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JP-2026514403-A - Oligonucleotide delivery carrier and method of use thereof

JP2026514403AJP 2026514403 AJP2026514403 AJP 2026514403AJP-2026514403-A

Abstract

This application relates to nucleic acids, and more particularly to oligonucleotide preparations comprising a target oligonucleotide and a non-target site covalently linked to the oligonucleotide, and to the pharmaceutical applications thereof.

Inventors

  • ガン ズーバオ
  • リ ロンチョン
  • カン ムーリン

Assignees

  • ラクティゲン セラピューティクス

Dates

Publication Date
20260511
Application Date
20240329
Priority Date
20230330

Claims (20)

  1. An oligonucleotide agent comprising a target oligonucleotide conjugated to a non-target site capable of promoting the delivery of the target oligonucleotide, wherein the non-target site comprises one or more covalently bonded structural units in series to form its backbone, with at least two adjacent structural units linked by phosphorothioate (PS) bonds, and each structural unit of the non-target site is selected from a chemical linker and a nucleotide, and the non-target site is It consists of one or more chemical linkers interposed between nucleotides, one or more nucleotides interposed between chemical linkers, a sequence of consecutive nucleotides and a sequence of chemical linkers continuously linked, or a sequence of chemical linkers that does not contain any nucleotides. An oligonucleotide agent in which at least one phosphodiester bond between two adjacent nucleotides, between two adjacent linkers, or between a nucleotide and an adjacent linker is replaced with a phosphorothioate (PS) bond, a mesylphosphoramide bond, or a boranophosphate bond.
  2. The aforementioned chemical linker a) L1 or S18 (Spacer-18 Linker) (1,1-Bis(4-methoxyphenyl)-1-phenyl-2,5,8,11,14,17-Hexaoxanonadecane-19-yl(2-cyanoethyl)diisopropylphosphorumidite), b) L4 or C6 (spacer-C6 linker) (6-(bis(4-methoxyphenyl)(phenyl)methoxy)hexyl(2-cyanoethyl)diisopropylphosphorumidite), c), L6(1,1-bis(4-methoxyphenyl)-1-phenyl-2,5,8,11,14-pentaoxahexadecane-16-yl (2-cyanoethyl)diisopropylphosphorumidite), d), L9 or S9 (spacer-9 linker) (2- (2- (2- (bis(4-methoxyphenyl) (phenyl)methoxy)ethoxy)ethoxy)ethyl (2-cyanoethyl)diisopropylphosphorumidite), e), L10 or C3 (spacer-C3 linker) (3-(bis(4-methoxyphenyl)(phenyl)methoxy)propyl(2-cyanoethyl)diisopropylphosphorumidite), f), L12(d spacer) ((2R, 3S)-2- ((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)tetrahydrofuran-3-yl(2-cyanoethyl)diisopropylphosphorumidite), g), L13 or C12 (spacer-C12 linker) (12- (bis(4-methoxyphenyl) (phenyl)methoxy)dodecyl (2-cyanoethyl)diisopropylphosphorumidite), h), L14 (spacer-L14 linker) (((1r, 4r)-4- ((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)cyclohexyl)methyl (d-cyanoethyl)diisopropylphosphorumidite), i) L15 (Spacer-L15 Linker) (4-(2-(Bis(4-methoxyphenyl)(phenyl)methoxy)ethyl)phenethyl(2-cyanoethyl)diisopropylphosphorumidite), j), L16 (spacer-L16 