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CN-122012496-A - Double-stranded ribonucleic acid for inhibiting CIDEB gene expression, and modifications, conjugates and uses thereof

CN122012496ACN 122012496 ACN122012496 ACN 122012496ACN-122012496-A

Abstract

The present disclosure relates to double-stranded ribonucleic acids and modifications, conjugates, and uses thereof for inhibiting CIDEB gene expression. In particular, the present disclosure relates to a double-stranded ribonucleic acid, double-stranded ribonucleic acid modification, double-stranded ribonucleic acid conjugate, prodrug, pharmaceutical composition and use for inhibiting CIDEB gene expression, and a method for inhibiting CIDEB gene expression in a cell. The double-stranded ribonucleic acid provided by the disclosure can be combined in cells to form an RNA-induced silencing complex (RISC), cut mRNA transcribed by CIDEB genes, efficiently and specifically inhibit CIDEB gene expression, is used for treating CIDEB gene-mediated diseases, and has important application prospects in clinical disease treatment.

Inventors

  • ZHAI BEIBEI
  • WANG SHUCHENG
  • HUANG HE
  • WANG XIAOJUN
  • NIU LIJIAO
  • WANG YAN
  • LIN GUOLIANG
  • CHAN YUNXIA
  • GENG YUXIAN

Assignees

  • 北京福元医药股份有限公司

Dates

Publication Date
20260512
Application Date
20251110
Priority Date
20241111

Claims (20)

