WO2022028457A1 - Arnsi pour inhiber l'expression du facteur xi de coagulation sanguine, et composition et utilisation médicale de celui-ci - Google Patents

Arnsi pour inhiber l'expression du facteur xi de coagulation sanguine, et composition et utilisation médicale de celui-ci Download PDF

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WO2022028457A1
WO2022028457A1 PCT/CN2021/110503 CN2021110503W WO2022028457A1 WO 2022028457 A1 WO2022028457 A1 WO 2022028457A1 CN 2021110503 W CN2021110503 W CN 2021110503W WO 2022028457 A1 WO2022028457 A1 WO 2022028457A1
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seq
nucleotide
sirna
antisense strand
nucleotides
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黄金宇
黄燕芬
刘楠
蔡国庆
尹科
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上海拓界生物医药科技有限公司
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
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    • A61K31/00—Medicinal preparations containing organic active ingredients
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    • A61K31/7088—Compounds having three or more nucleosides or nucleotides
    • A61K31/713—Double-stranded nucleic acids or oligonucleotides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
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    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing

Definitions

  • the present disclosure belongs to the field of biomedicine, and in particular relates to siRNA for inhibiting the expression of coagulation factor XI, compositions and medical uses thereof.
  • the blood circulatory system not only has a coagulation mechanism to prevent blood loss, but also has an anticoagulation mechanism to counteract improper intravascular embolism.
  • the dynamic balance between the two is the key to the normal body to maintain the state of blood flow in the body and prevent blood loss.
  • the coagulation process is a process in which a series of coagulation factors are activated by successive enzymatic hydrolysis, and finally thrombin is generated to form a fibrin clot. There are two activation pathways, exogenous and endogenous, in the human body.
  • the endogenous pathway includes: when the blood vessel wall is damaged, the subendothelial tissue is exposed, the negatively charged subendothelial collagen fibers are in contact with the coagulation factor, the factor XII is bound to it, and it is activated to factor XIIa with the participation of HK and PK; Factor XIIa activates factor XI to factor XIa in the presence of calcium ions independent of calcium ions; in the presence of calcium ions, activated factor XIa activates factor IX; IXa alone is quite ineffective in activating factor X, and it is much more potent than VIIIa. Combine to form a 1:1 complex, also known as the factor X enzyme complex; this reaction must also involve Ca 2+ and PL.
  • Exogenous pathways include: tissue factor (TF) released after tissue injury combines with calcium ions, factor VII (or factor VIIa) in plasma to form TF-Ca 2+ -FVII/FVIIa complex, which combines factor X Activated as factor Xa.
  • the two pathways converge upon activation of factor X.
  • Activated factor Xa and factor V form a prothrombin activator, which hydrolyzes prothrombin to thrombin, and thrombin converts fibrinogen to fibrin, and forms a fibrin clot under the action of platelets.
  • factor XI in the endogenous pathway is associated with the formation of venous thrombosis (JOOST CMMEIJERS et al., The New England Journal of Medicine, 2000, Vol. 342, No. 10, 696-701).
  • JOOST CMMEIJERS et al., The New England Journal of Medicine, 2000, Vol. 342, No. 10, 696-701.
  • Thromboembolism can lead to conditions such as deep vein thrombosis, pulmonary embolism, myocardial infarction, and stroke.
  • Anticoagulants can reduce the risk of thromboembolism.
  • Current anticoagulants such as warfarin, heparin and low molecular weight heparin (LMWH), coagulation factor X inhibitors, etc. have significant disadvantages, such as lack of predictability and specificity, Careful monitoring of patients is therefore required to prevent adverse side effects such as bleeding complications.
  • LMWH low molecular weight heparin
  • Careful monitoring of patients is therefore required to prevent adverse side effects such as bleeding complications.
  • RNA interference is an effective way to silence gene expression. According to statistics, more than 80% of the disease-related proteins in the human body cannot be targeted by current conventional small molecule drugs and biological macromolecular preparations, and belong to non-druggable proteins.
  • Antisense oligonucleotides (ASOs) and small interfering RNAs (siRNAs) are known strategies for silencing gene expression.
  • CN102245186A describes ASO targeting coagulation factor XI. There are no reports on siRNA against coagulation factor XI.
  • siRNA design Although some algorithms for siRNA design are known, such as mFOLD, these algorithms only consider the primary structure and roughly predicted secondary structure of mRNA, and do not consider the tertiary structure of mRNA and the interaction of mRNA with RNA-binding proteins for siRNA The effect of activity and selectivity, so only based on the existing algorithm is not enough to obtain siRNA with sufficient activity and selectivity.
  • the present disclosure provides mRNA and/or protein level compounds, compositions and methods capable of modulating factor XI.
  • the present disclosure provides an siRNA comprising a sense strand and an antisense strand, wherein the sense strand contains a nucleotide sequence A, the antisense strand contains a nucleotide sequence B, a nucleotide sequence A and a nucleotide sequence B is at least partially reverse complementary to form a double-stranded region;
  • nucleotide sequence A is equal to any one of the nucleotide sequences of the sense strand provided in Table 1, and there are no more than 4, no more than 3, no more than 2 or no more than each other. less than 1 nucleotide difference; and nucleotide sequence B is the same length as any of the antisense strand nucleotide sequences provided in Table 1, and there are no more than 4, no more than 3, No more than 2 or no more than 1 nucleotide difference.
  • the present disclosure also provides an siRNA capable of inhibiting the expression of coagulation factor XI, comprising a sense strand and an antisense strand, wherein the sense strand contains a nucleotide sequence A, the antisense strand contains a nucleotide sequence B, and the nucleoside Acid sequence A and nucleotide sequence B are at least partially reverse complementary to form a double-stranded region;
  • nucleotide sequence A is equal to any one of the nucleotide sequences of the sense strand provided in Table 1, and there are no more than 4, no more than 3, no more than 2 or no more than each other. less than 1 nucleotide difference; and nucleotide sequence B is the same length as any of the antisense strand nucleotide sequences provided in Table 1, and there are no more than 4, no more than 3, No more than 2 or no more than 1 nucleotide difference.
  • nucleotide sequence A there is no more than 1 nucleotide difference between nucleotide sequence A and any of the sense strand nucleotide sequences provided in Table 1; and/or said nucleotide sequence B and There is no more than 1 nucleotide difference between any of the antisense strand nucleotide sequences provided in Table 1.
  • nucleotide sequence A there is no more than 3 base mismatches between nucleotide sequence A and said nucleotide sequence B. In some embodiments, there is no more than 2 base mismatches between nucleotide sequence A and said nucleotide sequence B. In some embodiments, there is no more than 1 base mismatch between nucleotide sequence A and said nucleotide sequence B.
  • nucleotide sequence A there is no mismatch between nucleotide sequence A and said nucleotide sequence B.
  • the sense strand and the antisense strand each have a length of less than 30, less than 29, less than 28, less than 27, less than 26, less than 25, less than 24, less than 23, less than 22, less than 21, or less than 20 nucleotides.
  • the nucleotide sequence A is the nucleotide sequence set forth in any one of SEQ ID NOs: 1-23.
  • nucleotide sequence B is the nucleotide sequence set forth in any one of SEQ ID NOs: 24-46.
  • the present disclosure also provides an siRNA, which is the modified siRNA described above, wherein at least one nucleotide in the sense strand and/or the antisense strand is a modified nucleotide. In some embodiments, all nucleotides are modified nucleotides.
  • the modified nucleotides are independently selected from the group consisting of deoxy-nucleotides, 3'-terminal deoxy-thymidine nucleotides, 2'-O-methyl modified nucleotides, 2'- Fluorine modified nucleotides, 2'-deoxy-modified nucleotides, locked nucleotides, unlocked nucleotides, conformationally restricted nucleotides, restricted ethyl nucleotides, abasic nucleotides , 2'-amino-modified nucleotides, 2'-O-allyl-modified nucleotides, 2'-C-alkyl-modified nucleotides, 2'-hydroxy-modified nucleotides Acids, 2'-methoxyethyl-modified nucleotides, 2'-O-alkyl-modified nucleotides, morpholino nucleotides, phosphoramidates, nucle
  • the modified nucleotides are independently selected from the group consisting of: 2'-alkoxy modified nucleotides, 2'-substituted alkoxy modified nucleotides, 2'-alkyl modified nucleotides nucleotides, 2'-substituted alkyl-modified nucleotides, 2'-amino-modified nucleotides, 2'-substituted amino-modified nucleotides, 2'-fluoro-modified cores nucleotides, 2'-deoxynucleotides, 2'-deoxy-2'-fluoromodified nucleotides, 3'-deoxy-thymidine nucleotides, isonucleotides, LNA, ENA, cET, UNA and GNA.
  • the modified nucleotides are independently selected from the group consisting of: 2'-methoxy-modified nucleotides, 2'-fluoro-modified nucleotides, and 2'-deoxy-modified nucleotides .
  • a fluoro-modified nucleotide refers to a nucleotide formed by substituting the hydroxyl group at the 2' position of the ribosyl group of the nucleotide with fluorine.
  • the 2'-alkoxy-modified nucleotide is a methoxy-modified nucleotide (2'-OMe).
  • a 2'-substituted alkoxy-modified nucleotide eg, can be a 2'-O-methoxyethyl-modified nucleotide (2'-MOE).
  • 2'-amino modified nucleotides (2'- NH2 ).
  • At least one phosphate group in the sense and/or antisense strand is a phosphate group with a modifying group that imparts increased stability of the siRNA in a biological sample or environment .
  • the phosphate group having the modifying group is a phosphorothioate group.
  • a phosphorothioate group refers to a phosphodiester group in which a non-bridging oxygen atom is replaced by a sulfur atom.
  • the nucleotides at positions 2, 6, 9, 12 and 14 of the nucleotide sequence B are each independently a 2'-deoxynucleoside in the 5'-end to 3'-end direction Acid or 2'-fluoro modified nucleotides.
  • the nucleotides at positions 2, 4, 6, 9, 12, 14 and 18 of the nucleotide sequence B are each independently 2' in the 5' to 3' direction - Deoxynucleotides or 2'-fluoromodified nucleotides.
  • the nucleotides at positions 2, 4, 6, 9, 12, 14, 16 and 18 of the nucleotide sequence B are each independently, in the 5'-end to 3'-end direction 2'-deoxynucleotides or 2'-fluoromodified nucleotides.
  • the sense and antisense strands are the same or different in length, the sense strand being 19-23 nucleotides in length and the antisense strand 20-26 nucleotides in length.
  • the length ratio of the sense and antisense strands of the siRNA provided by the present disclosure can be 19/20, 19/21, 19/22, 19/23, 19/24, 19/25, 19/26, 20/20 , 20/21, 20/22, 20/23, 20/24, 20/25, 20/26, 21/20, 21/21, 21/22, 21/23, 21/24, 21/25, 21 /26, 22/20, 22/21, 22/22, 22/23, 22/24, 22/25, 22/26, 23/20, 23/21, 23/22, 23/23, 23/24 , 23/25 or 23/26.
  • the length ratio of the sense and antisense strands of the siRNA is 19/21, 21/23, or 23/
  • the phosphorothioate group is present in at least one position selected from the group consisting of:
  • the first nucleotide end in the 3'-5' direction of the sense strand
  • the first nucleotide end in the 3'-5' direction of the antisense strand
  • the sense strand binds to at least one targeting ligand.
  • the present disclosure also provides an siRNA comprising a sense strand and an antisense strand, wherein the sense strand contains a nucleotide sequence I, the antisense strand contains a nucleotide sequence II, and the nucleotide sequence I A double-stranded region is formed at least partially reverse-complementary to nucleotide sequence II, which is represented by the following formula:
  • each Na ' and Nb ' independently represents a modified nucleotide or an unmodified nucleotide, wherein the modifications on Na' and Nb ' are different; each X ' is independently Na ' or Nb ';Y' is Na ' or Nb '.
  • the present disclosure also provides an siRNA comprising a sense strand and an antisense strand, wherein the sense strand contains a nucleotide sequence I, the antisense strand contains a nucleotide sequence II, and the nucleotide sequence I A double-stranded region is formed at least partially reverse-complementary to nucleotide sequence II, which is represented by the following formula:
  • each N a ' independently represents a modified nucleotide or an unmodified nucleotide
  • each N b ' independently represents a 2'-fluoro-modified nucleotide or a 2'-deoxy-modified nucleus nucleotide
  • each X' is independently Na ' or Nb '
  • Y' is Na ' or Nb '.
  • the modified nucleotides are independently selected from the group consisting of: 2'-alkoxy modified nucleotides, 2'-substituted alkoxy modified nucleotides, 2'-alkyl modified nucleotides nucleotides, 2'-substituted alkyl-modified nucleotides, 2'-amino-modified nucleotides, 2'-substituted amino-modified nucleotides, 2'-fluoro-modified cores nucleotides, 2'-deoxynucleotides, 2'-deoxy-2'-fluoromodified nucleotides, 3'-deoxy-thymidine nucleotides, isonucleotides, LNA, ENA, cET, UNA and GNA.
  • the modified nucleotides are independently of each other 2'-methoxy modified nucleotides.
  • unmodified ()nucleotides refer to nucleotides consisting of naturally occurring nucleoside bases, sugar groups, and intranucleoside linkages.
  • the unmodified (or) nucleotides are RNA nucleotides (ie, beta-D-ribonucleosides) or DNA nucleotides (ie, beta-D-deoxyribonucleosides).
  • X' is Nb '.
  • Y' is Nb '.
  • the modification on Na' is different from the modification on Nb '; in some embodiments, Na ' is a 2'-methoxy-modified nucleotide.
  • nucleotide sequence I is represented by the formula:
  • each Na and Nb independently represent modified nucleotides or unmodified nucleotides, and the modifications on Na and Nb are different;
  • the modified nucleotides are independently selected from the group consisting of: 2'-alkoxy-modified nucleotides, 2'-substituted alkoxy-modified nucleotides, 2'-alkoxy-modified nucleotides base-modified nucleotides, 2'-substituted alkyl-modified nucleotides, 2'-amino-modified nucleotides, 2'-substituted amino-modified nucleotides, 2'-fluoro-modified nucleotides, 2'-deoxynucleotides, 2'-deoxy-2'-fluoromodified nucleotides, 3'-deoxy-thymidine nucleotides, isonucleotides, LNA, ENA, cET , UNA and GNA;
  • the modified nucleotides are independently selected from the group consisting of: 2'-methoxy-modified nucleotides, 2'-fluoro-modified nucleotides, and 2'-deoxy-modified cores Glycosides;
  • Na is a 2'-methoxy-modified nucleotide and Nb is a 2'-fluoro-modified nucleotide or a 2'-deoxy-modified nucleotide.
  • nucleotide sequence II is represented by the formula:
  • pMD-AS19 5'-NmNfNmNfNmNfNmNmNfNmNmNfNmNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3'.
