WO2023208106A1 - Compound having liver-targeting delivery effect and oligonucleotide conjugate thereof - Google Patents

Abstract

The present invention provides a compound represented by formula (I), or a prodrug, a tautomer, a stereoisomer, a solvate, an isotope derivative, or a pharmaceutically acceptable salt thereof. The compound of the present invention is a novel GalNAc derivative, which can form a chemical conjugate represented by formula (II) with an oligonucleotide, the chemical conjugate having a liver-targeting delivery effect.

Description

Compounds with liver-targeted delivery effects and oligonucleotide conjugates thereof

This application claims to have the prior rights submitted to the State Intellectual Property Office of China on April 29, 2022, with the patent application number 202210465744.5, and the invention title is "Compounds with liver-targeted delivery effects and oligonucleotide conjugates thereof" Priority interest in the application. The entirety of said prior application is incorporated into this application by reference.

Technical field

The present invention relates to the field of medical technology. Specifically, it relates to compounds with liver-targeted delivery effects and oligonucleotide conjugates thereof, their preparation methods and uses. This compound with liver-targeted delivery effects is a new type of compound. GalNAc derivatives.

Background technique

Asialoglycoprotein receptor (ASGPR) is an endocytic receptor specifically expressed in liver cells. It mainly exists on the cell membrane surface of liver parenchymal cells facing the sinusoidal space and is specific for sugar. . In recent years, breakthroughs have been made in liver-targeted delivery of nucleic acid drugs using the high-affinity ligand N-acetylgalactosamine (GalNAc) of ASGPR as a targeting molecule. Although this receptor has It has been discovered for many years, but liver-targeted drug delivery based on this receptor and its ligands is radiating continuous energy. Therefore, an in-depth understanding of the characteristics, properties, and mechanisms of ASGPR and its ligands requires innovative basic research, The design and development of new drugs are very important.

The main function of ASGPR is to remove asialoglycoprotein, apoptotic cells, and lipoproteins from the blood circulation system. In addition to asialoceruloplasmin, ASGPR can also bind to a variety of other asialoglycoproteins, such as erythropoietin, interferon, thyroglobulin, transferrin, hepatoglobulin, fetuin, and prothrombin etc., and these bindings have high specificity. At the same time, some studies have also found that ASGPR is a potential liver-specific receptor for the invasion of hepatitis B virus and Marburg virus. It is related to the infection of HepG2 cells or primary human urethral epithelial cells by Neisseria gonorrhoeae. , PHUECs), and may also be a major contributor to hemolysis in patients with alcoholic cirrhosis.

The amino acid sequence of ASGPR protein is highly conserved among various species. Human ASGPR is mainly expressed in liver parenchymal cells. After the GalNAc conjugate of the nucleic acid recognizes and binds to the ASGPR receptor, it will be rapidly endocytosed by the liver cells and form endosomes. Afterwards, the pH in the endosome drops, the GalNAc conjugate of the nucleic acid will dissociate from the ASGPR receptor, and the nucleic acid molecule will escape from the endosome. During the escape process, nucleic acid molecules will rapidly dissociate from GalNAc at the same time. About less than 1% of the nucleic acid molecules will escape from the double-layer lipid membrane structure of the endosome into the cytoplasm, and then go through a series of complex processes. Ultimately inducing a robust and sustained RNAi response.

Contents of the invention

The invention provides a class of compounds with liver-targeted delivery effects and oligonucleotide conjugates thereof or their prodrugs, tautomers, stereoisomers, solvates, isotope derivatives or their pharmaceutical Acceptable salts, this compound with liver-targeted delivery effect is a new type of GalNAc derivative, and also provides such compounds or their prodrugs, tautomers, stereoisomers, solvates, isotope derivatives or The pharmaceutically acceptable salts thereof are used in the preparation of medicaments, preferably in the preparation of medicaments for the treatment and prevention of physiological conditions or diseases caused by the expression of specific genes in liver cells.