  1. 1. A double-stranded ribonucleic acid that inhibits CIDEB gene expression, the double-stranded ribonucleic acid comprising a sense strand and an antisense strand, the sense strand being reverse-complementary and/or substantially reverse-complementary to the antisense strand to form a double-stranded region of the double-stranded ribonucleic acid; Wherein the sense strand comprises a sequence a that differs by no more than 3 nucleotides from at least 15 consecutive nucleotides in the target sequence, and the antisense strand comprises a sequence B that differs by no more than 3 nucleotides from the reverse complement of at least 15 consecutive nucleotides in the target sequence; The target sequence is selected from a nucleotide sequence shown in any one of SEQ ID NO 1-9 and a sequence consisting of at least 15 continuous nucleotides contained in any one of SEQ ID NO 1-9.
  2. 2. The double-stranded ribonucleic acid according to claim 1, wherein said target sequence is selected from the group consisting of the nucleotide sequences as set forth in any one of SEQ ID NOs 1, 4, 6 to 15, said sense strand comprises a sequence A consisting of at least 15 consecutive nucleotides of the nucleotide sequences as set forth in any one of SEQ ID NOs 1, 4, 6 to 15, and said antisense strand comprises a sequence B which is reverse-complementary and/or substantially reverse-complementary to the sequence consisting of at least 15 consecutive nucleotides of the nucleotide sequences as set forth in any one of SEQ ID NOs 1, 4, 6 to 15.
  3. 3. Double-stranded ribonucleic acid according to claim 1 or 2, wherein the sense strand consists of 15-28 nucleotides, preferably 19-25 nucleotides, more preferably 19-23 nucleotides, more preferably 19, 21 or 23 nucleotides, and/or The antisense strand consists of 15-28 nucleotides, preferably 19-25 nucleotides, more preferably 19-23 nucleotides, more preferably 19, 21 or 23 nucleotides.
  4. 4. A double-stranded ribonucleic acid according to claim 3, wherein the nucleotide sequence of the sense strand is a sequence A which differs by NO more than 1 nucleotide, preferably 19 to 25 consecutive nucleotides, more preferably 19 to 23 consecutive nucleotides, more preferably 19, 21 or 23 nucleotides, from a sequence consisting of 15 to 28 consecutive nucleotides in the nucleotide sequence shown in any one of SEQ ID NOs 1, 4, 6 to 15, and/or The nucleotide sequence of the antisense strand is a sequence B which differs by not more than 1 nucleotide, preferably 19 to 25 consecutive nucleotides, more preferably 19 to 23 consecutive nucleotides, still more preferably 19, 21 or 23 nucleotides from the reverse complement of a sequence consisting of 15 to 28 consecutive nucleotides in the nucleotide sequence shown in any one of SEQ ID NOs 1, 4, 6 to 15.
  5. 5. The double stranded ribonucleic acid of any one of claims 1 to 4, wherein the double stranded region is 15-25 nucleotides in length, preferably 19-23 nucleotides, more preferably 19, 21 or 23 nucleotides.
  6. 6. The double-stranded ribonucleic acid according to any one of claims 1 to 5, wherein, The sense strand and the antisense strand are complementary to form the double-stranded region, and the 3 '-end of the antisense strand has 1-2 protruding nucleotides extending out of the double-stranded region, and the 3' -end of the sense strand has a blunt end, or The sense strand is complementary to the antisense strand to form the double-stranded region, and the sense strand and the 3' -end of the antisense strand each have blunt ends.
  7. 7. The double-stranded ribonucleic acid of any one of claims 1 to 5, wherein the sense strand and the antisense strand are selected from the group consisting of: the sense strand comprises the sense strand of any one of the siRNAs shown in Table 1, table 1-1 herein, and the antisense strand comprises the antisense strand of the corresponding siRNA; preferably, the sense strand and the antisense strand are selected from the following combinations: The sense strand comprises the sense strand of any one of the siRNAs 64, 224, 282, 299, shown in Table 1, table 1-1 herein, and the antisense strand comprises the antisense strand of the corresponding siRNA.
  8. 8. The double-stranded ribonucleic acid of any one of claims 1 to 7, wherein each nucleotide in the sense strand is a modified nucleotide or an unmodified nucleotide independently of each other and/or each nucleotide in the antisense strand is a modified nucleotide or an unmodified nucleotide independently of each other.
  9. 9. The double-stranded ribonucleic acid of any one of claims 1 to 8, wherein any two nucleotides linked in the sense strand are linked by a phosphodiester or phosphorothioate linkage and/or any two nucleotides linked in the antisense strand are linked by a phosphodiester or phosphorothioate linkage.
  10. 10. The double-stranded ribonucleic acid of any one of claims 1 to 9, wherein the 5 'terminal nucleotide of the antisense strand is linked to a 5' phosphate group or a5 'phosphate derivative group, or the 5' terminal nucleotide of the antisense strand is not linked to a5 'phosphate group or a 5' phosphate derivative group.
  11. 11. The double-stranded ribonucleic acid of any one of claims 1 to 10, wherein neither the 5 'end nor the 3' end of the sense strand is linked to an inverted abasic deoxyribose residue, or the 3 'end nucleotide and/or the 5' end nucleotide of the sense strand is linked to an inverted abasic deoxyribose residue.