linker) (2- (1- (2- (bis(4-methoxyphenyl)(phenyl)methoxy)ethyl)cyclohexyl)ethyl (2-cyanoethyl)diisopropylphosphorumidite), k), C6x1((2S, 3S, 4S, 5S)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-5-methoxy-4-(pento-4-in-1-yloxy)tetrahydrofuran-3-yl(2-cyanoethyl)diisopropylphosphorumidite), l), C6x2((2S, 3S, 4S, 5S)-5-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-2-methoxy-4-(pento-4-in-1-yloxy)tetrahydrofuran-3-yl(2-cyanoethyl)diisopropylphosphorumidite), m), C6x5(2-((2-(bis(4-methoxyphenyl)(phenyl)methoxy)ethyl)(pento-4-in-1-yl)amino)ethyl(2-cyanoethyl)diisopropylphosphorumidite), n) , C6x7((9H-Fluoren-9-yl)methyl(4-((2S,4R)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-((bis(diisopropylamino)phosphanyl)oxy)pyrrolidine-1-yl)-4-oxobutyl)carbamate), o), L20 methyl 1-(5-(bis(4-methoxyphenyl)(phenyl)methoxy)pentyl)-2-(4-(((2-cyanoethoxy)(diisopropylamino)phosphanyl)oxy)butyl)-1H-benzo[d]imidazole-5-carboxylate, and The oligonucleotide agent according to claim 1, selected from p) and L42 6-((6-(bis(4-methoxyphenyl)(phenyl)methoxy)hexyl)disulfanyl)hexyl(2-cyanoethyl)diisopropylphosphoramidite.
  3. The chemical linker is a substituted or unsubstituted alkyl, a substituted or unsubstituted alkenyl, a substituted or unsubstituted alkynyl, an arylalkyl, an arylalkenyl, an arylalkynyl, a heteroarylalkyl, a heteroarylalkenyl, a heteroarylalkynyl, a heterocyclic alkyl, a heterocyclic alkenyl, a heterocyclic alkynyl, an aryl, a heteroaryl, a heterocyclic, a cycloalkyl, a cycloalkenyl, an alkylarylalkyl, an alkylarylalkenyl, an alkylarylalkynyl, an alkenylarylalkyl, an alkenylarylalkenyl, an alkenylarylalkynyl, an alkenylarylalkynyl, an alkenylarylalkynyl, an alkenylarylalkynyl, an alkenylarylalkynyl, an alkyl hetero A chemical group selected from arylalkynyl, alkenylheteroarylalkyl, alkenylheteroarylalkenyl, alkenylheteroarylalkynyl, alkenylheteroarylalkyl, alkenylheteroarylalkenyl, alkenylheteroarylalkynyl, alkylheterocyclicalkyl, alkylheterocyclicalkenyl, alkylheterocyclicalkenyl, alkylheterocyclicalkenyl, alkenylheterocyclicalkenyl, alkenylheterocyclicalkenyl, alkenylheterocyclicalkenyl, alkenylheterocyclicalkenyl, alkenylheterocyclicalkenyl, alkenylheterocyclicalkenyl, alkenylheterocyclicalkenyl, alkylaryl, alkenylaryl, alkenylaryl, alkylheteroaryl, alkenylheteroaryl, and alkynylheteroaryl, where one or more methylene groups are O, S, S(O), SO The oligonucleotide agent according to claim 1, wherein 2 , N(R') 2 , C(O), is interrupted or terminated by a cleavable linking group, a substituted or unsubstituted aryl, a substituted or unsubstituted heteroaryl, or a substituted or unsubstituted heterocyclic, where each R' is independently selected from hydrogen, a substituted or unsubstituted alkyl, aryl, aralkyl, alkylaryl, alkoxy, aryloxy, acyl, or an aliphatic group that may be linear or branched.
  4. The oligonucleotide agent according to any one of claims 1 to 3, wherein the non-target site is composed of m identical or different chemical linkers and n identical or different nucleotides, where m is an integer from 1 to 40 and n is an integer from 0 to 30.