  • nucleotide sequence II is represented by the formula:
  • pMD-AS18 5'-NmNfNmN DNA NmNfNmNmN DNA NmNmN DNA NmNfNmNfNmNmNmNmNm-3', wherein N DNA indicates that the nucleotide at this position is a 2'-deoxy-modified nucleotide.
  • nucleotide sequence I is represented by the formula:
  • pMD-SS3 5’-NmNmNmNmNfNmNfNfNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3’.
  • nucleotide sequence I and said nucleotide sequence II there is no more than 3 base mismatches between nucleotide sequence I and said nucleotide sequence II; in some embodiments, nucleotide sequence I and said nucleotide sequence There are no more than 2 base mismatches between sequence II; in some embodiments, there are no more than 1 base mismatch between nucleotide sequence I and said nucleotide sequence II; in In some embodiments, there is no mismatch between nucleotide sequence I and the nucleotide sequence II.
  • At least one phosphate group in the sense strand and/or antisense strand is a phosphate group with a modifying group.
  • the phosphate group having the modifying group is a phosphorothioate group.
  • the phosphorothioate group is present in at least one position selected from the group consisting of:
  • the first nucleotide end in the 3'-5' direction of the sense strand
  • the first nucleotide end in the 3'-5' direction of the antisense strand
  • nucleotide sequence II is represented by the formula:
  • MD-AS19 5'-NmsNfsNmNfNmNfNmNmNfNmNmNfNmNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmsNmsNm-3'.
  • nucleotide sequence II is represented by the formula:
  • MD-AS18 5'-NmsNfsNmN DNA NmNfNmNmN DNA NmNmN DNA NmNfNmNfNmNmNmsNm-3', where N DNA indicates that the nucleotide at this position is a 2'-deoxy-modified nucleotide.
  • nucleotide sequence I is represented by the formula:
  • MD-SS3 5'-NmsNmsNmNmNfNmNfNfNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm
  • the siRNA comprises one or two blunt ends.
  • blunt end or blunt end are used interchangeably and refer to the absence of unpaired nucleotides or nucleotide analogs at a given end of an siRNA, ie, no nucleotide overhangs. In most cases, an siRNA that is blunt-ended at both ends will be double-stranded over its entire length.
  • the siRNA comprises an overhang having 1 to 4 unpaired nucleotides, eg, an overhang of 2 or 3 unpaired nucleotides.
  • the siRNA comprises an overhang located at the 3'-end of the antisense strand of the siRNA.
  • the sense strand binds to at least one targeting ligand.
  • the present disclosure provides an siRNA comprising a sense strand and an antisense strand, wherein the sequence of the sense strand is shown in the formula:
  • MD-SS3 5'-NmsNmsNmNmNfNmNfNfNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm
  • MD-AS19 5'-NmsNfsNmNfNmNfNmNmNfNmNmNfNmNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmsNmsNm-3'.
  • the present disclosure also provides an siRNA capable of inhibiting the expression of a target gene, comprising a sense strand and an antisense strand, wherein the sense strand contains a segment of nucleotide sequence I, and the antisense strand contains a segment of nucleotide sequence II, so the The nucleotide sequence I and the nucleotide sequence II are at least partially reverse complementary to form a double-stranded region, and the nucleotide sequence II is represented by the following formula:
  • each N a ' independently represents a modified nucleotide or an unmodified nucleotide
  • each N b ' independently represents a 2'-fluoro-modified nucleotide or a 2'-deoxy-modified nucleus nucleotide
  • each X' is independently Na ' or Nb '
  • Y' is Na ' or Nb '.
  • the modified nucleotides are independently selected from the group consisting of: 2'-alkoxy-modified nucleotides, 2'-substituted alkoxy-modified nucleotides, 2'-alkoxy-modified nucleotides base-modified nucleotides, 2'-substituted alkyl-modified nucleotides, 2'-amino-modified nucleotides, 2'-substituted amino-modified nucleotides, 2'-fluoro-modified nucleotides, 2'-deoxynucleotides, 2'-deoxy-2'-fluoromodified nucleotides, 3'-deoxy-thymidine nucleotides, isonucleotides, LNA, ENA, cET , UNA and GNA.
  • the modified nucleotides are independently 2'-methoxy modified nucleotides.
  • the present disclosure also provides an siRNA comprising a sense strand and an antisense strand, wherein the sense strand contains a nucleotide sequence III, the antisense strand contains a nucleotide sequence IV, a nucleotide sequence III and a nucleotide sequence III.
  • Sequence IV is at least partially reverse complementary to form a double-stranded region, and nucleotide sequence IV is selected from the following sequences:
  • pMD-AS1 5’-NmNfNmNfNmNfNmNfNmNmNmNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNm-3’;
  • pMD-AS2 5'-NmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • pMD-AS4 5'-NmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • pMD-AS5 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • pMD-AS8 5'-NmNfNmNmNmNfNmNmNmNmNmN DNA NmN DNA NmNfNmNfNmNmNmNmNmNmNmNm-3';
  • pMD-AS9 5'-NmNfNmNmNmNfNmN DNA N DNA NmNmNmNmNfNmNfNmNmNmNmNmNmNm-3';
  • pMD-AS10 5'-NmNfNmNfNmNfNmNfNmNmN DNA NmNmN DNA NmNfNmNfNmN DNA NmNmNm-3';
  • pMD-AS11 5’-NmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3’;
  • pMD-AS12 5'-NmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • pMD-AS17 5'-NmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNm-3';
  • pMD-AS18 5'-NmNfNmN DNA NmNfNmNmN DNA NmNmN DNA NmNfNmNfNmNmNmNmNm-3';
  • pMD-AS19 5'-NmNfNmNfNmNfNmNmNfNmNmNfNmNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3'.
  • nucleotide sequence III is selected from the following sequences:
  • pMD-1 5'-NfNmNfNmNfNmNfNfNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNf-3';
  • pMD-2 5'-NmNmNmNmNmNmNmNfNfNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • pMD-4 5'-NmNmNfNmNfNmNfNfNfNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • pMD-6 5'-NmNmNmNmNfNmNfNfN DNA NmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • pMD-7 5'-NmNmNmNmNfNmN DNA NfNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • pMD-8 5'-NmNmNmNmNfNmNfN DNA NfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • pMD-10 5'-NmNmNmNmNmN DNA NmNfNfNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • pMD-11 5'-NmNmNmNmNmN DNA NmNfNfN DNA NmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • pMD-12 5'-NmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • the iRNA comprises a sense strand and an antisense strand selected from the group consisting of:
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3'
  • Antisense strand 5'-NmNfNmNfNmNfNmNfNmNfNmNmNmNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNm-3';
  • Antisense strand 5'-NmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmNmNmNmNmN DNA NmN DNA NmNfNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNmNfNmN DNA N DNA NmNmNmNmNfNmNfNmNmNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNfNmNfNmNfNmNmN DNA NmNmN DNA NmNfNmNfNmN DNA NmNmN m-3';
  • Antisense strand 5'-NmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNm-3';
  • Antisense strand 5'-NmNfNmN DNA NmNfNmNmN DNA NmNmN DNA NmNfNmNfNmNmNmNmNmNm-3';
  • Antisense strand 5'-NmNfNmNfNmNfNmNmNfNmNmNfNmNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNm-3'.
  • At least one phosphate group in the sense strand and/or antisense strand is a phosphate group with a modifying group
  • the phosphate group having the modifying group is a phosphorothioate group.
  • the phosphorothioate group is present in at least one position selected from the group consisting of:
  • the first nucleotide end in the 3'-5' direction of the sense strand
  • the first nucleotide end in the 3'-5' direction of the antisense strand
  • nucleotide sequence IV is selected from the following sequences:
  • MD-AS1 5'-NmsNfsNmNfNmNfNmNfNmNmNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNmsNfsNm-3';
  • MD-AS2 5'-NmsNfsNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmsNm-3';
  • MD-AS3 5’-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNfNmNfNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmsNm-3’;
  • MD-AS4 5'-NmsNfsNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmsNm-3';
  • MD-AS5 5’-NmsNfsNmNmNmNfNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmsNm-3’;
  • MD-AS6 5’-NmsNfsNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmsNm-3’;
  • MD-AS8 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmN DNA NmN DNA NmNfNmNfNmNmNmNmsNm-3';
  • MD-AS9 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmsNm-3';
  • MD-AS10 5'-NmsNfsNmNfNmNfNmNmNmNmN DNA NmNmN DNA NmNfNmNfNmN DNA NmsNmsNm-3';
  • MD-AS11 5'-NmsNfsNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmsNm-3';
  • MD-AS12 5'-NmsNfsNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmNmNmNmNmNmsNm-3';
  • MD-AS17 5'-NmsNfsNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNmNmNmNmNfNmNmNmNmNfsNm-3';
  • MD-AS18 5'-NmsNfsNmN DNA NmNfNmNmN DNA NmNmN DNA NmNfNmNmNmNmsNm-3';
  • nucleotide sequence III is selected from the following sequences:
  • MD-SS1 5'-NfsNmsNfNmNfNmNfNfNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfsNmsNf-3';
  • MD-SS2 5’-NmsNmsNmNmNmNmNfNfNfNmNmNmNmNmNmNmNmNmNmNmsNm-3’;
  • MD-SS3 5'-NmsNmsNmNmNfNmNfNfNfNmNmNmNmNmNmNmNmNmNmNmNmNmsNm-3';
  • MD-SS4 5'-NmsNmsNfNmNfNmNfNfNfNmNmNmNmNmNmNmNmNmNmNmNmsNm-3';
  • MD-SS5 5'-NmsNmsNmNmNmNfNfNfNfNmNmNmNmNmNmNmNmNmnsNmsNm-3';
  • MD-SS6 5'-NmsNmsNmNmNfNmNfNfN DNA NmNmNmNmNmnsNmsNm-3';
  • MD-SS7 5'-NmsNmsNmNmNfNmN DNA NfNfNmNmNmNmNmNmnsNmsNm-3';
  • MD-SS8 5'-NmsNmsNmNmNfNmNfN DNA NfNmNmNmNmNmNmNmNmNmsNmsNm-3';
  • MD-SS9 5'-NmsNmsNmNmNfNmN DNA NfN DNA NmNmNmNmNmNmnsNmsNm-3';
  • MD-SS10 5'-NmsNmsNmNmNmN DNA NmNfNfNfNmNmNmNmNmNmNmnsNmsNm-3';
  • MD-SS11 5'-NmsNmsNmNmNmNmNmNmNmNmsNm-3';
  • MD-SS12 5'-NmsNmsNmNmNmNmNmNmNmNmNmNmsNm-3';
  • MD-SS13 5’-NmsNmsNmNmNfNmNmNfNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmsNm-3’.
  • the siRNA comprises a sense strand and an antisense strand selected from the group consisting of:
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNfNmNfNmNfNmNmNfNmNmNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmsNfsNm-3';
  • Antisense strand 5'-NmsNfsNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmNmsNms Nm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNmNmNmNmNmNmNmNmNmNmNfNmNmNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmNmNmNmNmN DNA NmN DNA NmNfNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNmNfNmN DNA N DNA NmNmNmNmNfNmNfNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNfNmNfNmNmNmNmN DNA NmNmN DNA NmNfNmNfNmN DNA NmsNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNfNmNmNmNmNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfNmNmNfsNm-3';
  • Antisense strand 5'-NmsNfsNmN DNA NmNfNmNmN DNA NmNmN DNA NmNfNmNfNmNmNmsNm-3';
  • Antisense strand 5'-NmsNfsNmNfNmNfNmNmNfNmNmNfNmNmNfNmNfNmNfNmNfNmNfNmNfNmsNmsNm-3'.
  • the present disclosure also provides an siRNA conjugate comprising the above-mentioned siRNA and a conjugation group attached to the siRNA.
  • the conjugation group comprises a pharmaceutically acceptable targeting ligand and a linker, and the siRNA, the linker, and the targeting ligand are sequentially covalently or non-covalently linked.
  • the linker is attached to the 3' end of the sense strand of the siRNA.
  • a lipophilic group such as cholesterol can be introduced at the end of the siRNA sense strand.
  • Protein, vitamin E, etc. in order to facilitate the interaction with intracellular mRNA through the cell membrane composed of lipid bilayers.
  • siRNA can also be modified by non-covalent bonds, such as binding phospholipid molecules, polypeptides, cationic polymers, etc. through hydrophobic bonds or ionic bonds to increase stability and biological activity.
  • the present disclosure also provides a method for preparing the aforementioned siRNA or siRNA conjugate, comprising: synthesizing the aforementioned siRNA or siRNA conjugate; and purifying the siRNA or the siRNA conjugate.
  • the method comprises the steps of: (1) synthesis of oligoribonucleotides; (2) deprotection; (3) purification and isolation; (4) annealing.
  • step (1) comprises: using solid support mediated phosphoramidite chemistry on an RNA synthesizer (eg Dr. Oligo48 synthesizer (Biolytic)) to separately synthesize sense and antisense oligoribose cores 200 nanomolar (nmol) specification for synthesis; the coupling time for all phosphoramidite (50 mM acetonitrile solution) is 6 minutes (min), using 5-ethylthio-1H-tetrazole (ETT) as activation agent (0.6M acetonitrile solution), using 0.22M PADS dissolved in 1:1 volume ratio of acetonitrile and collidine solution as sulfurization reagent, sulfurization reaction time is 3 minutes (min), using iodopyridine/water solution as oxidant, Oxidation reaction time 2 minutes (min); According to whether the target product has 5'-phosphorothioate modification, select above-mentioned vulcanization reaction conditions or oxidation reaction conditions;
  • an RNA synthesizer e
  • Step (2) includes: after the solid-phase synthesis is completed, the oligoribonucleotide is cleaved from the solid support, and soaked in a 3:1 volume ratio of 28% ammonia water and ethanol solution at 50° C. for 16 hours. Then high-speed centrifugation, the supernatant was transferred to another centrifuge tube, concentrated and evaporated to dryness to obtain the crude oligonucleotide;
  • Step (3) includes: purifying the obtained crude oligonucleotide using C18 reverse chromatography, the mobile phase is 0.1M TEAA and acetonitrile, and using 3% trifluoroacetic acid solution to remove DMTr, collecting the target product and lyophilizing;
  • Step (4) includes: annealing the sense strand and antisense strand synthesized according to the above steps respectively according to an equimolar ratio, so that they form a double-stranded structure through hydrogen bonds, and finally dissolving the obtained double-stranded siRNA in 1 ⁇ PBS , and adjusted to the concentration required for the experiment.
  • the present disclosure provides a siRNA comprising a sense strand and an antisense strand forming a double-stranded region;
  • the sense strand comprises at least a nucleotide sequence that differs from any one of the sense strands in Table 1 by no more than 3 nucleotides 15 consecutive nucleotides;
  • the antisense strand comprises at least 15 consecutive nucleotides that differ from any antisense strand in Table 1 by no more than 3 nucleotide sequences;
  • the siRNA contains one or more modified Nucleotides.
