing from the nucleotide sequences shown in any of SEQ ID NO:208 to SEQ ID NO:276 at positions 1-19.

[0164] In some embodiments of this disclosure, the antisense strand contains no more than one nucleotide from positions 1 to 19 that differ from the nucleotide sequences shown in any of SEQ ID NO:208 to SEQ ID NO:276.

[0165] In some embodiments of this disclosure, the antisense strand comprises nucleotides 1-19 of any of the nucleotide sequences shown in SEQ ID NO:208 to SEQ ID NO:276.

[0166] In some embodiments of this disclosure, the antisense strand is selected from any of the nucleotide sequences shown in SEQ ID NO:SEQ ID NO:208 to SEQ ID NO:276.

[0167] In some embodiments of this disclosure, the double-stranded oligonucleotide is selected from the sense strand of any of the modified double strands 1 to 69 shown in Table 4, and nucleotides 1 to 19 of the antisense strand sequence.

[0168] In some embodiments of this disclosure, the modified double-stranded oligonucleotide is selected from:

[0169] The modified double strand 1 has an antisense strand sequence as shown in SEQ ID NO:208 and a sense strand sequence as shown in SEQ ID NO:139.

[0170] The modified double strand 2 has an antisense strand sequence as shown in SEQ ID NO:209 and a sense strand sequence as shown in SEQ ID NO:140.

[0171] The modified double strand 3 has an antisense strand sequence as shown in SEQ ID NO:210 and a sense strand sequence as shown in SEQ ID NO:141.

[0172] Modified double strand 4, wherein the antisense strand sequence of the modified double strand 4 is shown in SEQ ID NO:211 and the sense strand sequence is shown in SEQ ID NO:142;

[0173] The modified double strand 5 has an antisense strand sequence as shown in SEQ ID NO:212 and a sense strand sequence as shown in SEQ ID NO:143.

[0174] The modified double strand 6 has an antisense strand sequence as shown in SEQ ID NO:213 and a sense strand sequence as shown in SEQ ID NO:144.

[0175] The modified double strand 7, wherein the antisense strand sequence of the modified double strand 7 is shown in SEQ ID NO:214 and the sense strand sequence is shown in SEQ ID NO:145;

[0176] The modified double strand 8 has an antisense strand sequence as shown in SEQ ID NO:215 and a sense strand sequence as shown in SEQ ID NO:146.

[0177] The modified double strand 9 has an antisense strand sequence as shown in SEQ ID NO:216 and a sense strand sequence as shown in SEQ ID NO:147.

[0178] The modified double strand 10 has an antisense strand sequence as shown in SEQ ID NO:217 and a sense strand sequence as shown in SEQ ID NO:148.

[0179] The modified double strand 11 has an antisense strand sequence as shown in SEQ ID NO:218 and a sense strand sequence as shown in SEQ ID NO:149.

[0180] The modified double strand 12 has an antisense strand sequence as shown in SEQ ID NO:219 and a sense strand sequence as shown in SEQ ID NO:150.

[0181] The modified double strand 13 has an antisense strand sequence as shown in SEQ ID NO:220 and a sense strand sequence as shown in SEQ ID NO:151.

[0182] The modified double strand 14 has an antisense strand sequence as shown in SEQ ID NO:221 and a sense strand sequence as shown in SEQ ID NO:152.

[0183] The modified double strand 15 has an antisense strand sequence as shown in SEQ ID NO:222 and a sense strand sequence as shown in SEQ ID NO:153.

[0184] The modified double strand 16 has an antisense strand sequence as shown in SEQ ID NO:223 and a sense strand sequence as shown in SEQ ID NO:154.

[0185] The modified double strand 17 has an antisense strand sequence as shown in SEQ ID NO:224 and a sense strand sequence as shown in SEQ ID NO:155.

[0186] The modified double strand 18 has an antisense strand sequence as shown in SEQ ID NO:225 and a sense strand sequence as shown in SEQ ID NO:156.

[0187] The modified double strand 19 has an antisense strand sequence as shown in SEQ ID NO:226 and a sense strand sequence as shown in SEQ ID NO:157.

[0188] The modified double strand 20 has an antisense strand sequence as shown in SEQ ID NO:227 and a sense strand sequence as shown in SEQ ID NO:158.

[0189] The modified double strand 21 has an antisense strand sequence as shown in SEQ ID NO:228 and a sense strand sequence as shown in SEQ ID NO:159.

[0190] The modified double strand 22 has an antisense strand sequence as shown in SEQ ID NO:229 and a sense strand sequence as shown in SEQ ID NO:160.

[0191] The modified double strand 23 is shown in SEQ ID NO:230 and the positive strand sequence is shown in SEQ ID NO:161.

[0192] The modified double strand 24 has an antisense strand sequence as shown in SEQ ID NO:231 and a sense strand sequence as shown in SEQ ID NO:162.

[0193] The modified double strand 25 has an antisense strand sequence as shown in SEQ ID NO:232 and a sense strand sequence as shown in SEQ ID NO:163.

[0194] The modified double strand 26 has an antisense strand sequence as shown in SEQ ID NO:233 and a sense strand sequence as shown in SEQ ID NO:164.

[0195] The modified double strand 27 has an antisense strand sequence as shown in SEQ ID NO:234 ​​and a sense strand sequence as shown in SEQ ID NO:165.

[0196] The modified double strand 28 has an antisense strand sequence as shown in SEQ ID NO:235 and a sense strand sequence as shown in SEQ ID NO:166.

[0197] The modified double strand 29 has an antisense strand sequence as shown in SEQ ID NO:236 and a sense strand sequence as shown in SEQ ID NO:167.

[0198] The modified double strand 30 has an antisense strand sequence as shown in SEQ ID NO:237 and a sense strand sequence as shown in SEQ ID NO:168.

[0199] The modified double strand 31 has an antisense strand sequence as shown in SEQ ID NO:238 and a sense strand sequence as shown in SEQ ID NO:169.

[0200] The modified double strand 32 has an antisense strand sequence as shown in SEQ ID NO:239 and a sense strand sequence as shown in SEQ ID NO:170.

[0201] The modified double strand 33 has an antisense strand sequence as shown in SEQ ID NO:240 and a sense strand sequence as shown in SEQ ID NO:171.

[0202] The modified double strand 34 has an antisense strand sequence as shown in SEQ ID NO:241 and a sense strand sequence as shown in SEQ ID NO:172.

[0203] The modified double strand 35 has an antisense strand sequence as shown in SEQ ID NO:242 and a sense strand sequence as shown in SEQ ID NO:173.

[0204] The modified double strand 36 has an antisense strand sequence as shown in SEQ ID NO:243 and a sense strand sequence as shown in SEQ ID NO:174.

[0205] The modified double strand 37 has an antisense strand sequence as shown in SEQ ID NO:244 and a sense strand sequence as shown in SEQ ID NO:175.

[0206] The modified double strand 38 has an antisense strand sequence as shown in SEQ ID NO:245 and a sense strand sequence as shown in SEQ ID NO:176.

[0207] The modified double strand 39 has an antisense strand sequence as shown in SEQ ID NO:246 and a sense strand sequence as shown in SEQ ID NO:177.

[0208] The modified double strand 40 has an antisense strand sequence as shown in SEQ ID NO:247 and a sense strand sequence as shown in SEQ ID NO:178.

[0209] The modified double strand 41 has an antisense strand sequence as shown in SEQ ID NO:248 and a sense strand sequence as shown in SEQ ID NO:179.

[0210] The modified double strand 42 has an antisense strand sequence as shown in SEQ ID NO:249 and a sense strand sequence as shown in SEQ ID NO:180.

[0211] The modified double strand 43 has an antisense strand sequence as shown in SEQ ID NO:250 and a sense strand sequence as shown in SEQ ID NO:181.

[0212] The modified double strand 44 has an antisense strand sequence as shown in SEQ ID NO:251 and a sense strand sequence as shown in SEQ ID NO:182.

[0213] The modified double strand 45 has an antisense strand sequence as shown in SEQ ID NO:252 and a sense strand sequence as shown in SEQ ID NO:183.

[0214] The modified double strand 46 has an antisense strand sequence as shown in SEQ ID NO:253 and a sense strand sequence as shown in SEQ ID NO:184.

[0215] The modified double strand 47 has an antisense strand sequence as shown in SEQ ID NO:254 and a sense strand sequence as shown in SEQ ID NO:185.

[0216] The modified double strand 48 has an antisense strand sequence as shown in SEQ ID NO:255 and a sense strand sequence as shown in SEQ ID NO:186.

[0217] The modified double strand 49 has an antisense strand sequence as shown in SEQ ID NO:256 and a sense strand sequence as shown in SEQ ID NO:187.

[0218] The modified double strand 50 has an antisense strand sequence as shown in SEQ ID NO:257 and a sense strand sequence as shown in SEQ ID NO:188.

[0219] The modified double strand 51 has an antisense strand sequence as shown in SEQ ID NO:258 and a sense strand sequence as shown in SEQ ID NO:189.