  12. 12. The double-stranded ribonucleic acid of any one of claims 1 to 11, wherein the double-stranded ribonucleic acid is an siRNA for inhibiting CIDEB gene expression.
  13. 13. A double-stranded ribonucleic acid modification, which is a modification of a double-stranded ribonucleic acid according to any one of claims 1 to 12, said double-stranded ribonucleic acid modification comprising a chemical modification of at least one of: (1) Modification of at least one nucleotide in the sense strand, (2) Modification of the phosphodiester bond in at least one position in the sense strand, (3) Modification of at least one nucleotide in the antisense strand, (4) Modification of the phosphodiester bond at least one position in the antisense strand; Optionally, the nucleotide sequence of the antisense strand of the double-stranded ribonucleic acid modification comprises a sequence B and a sequence E, the 3' end of the sequence B in the antisense strand being linked to a sequence E consisting of 1-2 nucleotides, preferably a sequence E consisting of 1-2 thymidylate nucleotides; Alternatively, the sense strand and the antisense strand of the double-stranded ribonucleic acid modification are selected from the following sequence combinations: the nucleotide sequence of the sense strand is a sequence shown in a sequence A, and the nucleotide sequence of the antisense strand is a sequence shown in a sequence B; or the nucleotide sequence of the sense strand is a sequence shown in a sequence A, and the nucleotide sequence of the antisense strand is a sequence shown in a sequence B and a connecting sequence E.
  14. 14. The double-stranded ribonucleic acid modification of claim 13, where the modification of the nucleotide is selected from a2 '-fluoro modification, a 2' -alkoxy modification, a2 '-substituted alkoxy modification, a 2' -alkyl modification, a2 '-substituted alkyl modification, a 2' -deoxy modification, a nucleotide derivative modification, or a combination of any two or more thereof; Preferably, the modification of the nucleotide is selected from the group consisting of a 2'-F modification, a 2' -O-CH 3 modification, a 2'-O-CH 2 -CH 2 -O-CH 3 modification, a 2' -O-CH 2 -CH=CH 2 modification, a 2'-CH 2 -CH 2 -CH=CH 2 modification, a 2' -deoxy modification, a nucleotide derivative modification, or a combination of any two or more thereof; more preferably, the nucleotide derivative in the nucleotide derivative modification is selected from the group consisting of an iso nucleotide, LNA, ENA, cET, UNA, or GNA.
  15. 15. The double-stranded ribonucleic acid modification of claim 13 or 14, wherein, The ribonucleotides at the 9 th, 10 th, 11 th and 13 th positions in the sense strand are 2'-F modified ribonucleotides along the direction from the 3' end to the 5 'end, and the ribonucleotides at the other positions in the sense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 3 'end to the 5' end, the ribonucleotides at the 11 th, 12 th, 13 th and 15 th positions in the sense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the sense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 3 'end to the 5' end, the ribonucleotides at the 9 th, 11 th and 13 th positions in the sense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the sense strand are 2' -O-CH 3 modified ribonucleotides; or in the direction from the 3 'end to the 5' end, the ribonucleotides at the 5 th, 7 th, 9 th, 11 th and 13 th positions in the sense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the sense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 3 'end to the 5' end, the ribonucleotides at the 7 th, 9 th, 11 th and 13 th positions in the sense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the sense strand are 2' -O-CH 3 modified ribonucleotides; or in the direction from the 3 'end to the 5' end, the ribonucleotides at the 11 th, 12 th, 13 th and 17 th positions in the sense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the sense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 3 'end to the 5' end, the ribonucleotides at the 9 th, 11 th, 13 th and 17 th positions in the sense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the sense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 3 'end to the 5' end, the ribonucleotides at the 6 th, 11 th and 13 th positions in the sense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the sense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 3 '-end to the 5' -end, the ribonucleotides at the 11 th and 13 th positions in the sense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the sense strand are 2' -O-CH 3 modified ribonucleotides.
  16. 16. The double-stranded ribonucleic acid modification of any one of claims 13 to 15, wherein when neither the 3 'terminal nucleotide nor the 5' terminal nucleotide of the sense strand is attached to an inverted abasic deoxyribose residue, the sense strand comprises a phosphorothioate diester linkage at the positions shown below, in the 5 'terminal to 3' terminal direction: between nucleotide 1 and nucleotide 2 from the 5' -end of the sense strand, and Between nucleotide 2 and nucleotide 3 from the 5' end of the sense strand; When only the 3' terminal nucleotide of the sense strand is linked to an inverted abasic deoxyribose residue, the sense strand comprises a phosphorothioate linkage at the positions shown below, in the 5' terminal to 3' terminal direction: between nucleotide 1 and nucleotide 2 from the 5' end of the sense strand; Between nucleotide 2 and nucleotide 3 from the 5' -end of the sense strand, and Between the reverse abasic deoxyribose residue starting at the 3' end of the sense strand and nucleotide 1; when the 5 'and 3' terminal nucleotides of the sense strand are linked to inverted abasic deoxyribose residues, respectively, the sense strand comprises phosphorothioate linkages at the positions shown below, in the 5 'to 3' terminal direction: Between the reverse abasic deoxyribose residue starting at the 5' end of the sense strand and nucleotide 1; between nucleotide 1 and nucleotide 2 from the 5' -end of the sense strand, and The sense strand is between the 3' -end-initiated reverse abasic deoxyribose residue and nucleotide 1.