  • the antisense strand is at least partially reverse complementary to the target sequence to mediate RNA interference; in some embodiments, there are no more than 5, no more than 4, No more than 3, no more than 2, no more than 1 mismatch; in some embodiments, the antisense strand is fully reverse complementary to the target sequence.
  • the sense and antisense strands are at least partially reverse complementary to form a double-stranded region; in some embodiments, there are no more than 5, no more than 4, No more than 3, no more than 2, no more than 1 mismatch; in some embodiments, the sense and antisense strands are fully reverse complementary.
  • the sense strand of the siRNA contains three consecutive nucleotides that are 2'-fluoro-modified nucleotides.
  • the nucleotides at positions 2, 6, and 14 of the siRNA antisense strand of the present disclosure are each independently 2'-deoxynucleotides or 2'-deoxynucleotides in the 5'-end to 3'-end direction. '-Fluoromodified nucleotides.
  • the nucleotides at positions 2, 6, 14, and 16 of the antisense strand are each independently 2'-deoxynucleotides or 2'-fluoronucleotides in the 5'-end to 3'-end direction Substitute modified nucleotides.
  • the nucleotides at positions 2, 6, 9, 12, and 14 of the antisense strand are each independently 2'-deoxynucleotides or 2', in the 5'-end to 3'-end orientation - Fluorinated modified nucleotides.
  • the nucleotides at positions 2, 6, 10, 12, and 14 of the antisense strand are each independently 2'-deoxynucleotides or 2', in the 5'-end to 3'-end orientation - Fluorinated modified nucleotides.
  • the nucleotides at positions 2, 4, 6, 9, 12, 14, and 18 of the antisense strand are each independently a 2'-deoxynucleoside in the 5'-end to 3'-end orientation Acid or 2'-fluoro modified nucleotides.
  • the nucleotides at positions 2, 4, 6, 10, 12, 14, and 18 of the antisense strand are each independently a 2'-deoxynucleoside in the 5' to 3' direction Acid or 2'-fluoro modified nucleotides.
  • the nucleotides at positions 2, 4, 6, 9, 12, 14, 16, and 18 of the antisense strand are each independently 2'-deoxy, in the 5' to 3' direction Nucleotides or 2'-fluoromodified nucleotides.
  • the nucleotides at positions 2, 4, 6, 10, 12, 14, 16 and 18 of the antisense strand are each independently 2'-deoxy, in the 5' to 3' direction Nucleotides or 2'-fluoromodified nucleotides.
  • the nucleotides at positions 2, 4, 6, 9, 10, 12, 14, 16 and 18 of the antisense strand are each independently 2' in the 5' to 3' direction - Deoxynucleotides or 2'-fluoromodified nucleotides.
  • nucleotides 2, 6, and 14 of the antisense strand are each independently 2'-fluoro-modified nucleotides in the 5'-end to 3'-end direction;
  • the nucleotides at positions 2, 6, 14, and 16 of the antisense strand are each independently 2'-fluoro-modified nucleotides in the 5'-end to 3'-end direction;
  • the nucleotides at positions 2, 6, 12, and 14 of the antisense strand are each independently 2'-fluoro-modified nucleotides in the 5'-end to 3'-end direction;
  • the nucleotides at positions 2, 4, 6, 12, 14, 16 and 18 of the antisense strand are each independently 2'-fluoro-modified in the 5'-end to 3'-end direction nucleotides;
  • the nucleotides at positions 2, 4, 6, 9, 12, 14, 16, and 18 of the antisense strand are each independently 2'-fluoro in the 5'-end to 3'-end orientation Substitute modified nucleotides;
  • the nucleotides at positions 2, 4, 6, 10, 12, 14, 16 and 18 of the antisense strand are each independently 2'-fluoro, in the 5' to 3' direction Substitute modified nucleotides.
  • the sense and antisense strands each independently have 16 to 35, 16 to 34, 17 to 34, 17 to 33, 18 to 33, 18 to 32, 18 to 31, 18 to 30, 18 to 29, 18 to 28, 18 to 27, 18 to 26, 18 to 25, 18 to 24, 18 to 23, 19 to 25, 19 to 24, or 19 to 23 nucleotides.
  • the sense and antisense strands are the same or different in length, the sense strand is 19-23 nucleotides in length and the antisense strand is 19-26 nucleotides in length.
  • the length ratio of the sense and antisense strands of the siRNA provided by the present disclosure can be 19/20, 19/21, 19/22, 19/23, 19/24, 19/25, 19/26, 20/20 , 20/21, 20/22, 20/23, 20/24, 20/25, 20/26, 21/20, 21/21, 21/22, 21/23, 21/24, 21/25, 21 /26, 22/20, 22/21, 22/22, 22/23, 22/24, 22/25, 22/26, 23/20, 23/21, 23/22, 23/23, 23/24 , 23/25 or 23/26.
  • the length ratio of the sense and antisense strands of the siRNA is 19/21, 21/23, or 23
  • the siRNA comprises one or two blunt ends.
  • the siRNA comprises an overhang having 1 to 4 unpaired nucleotides, eg, 1, 2, 3, 4.
  • the siRNAs of the present disclosure comprise an overhang located at the 3' end of the antisense strand of the siRNA.
  • At least one additional nucleotide in the sense and/or antisense strand is a modified nucleotide.
  • the modified nucleotides are independently selected from the group consisting of deoxy-nucleotides, 3'-terminal deoxy-thymidine nucleotides, 2'-O-methyl modified nucleotides, 2'- Fluorine modified nucleotides, 2'-deoxy-modified nucleotides, locked nucleotides, unlocked nucleotides, conformationally restricted nucleotides, restricted ethyl nucleotides, abasic nucleotides , 2'-amino-modified nucleotides, 2'-O-allyl-modified nucleotides, 2'-C-alkyl-modified nucleotides, 2'-hydroxy-modified nucleotides Acids, 2'-methoxyethyl-modified nucleotides, 2'-O-alkyl-modified nucleotides, morpholino nucleotides, phosphoramidates, nucle
  • the modified nucleotides are independently selected from the group consisting of: 2'-alkoxy modified nucleotides, 2'-substituted alkoxy modified nucleotides, 2'-alkyl modified nucleotides nucleotides, 2'-substituted alkyl-modified nucleotides, 2'-amino-modified nucleotides, 2'-substituted amino-modified nucleotides, 2'-fluoro-modified cores nucleotides, 2'-deoxynucleotides, 2'-deoxy-2'-fluoromodified nucleotides, 3'-deoxy-thymidine nucleotides, isonucleotides, LNA, ENA, cET, UNA , GNA, or a nucleotide comprising a chemical modification of formula (I) of the present disclosure or a tautomer modification thereof.
  • a 2'-fluoromodified nucleotide refers to a nucleotide formed by substituting the hydroxyl group at the 2' position of the ribosyl of the nucleotide with fluorine.
  • the 2'-alkoxy-modified nucleotide is a 2'-methoxy-modified nucleotide (2'-OMe).
  • a 2'-substituted alkoxy-modified nucleotide for example, can be a 2'-O-methoxyethyl-modified nucleotide (2'-MOE) or a 2'-amino Modified Nucleotides (2'- NH2 ).
  • the modified nucleotides are independently selected from the group consisting of: 2'-methoxy-modified nucleotides, 2'-fluoro-modified nucleotides, 2'-deoxy-modified nucleotides Or the nucleotides comprising the chemical modification represented by the formula (I) of the present disclosure or its tautomer modification.
  • formula (I) is selected from
  • Y is selected from O, NH and S;
  • Each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H or C 1 -C 6 alkyl;
  • J 2 is H or C 1 -C 6 alkyl
  • Q1 is Q 2 is R 2 ; or Q 1 is R 2 and Q 2 is
  • J 1 is H or C 1 -C 6 alkyl
  • R 1 and R 2 are directly connected to form a ring
  • B is a base or a base analog
  • R1 is not H.
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • Formula (I) is selected from Formula (I-1):
  • Y is selected from O, NH and S;
  • Each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H or C 1 -C 6 alkyl;
  • Each J 1 , J 2 is independently H or C 1 -C 6 alkyl
  • R 1 and R 2 are directly connected to form a ring
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • Formula (I) is selected from Formula (I-2):
  • Y is selected from O, NH and S;
  • Each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H or C 1 -C 6 alkyl;
  • Each J 1 , J 2 is independently H or C 1 -C 6 alkyl
  • R 1 and R 2 are directly connected to form a ring
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H or C 1 -C 3 alkyl;
  • Each J 1 , J 2 is independently H or C 1 -C 3 alkyl
  • R1 and R2 are directly connected to form a ring.
  • B is a base or a base analog; for example, those selected from purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H, methyl, B radical, n-propyl or isopropyl;
  • Each J 1 , J 2 is independently H or methyl
  • R1 and R2 are directly connected to form a ring.
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H, methyl, B radical, n-propyl or isopropyl;
  • Each J 1 , J 2 is independently H or methyl
  • R1 and R2 are directly connected to form a ring.
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • Each J 1 , J 2 is independently H;
  • R 1 is selected from H, methyl and CH 2 OH;
  • R 2 is selected from H, OH, NH 2 , methyl and CH 2 OH;
  • R3 is selected from H, OH, NH2 , methyl and CH2OH ;
  • R 1 and R 2 are directly connected to form a ring
  • B is a base or a base analog; for example, those selected from purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • Each J 1 , J 2 is independently H;
  • R 1 is selected from H, methyl and CH 2 OH;
  • R 2 is selected from H, methyl and CH 2 OH;
  • R3 is selected from H, OH, NH2 , methyl and CH2OH ;
  • R 1 and R 2 are directly connected to form a ring
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • the chemical modification of formula (I) is selected from:
  • B is a base or a base analog; for example, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • the chemical modification represented by the formula (I) is selected from:
  • B is a base or a base analog; for example, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • nucleotides comprising the chemical modifications represented by formula (I) or tautomer modifications thereof are selected from the group consisting of cores comprising chemical modifications represented by formula (I') or tautomer modifications thereof Glycosides,
  • Y is selected from O, NH and S;
  • Each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H or C 1 -C 6 alkyl;
  • J 2 is H or C 1 -C 6 alkyl
  • Q1 ' is Q 2' is R 2 ; or Q 1' is R 2 and Q 2' is
  • J 1 is H or C 1 -C 6 alkyl
  • R 1 and R 2 are directly connected to form a ring
  • B is a base or a base analog
  • M is O or S
  • R1 is not H.
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • formula (I') is selected from formula (I'-1):
  • Y is selected from O, NH and S;
  • Each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H or C 1 -C 6 alkyl;
  • Each J 1 , J 2 is independently H or C 1 -C 6 alkyl
  • M is O or S
  • R 1 and R 2 are directly connected to form a ring
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • formula (I') is selected from formula (I'-2):
  • Y is selected from O, NH and S;
  • Each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H or C 1 -C 6 alkyl;
  • Each J 1 , J 2 is independently H or C 1 -C 6 alkyl
  • R 1 and R 2 are directly connected to form a ring
  • M is O or S
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • R1 is not H.
  • each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H or C 1 -C 3 alkyl;
  • Each J 1 , J 2 is independently H or C 1 -C 3 alkyl
  • R1 and R2 are directly connected to form a ring.
  • each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H, methyl, B radical, n-propyl or isopropyl;
  • Each J 1 , J 2 is independently H or methyl
  • R1 and R2 are directly connected to form a ring.
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • each X is independently selected from CR 4 (R 4 ′), S, NR 5 and NH-CO, wherein R 4 , R 4 ′, R 5 are each independently H, methyl, B radical, n-propyl or isopropyl;
  • Each J 1 , J 2 is independently H or methyl
  • R1 and R2 are directly connected to form a ring.
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • Each J 1 , J 2 is independently H;
  • R 1 is selected from H, methyl and CH 2 OH;
  • R 2 is selected from H, OH, NH 2 , methyl and CH 2 OH;
  • R3 is selected from H, OH, NH2 , methyl and CH2OH ;
  • R1 and R2 are directly connected to form a ring.
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • Each J 1 , J 2 is independently H;
  • R 1 is selected from H, methyl and CH 2 OH;
  • R 2 is selected from H, methyl and CH 2 OH;
  • R3 is selected from H, OH, NH2 , methyl and CH2OH ;
  • R1 and R2 are directly connected to form a ring.
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • the chemical modification represented by the formula (I') is selected from:
  • M is O or S
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • the chemical modification represented by the formula (I') is selected from:
  • M is O or S
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • the chemical modification represented by the formula (I') is selected from:
  • M is O or S
  • B is a base or a base analog; eg, those selected from the group consisting of purine bases, pyrimidine bases, indole, 5-nitroindole, and 3-nitropyrrole;
  • B is selected from the group consisting of adenine, guanine, isoguanine, hypoxanthine, xanthine, C2 modified purine, N8 modified purine, 2,6-diaminopurine, 6-dimethylaminopurine , 2-aminopurine, N6-alkyladenine, O6-alkylguanine, 7-deazapurine, cytosine, 5-methylcytosine, isocytosine, pseudocytosine, uracil, pseudouracil , 2-thiouridine, 4-thiouridine, C5-modified pyrimidine, thymine, indole, 5-nitroindole and 3-nitropyrrole;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitro indole and 3-nitropyrrole;
  • B is selected from the base at the position corresponding to the modified nucleotide of the antisense strand.
  • the chemical modification represented by the formula (I') includes but is not limited to:
  • adenine in their structure is replaced by a guanine, cytosine, uracil or thymine.
  • the modified nucleotides are located at positions 2 to 8 of the antisense strand in its 5' region.
  • the modified nucleotide is located at position 5, 6 or 7 in the 5' region of the antisense strand.
  • B when the chemical modification represented by formula (I) or its tautomer modification is at the 5th position in its 5' region, B is selected from adenine, guanine, 2,6-diaminopurine , 6-dimethylaminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitroindole and 3-nitropyrrole; preferably, B is when the antisense strand is in The base at the corresponding position in position 5 of its 5' region.
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethyl aminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitroindole and 3-nitropyrrole; preferably, B is the first position of the antisense strand in its 5' region The base at the corresponding position in position 6;
  • B is selected from adenine, guanine, 2,6-diaminopurine, 6-dimethyl aminopurine, 2-aminopurine, cytosine, uracil, thymine, indole, 5-nitroindole and 3-nitropyrrole; preferably, B is the first position of the antisense strand in its 5' region The base at the corresponding position in position 7.
  • all nucleotides are modified nucleotides.
  • the sense strand contains a nucleotide sequence (5'-3') of the formula:
  • each X is independently Na or Nb ;
  • each X' is independently Na ' or Nb '
  • Y' is Na' or Nb '
  • W' represents a 2'-methoxy-modified nucleotide or a nucleotide represented by formula (I)
  • the chemically modified nucleotides of , or their tautomers Na is a 2'-methoxy-modified nucleotide
  • N b is a 2'-fluoro-modified nucleotide.