[0220] The modified double strand 52 has an antisense strand sequence as shown in SEQ ID NO:259 and a sense strand sequence as shown in SEQ ID NO:190.

[0221] The modified double strand 53 has an antisense strand sequence as shown in SEQ ID NO:260 and a sense strand sequence as shown in SEQ ID NO:191.

[0222] The modified double strand 54 has an antisense strand sequence as shown in SEQ ID NO:261 and a sense strand sequence as shown in SEQ ID NO:192.

[0223] The modified double strand 55 has an antisense strand sequence as shown in SEQ ID NO:262 and a sense strand sequence as shown in SEQ ID NO:193.

[0224] The modified double strand 56 has an antisense strand sequence as shown in SEQ ID NO:263 and a sense strand sequence as shown in SEQ ID NO:194.

[0225] The modified double strand 57 has an antisense strand sequence as shown in SEQ ID NO:264 and a sense strand sequence as shown in SEQ ID NO:195.

[0226] The modified double strand 58 has an antisense strand sequence as shown in SEQ ID NO:265 and a sense strand sequence as shown in SEQ ID NO:196.

[0227] The modified double strand 59 has an antisense strand sequence as shown in SEQ ID NO:266 and a sense strand sequence as shown in SEQ ID NO:197.

[0228] The modified double strand 60 has an antisense strand sequence as shown in SEQ ID NO:267 and a sense strand sequence as shown in SEQ ID NO:198.

[0229] The modified double strand 61 has an antisense strand sequence as shown in SEQ ID NO:268 and a sense strand sequence as shown in SEQ ID NO:199.

[0230] The modified double strand 62 has an antisense strand sequence as shown in SEQ ID NO:269 and a sense strand sequence as shown in SEQ ID NO:200.

[0231] The modified double strand 63 has an antisense strand sequence as shown in SEQ ID NO:270 and a sense strand sequence as shown in SEQ ID NO:201.

[0232] The modified double strand 64 has an antisense strand sequence as shown in SEQ ID NO:271 and a sense strand sequence as shown in SEQ ID NO:202.

[0233] The modified double strand 65 has an antisense strand sequence as shown in SEQ ID NO:272 and a sense strand sequence as shown in SEQ ID NO:203.

[0234] The modified double strand 66 has an antisense strand sequence as shown in SEQ ID NO:273 and a sense strand sequence as shown in SEQ ID NO:204.

[0235] The modified double strand 67 has an antisense strand sequence as shown in SEQ ID NO:274 and a sense strand sequence as shown in SEQ ID NO:205.

[0236] The modified double strand 68 has an antisense strand sequence as shown in SEQ ID NO:275 and a sense strand sequence as shown in SEQ ID NO:206.

[0237] The modified double strand 69 has an antisense strand sequence as shown in SEQ ID NO:276 and a sense strand sequence as shown in SEQ ID NO:207.

[0238] Oligonucleotide conjugates

[0239] In a second aspect of this disclosure, an oligonucleotide conjugate is provided, comprising the double-stranded oligonucleotide described in the first aspect of this disclosure and a targeting ligand group.

[0240] In some embodiments of this disclosure, the targeting ligand group is a polypeptide.

[0241] In some embodiments of this disclosure, the targeting ligand group includes any of the polypeptide fragments shown in 1)-6) below:

[0242] 1)-X1(KKEEE) n -K m - where n = 1-5, m = 0 or 1 or 2, and X1 is selected from any L-α amino acid or L-β amino acid other than K and E;

[0243] 2)-CLPVASC-, preferably in cyclic peptide form

[0244] 3)-CYFQNC-; Preferred cyclic peptide form

[0245] 4)-KIDRI-;

[0246] 5)-IDRI-(-Ile-Asp-Arg-Ile-);

[0247] 6)-dXa-Ser-dXb-X2-dXc-Gly-Xd-Ile-Asp-Arg(Ak)-Ile-; where X2 is selected from any non-natural amino acid; dXa, dXb, dXc are selected from any D-type amino acid; Arg(Ak) is selected from arginine or alkylated arginine; and Xd is selected from Hyp or Pro.

[0248] In some embodiments of this disclosure, the targeting ligand group is selected from polypeptides comprising any of the amino acid sequence segments shown in a)-g).

[0249] a)-KKEEE-KKEEE-KKEEE-K-;;

[0250] b)-CKKEEE-KKEEE-KKEEE-K;

[0251] c)-CLPVASC-, preferably in cyclic peptide form

[0252] d)-KIDRI-;

[0253] e)-CYFQNCPRG-, preferably in cyclic peptide form

[0254] f)-DPhe-Ser-DHyp-Cha-DAla-Gly-Hyp-Ile-Asp-Arg(Me)-Ile-;

[0255] g)-DPhe-Ser-DPro-Cha-DAla-Gly-Pro-Ile-Asp-Arg-Ile-.

[0256] In some embodiments of this disclosure, the targeting ligand group is connected to the double-stranded oligonucleotide described in the first aspect of this disclosure via a linker.

[0257] In some embodiments of this disclosure, the structural unit of the targeting ligand group-linker in the oligonucleotide conjugate is selected from any of the structures shown in 1)-7) below:

[0258] 1)La-KKEEE-KKEEE-KKEEE-K-Laa;

[0259] 2)La-CKKEEE-KKEEE-KKEEE-K-Laa;

[0260] 3) La-CLPVASC-Laa; preferably a cyclic peptide.

[0261] 4)La-DPhe-Ser-DHyp-Cha-DAla-Gly-Hyp-Ile-Asp-Arg(Me)-Ile-Laa;

[0262] 5)La-CYFQNCPRG-Laa;

[0263] 6)La-DPhe-Ser-DPro-Cha-DAla-Gly-Pro-Ile-Asp-Arg-Ile-Laa;

[0264] 7)Lb-KIDRI-Laa.

[0265] In formulas 1)-6), either La or Laa is a linker and the other is a terminal blocking group, wherein the linker is a linking group containing at least one of a triazole group, a PEG unit, or an acyl group.

[0266] In some embodiments of this disclosure, La is a connector, the connector structure including -PEG2-CH2CH2CO- or triazole-PEG2-.

[0267] In formula 7), Lb is a linker containing a 6-10 membered aromatic ring or heteroaromatic ring, and Laa is independently selected from an amino group or an alkyl-substituted amino group, such as -NHCH3.

[0268] In some embodiments of this disclosure, the targeting ligand group is connected to the ends of the sense and / or antisense strands of the double-stranded oligonucleotide described in the first aspect of this disclosure via a linker.

[0269] In some embodiments of this disclosure, the structure of the targeting ligand group-linker in the double-stranded oligonucleotide conjugate is selected from any of the structures shown in a)-j), or a pharmaceutically acceptable salt thereof:

[0270] a)-La-KKEEE-KKEEE-KKEEE-K-Lc;

[0271] b)-La-CKKEEE-KKEEE-KKEEE-K-Lc;

[0272] c)-La-CLPVASC-Lc; preferably cyclic peptide form

[0273] d)-La-DPhe-Ser-DHyp-Cha-DAla-Gly-Hyp-Ile-Asp-Arg(Me)-Ile-Lc;

[0274] e)-La-CYFQNCPRG-Lc; preferably a cyclic peptide

[0275] f)-La-DPhe-Ser-DPro-Cha-DAla-Gly-Pro-Ile-Asp-Arg-Ile-Lc;

[0276]

[0277] Wherein, La is a linker, which is independently selected from any bond, -NH-, amide group or linking group containing at least one PEG unit.

[0278] In some embodiments of this disclosure, La is a linking group comprising at least two PEG units.

[0279] In some embodiments of this disclosure, La is selected from amide groups. n is 1-3, m is 0 or 1, and Z is -CO- (carbonyl) or -NH-.

[0280] Wherein, Lc is the terminal blocking group.

[0281] In some embodiments of this disclosure, Lc is a C-terminal blocking group.

[0282] In some embodiments of this disclosure, the Lc is selected from amino or alkyl-substituted amino groups.

[0283] In some embodiments of this disclosure, Lc is -NHCH3.

[0284] Pharmaceutical Composition

[0285] In a third aspect of this disclosure, a pharmaceutical composition is provided comprising the double-stranded oligonucleotides described in the first aspect of this disclosure and / or the double-stranded oligonucleotide conjugates described in the second aspect of this disclosure.

[0286] In some embodiments of this disclosure, the pharmaceutical composition further includes a pharmaceutically acceptable carrier or excipient.

[0287] Uses of inhibiting URAT1 gene expression

[0288] In a fourth aspect of this disclosure, the present disclosure provides the use of the double-stranded oligonucleotides described in the first aspect of this disclosure and / or the double-stranded oligonucleotide conjugates described in the second aspect of this disclosure and / or the pharmaceutical compositions described in the third aspect of this disclosure in the preparation of a medicament for inhibiting URAT1 gene expression.