  17. 17. The double-stranded ribonucleic acid modification of any one of claims 13 to 16, wherein, The ribonucleotides at the 2 nd, 6 th, 14 th and 16 th positions in the antisense strand are 2'-F modified ribonucleotides along the direction from the 5' end to the 3 'end, and the ribonucleotides at the other positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 5 'end to the 3' end, the ribonucleotides at the 2 nd, 6 th, 8 th, 9 th, 14 th and 16 th positions in the antisense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides; Or along the direction from the 5 'end to the 3' end, the ribonucleotides at the 2 nd position, the 14 th position and the 16 th position in the antisense strand are 2'-F modified ribonucleotides, the ribonucleotide at the 6 th position in the antisense strand is a nucleotide derivative GNA modified ribonucleotide, and the ribonucleotides at the rest positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 5 'end to the 3' end, the ribonucleotides at the 2 nd, 6 th, 14 th and 16 th positions in the antisense strand are 2'-F modified ribonucleotides, the ribonucleotide at the 7 th position in the antisense strand is a nucleotide derivative GNA modified ribonucleotide, and the ribonucleotides at the rest positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 5 'end to the 3' end, the ribonucleotides at the 2 nd, 3 rd, 4 th, 5 th, 7 th, 10 th and 14 th positions in the antisense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 5 'end to the 3' end, the ribonucleotides at the 2 nd, 3 rd, 4 th, 5 th, 7 th, 10 th and 14 th positions in the antisense strand are 2'-F modified ribonucleotides, the ribonucleotide at the 6 th position in the antisense strand is a nucleotide derivative GNA modified ribonucleotide, and the ribonucleotides at the rest positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 5 'end to the 3' end, the ribonucleotides at the 2 nd, 7 th, 10 th and 14 th positions in the antisense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 5 'end to the 3' end, the ribonucleotides at the 2 nd, 5 th, 7 th and 14 th positions in the antisense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 5 'end to the 3' end, the ribonucleotides at the 2 nd, 3 rd, 5 th, 7 th, 10 th, 12 th and 14 th positions in the antisense strand are 2'-F modified ribonucleotides, the ribonucleotide at the 6 th position in the antisense strand is a nucleotide derivative GNA modified ribonucleotide, and the ribonucleotides at the rest positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 5' end to the 3' end, the ribonucleotides at the 2 nd, 7 th and 14 th positions in the antisense strand are 2' -F modified ribonucleotides, the ribonucleotides at the 5 th and 12 th positions in the antisense strand are 2' -deoxidized modified nucleotides, and the ribonucleotides at the rest positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 5 'end to the 3' end, the ribonucleotides at the 2 nd, 7 th, 12 th and 14 th positions in the antisense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides; Or in the direction from the 5 'end to the 3' end, the ribonucleotides at the 2 nd, 6 th, 12 th and 14 th positions in the antisense strand are 2'-F modified ribonucleotides, and the ribonucleotides at the rest positions in the antisense strand are 2' -O-CH 3 modified ribonucleotides.
  18. 18. The double stranded ribonucleic acid modification of any one of claims 13 to 17, wherein the antisense strand comprises phosphorothioate linkages at positions shown below: between nucleotide 1 and nucleotide 2 from the 5' end of the antisense strand; between nucleotide 2 and nucleotide 3 from the 5' end of the antisense strand; Between nucleotide 1 and nucleotide 2 from the 3' end of the antisense strand; the antisense strand is between nucleotide 2 and nucleotide 3 from the 3' terminus.
  19. 19. The double stranded ribonucleic acid modification of any one of claims 13 to 18, wherein the sense strand and the antisense strand are selected from the group consisting of: the sense strand comprises the sense strand of any one of the double-stranded ribonucleic acid modifications shown in table 2 herein, and the antisense strand comprises the antisense strand of the corresponding siRNA modification; preferably, the sense strand and the antisense strand are selected from the following combinations: the sense strand comprises the sense strand of any one of the siRNA modifications shown in table 2 herein, and the antisense strand comprises the antisense strand of the corresponding siRNA modification.
  20. 20. A double-stranded ribonucleic acid conjugate, wherein the double-stranded ribonucleic acid conjugate comprises a double-stranded ribonucleic acid according to any one of claims 1 to 12, or a double-stranded ribonucleic acid modification according to any one of claims 13 to 19, and a conjugation group conjugated to the double-stranded ribonucleic acid or the double-stranded ribonucleic acid modification.