  • the antisense strand contains a nucleotide sequence (5'-3') of the formula:
  • each X' is independently Na ' or Nb '
  • Y' is Na' or Nb '
  • W' represents a 2'-methoxy-modified nucleotide or a nucleotide represented by formula (I)
  • the chemically modified nucleotides of , or their tautomers Na is a 2'-methoxy-modified nucleotide
  • N b is a 2'-fluoro-modified nucleotide.
  • the sense strand contains a nucleotide sequence of the formula:
  • Na is a 2'-methoxy-modified nucleotide
  • N b is a 2'-fluoro-modified nucleotide
  • the antisense strand contains a nucleotide sequence of the formula:
  • each X' is independently Na ' or Nb ', Y' is Na ' or Nb ';Na' is a 2'-methoxy-modified nucleotide, and Nb ' is 2' -Fluoro-modified nucleotide; W' represents a 2'-methoxy-modified nucleotide or a nucleotide modified by a chemical modification represented by formula (I) or a tautomer thereof.
  • the antisense strand contains a nucleotide sequence of the formula:
  • N a ' is 2'-methoxy-modified nucleotide
  • N b ' is 2'-fluoro-modified nucleotide
  • W' represents 2'-methoxy-modified nucleotide or formula The chemically modified nucleotide shown in (I) or its tautomer modified.
  • W' represents a 2'-methoxy-modified nucleotide or a nucleotide comprising a chemical modification represented by formula (I) or a tautomer modification thereof;
  • formula (I) is selected from:
  • B is selected from guanine, adenine, cytosine or uracil; in some specific embodiments, B is selected from the base at the corresponding position in the 7th position of the 5' region of the antisense strand.
  • formula (I) is selected from:
  • M is O or S; wherein: B is selected from guanine, adenine, cytosine or uracil; in some specific embodiments, B is selected from the antisense strand corresponding to the 7th position in its 5' region position of the base.
  • M is S. In some specific embodiments, M is O.
  • At least one phosphate group in the sense and/or antisense strand is a phosphate group with a modifying group that imparts increased stability of the siRNA in a biological sample or environment ;
  • the phosphate group with the modified group is a phosphorothioate group.
  • a phosphorothioate group refers to a phosphodiester group in which a non-bridging oxygen atom is replaced by a sulfur atom.
  • the phosphorothioate group is present in at least one position selected from the group consisting of:
  • the first nucleotide end in the 3'-5' direction of the sense strand
  • the first nucleotide end in the 3'-5' direction of the antisense strand
  • a plurality of phosphorothioate groups are included in the sense and/or antisense strands, the phosphorothioate groups being present in:
  • the first nucleotide end in the 3'-5' direction of the sense strand, and/or,
  • the sense strand is between the first nucleotide and the second nucleotide in the 3'-5' direction.
  • the sense strand has a phosphorothioate group at:
  • the first nucleotide end in the 3'-5' direction of the sense strand
  • the first nucleotide end of the sense strand in the 3'-5' direction.
  • the antisense strand has a phosphorothioate group at:
  • the sense strand is selected from a nucleotide sequence of the formula:
  • Nm represents any nucleotide modified by 2'-methoxy group, such as C, G, U, A, T modified by 2'-methoxy group
  • Nf represents any nucleotide modified by 2'-fluoro, For example, 2'-fluoro-modified C, G, U, A, T;
  • the antisense strand has a nucleotide sequence of the formula:
  • Nm' represents any nucleotide modified by 2'-methoxy, such as C, G, U, A, T modified by 2'-methoxy
  • Nf' represents any nucleotide modified by 2'-fluoro Acids such as 2'-fluoro-modified C, G, U, A, T;
  • W' represents a 2'-methoxy-modified nucleotide or a nucleotide modified by a chemical modification represented by formula (I) or a tautomer thereof;
  • formula (I) is selected from:
  • B is selected from guanine, adenine, cytosine or uracil, and in some embodiments is selected from the base of the antisense strand corresponding to position 7 in its 5' region.
  • formula (I) is selected from:
  • M is O or S; wherein: B is selected from guanine, adenine, cytosine or uracil; in some specific embodiments, B is selected from the antisense strand corresponding to the 7th position in its 5' region position of the base.
  • M is S. In some specific embodiments, M is O.
  • the sense strand comprises at least 15 contiguous nucleotides that differ from any of SEQ ID NO: 1 to SEQ ID NO: 23 by no more than 3 nucleotide sequences;
  • the antisense strand comprises at least 15 contiguous nucleotides that differ from any of SEQ ID NO: 24 to SEQ ID NO: 46 by no more than 3 nucleotide sequences;
  • the sense strand comprises at least 19 contiguous nucleotides that differ from any of SEQ ID NO: 1 to SEQ ID NO: 23 by no more than 3 nucleotide sequences; in some embodiments, The nucleotide sequences differ by no more than 1 nucleotide.
  • the antisense strand comprises at least 21 contiguous nucleotides that differ from any of SEQ ID NO: 24 to SEQ ID NO: 46 by no more than 3 nucleotide sequences; in some embodiments, The nucleotide sequences differ by no more than 1 nucleotide.
  • the sense strand nucleotide sequence is selected from the group consisting of: the nucleotide sequence of any one of EQ ID NO: 1 to SEQ ID NO: 23;
  • the antisense strand nucleotide sequence is selected from the group consisting of: the nucleotide sequence of any one of SEQ ID NO: 47 to SEQ ID NO: 69, wherein W' represents 2'-methoxy-modified Nucleotides, or nucleotides comprising chemical modifications represented by formula (I) or tautomer modifications thereof;
  • formula (I) is selected from:
  • B is selected from guanine, adenine, cytosine or uracil, and in some embodiments is selected from the base of the antisense strand corresponding to position 7 in its 5' region.
  • formula (I) is selected from:
  • M is O or S; wherein: B is selected from guanine, adenine, cytosine or uracil; in some specific embodiments, B is selected from the antisense strand corresponding to the 7th position in its 5' region position of the base.
  • M is S. In some specific embodiments, M is O.
  • the sense strand is selected from any one of SEQ ID NOs: 70-77, 86-93, 102-161, 282-293, 306-425;
  • the antisense strand is selected from any one of SEQ ID NOs: 78-85, 94-101, 162-261, 294-305;
  • the siRNA of the present disclosure is selected from,
  • the sense strand is selected from SEQ ID NO:142
  • the antisense strand is selected from SEQ ID NO:202, SEQ ID NO:222, or SEQ ID NO:242;
  • the sense strand is selected from SEQ ID NO: 143
  • the antisense strand is selected from SEQ ID NO: 203, SEQ ID NO: 223, or SEQ ID NO: 243;
  • the sense strand is selected from SEQ ID NO: 144, and the antisense strand is selected from SEQ ID NO: 204, SEQ ID NO: 224, or SEQ ID NO: 244;
  • the sense strand is selected from SEQ ID NO:145, and the antisense strand is selected from SEQ ID NO:205, SEQ ID NO:225, or SEQ ID NO:245;
  • the sense strand is selected from SEQ ID NO:146
  • the antisense strand is selected from SEQ ID NO:206, SEQ ID NO:226, or SEQ ID NO:246;
  • the sense strand is selected from SEQ ID NO: 147, and the antisense strand is selected from SEQ ID NO: 207, SEQ ID NO: 227, or SEQ ID NO: 247;
  • the sense strand is selected from SEQ ID NO:148, and the antisense strand is selected from SEQ ID NO:208, SEQ ID NO:228, or SEQ ID NO:248;
  • the sense strand is selected from SEQ ID NO:149
  • the antisense strand is selected from SEQ ID NO:209, SEQ ID NO:229, or SEQ ID NO:249;
  • the sense strand is selected from SEQ ID NO: 150
  • the antisense strand is selected from SEQ ID NO: 210, SEQ ID NO: 230, or SEQ ID NO: 250;
  • the sense strand is selected from SEQ ID NO:151
  • the antisense strand is selected from SEQ ID NO:211, SEQ ID NO:231, or SEQ ID NO:251;
  • the sense strand is selected from SEQ ID NO: 152
  • the antisense strand is selected from SEQ ID NO: 212, SEQ ID NO: 232, or SEQ ID NO: 252;
  • the sense strand is selected from SEQ ID NO: 153
  • the antisense strand is selected from SEQ ID NO: 213, SEQ ID NO: 233, or SEQ ID NO: 253;
  • the sense strand is selected from SEQ ID NO: 154, and the antisense strand is selected from SEQ ID NO: 214, SEQ ID NO: 234, or SEQ ID NO: 254;
  • the sense strand is selected from SEQ ID NO: 155
  • the antisense strand is selected from SEQ ID NO: 215, SEQ ID NO: 235, or SEQ ID NO: 255;
  • the sense strand is selected from SEQ ID NO: 156
  • the antisense strand is selected from SEQ ID NO: 216, SEQ ID NO: 236, or SEQ ID NO: 256;
  • the sense strand is selected from SEQ ID NO: 157
  • the antisense strand is selected from SEQ ID NO: 217, SEQ ID NO: 237, or SEQ ID NO: 257;
  • the sense strand is selected from SEQ ID NO: 158
  • the antisense strand is selected from SEQ ID NO: 218, SEQ ID NO: 238, or SEQ ID NO: 258;
  • the sense strand is selected from SEQ ID NO: 159
  • the antisense strand is selected from SEQ ID NO: 219, SEQ ID NO: 239, or SEQ ID NO: 259;
  • the sense strand is selected from SEQ ID NO: 160
  • the antisense strand is selected from SEQ ID NO: 220, SEQ ID NO: 240, or SEQ ID NO: 260;
  • the sense strand is selected from SEQ ID NO:161, and the antisense strand is selected from SEQ ID NO:221, SEQ ID NO:241, or SEQ ID NO:261;
  • the siRNA of the present disclosure is selected from,
  • the sense strand is selected from SEQ ID NO:346, and the antisense strand is selected from SEQ ID NO:202, SEQ ID NO:222, or SEQ ID NO:242;
  • the sense strand is selected from SEQ ID NO:347, and the antisense strand is selected from SEQ ID NO:203, SEQ ID NO:223, or SEQ ID NO:243;
  • the sense strand is selected from SEQ ID NO:348, and the antisense strand is selected from SEQ ID NO:204, SEQ ID NO:224, or SEQ ID NO:244;
  • the sense strand is selected from SEQ ID NO:349, and the antisense strand is selected from SEQ ID NO:205, SEQ ID NO:225, or SEQ ID NO:245;
  • the sense strand is selected from SEQ ID NO:350, and the antisense strand is selected from SEQ ID NO:206, SEQ ID NO:226, or SEQ ID NO:246;
  • the sense strand is selected from SEQ ID NO:351
  • the antisense strand is selected from SEQ ID NO:207, SEQ ID NO:227, or SEQ ID NO:247;
  • the sense strand is selected from SEQ ID NO:352
  • the antisense strand is selected from SEQ ID NO:208, SEQ ID NO:228, or SEQ ID NO:248;
  • the sense strand is selected from SEQ ID NO:353
  • the antisense strand is selected from SEQ ID NO:209, SEQ ID NO:229, or SEQ ID NO:249;
  • the sense strand is selected from SEQ ID NO:354, and the antisense strand is selected from SEQ ID NO:210, SEQ ID NO:230, or SEQ ID NO:250;
  • the sense strand is selected from SEQ ID NO:355, and the antisense strand is selected from SEQ ID NO:211, SEQ ID NO:231, or SEQ ID NO:251;
  • the sense strand is selected from SEQ ID NO:356, and the antisense strand is selected from SEQ ID NO:212, SEQ ID NO:232, or SEQ ID NO:252;
  • the sense strand is selected from SEQ ID NO:357
  • the antisense strand is selected from SEQ ID NO:213, SEQ ID NO:233, or SEQ ID NO:253;
  • the sense strand is selected from SEQ ID NO:358, and the antisense strand is selected from SEQ ID NO:214, SEQ ID NO:234, or SEQ ID NO:254;
  • the sense strand is selected from SEQ ID NO:359
  • the antisense strand is selected from SEQ ID NO:215, SEQ ID NO:235, or SEQ ID NO:255;
  • the sense strand is selected from SEQ ID NO:360, and the antisense strand is selected from SEQ ID NO:216, SEQ ID NO:236, or SEQ ID NO:256;
  • the sense strand is selected from SEQ ID NO:361, and the antisense strand is selected from SEQ ID NO:217, SEQ ID NO:237, or SEQ ID NO:257;
  • the sense strand is selected from SEQ ID NO:362, and the antisense strand is selected from SEQ ID NO:218, SEQ ID NO:238, or SEQ ID NO:258;
  • the sense strand is selected from SEQ ID NO:363, and the antisense strand is selected from SEQ ID NO:219, SEQ ID NO:239, or SEQ ID NO:259;
  • the sense strand is selected from SEQ ID NO:364, and the antisense strand is selected from SEQ ID NO:220, SEQ ID NO:240, or SEQ ID NO:260;
  • the sense strand is selected from SEQ ID NO:365, and the antisense strand is selected from SEQ ID NO:221, SEQ ID NO:241, or SEQ ID NO:261;
  • the siRNA of the present disclosure is selected from,
  • the sense strand is selected from SEQ ID NO:406, and the antisense strand is selected from SEQ ID NO:202, SEQ ID NO:222, or SEQ ID NO:242;
  • the sense strand is selected from SEQ ID NO:407
  • the antisense strand is selected from SEQ ID NO:203, SEQ ID NO:223, or SEQ ID NO:243;
  • the sense strand is selected from SEQ ID NO:408, and the antisense strand is selected from SEQ ID NO:204, SEQ ID NO:224, or SEQ ID NO:244;
  • the sense strand is selected from SEQ ID NO:409, and the antisense strand is selected from SEQ ID NO:205, SEQ ID NO:225, or SEQ ID NO:245;
  • the sense strand is selected from SEQ ID NO:410
  • the antisense strand is selected from SEQ ID NO:206, SEQ ID NO:226, or SEQ ID NO:246;
  • the sense strand is selected from SEQ ID NO:411, and the antisense strand is selected from SEQ ID NO:207, SEQ ID NO:227, or SEQ ID NO:247;
  • the sense strand is selected from SEQ ID NO:412, and the antisense strand is selected from SEQ ID NO:208, SEQ ID NO:228, or SEQ ID NO:248;
  • the sense strand is selected from SEQ ID NO:413, and the antisense strand is selected from SEQ ID NO:209, SEQ ID NO:229, or SEQ ID NO:249;
  • the sense strand is selected from SEQ ID NO:414, and the antisense strand is selected from SEQ ID NO:210, SEQ ID NO:230, or SEQ ID NO:250;
  • the sense strand is selected from SEQ ID NO:415
  • the antisense strand is selected from SEQ ID NO:211, SEQ ID NO:231, or SEQ ID NO:251;
  • the sense strand is selected from SEQ ID NO:416, and the antisense strand is selected from SEQ ID NO:212, SEQ ID NO:232, or SEQ ID NO:252;
  • the sense strand is selected from SEQ ID NO:417
  • the antisense strand is selected from SEQ ID NO:213, SEQ ID NO:233, or SEQ ID NO:253;
  • the sense strand is selected from SEQ ID NO:418, and the antisense strand is selected from SEQ ID NO:214, SEQ ID NO:234, or SEQ ID NO:254;
  • the sense strand is selected from SEQ ID NO:419, and the antisense strand is selected from SEQ ID NO:215, SEQ ID NO:235, or SEQ ID NO:255;
  • the sense strand is selected from SEQ ID NO:420, and the antisense strand is selected from SEQ ID NO:216, SEQ ID NO:236, or SEQ ID NO:256;
  • the sense strand is selected from SEQ ID NO:421, and the antisense strand is selected from SEQ ID NO:217, SEQ ID NO:237, or SEQ ID NO:257;
  • the sense strand is selected from SEQ ID NO:422, and the antisense strand is selected from SEQ ID NO:218, SEQ ID NO:238, or SEQ ID NO:258;
  • the sense strand is selected from SEQ ID NO:423, and the antisense strand is selected from SEQ ID NO:219, SEQ ID NO:239, or SEQ ID NO:259;
  • the sense strand is selected from SEQ ID NO:424, and the antisense strand is selected from SEQ ID NO:220, SEQ ID NO:240, or SEQ ID NO:260;
  • the sense strand is selected from SEQ ID NO:425, and the antisense strand is selected from SEQ ID NO:221, SEQ ID NO:241, or SEQ ID NO:261.