[0289] Uses in treating diseases

[0290] In a fifth aspect of this disclosure, the present disclosure provides the use of the double-stranded oligonucleotides described in the first aspect of this disclosure and / or the double-stranded oligonucleotide conjugates described in the second aspect of this disclosure and / or the pharmaceutical compositions described in the third aspect of this disclosure in the preparation of a medicament for treating and / or preventing pathological conditions or diseases associated with URAT1 expression.

[0291] In some embodiments of this disclosure, the pathological condition or disease is hyperuricemia or gout.

[0292] Methods to inhibit intracellular URAT1 expression

[0293] In a sixth aspect of this disclosure, this disclosure provides a method for inhibiting intracellular URAT1 expression, comprising administering to a subject the double-stranded oligonucleotides described in the first aspect of this disclosure and / or the double-stranded oligonucleotide conjugates described in the second aspect of this disclosure and / or the pharmaceutical compositions described in the third aspect of this disclosure.

[0294] In some embodiments of this disclosure, the subject is a human being.

[0295] Treatment methods for diseases

[0296] In a seventh aspect of this disclosure, this disclosure provides a method for preventing and / or treating hyperuricemia or gout, the method comprising administering to a subject an effective amount of the double-stranded oligonucleotides described in the first aspect of this disclosure and / or the double-stranded oligonucleotide conjugates described in the second aspect of this disclosure and / or the pharmaceutical compositions described in the third aspect of this disclosure.

[0297] In some embodiments of this disclosure, the subject is a human being.

[0298] The effective amount of the double-stranded oligonucleotides, double-stranded oligonucleotide conjugates, or pharmaceutical compositions described in this disclosure may vary depending on the administration method and the severity of the disease to be treated. Specifically, the effective amount can be determined by those skilled in the art based on various factors (e.g., through clinical trials). These factors include, but are not limited to: pharmacokinetic parameters of the active ingredient, such as bioavailability, metabolism, and half-life; the severity of the disease to be treated, the patient's weight, the patient's immune status, and the route of administration.

[0299] The medication may be administered to the subject via any suitable route known in the art, including but not limited to: oral or parenteral routes, including intravenous administration, intramuscular administration, subcutaneous administration, transdermal administration, airway administration (aerosol), pulmonary administration, nasal administration, rectal administration, and local administration (including oral and sublingual administration), such as intravenous administration.

[0300] The double-stranded oligonucleotides, double-stranded oligonucleotide conjugates, and pharmaceutical compositions disclosed herein can effectively inhibit URAT1 gene expression. The oligonucleotide conjugates and pharmaceutical compositions disclosed herein may help treat and / or prevent pathological conditions or diseases associated with URAT1 expression.

[0301] To make the objectives, technical solutions, and advantages of this disclosure clearer, the embodiments of this disclosure will be described in further detail below with reference to examples.

[0302] siRNA synthesis

[0303] Unless otherwise stated, all reagents, reagent consumables, and instruments used in this disclosure are commercially available. The main reagents and consumables are shown in Table 1, and the main instruments and equipment are shown in Table 2.

[0304] Table 1 Main Reagents and Consumables

[0305]

[0306]

[0307] Table 2 Main Instruments and Equipment

[0308] name Manufacturer High-speed refrigerated centrifuge Eppendorf NANODROP OneC Thermo Fisher Scientific Gradient PCR Amplification Instrument Eppendorf Real-time PCR instrument ABI StepOne Plus Inverted microscope OLYMPUS Biosafety cabinet ESCO Cell incubator Thermo Scientific Gel Imaging Shanghai Tianneng Life Science Co., Ltd. Electrophoresis apparatus Beijing Liuyi Instrument Factory

[0309] siRNA sequence design

[0310] The disclosed siRNA sequences target all transcripts and predicted transcripts of the URAT1 gene (NM_144585.4, NM_153378.3, NM_001276326.2, NM_001276327.2, XM_006718430.5, XM_006718431.5, XM_054367604.1, XM_054367605.1). The URAT1 gene sequence information is obtained from the NCBI gene database (https: / / www.ncbi.nlm.nih.gov / gene / ).

[0311] Preparation Example 1: Synthesis of siRNA

[0312] (1-1) Synthesis of the Chain of Justice (SS)

[0313] The phosphoramidite solid-phase synthesis method involves sequentially linking nucleoside monomers along the 3'-5' direction according to the nucleotide sequence. Each linking of a nucleoside monomer involves four steps: deprotection, coupling, capping, and oxidation or sulfidation. The synthetic conditions are given below:

[0314] The nucleoside monomer was prepared into an acetonitrile solution with a concentration of 0.1 M.

[0315] The deprotection reaction conditions were the same for each step. The deprotection reaction conditions were: temperature 25℃, reaction time 70 seconds, deprotection reagent was a dichloromethane solution of dichloroacetic acid (3% by volume), and the molar ratio of dichloroacetic acid to the 4,4'-dimethoxytriphenylmethyl protecting group on the solid support was 5:1.

[0316] The conditions for each coupling reaction were identical. The coupling reaction conditions were as follows: temperature 25℃, molar ratio of nucleic acid sequence to nucleoside monomer on the solid-phase support 1:10, molar ratio of nucleic acid sequence to coupling reagent on the solid-phase support 1:65, reaction time 600 seconds, coupling reagent 0.5M acetonitrile solution of 5-ethylthio-1H-tetrazole, and thioreagent 0.2mol / L acetonitrile / pyridine mixed solution of hydrogenated xanthanin (acetonitrile and pyridine volume ratio 1:1).

[0317] The conditions for each capping reaction were identical. The conditions for the capping reaction were: temperature 25℃; reaction time 2 minutes; the capping reagent solution was a 1:1 molar ratio of Cap1 and Cap2, where Cap1 was a 20% (v / v) N-methylimidazole pyridine / acetonitrile mixture with a pyridine to acetonitrile volume ratio of 3:5, and Cap2 was a 20% (v / v) acetic anhydride acetonitrile solution; the molar ratio of N-methylimidazole in Cap1 and acetic anhydride in Cap2 to the nucleic acid sequence linked on the solid-phase support was 1:1:1.

[0318] The conditions for each oxidation reaction were identical. The oxidation reaction conditions were: temperature 25°C; reaction time 3 seconds; oxidizing agent concentration of 0.05M iodine solution, with a molar ratio of iodine to the nucleic acid sequence linked on the solid support in the coupling reaction of 30:1; the oxidation reaction was carried out in a water / pyridine mixed solvent (water to pyridine volume ratio 1:9). The sulfidation reaction conditions were: temperature 25°C; reaction time 360 ​​seconds; thioreagent concentration of 0.2M hydroflavin in pyridine solution, with a molar ratio of thioreagent to the nucleic acid sequence linked on the solid support in the coupling reaction of 4:1; the thioreagent reaction was carried out in a water / pyridine mixed solvent (water to pyridine volume ratio 1:9).

[0319] After the last nucleoside monomer was ligated, the nucleic acid sequence ligated on the solid-phase support was sequentially cut, deprotected, purified, and desalted, and then freeze-dried to obtain the positive strand, wherein:

[0320] The cleavage and deprotection conditions were as follows: The synthesized nucleotide sequence linked to a solid-phase support was added to 25% (w / w) ammonia solution at a concentration of 0.5 mL / μmol. The reaction was carried out at 55 °C for 16 hours. The solvent was removed, and the solution was concentrated to dryness under vacuum. After ammonia treatment, the product was dissolved in 0.4 mL / μmol N-methylpyrrolidone relative to the amount of single-stranded nucleic acid. Subsequently, 0.3 mL / μmol triethylamine and 0.6 mL / μmol triethylamine trifluoride were added to deprotect the 2'-O-TBDMS protection on the ribose.

[0321] Purification and desalting conditions: Nucleic acid purification was performed using a preparative ion chromatography column (Source 15Q) with a NaCl gradient elution. Specifically: eluent 1 was 20 mM sodium phosphate (pH = 8.1), and the solvent was a water / acetonitrile mixture (water to acetonitrile volume ratio of 9:1); eluent 2 was 1.5 M sodium chloride, 20 mM sodium phosphate (pH = 8.1), and the solvent was a water / acetonitrile mixture (water to acetonitrile volume ratio of 9:1); the elution gradient was eluent 1: eluent 2 = (100:0) - (50:50). The product eluates were collected and combined, and desalting was performed using a reverse chromatographic purification column. Desalting conditions included using a dextran gel column (g25 dextran gel) and elution with deionized water.

[0322] Detection: Purity was determined using ion exchange chromatography (IEX-HPLC); molecular weight was determined using liquid chromatography-mass spectrometry (LC-MS). The measured molecular weight was compared with the theoretical value. If the measured value and the theoretical value were consistent, it indicated that the siRNA positive strand had been obtained.

[0323] (1-2) Synthesis of antisense strand (AS)

[0324] The phosphoramidite solid-phase synthesis method involves linking nucleoside monomers one by one along the 3'-5' direction according to the nucleotide sequence. Each linking of a nucleoside monomer includes four steps: deprotection, coupling, capping, and oxidation or sulfidation. The deprotection, coupling, capping, oxidation or sulfidation reaction conditions, cleavage and deprotection conditions, purification and desalting conditions in the solid-phase synthesis of the antisense strand are the same as those in step (1-1) for the synthesis of the sense strand.