  5. The non-target region is composed of a sequence of consecutively linked chemical linkers represented by the equation (linker1) x - (linker2) y , where linker1 is the first chemical linker, linker2 is a second chemical linker different from the first chemical linker, and x and y are integers satisfying 0 < x + y < 50. For example, the non-target site is S9, (S9) 2 , (S9) 3 , (S9) 4 , (S9) 5 , (S9) 6 , (S9) 7 , (S9) 8 , (S9) 9 , (S9) 10 , (S9) 11 , (S9) 12 , (S9) 13 , (S9) 14 , (S9) 15 , (S9) 16 , (S9) 17 , (S9) 18 , (S9) 19 , (S9) 20 , (S9) 21 , (S9) 22 , (S9) 23 , (S9) 24 , (S9) 25 , (S9) 26 , (S9) 27 , (S9) 28 , (S9) 29 (S9) 30 , L10, (L10) 2 , (L10) 3 , (L10) 4 , (L10) 5 , (L10) 6 , (L10) 7 , (L10) 8 , (L10) 9 , (L10) 10 , (L10) 11 , (L10) 12 , (L10) 13 , (L10) 14 , (L10) 15 , (L10) 16 , (L10) 17 , (L10) 18 , (L10) 19 , (L10) 20 , (L10) 21 , (L10) 22 , (L10) 23 , (L10) 24 , (L10) 25 , (L10) 26 , (L10) 27 , (L10) 28 , (L10) 29 , (L10) 30 , L12, (L12)2, (L12)3, (L12)4, (L12)5, (L12)6, (L12)7, (L12)8, (L12)9, (L12)10, (L12)11, (L12)12, (L12)13, (L12)14, (L12)15, (L12)1 6, (L12)17, (L12)18, (L12)19, (L12)20, (L12)21, (L12)22, (L12)23, (L12)24, (L12)25, (L12)26, (L12)27, (L12)28, (L12)29, (L12)30, S9-L10, S9-(L10) 2 , S9-(L10) 3 , S9-(L10) 4 , S9-(L10) 5 , S9-(L10) 6 , S9-(L10) 7 , S9-(L10) 8 , S9-(L10) 9 , S9-(L10) 10 , S9-(L10) 11 , S9-(L10) 12 , S9-(L10) 13 , S9-(L10) 14 , S9-(L10) 15 , S9-(L10) 16 , S9-(L10) 17 , S9-(L10) 18 , S9-(L10) 19 , S9-(L10) 20 , S9-(L10) 21 , S9-(L10) 22 , S9-(L10) 23 , S9-(L10) 24 , S9-(L10) 25 , S9-(L10) 26 , S9-(L10) 27 , S9-(L10) 28 , S9-(L10) 29 , S9-(L10) 30 , S9-L12, S9-(L12) 2 , S9-(L12) 3 , S9-(L12) 4 , S9-(L12) 5 , S9-(L12) 6 , S9-(L12) 7 , S9-(L12) 8 , S9-(L12) 9 , S9-(L12) 10 , S9-(L12) 11 , S9-(L12) 12 , S9-(L12) 13 , S9-(L12) 14 , S9-(L12) 15 , S9-(L12) 16 , S9-(L12) 17 , S9-(L12) 18 , S9-(L12) 19 , S9-(L12) 20 , S9-(L12) 21 , S9-(L12) 22 , S9-(L12) 23 , S9-(L12) 24 , S9-(L12) 25 , S9-(L12) 26 , S9-(L12) 27 , S9-(L12) 28 , S9-(L12) 29 , S9-(L12) 30 , L20, (L20) 2 , (L20) 3 , (L20) 4 , (L20) 5 , (L20) 6 , (L20) 7 , (L20) 8 , (L20) 9 , (L20) 10 , (L20) 11 , (L20) 12 , (L20) 13 , (L20) 14 , (L20) 15 , (L20) 16 , (L20) 17 , (L20) 18 , (L20) 19 , (L20) 20 , (L20) 21 , (L20) 22 , (L20) 23 , (L20) 24 , (L20) 25 , (L20) 26 , (L20) 27 , (L20) 28 , (L20) 29 , (L20) 30 , L42, (L42) 2 , (L42) 3 , (L42) 4 , (L42) 5 , (L42) 6 , (L42) 7 , (L42) 8 , (L42) 9 , (L42) 10 , (L42) 11 , (L42) 12 , (L42) 13 , (L42) 14 , (L42) 15 , (L42) 16 , (L42) 17 , (L42) 18 , (L42) 19 , (L42) 20 , (L42) 21 , (L42) 22 , (L42) 23 , (L42) 24 , (L42) 25 , (L42) 26 , (L42) 27 , (L42) 28 , (L42) 29 , (L42) 30 , L20-L12, L20-(L12) 2 , L20-(L12) 3 , L20-(L12) 4 , L20-(L12) 5 , L20-(L12) 6 , L20-(L12) 7 , L20-(L12) 8 , L20-(L12) 9 , L20-(L12) 10 , L20-(L12) 11 , L20-(L12) 12 , L20-(L12) 13 , L20-(L12) 14 , L20-(L12) 15 , L20-(L12) 16 , L20-(L12) 17 , L20-(L12) 18 , L20-(L12) 19 , L20-(L12) 20 , L20-(L12) 21 , L20-(L12) 22 , L20-(L12) 23 , L20-(L12) 24 , L20-(L12) 25 , L20-(L12) 26 , L20-(L12) 27 , L20-(L12) 28 , L20-(L12) 29 , L20-(L12) 30 , or L42-L12, L42-(L12) 2 , L42-(L12) 3 , L42-(L12) 4 , L42-(L12) 5 , L42-(L12) 6 , L42-(L12) 7 , L42-(L12) 8 , L42-(L12) 9 , L42-(L12) 10 , L42-(L12) 11 , L42-(L12) 12 , L42-(L12) 13 , L42-(L12) 14 , L42-(L12) 15 , L42-(L12) 16 , L42-(L12) 17 , L42-(L12) 18 , L42-(L12) 19 , L42-(L12) 20 , L42-(L12) 21 , L42-(L12) 22 , L42-(L12) 23 , L42-(L12) 24 , L42-(L12) 25 , L42-(L12) 26 , L42-(L12) 27 , L42-(L12) 28 , L42-(L12) 29 , or L42-(L12) 30 , It consists of one of the following: Hereinafter, at least one phosphodiester bond between two adjacent linkers is substituted with a phosphorothioate (PS) bond, a mesylphosphoramidate bond, or a boranophosphate bond, as described in any one of claims 1 to 4.
  6. The oligonucleotide agent according to any one of claims 1 to 5, wherein the non-target site contains 1 to about 50, about 2 to about 48, about 3 to about 46, about 4 to about 44, about 5 to about 42, about 6 to about 40, about 7 to about 38, about 8 to about 36, about 9 to about 34, about 10 to about 32, about 11 to about 30, about 12 to about 28, about 13 to about 26, about 14 to about 24, about 15 to about 22, about 16 to about 20, or about 17 to about 18 phosphorothioate (PS) bonds in its skeleton.
  7. The oligonucleotide agent according to any one of claims 1 to 5, wherein the non-target site contains at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, or 40 or more phosphorothioate (PS) bonds in its skeleton.
  8. The oligonucleotide agent according to any one of claims 1 to 7, wherein the target oligonucleotide is an antisense oligonucleotide, or a double-stranded oligonucleotide (e.g., siRNA or saRNA) comprising a sense strand and an antisense strand.
  9. The oligonucleotide agent according to claim 8, wherein the non-target site is conjugated to the sense strand or antisense strand of the double-stranded oligonucleotide.
  10. The oligonucleotide agent according to any one of claims 1 to 9, wherein, if a nucleotide is present at the non-target site, the nucleotide is either an unchemically modified nucleotide or at least one nucleotide is a chemically modified nucleotide.
  11. The oligonucleotide agent according to any one of claims 1 to 10, wherein all nucleotides of the target oligonucleotide are unchemically modified nucleotides, or at least one nucleotide is chemically modified, or at least one phosphodiester bond between two adjacent nucleotides in the target oligonucleotide is substituted with a phosphorothioate (PS) bond, a mesylphosphoramidate bond, or a boranophosphate bond.
  12. The aforementioned chemically modified nucleotide a) Modification of the 2'-OH group of ribose in a nucleotide, b) Modification or unmodification of the base portion on the nucleoside ring in the nucleotide, c) that one nucleotide is a locked nucleic acid or a cross-linked nucleic acid, and d) Each nucleotide is a deoxyribonucleotide (DNA), The oligonucleotide agent according to claim 10 or 11, comprising one or more of the above.