  • the siRNA described above when contacted with cells expressing the target gene, is screened for activity by, for example, psiCHECK and luciferase reporter gene assays, other methods such as PCR or branched DNA (bDNA) based methods, or protein-based Methods, as determined by immunofluorescence assays such as Western Blot or flow cytometry, the above siRNAs inhibit the expression of the target gene by at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30% , at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%.
  • psiCHECK and luciferase reporter gene assays other methods
  • the siRNA described above when contacted with cells expressing the target gene, is screened for activity by, for example, psiCHECK and luciferase reporter gene assays, other methods such as PCR or branched DNA (bDNA) based methods, or protein-based Methods, such as immunofluorescence analysis, such as Western Blot or flow cytometry, the percentage of residual expression of target gene mRNA caused by the above siRNA is not higher than 99%, not higher than 95%, not higher than 90%, not higher at 85%, no more than 80%, no more than 75%, no more than 70%, no more than 65%, no more than 60%, no more than 55%, no more than 50%, no more than 45% %, no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, or no more than 10%.
  • psiCHECK and luciferase reporter gene assays other methods such as PCR or branched DNA (bDNA) based methods,
  • siRNAs comprising chemical modifications of the present disclosure when contacted with cells expressing the target gene, are screened for, for example, psiCHECK activity and luciferase reporter gene assays, others such as PCR or branched DNA (bDNA) based methods , or protein-based methods, such as assayed by immunofluorescence assays, such as Western Blot, or flow cytometry, comprising chemical modifications of the present disclosure, such as siRNAs comprising chemical modifications of formula (I) while maintaining target activity , reduces off-target activity by at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, or at least 75%.
  • psiCHECK activity and luciferase reporter gene assays others such as PCR or branched DNA (bDNA) based methods , or protein-based methods, such as assayed by immunofluorescence assays, such
  • siRNAs comprising chemical modifications of the present disclosure when contacted with cells expressing the target gene, are screened for, for example, psiCHECK activity and luciferase reporter gene assays, others such as PCR or branched DNA (bDNA) based methods , or protein-based methods, such as assayed by immunofluorescence assays, such as Western Blot, or flow cytometry, comprising chemical modifications of the present disclosure, such as siRNAs comprising chemical modifications of formula (I) that reduce on-target activity by up to 20% %, up to 19%, up to 15%, up to 10%, up to 5%, or more than 1% while reducing off-target activity by at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70% or at least 75%.
  • psiCHECK activity and luciferase reporter gene assays others such as PCR or branched DNA (
  • siRNAs comprising chemical modifications of the present disclosure when contacted with cells expressing the target gene, are screened for, for example, psiCHECK activity and luciferase reporter gene assays, others such as PCR or branched DNA (bDNA) based methods , or a protein-based method, as determined by an immunofluorescence assay, such as Western Blot, or flow cytometry, comprising a chemical modification of the present disclosure, such as an siRNA comprising a chemical modification of formula (I) that increases on-target activity by at least 1 %, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, At least 65%, at least 70%, at least 75%, or at least 80% while reducing off-target activity by at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%
  • the present disclosure also provides an siRNA conjugate comprising any one of the siRNAs described above and a targeting ligand linked to the siRNA.
  • the siRNA and the targeting ligand are covalently or non-covalently linked.
  • the targeting ligand targets the liver; in some embodiments, the targeting ligand binds the asialoglycoprotein receptor (ASGPR); in some embodiments, the targeting ligand comprises A cluster of galactose or a cluster of galactose derivatives selected from N-acetyl-galactosamine, N-trifluoroacetylgalactosamine, N-propionylgalactosamine, N-n-butyryl Galactosamine or N-isobutyrylgalactosamine.
  • the targeting ligand is attached to the 3' end of the sense strand of the siRNA.
  • a lipophilic group such as cholesterol can be introduced at the end of the sense strand of siRNA.
  • Protein, vitamin E, etc. in order to facilitate the interaction with intracellular mRNA through the cell membrane composed of lipid bilayers.
  • siRNA can also be modified by non-covalent bonds, such as binding phospholipid molecules, polypeptides, cationic polymers, etc. through hydrophobic bonds or ionic bonds to increase stability and biological activity.
  • the targeting ligand is attached to the end of the siRNA through a phosphate group, phosphorothioate group, or phosphonate group. In some embodiments, the targeting ligand is indirectly attached to the end of the siRNA through a phosphate group, phosphorothioate group, or phosphonate group. In some embodiments, the targeting ligand is directly attached to the end of the siRNA through a phosphate, phosphorothioate, or phosphonic acid group. In some embodiments, the targeting ligand is directly attached to the end of the siRNA through a phosphate or phosphorothioate group. In some embodiments, the targeting ligand is directly attached to the 3' end of the siRNA sense strand through a phosphate or phosphorothioate group.
  • the targeting ligand structure is shown in the following formula (II),
  • T is the targeting moiety
  • E is the branching group
  • L1 is the linker moiety
  • L2 is the tethering moiety between the targeting moiety and the branching group, wherein i is selected from an integer from 1 to 10, such as 1, 2 , 3, 4, 5, 6, 7, 8, 9, 10.
  • i is selected from an integer from 2 to 8.
  • i is selected from an integer from 3 to 5.
  • L 1 is N
  • R 9 and R 10 are each independently selected from -S-, -NH-, -O-, -C(O)-, -OC(O)-, -C(O)O-, -NHC(O)- , -C(O)NH-, -CH2- , -CH2NH-, -CH2O- , -NH-C(O) -CH2- , -C(O) -CH2 - NH-, -NH(CO)NH-, 3-12-membered heterocyclic group, the -CH 2 - is optionally substituted with a substituent selected from halogen, alkyl, alkoxy, alkylamino, and the alkyl is optionally further is substituted with a substituent selected from hydroxy, amino, halogen;
  • R 11 is selected from deuterium, halogen, alkyl, amino, cyano, nitro, alkenyl, alkynyl, carboxyl, hydroxyl, mercapto, alkylmercapto, alkoxy, alkylamino, -C(O)-alkyl, -C(O)-O-Alkyl, -CONH 2 , -CONH-Alkyl, -OC(O)-Alkyl, -NH-C(O)-Alkyl, -S(O)O-Alkyl , -S(O)ONH 2 , -S(O)ONH-alkyl, said alkyl, alkenyl, alkynyl, alkylmercapto, alkyloxy, -C(O)-alkyl, - C(O)-O-Alkyl, -CONH-Alkyl, -OC(O)-Alkyl, -NH-C(
  • the k is selected from 0, 1, 2, 3, 4;
  • the j is selected from integers from 1 to 20 (eg 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20).
  • L 1 is N
  • R 11 is selected from deuterium, halogen, alkyl, amino, cyano, nitro, alkenyl, alkynyl, carboxyl, hydroxyl, mercapto, alkylmercapto, alkoxy, alkylamino, -C(O)-alkyl , -C(O)-O-Alkyl, -CONH 2 , -CONH-Alkyl, -OC(O)-Alkyl, -NH-C(O)-Alkyl, -S(O)O-Alkyl base, -S(O)ONH 2 , -S(O)ONH-alkyl, said alkyl, alkenyl, alkynyl, carboxyl, alkylmercapto, alkyloxy, -C(O)-alkane Alkyl, -C(O)-O-Alkyl, -CONH-Alkyl, -OC(O)-Al,
  • the k is selected from 0, 1, 2, 3, 4;
  • L 1 is N
  • L 1 is N
  • L 1 is N
  • E in the targeting ligand is
  • the R 12 , R 13 , R 14 and R 15 are each independently selected from -C(O)NH-, -C(O)-, the carbonyl group is optionally further substituted by an alkyl group, and the alkyl group optionally further substituted by the group consisting of alkyl, hydroxy, -C(O)O-, -C(O)O-alkyl-, -C(O)NH-;
  • the X 2 , X 3 , X 4 and X 5 are each independently an integer selected from 0 to 10 (eg, 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10).
  • E in the targeting ligand is
  • R 12 , R 13 , R 14 and R 15 are each independently selected from -C(O)NH-, -C(O)-, said -C(O)NH-, -C(O)- Optionally further substituted with an alkyl group optionally further selected from the group consisting of alkyl, hydroxyl, -C(O)O-, -C(O)O-alkyl-, -C(O)NH- replace;
  • the X 2 , X 3 , X 4 and X 5 are each independently an integer selected from 0 to 10 (eg, 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10).
  • E in the targeting ligand is
  • the R 12 , R 13 , R 14 and R 15 are each independently selected from -C(O)NH-, -C(O)-, Substituents substituted by , the X 2 , X 3 , X 4 and X 5 are each independently selected from an integer from 0 to 10 (for example, 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10).
  • E in the targeting ligand is
  • E in the targeting ligand is selected from
  • E in the targeting ligand is selected from
  • E in the targeting ligand is selected from
  • E in the targeting ligand is
  • L 1 is selected from the following structures:
  • R 9 and R 10 are each independently selected from -S-, -NH-, -O-, -S-, -C(O)-, -OC(O)-, -C(O)O-, -NHC (O)-, -C(O)NH-, -CH2- , -CH2NH-, -CH2O- , -NH-C(O) -CH2- , -C (O) -CH2 -NH-, -NH(CO)NH-, 3-12-membered heterocyclic group, the -CH 2 - is optionally substituted with a substituent selected from halogen, alkyl, alkoxy, alkylamino, the alkane The group is optionally further substituted with a substituent selected from hydroxy, amino, halogen;
  • R 11 is selected from deuterium, halogen, alkyl, amino, cyano, nitro, alkenyl, alkynyl, carboxyl, hydroxyl, mercapto, alkylmercapto, alkoxy, alkylamino, -C(O)-alkyl, -C(O)-O-Alkyl, -CONH 2 , -CONH-Alkyl, -OC(O)-Alkyl, -NH-C(O)-Alkyl, -S(O)O-Alkyl , -S(O)ONH 2 , -S(O)ONH-alkyl, said alkyl, alkenyl, alkynyl, alkylmercapto, alkyloxy, -C(O)-alkyl, - C(O)-O-Alkyl, -CONH-Alkyl, -OC(O)-Alkyl, -NH-C(
  • the k is selected from 0, 1, 2, 3, 4;
  • Said j is selected from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20.
  • E in the targeting ligand is selected from:
  • L1 is selected from :
  • E in the targeting ligand is selected from:
  • L1 is selected from : That is, EL 1 is
  • E in the targeting ligand is selected from:
  • L1 is selected from : That is, EL 1 is
  • E in the targeting ligand is selected from L1 is selected from That is, EL 1 is
  • E in the targeting ligand is selected from L1 is selected from That is, EL 1 is
  • L 2 is the tethering moiety between the targeting moiety and the branching group, and L 2 plays the role of linking and spacing between the branching group and each targeting moiety.
  • one end of L2 is directly attached to the targeting ligand and the other end is directly attached to the branching group E.
  • one end of L 2 is directly attached to the targeting ligand and the other end is indirectly attached to the branching group E.
  • one end of L2 is indirectly linked to the targeting ligand and the other end is indirectly linked to the branching group E.
  • the targeting ligands disclosed herein include 2 L2 and 2 targeting moieties.
  • the targeting ligands disclosed herein include 3 L2 and 3 targeting moieties.
  • the targeting ligands disclosed herein include 4 L2 and 4 targeting moieties.
  • the targeting ligands disclosed herein include multiple L 2s and multiple targeting moieties.
  • L2 in the present disclosure is selected from 1 or a combination of 2-20 covalently linked groups (eg, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20):
  • Substituted or unsubstituted cycloalkyl eg cyclohexyl, cyclopropyl, cyclobutyl, cyclopentyl, cycloheptyl, cyclooctyl, etc.
  • substituted or unsubstituted cycloalkenyl eg cyclohexene base, cyclobutenyl, cyclopentenyl, cycloheptenyl, cyclooctenyl, cyclohexadienyl, cyclopentadienyl, cycloheptadienyl, cyclooctadienyl, etc.
  • substituted or unsubstituted aryl eg phenyl, naphthyl, binaphthyl, anthracenyl, etc.
  • substituted or unsubstituted heteroaryl eg pyridyl, pyrimidinyl,
  • there are from 1 to 20 L2 in the present disclosure eg, 1, 2 , 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16 , 17, 18, 19 or 20
  • 1 to 20 L2 in the present disclosure (eg, 1, 2 , 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16 , 17, 18, 19 or 20) optional from
  • the targeting ligand comprises L 2 having the structure shown below,
  • x6 is an integer from 1 to 20 (eg, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20).
  • the targeting ligand comprises L2 having the structure shown below,
  • the targeting ligand comprises L2 having the structure shown below,
  • the targeting ligand comprises L2 having the structure shown below,
  • the targeting ligand comprises L2 having the structure shown below,
  • the targeting ligand comprises L2 having the structure shown below,
  • the targeting ligand comprises L2 having the structure shown below,
  • x 7 is an integer from 1 to 20 (eg, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20), and Z is
  • the targeting ligand has L2 of the structure shown below,
  • the targeting ligand has L2 of the structure shown below,
  • x8 is an integer from 1 to 20 (eg, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16 , 17, 18, 19 or 20), and Z is
  • the targeting ligand has L2 of the structure shown below,
  • x 9 and X 10 are each independently selected from integers from 1 to 20 (eg, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20), and Z is
  • the targeting ligand has L2 of the structure shown below,
  • the targeting ligand has L2 of the structure shown below, wherein x7 and X8 are each independently selected from integers from 1 to 20 (eg, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16 , 17, 18, 19, or 20), and Z is
  • the targeting ligand has the following structure:
  • the targeting ligand has the following structure:
  • the targeting ligand has the following structure:
  • the targeting ligand has the following structure:
  • the targeting moiety of the targeting ligand is comprised of one or more targeting groups or targeting moieties that assist in directing delivery of the therapeutic agent attached thereto to the desired target site.