[0325] Detection: Purity was determined using ion exchange chromatography (IEX-HPLC); molecular weight was determined using liquid chromatography-mass spectrometry (LC-MS). The measured molecular weight was compared with the theoretical value. If the measured value and the theoretical value were consistent, it indicated that the siRNA antisense strand had been obtained.

[0326] (1-3) Synthesis of siRNA

[0327] The sense strand synthesized in step (1-1) and the antisense strand synthesized in step (1-2) were mixed in an equimolar ratio, dissolved in water for injection, and heated to 95°C. The mixture was then slowly cooled to room temperature and kept at room temperature for 10 minutes to allow the sense and antisense strands to form a double-stranded structure through hydrogen bonds, thereby obtaining siRNA with the sense and antisense strands shown in Table 4.

[0328] Table 3. Nucleotide sequence information of unmodified siRNA

[0329]

[0330]

[0331]

[0332] Table 4. Modified siRNA sequence information

[0333]

[0334]

[0335]

[0336]

[0337]

[0338] Unless otherwise specified, the base composition and modifications described in this disclosure have the following meanings: uppercase letters A, U, G, C, and T represent the base composition of nucleotides; lowercase letter m indicates that the nucleotide adjacent to the left of letter m is a nucleotide modified with 2'-O-methyl; lowercase letter f indicates that the nucleotide adjacent to the left of letter f is a nucleotide modified with 2'-fluoro; (moe) indicates that the nucleotide adjacent to the left of (moe) is a nucleotide modified with 2'-O-methoxyethyl; and lowercase letter s indicates that the two nucleotides adjacent to the left and right of letter s are connected by a phosphate thioester bond.

[0339] The structural formula of the nucleotide modified with 2'-O-methyl is

[0340] The structural formula of the 2'-fluorinated nucleotide is

[0341] The structural formula of the nucleotide modified with 2'-O-methoxyethyl is:

[0342] Where Base represents the nucleobases A, U, G, C, and T. Q represents a hydroxyl or thiol group.

[0343] Biological testing experiments

[0344] Unless otherwise stated, all siRNA sequences used in this disclosure were synthesized by Suzhou Xuanjing Biotechnology Co., Ltd.; all PCR primers used in this disclosure were synthesized by Beijing Qingke Biotechnology Co., Ltd.

[0345] in vitro activity assessment of siRNA sequences

[0346] Vero cells (gifted by Beijing Institute of Technology) were cultured in complete medium (DMEM medium containing 10% fetal bovine serum and 1% penicillin-dextrose antibiotics) at 5% CO2 and 37°C. After reaching 80% confluence, the cells were digested with trypsin and resuspended. The resuspended cells were then adjusted to a density of 0.6 × 10⁻⁶ cells / mL using complete medium. 5 / wells were inoculated into 24-well plates and transfected after 24 hours of incubation.

[0347] Preparation of siRNA transfection mixture: Prepare a 20 μM stock solution of siRNA with PBS. Then, add 1.5 μL of the siRNA stock solution to each well and dilute it with 48.5 μL of Opti-MEM medium to prepare 50 μL of siRNA working solution. Add 3 μL of the stock solution to each well. The RNAiMAX transfection reagent was diluted in 47 μl of Opti-MEM medium and incubated at room temperature for 5 minutes to prepare a 50 μl working solution for transfection. Then, 50 μl of siRNA working solution was mixed with the 50 μl working solution of the transfection reagent to prepare the siRNA transfection mixture. For the MOCK group, 50 μl of Opti-MEM medium was mixed with 50 μl of the working solution of the transfection reagent.

[0348] On the day of transfection, replace 500 μl / well of Opti-MEM medium in each well of the 24-well plate, and add 100 μl of the corresponding siRNA transfection mixture or MOCK group transfection mixture to each well, with a final siRNA concentration of 50 nM / well. After culturing for another 24 hours, extract total RNA using a chemical extraction method. Specifically, after removing the medium from each well, add 1 mL of Trizol solution, lyse the cells by pipetting, transfer the lysate to a 1.5 mL centrifuge tube, add 200 μL of chloroform, vortex to mix, and incubate at room temperature for 3 min. Centrifuge at 12000 rpm for 10 min at 4°C. Add 400 μL of the supernatant to a centrifuge tube containing 400 μL of isopropanol, mix, and incubate at room temperature for 10 min. Centrifuge at 12000 rpm for 10 min at 4°C, and discard the supernatant. Add 1 mL of 75% ethanol, invert the centrifuge tube, and wash the precipitate. Centrifuge at 12000 rpm for 5 min at 4°C, remove the supernatant, air dry at room temperature, and extract total RNA.

[0349] Take 1 μg of total RNA, and use a reverse transcription kit (Promega, Reverse Transcription System, A3500) with Oligo(dT)15 reverse transcription primers. Prepare a 20 μL reverse transcription system according to the instructions of the reverse transcription kit and complete the reverse transcription reaction. After the reaction, add 80 μL of RNase-free water to the reverse transcription system to obtain a cDNA solution. Then, use a real-time quantitative PCR kit (ABI, SYBR). TM Select MasterMix (Catalog number: 4472908) was used to detect the expression level of target gene mRNA in Vero cells. In this real-time quantitative PCR method, primers targeting the target gene and primers targeting the internal reference gene were used to detect the target gene and the internal reference gene, respectively. A 20 μL Real-time PCR reaction system was prepared for each PCR detection well according to the instructions of the real-time quantitative PCR kit. Each reaction system contained 5 μL of cDNA solution obtained from the above reverse transcription reaction and 10 μL of SYBR Green. TMSelect Master Mix, 0.5 μL of 10 μM upstream primer, 0.5 μL of 10 μM downstream primer, and 4 μL of RNase-Free H2O. Place the prepared reaction mixture in a real-time quantitative PCR instrument (ABI, StepOnePlus). TM Real-time PCR amplification was performed using a three-step method. The amplification program was: 95℃ pre-denaturation for 10 min, followed by 95℃ denaturation for 30 s, 60℃ annealing for 30 s, and 72℃ extension for 30 s. This denaturation, annealing, and extension process was repeated for 40 cycles. In this real-time quantitative PCR method, the ΔΔCt method was used to calculate the relative quantitative levels and inhibition rates of the target gene mRNA in each test group. The calculation method is as follows:

[0350] ΔCt(test group) = Ct(target gene in test group) - Ct(internal reference gene in test group)

[0351] ΔCt(control group) = Ct(target gene in control group) - Ct(internal reference gene in control group)

[0352] ΔCt(test group) = ΔCt(test group) - ΔCt(control group average)

[0353] ΔCt(control group) = ΔCt(control group) - ΔCt(control group average)

[0354] Here, ΔCt (control group average) is the arithmetic mean of ΔCt (control group) from several replicates in the MOCK group. Therefore, each cell well in both the test and control groups corresponds to a ΔCt value.

[0355] The relative expression level of the target gene mRNA in the test group was 2. -ΔΔCt (Test group) × 100%

[0356] Using the control group as a baseline, the expression level of the target gene mRNA in the test group was normalized, and the expression level of the target gene mRNA in the control group was defined as 100%.

[0357] The inhibition rate of target gene mRNA expression in the test group = (1 - relative expression level of target gene mRNA in the test group) × 100%

[0358] Unless otherwise stated, in vivo activity data are presented in [year]. The experimental data were all plotted and analyzed using GraphPadprism 8.0 software.

[0359] Example 1: siRNA sequence targeting the urate transporter 1 gene in Vero cells. Evaluation of the inhibitory activity of 1;URAT1)

[0360] This embodiment uses an in vitro cell transfection method to evaluate the inhibitory activity of URAT1-targeting siRNA sequences on Vero cells. Vero cells grown to 80% confluency were digested, plated in 24-well plates, cultured overnight, and then transfected. A mock control group was also included. The final siRNA transfection concentration was 50 nM / well. 24 hours post-transfection, RNA extraction, reverse transcription, and Real-time PCR were performed as described above. The relative quantification of target gene mRNA in each test group was calculated using the aforementioned ΔΔCt method.

[0361] Table 5. Sequence list of primers used in Example 1

[0362]

[0363] The results of Example 1 show that all designed sequences can effectively reduce the expression of URAT1 mRNA, among which RX591008, RX591011, RX591013, RX591014, RX591017, RX591018, RX591019, RX591022 and RX591024 sequences have relatively better gene repression levels (Table 6).

[0364] Table 6 shows the inhibitory activity of Vero cells on the target genes after administration of the siRNA conjugate described in this embodiment.

[0365]

[0366]

[0367]

[0368] The above specific embodiments are merely illustrative of the present invention and do not represent a limitation thereof. For those skilled in the art, various modifications and improvements can be made without departing from the spirit and substance of this disclosure, and these modifications and improvements are also considered to be within the scope of protection of this disclosure.