  13. The oligonucleotide agent according to claim 12, wherein the chemically modified nucleotide has a 2'-OH ribose modification selected from 2'-fluoro-2'-deoxynucleoside (2'-F) modification, 2'-O-methyl (2'-O-Me) modification, and 2'-O-(2-methoxyethyl) (2'-O-MOE) modification.
  14. If a nucleotide is present at the non-target site, the nucleotide is selected from the group consisting of RNA, DNA, cross-linked nucleic acid (BNA), locked nucleic acid (LNA), and peptide nucleic acid (PNA), according to any one of claims 1 to 13.
  15. The oligonucleotide agent according to any one of claims 10 or 11, wherein the at least one chemically modified nucleotide is a nucleotide to which a 5'-phosphate group, 5-methylcytosine, or 5'-(E)-vinylphosphonate has been added.
  16. The oligonucleotide agent according to any one of claims 1 to 15, wherein the target oligonucleotide and the non-target site are directly conjugated, for example, via a phosphorothioate (PS) bond.
  17. The oligonucleotide agent according to claim 16, wherein the terminal unit or internal unit of the non-target site is conjugated to the target oligonucleotide.
  18. The oligonucleotide agent according to any one of claims 1 to 17, wherein the non-target site is conjugated to the 3' end, 5' end, both the 3' and 5' ends, or one internal nucleotide of the sense strand or antisense strand of the double-stranded oligonucleotide.
  19. The aforementioned non-target site is 1) A single chemical linker, which does not contain nucleotides, is conjugated to the terminal of the target oligonucleotide via a phosphorothioate (PS) bond. 2) A sequence of nucleotide-free, sequentially linked chemical linkers conjugated to one end of the target oligonucleotide, for example, via 1 to 20 deoxyribonucleotides (DNA) of any choice. 3) Two sequences of nucleotide-free, sequentially linked chemical linkers, conjugated to both ends of each target oligonucleotide, for example, via 1 to 20 deoxyribonucleotides (DNA) as optional, and 4) A sequence comprising one or more consecutive nucleotide sequences and a sequence of consecutively linked chemical linkers, wherein the nucleotides are interposed between the chemical linkers (preferably, each nucleotide is interposed between two chemical linkers, or one or more consecutive nucleotide sequences are interposed between chemical linkers). Selected from, Herein, at least one phosphodiester bond between two adjacent nucleotides, between two adjacent linkers, or between a nucleotide and an adjacent linker is substituted with a phosphorothioate (PS) bond, as described in any one of claims 1 to 18.
  20. The oligonucleotide agent according to any one of claims 1 to 19, wherein the internal nucleotides of the sense strand or antisense strand of the double-stranded oligonucleotide are substituted with a linking component, and the non-target site is conjugated to the linking component.