  • the targeting moiety can bind to a cell or cellular receptor and initiate endocytosis to facilitate entry of the therapeutic agent into the cell.
  • Targeting moieties can include compounds that have affinity for cell receptors or cell surface molecules or antibodies.
  • Various targeting ligands containing targeting moieties can be linked to therapeutic agents and other compounds to target the agents to cells and specific cellular receptors.
  • types of targeting moieties include carbohydrates, cholesterol, and cholesteryl groups or steroids.
  • Targeting moieties that can bind to cellular receptors include carbohydrates such as galactose, galactose derivatives (eg N-acetyl-galactosamine, N-trifluoroacetylgalactosamine, N-propionylgalactosamine, N-n-butyrylgalactosamine, N-isobutyrylgalactosamine), mannose and mannose derivatives).
  • Targeting moieties known to bind the asialoglycoprotein receptor are particularly useful for directing delivery of oligomeric compounds to the liver.
  • the asialoglycoprotein receptor is abundantly expressed on liver cells (hepatocytes).
  • Cell receptor targeting moieties targeting ASCPR include galactose and galactose derivatives. Specifically, clusters of galactose derivatives, including clusters consisting of 2, 3, 4, or more than 4 N-acetyl-galactosamines (GalNAc or NAG), can promote the uptake of certain compounds in hepatocytes.
  • GalNAc clusters coupled to oligomeric compounds were used to direct the composition to the liver, where N-acetyl-galactosamine sugars were able to bind to asialoglycoprotein receptors on the surface of liver cells. Binding of the asialoglycoprotein receptor is thought to initiate receptor-mediated endocytosis, thereby facilitating the entry of compounds into the cell interior.
  • a targeting ligand may include 2, 3, 4, or more than 4 targeting moieties.
  • the targeting ligands disclosed herein can include 1, 2 , 3, 4, or more than 4 targeting moieties linked to a branching group through L2.
  • the targeting ligand is in the form of a galactose cluster.
  • each targeting moiety includes a galactosamine derivative, which is N-acetyl-galactosamine.
  • Other sugars that can be used as targeting moieties and have affinity for the asialoglycoprotein receptor can be selected from galactose, galactosamine, N-formyl-galactosamine, N-acetyl-galactosamine, N-propionyl-galactosamine, N-n-butyryl-galactosamine and N-isobutyryl-galactosamine, etc.
  • the targeting ligands of the present disclosure include N-acetylgalactosamine as a targeting moiety
  • the targeting ligand includes three terminal galactosamine or galactosamine derivatives (such as N-acetyl-galactosamine), each of which has an affinity for the sialoglycoprotein receptor.
  • the targeting ligand includes a three-terminal N-acetyl-galactosamine (GalNAc or NAG) as a targeting moiety.
  • the targeting ligand includes four terminal galactosamine or galactosamine derivatives (such as N-acetyl-galactosamine), each of which has an affinity for the asialoglycoprotein receptor .
  • the targeting ligand includes four terminal N-acetyl-galactosamine (GalNAc or NAG) as targeting moieties.
  • the targeting ligands provided by the present disclosure have the following structures,
  • the targeting ligands provided by the present disclosure have the following structures,
  • the targeting ligands provided by the present disclosure have the following structures,
  • the targeting ligands provided by the present disclosure have the following structures,
  • the siRNA of the present disclosure is linked to a targeting ligand of the present disclosure to form a siRNA conjugate having the following,
  • T is the targeting moiety
  • E is the branching group
  • L1 is the linker moiety
  • L2 is the tethering moiety between the targeting moiety and the branching group
  • x is an integer from 1 to 10
  • D is the targeting coagulation factor XI siRNA.
  • D is an siRNA targeting factor XI.
  • D is any siRNA of the present disclosure.
  • the L1 is linked to the 3' end of the sense strand of the siRNA.
  • the targeting ligand is attached to the end of the siRNA through a phosphate group, phosphorothioate group, or phosphonate group.
  • the targeting ligand is indirectly attached to the end of the siRNA through a phosphate group, phosphorothioate group, or phosphonate group.
  • the targeting ligand is directly attached to the end of the siRNA through a phosphate, phosphorothioate, or phosphonic acid group.
  • the targeting ligand is directly attached to the end of the siRNA through a phosphate group, a phosphorothioate group.
  • the targeting ligand is directly attached to the 3' end of the siRNA sense strand through a phosphate group, a phosphorothioate group.
  • the present disclosure provides an siRNA conjugate
  • D is siRNA targeting coagulation factor XI.
  • D is an siRNA targeting factor XI; in some embodiments, D is any siRNA of the disclosure.
  • the targeting ligand is directly attached to the 3' end of the siRNA sense strand through a phosphate group or a phosphorothioate group.
  • the present disclosure provides an siRNA conjugate
  • D is an siRNA targeting factor XI; in some embodiments, in some embodiments, D is any siRNA of the disclosure.
  • the targeting ligand is directly attached to the 3' end of the siRNA sense strand through a phosphate group or a phosphorothioate group.
  • the present disclosure provides an siRNA conjugate
  • D is an siRNA targeting coagulation factor XI; in some embodiments, D is any siRNA of the present disclosure.
  • the targeting ligand is directly attached to the 3' end of the siRNA sense strand through a phosphate group or a phosphorothioate group.
  • the present disclosure provides an siRNA conjugate
  • D is an siRNA targeting coagulation factor XI; in some embodiments, D is any siRNA of the present disclosure.
  • the targeting ligand is directly attached to the 3' end of the siRNA sense strand through a phosphate group or a phosphorothioate group.
  • the L1 and D are linked through a phosphate group, a phosphorothioate group, or a phosphonic acid group.
  • the L 1 and the 3' end of the D sense strand are linked by a phosphate group, a phosphorothioate group, or a phosphonic acid group.
  • the L 1 and the 3' end of the D sense strand are directly linked through a phosphate group or a phosphorothioate group.
  • the L1 is indirectly linked to the 3' end of the D sense strand through a phosphate group or a phosphorothioate group.
  • the siRNA conjugate sense strand is selected from the group consisting of: any one of SEQ ID NO:262 to SEQ ID NO:281.
  • the siRNA conjugate antisense strand is selected from the group consisting of: any one of SEQ ID NO:202 to SEQ ID NO:261.
  • the siRNA conjugate is selected from:
  • the sense strand is selected from SEQ ID NO:262, and the antisense strand is selected from SEQ ID NO:202, SEQ ID NO:222, or SEQ ID NO:242;
  • the sense strand is selected from SEQ ID NO:263, and the antisense strand is selected from SEQ ID NO:203, SEQ ID NO:223, or SEQ ID NO:243;
  • the sense strand is selected from SEQ ID NO:264, and the antisense strand is selected from SEQ ID NO:204, SEQ ID NO:224, or SEQ ID NO:244;
  • the sense strand is selected from SEQ ID NO:265, and the antisense strand is selected from SEQ ID NO:205, SEQ ID NO:225, or SEQ ID NO:245;
  • the sense strand is selected from SEQ ID NO:266, and the antisense strand is selected from SEQ ID NO:206, SEQ ID NO:226, or SEQ ID NO:246;
  • the sense strand is selected from SEQ ID NO:267
  • the antisense strand is selected from SEQ ID NO:207, SEQ ID NO:227, or SEQ ID NO:247;
  • the sense strand is selected from SEQ ID NO:268, and the antisense strand is selected from SEQ ID NO:208, SEQ ID NO:228, or SEQ ID NO:248;
  • the sense strand is selected from SEQ ID NO:269
  • the antisense strand is selected from SEQ ID NO:209, SEQ ID NO:229, or SEQ ID NO:249;
  • the sense strand is selected from SEQ ID NO:270, and the antisense strand is selected from SEQ ID NO:210, SEQ ID NO:230, or SEQ ID NO:250;
  • the sense strand is selected from SEQ ID NO:271, and the antisense strand is selected from SEQ ID NO:211, SEQ ID NO:231, or SEQ ID NO:251;
  • the sense strand is selected from SEQ ID NO:272, and the antisense strand is selected from SEQ ID NO:212, SEQ ID NO:232, or SEQ ID NO:252;
  • the sense strand is selected from SEQ ID NO:273, and the antisense strand is selected from SEQ ID NO:213, SEQ ID NO:233, or SEQ ID NO:253;
  • the sense strand is selected from SEQ ID NO:274, and the antisense strand is selected from SEQ ID NO:214, SEQ ID NO:234, or SEQ ID NO:254;
  • the sense strand is selected from SEQ ID NO:275, and the antisense strand is selected from SEQ ID NO:215, SEQ ID NO:235, or SEQ ID NO:255;
  • the sense strand is selected from SEQ ID NO:276, and the antisense strand is selected from SEQ ID NO:216, SEQ ID NO:236, or SEQ ID NO:256;
  • the sense strand is selected from SEQ ID NO:277
  • the antisense strand is selected from SEQ ID NO:217, SEQ ID NO:237, or SEQ ID NO:257;
  • the sense strand is selected from SEQ ID NO:278, and the antisense strand is selected from SEQ ID NO:218, SEQ ID NO:238, or SEQ ID NO:258;
  • the sense strand is selected from SEQ ID NO:279, and the antisense strand is selected from SEQ ID NO:219, SEQ ID NO:239, or SEQ ID NO:259;
  • the sense strand is selected from SEQ ID NO:280, and the antisense strand is selected from SEQ ID NO:220, SEQ ID NO:240, or SEQ ID NO:260;
  • the sense strand is selected from SEQ ID NO:281, and the antisense strand is selected from SEQ ID NO:221, SEQ ID NO:241, or SEQ ID NO:261.
  • compositions comprising a conjugate as claimed above, and one or more pharmaceutically acceptable excipients such as a carrier, vehicle, diluent, and/or delivery polymer thing.
  • siRNA or siRNA conjugates of the present disclosure such as encapsulation in liposomes, microparticles, microcapsules, recombinant cells capable of expressing the compound, receptor-mediated intracellular Endocytosis, construction of nucleic acid as part of a retrovirus or other vector.
  • Another aspect of the present disclosure provides use of the above conjugate or a composition containing the conjugate in the manufacture of a medicament for treating a disease in a subject, in some embodiments selected from liver-derived diseases.
  • Another aspect of the present disclosure provides a method of treating a disease in a subject, comprising administering to the subject the above-described conjugate, or composition.
  • Another aspect of the present disclosure provides a method of inhibiting mRNA expression in a subject, the method comprising administering to the subject the above-described conjugate, or composition.
  • Another aspect of the present disclosure provides a method of delivering an expression-inhibiting oligomeric compound to the liver in vivo, administering the above-described conjugate, or composition, to a subject.
  • the conjugates, compositions and methods disclosed herein can reduce the level of a target mRNA in a cell, cell population, cell population, tissue or subject, comprising: administering to the subject a therapeutically effective amount of an expression-inhibiting oligomer described herein
  • the expression-inhibiting oligomer is linked to a targeting ligand, thereby inhibiting the expression of the target mRNA in the subject.
  • the subject has been previously identified as having pathogenic upregulation of the target gene in the targeted cell or tissue.
  • a subject as described in the present disclosure refers to a subject suffering from a disease or disorder that would benefit from reduction or inhibition of target mRNA expression.
  • Delivery can be by local administration (eg, direct injection, implantation, or topical administration), systemic administration, or subcutaneous, intravenous, intraperitoneal, or parenteral routes, including intracranial (eg, intraventricular, parenchymal) intramuscular, transdermal, airway (aerosol), nasal, oral, rectal, or topical (including buccal and sublingual) administration.
  • local administration eg, direct injection, implantation, or topical administration
  • systemic administration eg., systemic administration, or subcutaneous, intravenous, intraperitoneal, or parenteral routes, including intracranial (eg, intraventricular, parenchymal) intramuscular, transdermal, airway (aerosol), nasal, oral, rectal, or topical (including buccal and sublingual) administration.
  • compositions provided by the present disclosure may be administered by injection, eg, intravenous, intramuscular, intradermal, subcutaneous, intraduodenal, or intraperitoneal injection.
  • the conjugate can be packaged in a kit.
  • the present disclosure also provides a pharmaceutical composition comprising the siRNA or siRNA conjugate of the present disclosure.
  • the pharmaceutical composition may further include pharmaceutically acceptable adjuvants and/or adjuvants, and the adjuvants may be one or more various formulations or compounds conventionally used in the art.
  • the pharmaceutically acceptable adjuvant may include at least one of pH buffering agents, protecting agents and osmotic pressure adjusting agents.
  • the siRNA conjugates or pharmaceutical compositions described above when contacted with cells expressing the target gene, are screened for, for example, psiCHECK activity and luciferase reporter gene assays, others such as PCR or based on branched DNA (bDNA)
  • the above-described siRNA conjugate or pharmaceutical composition inhibits the expression of the target gene by at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75% , at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%.
  • the siRNA conjugates or pharmaceutical compositions described above when contacted with cells expressing the target gene, are screened for, for example, psiCHECK activity and luciferase reporter gene assays, others such as PCR or based on branched DNA (bDNA) method, or a protein-based method, such as immunofluorescence assay, such as Western Blot or flow cytometry, the residual expression percentage of target gene mRNA caused by the above-mentioned siRNA conjugate or pharmaceutical composition is not higher than 99%, not higher than 95%, not higher than 90%, not higher than 85%, not higher than 80%, not higher than 75%, not higher than 70%, not higher than 65%, not higher than 60%, not higher over 55%, up to 50%, up to 45%, up to 40%, up to 35%, up to 30%, up to 25%, up to 20%, up to 15 %, or not higher than 10%.
  • psiCHECK activity and luciferase reporter gene assays others such as PCR or based on
  • the siRNA conjugate or pharmaceutical composition when contacted with cells expressing the target gene, is screened for, for example, psiCHECK activity and luciferase reporter gene assays, others such as PCR or branched DNA (bDNA) based assays.
  • Methods, or protein-based methods, such as immunofluorescence assays, such as Western Blot, or flow cytometry the siRNA conjugate reduces off-target activity by at least 20%, at least 25% while maintaining on-target activity , at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, or at least 75%.
  • the siRNA conjugate or pharmaceutical composition when contacted with cells expressing the target gene, is screened for, for example, psiCHECK activity and luciferase reporter gene assays, others such as PCR or branched DNA (bDNA) based assays.
  • the siRNA conjugate reduces on-target activity by up to 20%, up to 19%, up to 15%, up to 10% , at least 5% or more than 1% while reducing off-target activity by at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60% %, at least 65%, at least 70%, or at least 75%.