Description

Double-stranded ribonucleic acid for inhibiting CIDEB gene expression, and modifications, conjugates and uses thereof Technical Field The present disclosure is in the field of biological medicine, and in particular, relates to a double-stranded ribonucleic acid, a double-stranded ribonucleic acid modification, a double-stranded ribonucleic acid conjugate, a prodrug, a pharmaceutical composition and use for inhibiting CIDEB gene expression, and a method for inhibiting CIDEB gene expression in a cell. Background Cell death-inducing DFFA (DNA Fragmentation Factor A, DNA fragmentation factor alpha subunit) -like effector b (CIDEB) is a member of the CIDE protein family, expressed mainly in the liver and small intestine, a protein associated with the endoplasmic reticulum (endoplasmic reticulum, abbreviated ER) and lipid droplets (lipid droplets, abbreviated LD). The expression of CIDEB protein induces cell death, and its physiological function is related to various lipid metabolism, in particular, can promote the lipidation and maturation of very low density lipoprotein (very low density lipoprotein, abbreviated as VLDL) particles, and plays a key role in maintaining liver lipid homeostasis. CIDEB protein can promote the absorption and storage of lipid in liver and intestinal tract, and over-expression of CIDEB protein can cause hepatocyte damage and lipid metabolism imbalance, and the process is extremely easy to cause liver steatosis. In summary, by inhibiting CIDEB gene expression in a patient, disorders associated with metabolic disorders such as liver fibrosis, hepatitis b, hepatitis c, nonalcoholic steatohepatitis, atherosclerosis, cardiovascular disease, ulcerative colitis, clear cell renal cell carcinoma, obesity, diabetes, liver steatosis, metabolic disorders can be prevented and treated. At present, no related medicine only aiming at the expression of the genes is marketed, so that the development of the medicine targeting CIDEB targets has important value. Disclosure of Invention Problems to be solved by the invention The present disclosure aims to provide double-stranded ribonucleic acid, double-stranded ribonucleic acid modifications, double-stranded ribonucleic acid conjugates, prodrugs, pharmaceutical compositions, which are effective for RNA-induced silencing complex (RISC) -mediated cleavage of RNA transcripts of CIDEB genes, thereby being capable of selectively and effectively inhibiting CIDEB gene expression, and achieving the purpose of disease treatment. In view of the problems in the prior art, there is a need to develop more CIDEB inhibitors for the treatment of CIDEB-related diseases including liver fibrosis, hepatitis b, hepatitis c, non-alcoholic steatohepatitis, atherosclerosis, cardiovascular diseases, ulcerative colitis, clear cell renal cell carcinoma, obesity, diabetes, liver steatosis, metabolic disorders, etc., as well as other unidentified related conditions, pathologies or syndromes. The present disclosure aims at providing a series of double-stranded ribonucleic acid, double-stranded ribonucleic acid modifier, double-stranded ribonucleic acid conjugate, prodrug and pharmaceutical composition for inhibiting CIDEB gene expression, which can inhibit CIDEB gene expression and has important application prospects in clinical disease treatment. Solution for solving the problem [1] A double-stranded ribonucleic acid that inhibits CIDEB gene expression, said double-stranded ribonucleic acid comprising a sense strand and an antisense strand, said sense strand being reverse-complementary and/or substantially reverse-complementary to said antisense strand forming a double-stranded region of said double-stranded ribonucleic acid; Wherein the sense strand comprises a sequence a that differs by no more than 3 nucleotides from at least 15 consecutive nucleotides in the target sequence, and the antisense strand comprises a sequence B that differs by no more than 3 nucleotides from the reverse complement of at least 15 consecutive nucleotides in the target sequence; The target sequence is selected from a nucleotide sequence shown in any one of SEQ ID NO 1-9 and a sequence consisting of at least 15 continuous nucleotides contained in any one of SEQ ID NO 1-9. [2] A double-stranded ribonucleic acid according to [1], wherein the target sequence is selected from the nucleotide sequences shown in any one of SEQ ID NOs 1, 4, 6 to 15, the sense strand comprises a sequence A consisting of at least 15 consecutive nucleotides in the nucleotide sequence shown in any one of SEQ ID NOs 1, 4, 6 to 15, and the antisense strand comprises a sequence B consisting of at least 15 consecutive nucleotides in the nucleotide sequence shown in any one of SEQ ID NOs 1, 4, 6 to 15, which is reverse-complementary and/or substantially reverse-complementary. [3] The double-stranded ribonucleic acid according to [1] or [2], wherein the sense strand consists of 15 to 28 nucleotides, preferably 19 to 25 nucleotides, more pref