Claims

1. A double-stranded oligonucleotide for inhibiting URAT1 gene expression, characterized in that: The double-stranded oligonucleotide comprises a sense strand and an antisense strand, the sense strand and the antisense strand being 17 to 30 nucleotides in length, and the sense strand and the antisense strand being at least partially complementary; wherein the antisense strand comprises at least 17 consecutive nucleotides whose first 19 nucleotides differ from the nucleotide sequences shown in any of SEQ ID NO:70 to SEQ ID NO:138 by no more than 3 nucleotides.

2. The double-stranded oligonucleotide according to claim 1, characterized in that: The positive strand comprises at least 17 consecutive nucleotides that differ from the nucleotide sequences shown in any of SEQ ID NO:1 to SEQ ID NO:69 by no more than 3 nucleotides; Optionally, the antisense strand is 21 to 23 nucleotides in length; the sense strand is 19 to 21 nucleotides in length.

3. The double-stranded oligonucleotide according to claim 1, characterized in that: The antisense strand contains no more than 3 nucleotides different from the nucleotides at positions 1-19 of any of the nucleotide sequences shown in SEQ ID NO:70 to SEQ ID NO:138; Optionally, the antisense strand contains no more than two nucleotides differing from the nucleotide sequences shown in any of SEQ ID NO:70 to SEQ ID NO:138 in positions 1-19; Optionally, the antisense strand contains no more than one nucleotide from positions 1 to 19 that differ from the nucleotide sequences shown in any of SEQ ID NO:70 to SEQ ID NO:138; Optionally, the antisense strand comprises nucleotides 1-19 of any of the nucleotide sequences shown in SEQ ID NO:70 to SEQ ID NO:138; Optionally, the antisense strand is selected from any of the nucleotide sequences shown in SEQ ID NO:70 to SEQ ID NO:

138.

4. The double-stranded oligonucleotide according to any one of claims 1-3, characterized in that: The sense and antisense strands have a mismatch of no more than 3 nucleotides; Optionally, the sense strand and the antisense strand may have a mismatch of no more than 2 or 1 nucleotides; Optionally, the justice chain and the antisense chain are completely complementary.

5. The double-stranded oligonucleotide according to any one of claims 1 to 4, characterized in that: The double-stranded oligonucleotide is selected from the sense strand sequence of any double-stranded oligomer 1 to double-stranded oligomer 69 shown in Table 3, and the 1st to 19th nucleotide sequences of the antisense strand sequence; Optionally, the double-stranded oligonucleotide is selected from: The double-stranded body 1, wherein the antisense strand sequence of the double-stranded body 1 is shown in SEQ ID NO:70 and the sense strand sequence is shown in SEQ ID NO:1; Double strand 2, wherein the antisense strand sequence of the double strand 2 is shown in SEQ ID NO:71 and the sense strand sequence is shown in SEQ ID NO:2; The double-stranded body 3, wherein the antisense strand sequence of the double-stranded body 3 is shown in SEQ ID NO:72 and the sense strand sequence is shown in SEQ ID NO:3; The double-stranded body 4, wherein the antisense strand sequence of the double-stranded body 4 is shown in SEQ ID NO:73 and the sense strand sequence is shown in SEQ ID NO:4; The double-stranded body 5, wherein the antisense strand sequence of the double-stranded body 5 is shown in SEQ ID NO:74 and the sense strand sequence is shown in SEQ ID NO:5; The double-stranded body 6, wherein the antisense strand sequence of the double-stranded body 6 is shown in SEQ ID NO:75 and the sense strand sequence is shown in SEQ ID NO:6; The double-stranded body 7, wherein the antisense strand sequence of the double-stranded body 7 is shown in SEQ ID NO:76 and the sense strand sequence is shown in SEQ ID NO:7; The double-stranded body 8, wherein the antisense strand sequence of the double-stranded body 8 is shown in SEQ ID NO:77 and the sense strand sequence is shown in SEQ ID NO:8; The double-stranded body 9, wherein the antisense strand sequence of the double-stranded body 9 is shown in SEQ ID NO:78 and the sense strand sequence is shown in SEQ ID NO:9; The double-stranded body 10, wherein the antisense strand sequence of the double-stranded body 10 is shown in SEQ ID NO:79 and the sense strand sequence is shown in SEQ ID NO:10; The double-stranded body 11, wherein the antisense strand sequence of the double-stranded body 11 is shown in SEQ ID NO:80 and the sense strand sequence is shown in SEQ ID NO:11; The double-stranded body 12, wherein the antisense strand sequence of the double-stranded body 12 is shown in SEQ ID NO:81 and the sense strand sequence is shown in SEQ ID NO:12; The double-stranded body 13, wherein the antisense strand sequence of the double-stranded body 13 is shown in SEQ ID NO:82 and the sense strand sequence is shown in SEQ ID NO:13; The double-stranded body 14, wherein the antisense strand sequence of the double-stranded body 14 is shown in SEQ ID NO:83 and the sense strand sequence is shown in SEQ ID NO:14; The double-stranded body 15, wherein the antisense strand sequence of the double-stranded body 15 is shown in SEQ ID NO:84 and the sense strand sequence is shown in SEQ ID NO:15; The double-stranded body 16, wherein the antisense strand sequence of the double-stranded body 16 is shown in SEQ ID NO:85 and the sense strand sequence is shown in SEQ ID NO:16; The double-stranded body 17, wherein the antisense strand sequence of the double-stranded body 17 is shown in SEQ ID NO:86 and the sense strand sequence is shown in SEQ ID NO:17; The double-stranded body 18, wherein the antisense strand sequence of the double-stranded body 18 is shown in SEQ ID NO:87 and the sense strand sequence is shown in SEQ ID NO:18; The double-stranded body 19, wherein the antisense strand sequence of the double-stranded body 19 is shown in SEQ ID NO:88 and the sense strand sequence is shown in SEQ ID NO:19; The double-stranded body 20, wherein the antisense strand sequence of the double-stranded body 20 is shown in SEQ ID NO:89 and the sense strand sequence is shown in SEQ ID NO:20; The double-stranded body 21, wherein the antisense strand sequence of the double-stranded body 21 is shown in SEQ ID NO:90 and the sense strand sequence is shown in SEQ ID NO:21; The double-stranded body 22, wherein the antisense strand sequence of the double-stranded body 22 is shown in SEQ ID NO:91 and the sense strand sequence is shown in SEQ ID NO:22; The double-stranded body 23, wherein the antisense strand sequence of the double-stranded body 23 is shown in SEQ ID NO:92 and the sense strand sequence is shown in SEQ ID NO:23; The double-stranded body 24, wherein the antisense strand sequence of the double-stranded body 24 is shown in SEQ ID NO:93 and the sense strand sequence is shown in SEQ ID NO:24; The double-stranded body 25, wherein the antisense strand sequence of the double-stranded body 25 is shown in SEQ ID NO:94 and the sense strand sequence is shown in SEQ ID NO:25; The double-stranded body 26, wherein the antisense strand sequence of the double-stranded body 26 is shown in SEQ ID NO:95 and the sense strand sequence is shown in SEQ ID NO:26; The double-stranded body 27, wherein the antisense strand sequence of the double-stranded body 27 is shown in SEQ ID NO:96 and the sense strand sequence is shown in SEQ ID NO:27; The double-stranded body 28, wherein the antisense strand sequence of the double-stranded body 28 is shown in SEQ ID NO:97 and the sense strand sequence is shown in SEQ ID NO:28; The double-stranded body 29, wherein the antisense strand sequence of the double-stranded body 29 is shown in SEQ ID NO:98 and the sense strand sequence is shown in SEQ ID NO:29; The double-stranded body 30, wherein the antisense strand sequence of the double-stranded body 30 is shown in SEQ ID NO:99 and the sense strand sequence is shown in SEQ ID NO:30; The double-stranded body 31, wherein the antisense strand sequence of the double-stranded body 31 is shown in SEQ ID NO:100 and the sense strand sequence is shown in SEQ ID NO:31; The double-stranded body 32, wherein the antisense strand sequence of the double-stranded body 32 is shown in SEQ ID NO:101 and the sense strand sequence is shown in SEQ ID NO:32; The double-stranded body 33, wherein the antisense strand sequence of the double-stranded body 33 is shown in SEQ ID NO:102 and the sense strand sequence is shown in SEQ ID NO:33; The double-stranded body 34, wherein the antisense strand sequence of the double-stranded body 34 is shown in SEQ ID NO:103 and the sense strand sequence is shown in SEQ ID NO:34; The double-stranded body 35, wherein the antisense strand sequence of the double-stranded body 35 is shown in SEQ ID NO:104 and the sense strand sequence is shown in SEQ ID NO:35; The double-stranded body 36, wherein the antisense strand sequence of the double-stranded body 36 is shown in SEQ ID NO:105 and the sense strand sequence is shown in SEQ ID NO:36; The double-stranded body 37, wherein the antisense strand sequence of the double-stranded body 37 is shown in SEQ ID NO:106 and the sense strand sequence is shown in SEQ ID NO:37; The double-stranded body 38, wherein the antisense strand sequence of the double-stranded body 38 is shown in SEQ ID NO:107 and the sense strand sequence is shown in SEQ ID NO:38; The double-stranded body 39, wherein the antisense strand sequence of the double-stranded body 39 is shown in SEQ ID NO:108 and the sense strand sequence is shown in SEQ ID NO:39; The double-stranded body 40, wherein the antisense strand sequence of the double-stranded body 40 is shown in SEQ ID NO:109 and the sense strand sequence is shown in SEQ ID NO:40; The double-stranded body 41, wherein the antisense strand sequence of the double-stranded body 41 is shown in SEQ ID NO:110 and the sense strand sequence is shown in SEQ ID NO:41; The double-stranded body 42, wherein the antisense strand sequence of the double-stranded body 42 is shown in SEQ ID NO:111 and the sense strand sequence is shown in SEQ ID NO:42; The double-stranded body 43, wherein the antisense strand sequence of the double-stranded body 43 is shown in SEQ ID NO:112 and the sense strand sequence is shown in SEQ ID NO:43; The double-stranded body 44, wherein the antisense strand sequence of the double-stranded body 44 is shown in SEQ ID NO:113 and the sense strand sequence is shown in SEQ ID NO:44; The double-stranded body 45, wherein the antisense strand sequence of the double-stranded body 45 is shown in SEQ ID NO:114 and the sense strand sequence is shown in SEQ ID NO:45; The double-stranded body 46, wherein the antisense strand sequence of the double-stranded body 46 is shown in SEQ ID NO:115 and the sense strand sequence is shown in SEQ ID NO:46; The double-stranded body 47, wherein the antisense strand sequence of the double-stranded body 47 is shown in SEQ ID NO:116 and the sense strand sequence is shown in SEQ ID NO:47; The double-stranded body 48, wherein the antisense strand sequence of the double-stranded body 48 is shown in SEQ ID NO:117 and the sense strand sequence is shown in SEQ ID NO:48; The double-stranded body 49, wherein the antisense strand sequence of the double-stranded body 49 is shown in SEQ ID NO:118 and the sense strand sequence is shown in SEQ ID NO:49; The double-stranded body 50, wherein the antisense strand sequence of the double-stranded body 50 is shown in SEQ ID NO:119 and the sense strand sequence is shown in SEQ ID NO:50; The double-stranded body 51, wherein the antisense strand sequence of the double-stranded body 51 is shown in SEQ ID NO:120 and the sense strand sequence is shown in SEQ ID NO:51; The double-stranded body 52, wherein the antisense strand sequence of the double-stranded body 52 is shown in SEQ ID NO:121 and the sense strand sequence is shown in SEQ ID NO:52; The double-stranded body 53, wherein the antisense strand sequence of the double-stranded body 53 is shown in SEQ ID NO:122 and the sense strand sequence is shown in SEQ ID NO:53; The double-stranded body 54, wherein the antisense strand sequence of the double-stranded body 54 is shown in SEQ ID NO:123 and the sense strand sequence is shown in SEQ ID NO:54; The double-stranded body 55, wherein the antisense strand sequence of the double-stranded body 55 is shown in SEQ ID NO:124 and the sense strand sequence is shown in SEQ ID NO:55; The double-stranded body 56, wherein the antisense strand sequence of the double-stranded body 56 is shown in SEQ ID NO:125 and the sense strand sequence is shown in SEQ ID NO:56; The double-stranded body 57, wherein the antisense strand sequence of the double-stranded body 57 is shown in SEQ ID NO:126 and the sense strand sequence is shown in SEQ ID NO:57; The double-stranded body 58, wherein the antisense strand sequence of the double-stranded body 58 is shown in SEQ ID NO:127 and the sense strand sequence is shown in SEQ ID NO:58; The double-stranded body 59, wherein the antisense strand sequence of the double-stranded body 59 is shown in SEQ ID NO:128 and the sense strand sequence is shown in SEQ ID NO:59; The double-stranded body 60, wherein the antisense strand sequence of the double-stranded body 60 is shown in SEQ ID NO:129 and the sense strand sequence is shown in SEQ ID NO:60; The double-stranded body 61, wherein the antisense strand sequence of the double-stranded body 61 is shown in SEQ ID NO:130 and the sense strand sequence is shown in SEQ ID NO:61; The double-stranded body 62, wherein the antisense strand sequence of the double-stranded body 62 is shown in SEQ ID NO:131 and the sense strand sequence is shown in SEQ ID NO:62; The double-stranded body 63, wherein the antisense strand sequence of the double-stranded body 63 is shown in SEQ ID NO:132 and the sense strand sequence is shown in SEQ ID NO:63; The double-stranded body 64, wherein the antisense strand sequence of the double-stranded body 64 is shown in SEQ ID NO:133 and the sense strand sequence is shown in SEQ ID NO:64; The double-stranded body 65, wherein the antisense strand sequence of the double-stranded body 65 is shown in SEQ ID NO:134 and the sense strand sequence is shown in SEQ ID NO:65; The double-stranded body 66, wherein the antisense strand sequence of the double-stranded body 66 is shown in SEQ ID NO:135 and the sense strand sequence is shown in SEQ ID NO:66; The double-stranded body 67, wherein the antisense strand sequence of the double-stranded body 67 is shown in SEQ ID NO:136 and the sense strand sequence is shown in SEQ ID NO:67; The double-stranded body 68, wherein the antisense strand sequence of the double-stranded body 68 is shown in SEQ ID NO:137 and the sense strand sequence is shown in SEQ ID NO:68; The double strand 69, wherein the antisense strand sequence of the double strand 69 is shown in SEQ ID NO:138 and the sense strand sequence is shown in SEQ ID NO:

69.

6. The double-stranded oligonucleotide according to any one of claims 1 to 5, characterized in that: The double-stranded oligonucleotide contains at least one modified nucleotide; Optionally, each nucleotide in the double-stranded oligonucleotide is a modified nucleotide; Optionally, the modified nucleotide is selected from: 2'-halogenated nucleotides, 2'-deoxygenated nucleotides, 2'-O-alkylated nucleotides, and 2'-O-(CH2). n -OMe modified nucleotides (n selected from 1 or 2), 2'-amino modified nucleotides, nucleotides modified with 2',2'-[halogen, halogen] disubstituted or 2',2'-[halogen, alkyl] disubstituted groups, debased nucleotides, or nucleotide-like nucleotides; Optionally, the modified nucleotide is selected from: 2'-fluoro-modified nucleotides, 2'-deoxy-modified nucleotides, 2'-O-methyl-modified nucleotides, 2'-O-methoxyethyl-modified nucleotides, and 2',2'-[fluoro,methyl]di-substituent-modified nucleotides.

7. The double-stranded oligonucleotide according to claim 6, characterized in that: The antisense strand of the double-stranded oligonucleotide is 21 nucleotides long, and the sense strand is 19 nucleotides long, and each nucleotide in the double-stranded oligonucleotide is a modified nucleotide; wherein: In the direction from the 5' end to the 3' end, at least four nucleotides at positions 2, 6, 9, 14, and 16 of the antisense strand are selected from 2'-F modified nucleotides, and the nucleotides at the remaining positions are selected from 2'-methoxy modified nucleotides or 2'-O-methoxyethyl modified nucleotides; at least three nucleotides at positions 7-10 of the sense strand are selected from 2'-F modified nucleotides, and the remaining positions are 2'-methoxy modified nucleotides. Optionally, in the direction from the 5' end to the 3' end, the nucleotides at positions 2, 6, 9, 14, and 16 of the antisense strand are selected from 2'-F modified nucleotides, the nucleotide at position 15 is selected from 2'-O-methoxyethyl modified nucleotides or 2'-methoxy modified nucleotides, and the nucleotides at the remaining positions are 2'-methoxy modified nucleotides or other modified nucleotides besides the above modifications; the nucleotides at positions 7-10 of the sense strand are selected from 2'-F modified nucleotides, and the nucleotides at the remaining positions are selected from 2'-methoxy modified nucleotides or other modified nucleotides besides the above modifications; Optionally, in the direction from the 5' end to the 3' end, the nucleotides at positions 2, 6, 9, 14, and 16 of the antisense strand are selected from 2'-F modified nucleotides, the nucleotide at position 15 is selected from 2'-O-methoxyethyl modified nucleotides, and the nucleotides at the remaining positions are 2'-methoxy modified nucleotides; the nucleotides at positions 7-10 of the sense strand are selected from 2'-F modified nucleotides, and the nucleotides at the remaining positions are selected from 2'-methoxy modified nucleotides. Optionally, the double-stranded oligonucleotide has a modified phosphate group selected from thiophosphate groups, and the thiophosphate group is present at at least one of the following positions: 1) between the first and second nucleotides at the 5' end of the sense strand, and / or between the second and third nucleotides; 2) between the first and second nucleotides at the 3' end of the sense strand, and / or between the second and third nucleotides; 3) between the first and second nucleotides at the 5' end of the antisense strand, and / or between the second and third nucleotides; 4) between the first and second nucleotides at the 3' end of the antisense strand, and / or between the second and third nucleotides, and / or between the third and fourth nucleotides; 5) between the tenth and eleventh nucleotides at the 5' end of the antisense strand.