  • the siRNA conjugate or pharmaceutical composition when contacted with cells expressing the target gene, is screened for, for example, psiCHECK activity and luciferase reporter gene assays, others such as PCR or branched DNA (bDNA) based assays.
  • the siRNA conjugate increases on-target activity by at least 1%, at least 5%, at least 10%, at least 15% , at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75% or at least 80% while reducing off-target activity by at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, At least 70% or at least 75%.
  • the present disclosure also provides a cell comprising the siRNA or siRNA conjugate of the present disclosure.
  • the present disclosure also provides a kit comprising the siRNA or siRNA conjugate of the present disclosure.
  • the present disclosure also provides a method for silencing a target gene or mRNA of a target gene in a cell, the method comprising the step of introducing into the cell an siRNA, siRNA conjugate and/or pharmaceutical composition according to the present disclosure.
  • the present disclosure also provides a method for silencing a target gene or mRNA of a target gene in a cell in vivo or in vitro, the method comprising introducing into the cell an siRNA, siRNA conjugate and/or pharmaceutical composition according to the present disclosure steps in .
  • the present disclosure also provides a method for inhibiting a target gene or mRNA expression of a target gene, the method comprising administering to a subject in need thereof an effective amount or an effective dose of the siRNA, siRNA conjugate and /or pharmaceutical composition.
  • administration is by means of administration including intramuscular, intrabronchial, intrapleural, intraperitoneal, intraarterial, lymphatic, intravenous, subcutaneous, cerebrospinal, or combinations thereof.
  • the effective amount or effective dose of the siRNA, siRNA conjugate and/or pharmaceutical composition is about 0.001 mg/kg body weight to about 200 mg/kg body weight, about 0.01 mg/kg body weight to about 100 mg/kg body weight Or about 0.5 mg/kg body weight to about 50 mg/kg body weight.
  • the target gene is the factor XI gene.
  • the present disclosure also provides a pharmaceutical composition comprising the siRNA or siRNA conjugate of the present disclosure.
  • the pharmaceutical composition may further include pharmaceutically acceptable adjuvants and/or adjuvants, and the adjuvants may be one or more various formulations or compounds conventionally used in the art.
  • the pharmaceutically acceptable adjuvant may include at least one of pH buffering agents, protecting agents and osmotic pressure adjusting agents.
  • the siRNA conjugates or pharmaceutical compositions described above when contacted with cells expressing the target gene, are detected by, for example: PCR or branched DNA (bDNA) based methods, or protein based methods such as immunofluorescence assays
  • the above-mentioned siRNA conjugate or pharmaceutical composition inhibits the expression of the target gene by at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, as determined by Western Blot or flow cytometry, for example , at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%.
  • the siRNA conjugates or pharmaceutical compositions described above when contacted with cells expressing the target gene, are detected by, for example: PCR or branched DNA (bDNA) based methods, or protein based methods such as immunofluorescence assays
  • the remaining expression percentage of target gene mRNA caused by the above-mentioned siRNA conjugate or pharmaceutical composition is not higher than 99%, not higher than 95%, not higher than 90%, not high at 85%, no more than 80%, no more than 75%, no more than 70%, no more than 65%, no more than 60%, no more than 55%, no more than 50%, no more than 45% %, no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, or no more than 10%.
  • the present disclosure also provides a cell comprising the siRNA or siRNA conjugate of the present disclosure.
  • the present disclosure also provides a kit comprising the siRNA or siRNA conjugate of the present disclosure.
  • the present disclosure also provides an siRNA, a siRNA conjugate, or a pharmaceutical composition comprising the same, for use in the treatment and/or prevention of a subject suffering from a coagulation factor XI-related disease.
  • the present disclosure also provides use of an siRNA, siRNA conjugate or pharmaceutical composition according to the present disclosure in the manufacture of a medicament for the treatment and/or prevention of a subject suffering from a factor XI-related disease.
  • the present disclosure also provides a method of treating/preventing a coagulation factor XI-related disease, comprising administering to a subject in need thereof a therapeutically and/or prophylactically effective amount of the siRNA, siRNA conjugate or siRNA according to the present disclosure pharmaceutical composition.
  • administration is by means of administration including intramuscular, intrabronchial, intrapleural, intraperitoneal, intraarterial, lymphatic, intravenous, subcutaneous, cerebrospinal, or combinations thereof.
  • the factor XI-related disorder is a thromboembolic complication, particularly deep vein thrombosis, pulmonary embolism, myocardial infarction, or stroke.
  • the individual is at risk for blood clotting disorders, including, but not limited to, infarction, thrombosis, embolism, and thromboembolism, such as, for example, deep vein thrombosis, pulmonary embolism, myocardial infarction, and stroke.
  • the individual is identified as in need of anticoagulation therapy.
  • disorders include, but are not limited to, those undergoing major orthopaedic surgery (eg, hip/knee replacement or hip fracture surgery) and patients requiring long-term treatment such as those experiencing atrial fibrillation to prevent stroke.
  • the present disclosure also provides a method of inhibiting FXI gene expression in a cell.
  • the method includes contacting a cell with an siRNA, siRNA conjugate, or a pharmaceutical composition comprising the same of the present disclosure, and maintaining for a time sufficient to achieve degradation of the mRNA transcript of the FXI gene, thereby inhibiting the expression of the FXI gene in the cell.
  • the present disclosure also provides a method for silencing a target gene or mRNA of a target gene in a cell, the method comprising the step of introducing into the cell an siRNA, siRNA conjugate according to the present disclosure, or a pharmaceutical composition according to the present disclosure .
  • the present disclosure also provides a method for silencing a target gene or mRNA of a target gene in a cell in vivo or in vitro, the method comprising introducing an siRNA, a siRNA conjugate according to the present disclosure, or a pharmaceutical composition according to the present disclosure steps in the cell.
  • the present disclosure also provides a method for inhibiting a target gene or mRNA expression of a target gene, the method comprising administering to a subject in need thereof an siRNA, an siRNA conjugate, or a pharmaceutical composition comprising the same according to the present disclosure .
  • the target gene is the human factor XI gene, TTR gene.
  • the present disclosure also provides an siRNA or siRNA conjugate, characterized in that a base T is used to replace one or more bases U of any siRNA or siRNA conjugate of the present disclosure, such as 1, 2, 3 1, 3, 5, 6, 7, 8, 9, 10.
  • the pharmaceutically acceptable salts of the compounds described in the present disclosure are selected from inorganic salts or organic salts, and the compounds described in the present disclosure can react with acidic or basic substances to form corresponding salts.
  • the compounds of the present disclosure may exist in particular geometric or stereoisomeric forms.
  • This disclosure contemplates all such compounds, including cis and trans isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers isomers, (D)-isomers, (L)-isomers, and racemic and other mixtures thereof, such as enantiomerically or diastereomerically enriched mixtures, all of which belong to within the scope of this disclosure.
  • Additional asymmetric carbon atoms may be present in substituents such as alkyl. All such isomers, as well as mixtures thereof, are included within the scope of this disclosure.
  • tautomer or "tautomeric form” refers to structural isomers of different energies that are interconvertible via a low energy barrier.
  • proton tautomers also known as proton tautomers
  • proton transfer such as keto-enol and imine-enamine, lactam-lactam isomerizations .
  • An example of a lactam-lactam equilibrium is between A and B as shown below.
  • the compounds of the present disclosure may be asymmetric, eg, have one or more stereoisomers. Unless otherwise specified, all stereoisomers include, such as enantiomers and diastereomers.
  • Compounds of the present disclosure containing asymmetric carbon atoms can be isolated in optically pure or racemic forms. Optically pure forms can be resolved from racemic mixtures or synthesized by using chiral starting materials or chiral reagents.
  • Optically active (R)- and (S)-isomers can be prepared by chiral synthesis or chiral reagents or other conventional techniques. If one enantiomer of a compound of the present disclosure is desired, it can be prepared by asymmetric synthesis or derivatization with a chiral auxiliary, wherein the resulting mixture of diastereomers is separated and the auxiliary group is cleaved to provide pure desired enantiomer.
  • a diastereomeric salt is formed with an appropriate optically active acid or base, followed by conventional methods known in the art
  • the diastereoisomers were resolved and the pure enantiomers recovered.
  • separation of enantiomers and diastereomers is usually accomplished by the use of chromatography employing a chiral stationary phase, optionally in combination with chemical derivatization (eg, from amines to amino groups) formate).
  • the present disclosure also includes certain isotopically-labeled compounds of the present disclosure which are identical to those described herein, except that one or more atoms have been replaced by an atom having an atomic weight or mass number different from that normally found in nature.
  • isotopes that can be incorporated into the compounds of the present disclosure include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, iodine, and chlorine, such as 2H, 3H , 11C , 13C , 14C , 13 , respectively N, 15 N, 15 O, 17 O, 18 O, 31 P, 32 P, 35 S, 18 F, 123 I, 125 I and 36 Cl and the like.
  • deuterium when a position is specifically designated as deuterium (D), the position is understood to have an abundance of deuterium (ie, at least 1000 times greater than the natural abundance of deuterium (which is 0.015%)) % of deuterium incorporated).
  • Exemplary compounds having natural abundance greater than deuterium may be at least 1000 times more abundant deuterium, at least 2000 times more abundant deuterium, at least 3000 times more abundant deuterium, at least 4000 times more abundant deuterium, at least 4000 times more abundant 5000 times more abundant deuterium, at least 6000 times more abundant deuterium or more abundant deuterium.
  • the present disclosure also includes compounds of Formula I in various deuterated forms.
  • Each available hydrogen atom attached to a carbon atom can be independently replaced by a deuterium atom.
  • Those skilled in the art can synthesize compounds of formula I in deuterated form with reference to the relevant literature.
  • Commercially available deuterated starting materials can be used in preparing deuterated forms of the compounds of formula I, or they can be synthesized using conventional techniques using deuterated reagents including, but not limited to, deuterated borane, trideuterated borane Tetrahydrofuran solution, deuterated lithium aluminum hydride, deuterated iodoethane and deuterated iodomethane, etc.
  • the bond Indicates an unspecified configuration, i.e. if a chiral isomer exists in the chemical structure, the bond can be or both Two configurations.
  • the bond no configuration is specified, i.e. the bond The configuration can be E or Z, or both E and Z configurations.
  • Factor XI nucleic acid refers to any nucleic acid encoding Factor XI.
  • a factor XI nucleic acid includes a DNA sequence encoding factor XI such as a "factor XI gene," an RNA sequence transcribed from DNA encoding factor XI (including intron- and exon-containing genomic DNA), and mRNA sequence encoding factor XI.
  • Factor XI gene is any of the following sequences: GENBANK Accession No.
  • Vector XI mRNA refers to mRNA encoding Factor XI protein.
  • the sense strand (also known as SS, SS or sense strand) of an siRNA refers to the strand comprising the same or substantially the same sequence as the target mRNA sequence;
  • the antisense strand (also called AS or AS strand) of an siRNA refers to the strand having a sequence complementary to the target mRNA sequence.
  • capital letters C, G, U, A, T represent the base composition of nucleotides; lower case letter d represents that the adjacent nucleotide to the right of the letter d is deoxyribose Nucleotides; lowercase letter m indicates that a nucleotide adjacent to the left of the letter m is a methoxy-modified nucleotide; lowercase letter f indicates that a nucleotide adjacent to the left of the letter f is a fluorine modified The lowercase letter s indicates that the two nucleotides adjacent to the letter s are connected by phosphorothioate groups.
  • fluoro-modified nucleotide refers to a nucleotide formed by substituting the hydroxyl group at the 2' position of the ribosyl of a nucleotide with fluorine
  • non-fluoro-modified nucleotide refers to a nucleoside Nucleotides or nucleotide analogs formed by the substitution of the hydroxyl group at the 2' position of the ribosyl group of an acid by a non-fluorine group.
  • Nucleotide analog refers to a nucleic acid that can replace a nucleotide, but is structurally different from adenine A group of ribonucleotides, guanine ribonucleotides, cytosine ribonucleotides, uracil ribonucleotides, or thymidine ribonucleotides. Such as isonucleotides, bridged nucleotides (bridged nucleic acid, BNA for short) or acyclic nucleotides.
  • the methoxy-modified nucleotide refers to a nucleotide formed by replacing the 2'-hydroxyl group of the ribosyl group with a methoxy group.
  • Isonucleotides refer to compounds formed by changing the position of the base in the nucleotide on the ribose ring.
  • the isonucleotide may be a compound formed by moving the base from the 1'-position to the 2'-position or the 3'-position of the ribose ring.
  • BNA refers to constrained or inaccessible nucleotides.
  • the BNA may contain a five-membered, six-membered, or seven-membered ring bridged structure with "fixed" C3'-endoglycan constriction.
  • the bridge is typically incorporated at the 2'-,4'-position of the ribose sugar to provide a 2',4'-BNA nucleotide.
  • the BNA can be an LNA, ENA, cET BNA, or the like.
  • Acyclic nucleotides are a class of nucleotides formed by opening the sugar ring of a nucleotide.
  • an acyclic nucleotide can be an unlocked nucleic acid (UNA) or a glycerol nucleic acid (GNA).
  • nucleotide difference between a nucleotide sequence and another nucleotide sequence, which means that the base type of the nucleotide at the same position has changed in the former compared with the latter, For example, when one nucleotide base is A in the latter, and the corresponding nucleotide base at the same position in the former is U, C, G or T, it is regarded as the one of the two nucleotide sequences. There is a nucleotide difference at this position. In some embodiments, when an abasic nucleotide or its equivalent is substituted for a nucleotide at the original position, a nucleotide difference may also be considered to have occurred at that position.
  • the term "at least 15 contiguous nucleotides that differ by no more than 3 nucleotide sequences from any of the sense strands in Table 1" is intended to mean the sense strands of the siRNAs described herein
  • the strand comprises at least 15 contiguous nucleotides of any of the sense strands in Table 1, or differs by no more than 3 nucleotide sequences from at least 15 contiguous nucleotides of any of the sense strands in Table 1, optionally, They differ by no more than 2 nucleotide sequences, optionally, by 1 nucleotide sequence.
  • the siRNA sense strands described herein comprise at least 16 contiguous nucleotides of any of the sense strands in Table 1, or differ by no more than 3 from at least 16 contiguous nucleotides of any of the sense strands in Table 1 nucleotide sequences, optionally, differ by no more than 2 nucleotide sequences, optionally, differ by 1 nucleotide sequence;
  • the siRNA sense strands described herein comprise at least 17 contiguous nucleotides of any one of the sense strands in Table 1, or differ by no more than 3 from at least 17 contiguous nucleotides of any one of the sense strands in Table 1 nucleotide sequences, optionally, differ by no more than 2 nucleotide sequences, optionally, differ by 1 nucleotide sequence;
  • the siRNA sense strands described herein comprise at least 18 contiguous nucleotides of any one of the sense strands in Table 1, or differ by no more than 3 from at least 18 contiguous nucleotides of any one of the sense strands in Table 1 nucleotide sequences, optionally, differ by no more than 2 nucleotide sequences, optionally, differ by 1 nucleotide sequence;
  • the siRNA sense strands described herein comprise all 19 contiguous nucleotides of any one of the sense strands in Table 1, or differ by no more than 3 from all 19 contiguous nucleotides of any one of the sense strands in Table 1
  • the nucleotide sequences optionally, differ by no more than 2 nucleotide sequences, optionally, by 1 nucleotide sequence.