Description

This application relates to the technical field of nucleic acids, and more particularly to oligonucleotide agents comprising double-stranded RNA (dsRNA, duplex) and a non-target site covalently conjugated to the dsRNA, and to the pharmaceutical applications of the same. (Cross-reference with related applications) This application claims priority to the filing date of the provisional patent application "PCT/CN2023/085065," filed on March 30, 2023, and its disclosures are incorporated herein by reference in their entirety. (Array List) This application includes a sequence listing submitted electronically in computer-readable format, which is incorporated herein by reference in its entirety. Oligonucleotides are an innovative class of therapeutic agents that have attracted significant attention in the scientific community due to their flexibility in addressing a wide range of diseases through diverse mechanisms of action (MOAs). This therapeutic category is broadly divided into single-stranded antisense oligonucleotides (ASOs) and double-stranded RNA molecules (dsRNAs), which, while having different applications, share similar chemical compositions. Small interfering RNAs (siRNAs) and small activating RNAs (saRNAs), including dsRNA molecules, are representative analogues within the dsRNA subclass. These molecules exhibit chemical similarities in their structural framework, length, the strategies used for chemical modification, and terminal structures. Functionally, both siRNAs and saRNAs undergo a series of biological processes necessary for their mechanism of action. These include intracellular uptake, evasion from endosomal or lysosomal pathways, and interaction with the Argonaut (AGO) protein in the cytoplasm. This interaction cleaves and releases the non-functional "passenger" strand, while the functional "guide" strand remains bound to AGO and exerts its mechanism of action. This common biological process suggests that siRNA and saRNA delivery technologies are mutually interchangeable, potentially providing a unified approach to the delivery of these therapeutic molecules. Compared to ASOs, which can independently interact with intracellular uptake mechanisms via targeted chemical modifications, dsRNA molecules such as siRNA and saRNA require the support of a drug delivery system (DDS) to effectively reach target cells and exert therapeutic effects. Currently, various dsRNA DDS platforms, including polymer-based, lipid-based, and conjugate-based systems, are being developed for dsRNA delivery. However, the efficiency of these systems in reaching specific target organs, tissues, and cells remains limited, highlighting the need for innovative and advanced delivery solutions. (overview) This application provides a novel oligonucleotide agent or oligonucleotide agent conjugate comprising a target oligonucleotide (e.g., double-stranded RNA) and a non-target site conjugated to the target oligonucleotide. The oligonucleotide agent constitutes a system having "self-delivery" properties. Surprisingly, the inventors have found that when the non-target site disclosed herein is conjugated to a target oligonucleotide (including double-stranded RNA, siRNA, or saRNA), favorable in vivo distribution and high in vivo activity are obtained in local administration to selected tissues and systemic delivery to multiple organs and tissues such as the liver, muscle, lungs, kidneys, bladder, brain, spinal cord, heart, eyes, and spleen. In some embodiments, the following oligonucleotide agents are provided: (a) a double-stranded oligonucleotide comprising a sense strand and an antisense strand, wherein the antisense strand is complementary to the target nucleic acid; and (b) a non-target site comprising one or more components covalently linked by at least one phosphorothioate bond, wherein the double-stranded oligonucleotide is conjugated to the non-target site to form the oligonucleotide agent. In some embodiments, the double-stranded oligonucleotide is siRNA or saRNA. The non-target sites disclosed herein may comprise one or more identical or different components (or "units"), which are covalently linked in series to form the framework of the non-target site. The non-target site may have a linear or branched structure. In some embodiments, the components may be substituted or unsubstituted alkyl, aralkyl, alkoxy, aryloxy, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, arylalkyl, arylalkenyl, arylalkynyl, heteroarylalkyl, heteroarylalkenyl, heteroarylalkynyl, heterocyclic alkyl, heterocyclic alkenyl, heterocyclic alkynyl, aryl, heteroaryl, heterocyclic, cycloalkyl, cycloalkenyl, alkylarylalkyl, alkylarylalkenyl, alkylarylalkynyl, alkenylarylalkyl, alkenylarylalkenyl, alkenylarylalkynyl, alkynylarylalkyl, alkynylarylalkenyl, alkynylarylalkynyl, alkylheteroarylalkyl, alkylheteroarylalkyl Selected from teloarylalkenyl, alkylheteroarylalkynyl, alkenylheteroarylalkyl, alkenylheteroarylalkenyl, alkenylhete