8. The double-stranded oligonucleotide according to claim 7, characterized in that: The antisense strand comprises at least 17 consecutive nucleotides whose nucleotides from positions 1 to 19 differ by no more than 3 nucleotides from any of the nucleotide sequences shown in SEQ ID NO:208 to SEQ ID NO:276; the sense strand comprises at least 17 consecutive nucleotides whose nucleotide sequences differ by no more than 3 nucleotides from any of the nucleotide sequences shown in SEQ ID NO:139 to SEQ ID NO:

207. Optionally, the antisense strand contains no more than 3 nucleotides different from the nucleotides at positions 1-19 of any of the nucleotide sequences shown in SEQ ID NO:208 to SEQ ID NO:276; Optionally, the antisense strand contains no more than two nucleotides differing from the nucleotide sequences shown in any of SEQ ID NO:208 to SEQ ID NO:276 in positions 1-19; Optionally, the antisense strand contains no more than one nucleotide from positions 1 to 19 that differ from the nucleotide sequences shown in any of SEQ ID NO:208 to SEQ ID NO:276; Optionally, the antisense strand comprises nucleotides 1-19 of any of the nucleotide sequences shown in SEQ ID NO:208 to SEQ ID NO:276; Optionally, the antisense strand is selected from any of the nucleotide sequences shown in SEQ ID NO: SEQ ID NO: 208 to SEQ ID NO: 276; Optionally, the double-stranded oligonucleotide is selected from the sense strand of any of the modified double strands 1 to 69 shown in Table 4, and the first to 19 nucleotides in the antisense strand sequence; Optionally, the double-stranded oligonucleotide is selected from: The modified double strand 1 has an antisense strand sequence as shown in SEQ ID NO:208 and a sense strand sequence as shown in SEQ ID NO:

139. The modified double strand 2 has an antisense strand sequence as shown in SEQ ID NO:209 and a sense strand sequence as shown in SEQ ID NO:

140. The modified double strand 3 has an antisense strand sequence as shown in SEQ ID NO:210 and a sense strand sequence as shown in SEQ ID NO:

141. Modified double strand 4, wherein the antisense strand sequence of the modified double strand 4 is shown in SEQ ID NO:211 and the sense strand sequence is shown in SEQ ID NO:142; The modified double strand 5 has an antisense strand sequence as shown in SEQ ID NO:212 and a sense strand sequence as shown in SEQ ID NO:

143. The modified double strand 6 has an antisense strand sequence as shown in SEQ ID NO:213 and a sense strand sequence as shown in SEQ ID NO:

144. The modified double strand 7, wherein the antisense strand sequence of the modified double strand 7 is shown in SEQ ID NO:214 and the sense strand sequence is shown in SEQ ID NO:145; The modified double strand 8 has an antisense strand sequence as shown in SEQ ID NO:215 and a sense strand sequence as shown in SEQ ID NO:

146. The modified double strand 9 has an antisense strand sequence as shown in SEQ ID NO:216 and a sense strand sequence as shown in SEQ ID NO:

147. The modified double strand 10 has an antisense strand sequence as shown in SEQ ID NO:217 and a sense strand sequence as shown in SEQ ID NO:

148. The modified double strand 11 has an antisense strand sequence as shown in SEQ ID NO:218 and a sense strand sequence as shown in SEQ ID NO:

149. The modified double strand 12 has an antisense strand sequence as shown in SEQ ID NO:219 and a sense strand sequence as shown in SEQ ID NO:

150. The modified double strand 13 has an antisense strand sequence as shown in SEQ ID NO:220 and a sense strand sequence as shown in SEQ ID NO:

151. The modified double strand 14 has an antisense strand sequence as shown in SEQ ID NO:221 and a sense strand sequence as shown in SEQ ID NO:

152. The modified double strand 15 has an antisense strand sequence as shown in SEQ ID NO:222 and a sense strand sequence as shown in SEQ ID NO:

153. The modified double strand 16 has an antisense strand sequence as shown in SEQ ID NO:223 and a sense strand sequence as shown in SEQ ID NO:

154. The modified double strand 17 has an antisense strand sequence as shown in SEQ ID NO:224 and a sense strand sequence as shown in SEQ ID NO:

155. The modified double strand 18 has an antisense strand sequence as shown in SEQ ID NO:225 and a sense strand sequence as shown in SEQ ID NO:

156. The modified double strand 19 has an antisense strand sequence as shown in SEQ ID NO:226 and a sense strand sequence as shown in SEQ ID NO:

157. The modified double strand 20 has an antisense strand sequence as shown in SEQ ID NO:227 and a sense strand sequence as shown in SEQ ID NO:

158. The modified double strand 21 has an antisense strand sequence as shown in SEQ ID NO:228 and a sense strand sequence as shown in SEQ ID NO:

159. The modified double strand 22 has an antisense strand sequence as shown in SEQ ID NO:229 and a sense strand sequence as shown in SEQ ID NO:

160. The modified double strand 23 is shown in SEQ ID NO:230 and the positive strand sequence is shown in SEQ ID NO:

161. The modified double strand 24 is shown in SEQ ID NO:231 and the positive strand sequence is shown in SEQ ID NO:

162. The modified double strand 25 has an antisense strand sequence as shown in SEQ ID NO:232 and a sense strand sequence as shown in SEQ ID NO:

163. The modified double strand 26 has an antisense strand sequence as shown in SEQ ID NO:233 and a sense strand sequence as shown in SEQ ID NO:

164. The modified double strand 27 has an antisense strand sequence as shown in SEQ ID NO:234 ​​and a sense strand sequence as shown in SEQ ID NO:

165. The modified double strand 28 has an antisense strand sequence as shown in SEQ ID NO:235 and a sense strand sequence as shown in SEQ ID NO:

166. The modified double strand 29 has an antisense strand sequence as shown in SEQ ID NO:236 and a sense strand sequence as shown in SEQ ID NO:

167. The modified double strand 30 has an antisense strand sequence as shown in SEQ ID NO:237 and a sense strand sequence as shown in SEQ ID NO:

168. The modified double strand 31 has an antisense strand sequence as shown in SEQ ID NO:238 and a sense strand sequence as shown in SEQ ID NO:

169. The modified double strand 32 has an antisense strand sequence as shown in SEQ ID NO:239 and a sense strand sequence as shown in SEQ ID NO:

170. The modified double strand 33 has an antisense strand sequence as shown in SEQ ID NO:240 and a sense strand sequence as shown in SEQ ID NO:

171. The modified double strand 34 has an antisense strand sequence as shown in SEQ ID NO:241 and a sense strand sequence as shown in SEQ ID NO:

172. The modified double strand 35 has an antisense strand sequence as shown in SEQ ID NO:242 and a sense strand sequence as shown in SEQ ID NO:

173. The modified double strand 36 has an antisense strand sequence as shown in SEQ ID NO:243 and a sense strand sequence as shown in SEQ ID NO:

174. The modified double strand 37 has an antisense strand sequence as shown in SEQ ID NO:244 and a sense strand sequence as shown in SEQ ID NO:

175. The modified double strand 38 has an antisense strand sequence as shown in SEQ ID NO:245 and a sense strand sequence as shown in SEQ ID NO:

176. The modified double strand 39 has an antisense strand sequence as shown in SEQ ID NO:246 and a sense strand sequence as shown in SEQ ID NO:

177. The modified double strand 40 has an antisense strand sequence as shown in SEQ ID NO:247 and a sense strand sequence as shown in SEQ ID NO:

178. The modified double strand 41 has an antisense strand sequence as shown in SEQ ID NO:248 and a sense strand sequence as shown in SEQ ID NO:

179. The modified double strand 42 has an antisense strand sequence as shown in SEQ ID NO:249 and a sense strand sequence as shown in SEQ ID NO:

180. The modified double strand 43 has an antisense strand sequence as shown in SEQ ID NO:250 and a sense strand sequence as shown in SEQ ID NO:

181. The modified double strand 44 has an antisense strand sequence as shown in SEQ ID NO:251 and a sense strand sequence as shown in SEQ ID NO:

182. The modified double strand 45 has an antisense strand sequence as shown in SEQ ID NO:252 and a sense strand sequence as shown in SEQ ID NO:

183. The modified double strand 46 has an antisense strand sequence as shown in SEQ ID NO:253 and a sense strand sequence as shown in SEQ ID NO:

184. The modified double strand 47 has an antisense strand sequence as shown in SEQ ID NO:254 and a sense strand sequence as shown in SEQ ID NO:

185. The modified double strand 48 has an antisense strand sequence as shown in SEQ ID NO:255 and a sense strand sequence as shown in SEQ ID NO:

186. The modified double strand 49 has an antisense strand sequence as shown in SEQ ID NO:256 and a sense strand sequence as shown in SEQ ID NO:

187. The modified double strand 50 has an antisense strand sequence as shown in SEQ ID NO:257 and a sense strand sequence as shown in SEQ ID NO:

188. The modified double strand 51 has an antisense strand sequence as shown in SEQ ID NO:258 and a sense strand sequence as shown in SEQ ID NO:

189. The modified double strand 52 has an antisense strand sequence as shown in SEQ ID NO:259 and a sense strand sequence as shown in SEQ ID NO:

190. The modified double strand 53 has an antisense strand sequence as shown in SEQ ID NO:260 and a sense strand sequence as shown in SEQ ID NO:

191. The modified double strand 54 has an antisense strand sequence as shown in SEQ ID NO:261 and a sense strand sequence as shown in SEQ ID NO:

192. The modified double strand 55 has an antisense strand sequence as shown in SEQ ID NO:262 and a sense strand sequence as shown in SEQ ID NO:

193. The modified double strand 56 has an antisense strand sequence as shown in SEQ ID NO:263 and a sense strand sequence as shown in SEQ ID NO:

194. The modified double strand 57 has an antisense strand sequence as shown in SEQ ID NO:264 and a sense strand sequence as shown in SEQ ID NO:

195. The modified double strand 58 has an antisense strand sequence as shown in SEQ ID NO:265 and a sense strand sequence as shown in SEQ ID NO:

196. The modified double strand 59 has an antisense strand sequence as shown in SEQ ID NO:266 and a sense strand sequence as shown in SEQ ID NO:

197. The modified double strand 60 has an antisense strand sequence as shown in SEQ ID NO:267 and a sense strand sequence as shown in SEQ ID NO:

198. The modified double strand 61 has an antisense strand sequence as shown in SEQ ID NO:268 and a sense strand sequence as shown in SEQ ID NO:

199. The modified double strand 62 has an antisense strand sequence as shown in SEQ ID NO:269 and a sense strand sequence as shown in SEQ ID NO:

200. The modified double strand 63 has an antisense strand sequence as shown in SEQ ID NO:270 and a sense strand sequence as shown in SEQ ID NO:

201. The modified double strand 64 has an antisense strand sequence as shown in SEQ ID NO:271 and a sense strand sequence as shown in SEQ ID NO:

202. The modified double strand 65 has an antisense strand sequence as shown in SEQ ID NO:272 and a sense strand sequence as shown in SEQ ID NO:

203. The modified double strand 66 has an antisense strand sequence as shown in SEQ ID NO:273 and a sense strand sequence as shown in SEQ ID NO:

204. The modified double strand 67 has an antisense strand sequence as shown in SEQ ID NO:274 and a sense strand sequence as shown in SEQ ID NO:

205. The modified double strand 68 has an antisense strand sequence as shown in SEQ ID NO:275 and a sense strand sequence as shown in SEQ ID NO:

206. The modified double strand 69 has an antisense strand sequence as shown in SEQ ID NO:276 and a sense strand sequence as shown in SEQ ID NO:

207.

9. An oligonucleotide conjugate, characterized in that: It comprises the double-stranded oligonucleotide as described in any one of claims 1 to 8 and a targeting ligand group; Optionally, the targeting ligand group is a polypeptide, and includes any of the polypeptide fragments shown in 1)-6) below: 1)-X1(KKEEE) n -K m - where n = 1-5, m = 0 or 1 or 2, and X1 is selected from any L-α amino acid or L-β amino acid other than K and E; 2)-CLPVASC-, preferably in cyclic peptide form 3)-CYFQNC-, preferably in cyclic peptide form 4)-KIDRI-; 5)-IDRI-(-Ile-Asp-Arg-Ile-); 6)-dXa-Ser-dXb-X2-dXc-Gly-Xd-Ile-Asp-Arg(Ak)-Ile-; where X2 is selected from any non-natural amino acid; dXa, dXb, dXc are selected from any D-type amino acid; Arg(Ak) is selected from arginine or alkylated arginine; Xd is selected from Hyp or Pro; Preferably, the targeting ligand group is selected from polypeptides containing any of the amino acid sequence segments shown in a)-g). a)-KKEEE-KKEEE-KKEEE-K-;; b)-CKKEEE-KKEEE-KKEEE-K; c)-CLPVASC-, preferably in cyclic peptide form d)-KIDRI-; e)-CYFQNCPRG-, preferably in cyclic peptide form f)-DPhe-Ser-DHyp-Cha-DAla-Gly-Hyp-Ile-Asp-Arg(Me)-Ile-; g)-DPhe-Ser-DPro-Cha-DAla-Gly-Pro-Ile-Asp-Arg-Ile-.

10. The oligonucleotide conjugate according to claim 9, characterized in that, The targeting ligand group is connected to the double-stranded oligonucleotide of any one of claims 1 to 8 via a linker; Optionally, the structural unit of the targeting ligand group-connector in the conjugate is selected from any of the structures shown in 1)-7) below: 1)La-KKEEE-KKEEE-KKEEE-K-Laa; 2)La-CKKEEE-KKEEE-KKEEE-K-Laa; 3) La-CLPVASC-Laa, preferably a cyclic peptide. 4)La-DPhe-Ser-DHyp-Cha-DAla-Gly-Hyp-Ile-Asp-Arg(Me)-Ile-Laa; 5)La-CYFQNCPRG-Laa; 6)La-DPhe-Ser-DPro-Cha-DAla-Gly-Pro-Ile-Asp-Arg-Ile-Laa; 7) Lb-KIDRI-Laa; In formulas 1)-6), either La or Laa is a linker and the other is a terminal blocking group, wherein the linker is a linking group containing at least one of a triazole group, a PEG unit, or an acyl group. Optionally, La is a connector, and the connector structure includes -PEG2-CH2CH2CO- or -triazolyl-PEG2-; In formula 7), Lb is a linking group containing a 6-10 membered aromatic ring or heteroaromatic ring, and Laa is independently selected from an amino group or an alkyl-substituted amino group, such as -NHCH3.

11. The double-stranded oligonucleotide conjugate according to claim 10, characterized in that: The targeting ligand group is connected to the ends of the sense and / or antisense strands of the double-stranded oligonucleotide according to any one of claims 1 to 8 via a linker.

12. The double-stranded oligonucleotide conjugate according to claim 10 or 11, characterized in that: The structure of the targeting ligand group-linker in the double-stranded oligonucleotide conjugate is selected from any of the structures shown in a)-j), or a pharmaceutically acceptable salt thereof: a)-La-KKEEE-KKEEE-KKEEE-K-Lc; b)-La-CKKEEE-KKEEE-KKEEE-K-Lc; c)-La-CLPVASC-Lc; preferably cyclic peptide form d)-La-DPhe-Ser-DHyp-Cha-DAla-Gly-Hyp-Ile-Asp-Arg(Me)-Ile-Lc; e)-La-CYFQNCPRG-Lc; preferably a cyclic peptide f)-La-DPhe-Ser-DPro-Cha-DAla-Gly-Pro-Ile-Asp-Arg-Ile-Lc; g) h) i) j) Wherein, La is a linker, which is independently selected from any bond, -NH-, amide group or linking group containing at least one PEG unit; Optionally, La is a linking group comprising at least two PEG units; Optionally, La is selected from amide groups, or n is 1-3, m is 0 or 1, and Z is -CO- (carbonyl) or -NH-. Wherein, Lc is a terminal blocking group; optionally, it is a C-terminal blocking group, wherein Lc is selected from amino or alkyl-substituted amino groups, preferably -NHCH3.

13. A pharmaceutical composition, characterized in that: The composition comprises the double-stranded oligonucleotide as described in any one of claims 1 to 8 and / or the double-stranded oligonucleotide conjugate as described in any one of claims 9 to 12.

14. Use of the double-stranded oligonucleotide of any one of claims 1 to 8 and / or the double-stranded oligonucleotide conjugate of any one of claims 9 to 12 and / or the pharmaceutical composition of claim 13 in the preparation of a medicament for inhibiting URAT1 gene expression.

15. Use of the double-stranded oligonucleotide of any one of claims 1 to 8 and / or the double-stranded oligonucleotide conjugate of any one of claims 9 to 12 and / or the composition of claim 13 in the preparation of a medicament for treating and / or preventing pathological conditions or diseases associated with URAT1 expression.

16. The use according to claim 15, characterized in that: The pathological condition or disease is hyperuricemia or gout.

17. A method for inhibiting intracellular URAT1 expression, characterized in that, It includes administering to a subject the double-stranded oligonucleotide of any one of claims 1 to 8 and / or the double-stranded oligonucleotide conjugate of any one of claims 9 to 12 and / or the composition of claim 13; optionally, the subject is a human.

18. A method for preventing and / or treating hyperuricemia or gout, the method comprising administering to a subject the double-stranded oligonucleotide of any one of claims 1 to 8 and / or the double-stranded oligonucleotide conjugate of any one of claims 9 to 12 and / or the composition of claim 13.

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