  • the term "at least 15 contiguous nucleotides that differ by no more than 3 nucleotide sequences from any of the antisense strands in Table 1" is intended to mean the The siRNA antisense strand comprises at least 15 contiguous nucleotides of any antisense strand in Table 1, or differs by no more than 3 nucleotide sequences from at least 15 contiguous nucleotides of any antisense strand in Table 1 , optionally, differ by no more than 2 nucleotide sequences, optionally, differ by 1 nucleotide sequence.
  • the siRNA antisense strands described herein comprise or differ from at least 16 contiguous nucleotides of any antisense strand in Table 1, or differ from at least 16 contiguous nucleotides of any antisense strand in Table 1 no more than 3 nucleotide sequences, optionally, differing by no more than 2 nucleotide sequences, optionally, differing by 1 nucleotide sequence;
  • the siRNA antisense strands described herein comprise or differ from at least 17 contiguous nucleotides of any antisense strand in Table 1, or differ from at least 17 contiguous nucleotides of any antisense strand in Table 1 no more than 3 nucleotide sequences, optionally, differing by no more than 2 nucleotide sequences, optionally, differing by 1 nucleotide sequence;
  • the siRNA antisense strands described herein comprise or differ from at least 18 contiguous nucleotides of any antisense strand in Table 1, or differ from at least 18 contiguous nucleotides of any antisense strand in Table 1 no more than 3 nucleotide sequences, optionally, differing by no more than 2 nucleotide sequences, optionally, differing by 1 nucleotide sequence;
  • the siRNA antisense strands described herein comprise or differ from at least 19 contiguous nucleotides of any antisense strand in Table 1, or differ from at least 19 contiguous nucleotides of any antisense strand in Table 1 no more than 3 nucleotide sequences, optionally, differing by no more than 2 nucleotide sequences, optionally, differing by 1 nucleotide sequence;
  • the siRNA antisense strands described herein comprise at least 20 contiguous nucleotides of any antisense strand as in Table 1, or differ from at least 20 contiguous nucleotides of any antisense strand in Table 1 no more than 3 nucleotide sequences, optionally, differing by no more than 2 nucleotide sequences, optionally, differing by 1 nucleotide sequence;
  • siRNA antisense strands described herein comprise, or differ from, all 21 contiguous nucleotides of any antisense strand in Table 1 No more than 3 nucleotide sequences, optionally, no more than 2 nucleotide sequences, optionally 1 nucleotide sequence.
  • the terms "complementary” or “reverse complementary” are used interchangeably and have the meaning well known to those skilled in the art, that is, in a double-stranded nucleic acid molecule, the bases of one strand are The bases on the strand pair up in a complementary fashion.
  • the purine base adenine (A) is always paired with the pyrimidine base thymine (T) (or uracil (U) in RNA);
  • the purine base guanine (C) is always paired with the pyrimidine base Cytosine (G) pairs.
  • Each base pair consists of a purine and a pyrimidine.
  • mismatch in the art means that in a double-stranded nucleic acid, the bases at corresponding positions are not paired in complementary form.
  • inhibitor As used herein, the term “inhibit”, is used interchangeably with “reduce,” “silence,” “down-regulate,” “repression,” and other similar terms, and includes any level of inhibition.
  • the term "inhibiting factor XI expression” includes inhibiting the expression of the factor XI gene and variants (eg, naturally occurring variants) or mutants of the factor XI gene, inhibiting the expression of factor XI mRNA, And/or inhibit the expression of coagulation factor XI protein.
  • the factor XI gene can be a wild-type human factor XI gene, a mutant human factor XI gene, or a transgenic human factor XI gene in the context of genetically manipulated cells, cell populations, or organisms.
  • Inhibition of factor XI gene expression includes inhibition of factor XI gene at any level, such as at least partial inhibition of factor XI gene expression, such as inhibition of at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, At least about 97%, at least about 98%, or at least about 99%.
  • at least partial inhibition of factor XI gene expression such as inhibition of at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least
  • Factor XI gene expression can be assessed based on the level of any variable associated with factor XI gene expression, such as factor XI mRNA levels or factor XI protein levels. Inhibition can be assessed by a reduction in absolute or relative levels of one or more of these variables compared to control levels.
  • the control level can be any type of control level used in the art, such as a pre-dose baseline level or from a similar untreated or controlled (eg buffer only control or inert control) treated subject, cell , or the level determined by the sample.
  • the residual expression of mRNA can be used to characterize the degree of inhibition of target gene expression by siRNA, for example, the residual expression of mRNA is not higher than 99%, not higher than 95%, not higher than 90%, not higher than 85%, not higher than higher than 80%, not higher than 75%, not higher than 70%, not higher than 65%, not higher than 60%, not higher than 55%, not higher than 50%, not higher than 45%, not higher than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, or no more than 10%.
  • the "compounds”, “ligands”, “nucleic acid ligand conjugates”, and “nucleic acids” of the present disclosure can be independently expressed as salts, mixed salts or non-salts (eg, free acids or free bases). form exists. When present in the form of a salt or mixed salts, it can be a pharmaceutically acceptable salt.
  • pharmaceutically acceptable salt includes pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.
  • “Pharmaceutically acceptable acid addition salts” refers to salts with inorganic or organic acids that retain the biological effectiveness of the free base without other side effects.
  • Inorganic acid salts include but are not limited to hydrochloride, hydrobromide, sulfate, nitrate, phosphate, etc.; organic acid salts include but are not limited to formate, acetate, 2,2-dichloroacetate , trifluoroacetate, propionate, caproate, caprylate, caprate, undecylenate, glycolate, gluconate, lactate, sebacate, hexamethylene Acid, glutarate, malonate, oxalate, maleate, succinate, fumarate, tartrate, citrate, palmitate, stearate, oleate , cinnamate, laurate, malate, glutamate, pyroglutamate, aspartate, benzoate, mesylate, benzenesulfonate, p-tol
  • “Pharmaceutically acceptable base addition salts” refers to salts with inorganic or organic bases that retain the biological availability of the free acid without other adverse effects. Salts derived from inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like. Preferred inorganic salts are ammonium, sodium, potassium, calcium and magnesium salts, preferably sodium.
  • Salts derived from organic bases include, but are not limited to, the following: primary, secondary and tertiary amines, substituted amines, including natural substituted amines, cyclic amines, and basic ion exchange resins , such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, triethanolamine, dimethylethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, bicyclic Hexylamine, lysine, arginine, histidine, caffeine, procaine, choline, betaine, ethylenediamine, glucosamine, methylglucamine, theobromine, purine, piperazine, piperazine pyridine, N-ethylpiperidine, polyamine resin, etc.
  • Preferred organic bases include isopropylamine, diethylamine, ethanolamine, trimethylamine, dicyclohe
  • an “effective amount” or “effective dose” refers to the amount of a drug, compound, or pharmaceutical composition necessary to obtain any one or more beneficial or desired therapeutic results.
  • beneficial or desired results include elimination or reduction of risk, reduction in severity, or delay in onset of disorders, including biochemical, tissue academic and/or behavioral symptoms.
  • beneficial or desired outcomes include clinical outcomes, such as reducing the incidence of or ameliorating one or more symptoms of the various target gene, target mRNA or target protein-related disorders of the present disclosure, reducing treatment disorders
  • the dosage of the other agent is required to enhance the therapeutic effect of the other agent, and/or delay the progression of the target gene, target mRNA or target protein-related disorder of the present disclosure in the patient.
  • the effective amount or effective dose of siRNA is about 0.001 mg/kg body weight to about 200 mg/kg body weight, about 0.01 mg/kg body weight to about 100 mg/kg body weight, or about 0.5 mg/kg body weight to about 50 mg/kg body weight. kg body weight.
  • a “pharmaceutical composition” comprises the siRNA or siRNA conjugate of the present disclosure and a pharmaceutically acceptable adjuvant and/or adjuvant, which may be one or more various formulations or compounds conventionally used in the art.
  • the pharmaceutically acceptable adjuvant may include at least one of pH buffering agents, protective agents and osmotic pressure regulators.
  • subject As used herein, “subject”, “patient”, “subject” or “individual” are used interchangeably and include humans or non-human animals, eg, mammals, eg, humans or monkeys.
  • the siRNA provided by the present disclosure can be obtained by conventional preparation methods in the art (eg, solid-phase synthesis and liquid-phase synthesis methods). Among them, solid-phase synthesis has already had commercial customized services.
  • Modified nucleotide groups can be introduced into siRNAs described in the present disclosure by using nucleoside monomers with corresponding modifications, methods of preparing nucleoside monomers with corresponding modifications, and introduction of modified nucleotide groups Methods for siRNA are also well known to those skilled in the art.
  • chemical modification includes all alterations of nucleotides by chemical means, such as the addition or removal of chemical moieties, or the substitution of one chemical moiety for another.
  • base includes any known DNA and RNA bases, base analogs such as purines or pyrimidines, which also includes the natural compounds adenine, thymine, guanine, cytosine, uracil, inosine and natural analogues.
  • base analogs are generally purine or pyrimidine bases, excluding common bases: guanine (G), cytosine (C), adenine (A), thymine (T), and uracil ( U).
  • bases include hypoxanthine (I), xanthine (X), 3 ⁇ -D-ribofuranosyl-(2,6-diaminopyrimidine) (K), 3- ⁇ -D-ribofuranose yl-(1-methyl-pyrazolo[4,3-d]pyrimidine-5,7(4H,6H)-dione) (P), isocytosine (iso-C), isoguanine (iso -G), 1- ⁇ -D-ribofuranosyl-(5-nitroindole), 1- ⁇ -D-ribofuranosyl-(3-nitropyrrole), 5-bromouracil, 2-amino Purine, 4-thio-dT, 7-(2-thienyl)-
  • universal base refers to a heterocyclic moiety located at the 1' position of a nucleotide sugar moiety in a modified nucleotide or an equivalent position in a nucleotide sugar moiety substitution, which heterocyclic moiety is present when present on a nucleic acid duplex.
  • more than one base can be positioned relative to one another without altering the duplex structure (eg, the structure of the phosphate backbone).
  • the universal base does not disrupt the ability of the single-stranded nucleic acid in which it resides to form a duplex with the target nucleic acid.
  • a single-stranded nucleic acid containing a universal base to form a duplex with a target nucleic acid can be determined by methods apparent to those skilled in the art (eg, UV absorbance, circular dichroism, gel shift, single-stranded nuclease sensitivity, etc. ). Additionally, the conditions under which duplex formation is observed can be varied to determine duplex stability or formation, eg, temperature, such as melting temperature (Tm), correlates with nucleic acid duplex stability.
  • Tm melting temperature
  • the Tm of the duplex formed by the single-stranded nucleic acid containing the universal base with the target nucleic acid is lower than that of the duplex formed with the complementary nucleic acid.
  • the Tm of the duplex formed by the single-stranded nucleic acid containing the universal base and the target nucleic acid is higher than that of the single-stranded nucleic acid with the mismatched base. nucleic acid duplexes.
  • Some universal bases can be formed by base pairing between universal bases and all bases guanine (G), cytosine (C), adenine (A), thymine (T) and uracil (U) hydrogen bonds for base pairing.
  • Universal bases are not bases that only form base pairs with a single complementary base.
  • a universal base may not hydrogen bond, form one hydrogen bond, or form more than one hydrogen bond with each of the G, C, A, T, and U opposing it on the opposite strand of the duplex.
  • the universal base does not interact with the opposite base on the opposite strand of the duplex.
  • base pairing with universal bases does not alter the double helix structure of the phosphate backbone.
  • Universal bases can also interact with bases in adjacent nucleotides on the same nucleic acid strand via stacking interactions. This stacking interaction can stabilize the duplex, especially if the universal base does not form any hydrogen bonds with bases positioned opposite it on the opposite strand of the duplex.
  • Non-limiting examples of universal binding nucleotides include inosine, 1-beta-D-ribofuranosyl-5-nitroindole, and/or 1-beta-D-ribofuranosyl-3-nitropyrrole.
  • blunt end or blunt end are used interchangeably and refer to the absence of unpaired nucleotides or nucleotide analogs at a given end of an siRNA, ie, no nucleotide overhangs. In most cases, an siRNA that is blunt-ended at both ends will be double-stranded over its entire length.

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Abstract

L'invention concerne un ARNsi pour inhiber l'expression du facteur XI de coagulation sanguine, ainsi qu'une composition et son utilisation médicale. En particulier, l'ARNsi comprend un brin sens et un brin antisens, le brin sens contenant une séquence nucléotidique A et le brin antisens contenant une séquence nucléotidique B, la séquence nucléotidique A et la séquence nucléotidique B étant au moins partiellement inversement complémentaires pour former une région double brin. La séquence nucléotidique A et la séquence nucléotidique du brin sens présentée dans le tableau 1 ne diffèrent pas de plus de 3 nucléotides l'une de l'autre, et la séquence nucléotidique B et la séquence nucléotidique du brin antisens présentée dans le tableau 1 ne diffèrent pas de plus de 3 nucléotides l'une de l'autre. L'invention concerne en outre une composition pharmaceutique, une cellule ou un kit contenant l'ARNsi, et l'utilisation de l'ARNsi dans la préparation d'un médicament pour traiter et/ou prévenir un sujet ayant des maladies liées au facteur XI de coagulation sanguine.
PCT/CN2021/110503 2020-08-04 2021-08-04 Arnsi pour inhiber l'expression du facteur xi de coagulation sanguine, et composition et utilisation médicale de celui-ci WO2022028457A1 (fr)

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WO2024046297A1 (fr) * 2022-09-02 2024-03-07 北京福元医药股份有限公司 Arnsi pour inhiber l'expression du gène du récepteur de l'asialoglycoprotéine et conjugué, composition pharmaceutique et utilisation associée
WO2024061185A1 (fr) * 2022-09-20 2024-03-28 上海舶望制药有限公司 Réactif d'arni spécifiquement modifié et composition
WO2024073363A3 (fr) * 2022-09-26 2024-05-23 Sirius Therapeutics, Inc. Molécules d'acide polynucléique pour inhiber l'expression de fxi, compositions pharmaceutiques et leurs utilisations
CN117881783A (zh) * 2023-02-17 2024-04-12 苏州时安生物技术有限公司 一种用于抑制细胞程序性死亡-配体1基因表达的siRNA、其缀合物和药物组合物及用途

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