Specifically, the present invention provides a compound represented by formula (I), or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, which Has the following structure:

where each R is independently selected from:

Y ₁ , Y ₂ , Y ₃ , Y ₄ , Y ₅ , Y ₆ are each independently selected from -O-, -S-, -N(G ₁ )-, -C(G ₁ ) ₂ -; G ₁ is each The second occurrence is independently selected from hydrogen, halogen, hydroxyl, amino, carboxyl, or C _1-10 alkyl optionally substituted by one or more of halogen, hydroxyl, amino, carboxyl, C _1-6 alkyl Base, C _1-10 alkoxy group, -N(C _1-10 alkyl) ₂ ;

Z ₁ , Z ₂ , Z ₃ and Z ₄ are each independently selected from C, N ⁺ and Si;

Each occurrence of A ₁ and A ₂ is independently selected from hydrogen, or acetyl, propionyl, benzoyl, benzyl, which may be optionally substituted by one or more of halogen, hydroxyl, amino, and carboxyl. benzyloxycarbonyl;

_{X 1 , X 2 , and _ _ _ _} G ₃ ) ₂ C(G ₃ ) ₂ S) _m -, or C _2-6 alkenyl, C _2-6 alkynyl, C _3-10 cycloalkyl optionally substituted by one or more G ₃ , C _3-10 cycloalkenyl, 3-10 membered heterocyclyl, C _6-14 aryl, 5-12 membered heteroaryl;

Each occurrence of G ₃ is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, oxo, C _1-6 alkyl, C _1-6 alkoxy, or by deuterium, halogen, hydroxyl, amino One or more optionally substituted C _1-6 alkyl, C _1-6 alkoxy;

X ₄ is independently selected from bonds, -(C(G ₄ ) ₂ ) _a -, -(C(G ₄ ) ₂ C(G ₃ ) ₂ O) _b -, -(C(G ₄ ) ₂ C(G ₃ ) ₂ S) _b -, or C _2-6 alkenyl, C _2-6 alkynyl, C _3-10 cycloalkyl, C _3-10 cycloalkenyl optionally substituted by one or more G ₄ , 3-10 membered heterocyclyl, C _6-14 aryl, 5-12 membered heteroaryl;

Each occurrence of G ₄ is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, oxo, C _1-6 alkyl, C _1-6 alkoxy, or by deuterium, halogen, hydroxyl, amino One or more optionally substituted C _1-6 alkyl, C _1-6 alkoxy;

n and a are independently selected from the integers from 0 to 24 each time they appear, and m and b are independently selected from the integers from 0 to 12 each time they appear;

J ₁ , J ₂ , J ₃ , and J ₄ are each independently selected from the group consisting of bonds, -O-, -S-, -NH-, -C(O)-, -C(S)-, -S(O)- , -S(O) ₂ -, -NHC(O)-, -C(O)NH-, -C(O)O-, -OC(O)-, -SS-, -N(G ₅ )C (O)-, -C(O)N(G ₅ )-, -N(G ₅ )-, -C(G ₅ ) ₂ -, -Si(G ₅ ) ₂ -, -P(O)(OH )-, -P(O)O-,

Each occurrence of G ₅ is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, carboxyl, -BH ₂ , -B(OH) ₂ , -N(C _1-6 alkyl) ₂ , C _1-12 alkyl group, C _1-12 alkoxy group, _or C 1-12 alkyl group, C _1-12 alkoxy group, C _3-10 ring optionally substituted by one or more of deuterium, halogen, hydroxyl, and amino group Alkyl, C _3-10 cycloalkenyl, 3-10 membered heterocyclyl, C _6-14 aryl, 5-12 membered heteroaryl;

Each occurrence of J ₅ and J ₆ is independently selected from the group consisting of bonds, -O-, -S-, -NH-, -C(O)-, -C(S)-, -S(O)-, - S(O) ₂ -, -NHC(O)-, -C(O)NH-, -C(O)O-, -OC(O)-, -SS-, -N(G ₆ )C(O )-, -C(O)N(G ₆ )-, -N(G ₆ )-, -C(G ₆ ) ₂ -, -Si(G ₆ ) ₂ -, -P(O)(OH)- ,-P(O)O-,

Each occurrence of G ₆ is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, carboxyl, -BH ₂ , -B(OH) ₂ , -N(C _1-6 alkyl) ₂ , C _1-12 alkyl group, C _1-12 alkoxy group, _or C 1-12 alkyl group, C _1-12 alkoxy group, C _3-10 ring optionally substituted by one or more of deuterium, halogen, hydroxyl, and amino group Alkyl, C _3-10 cycloalkenyl, 3-10 membered heterocyclyl, C _6-14 aryl, 5-12 membered heteroaryl;

_{Each occurrence of _ _} , C _6-14 aryl, 5-12 membered heteroaryl; the C _1-6 alkyl, C _1-6 alkoxy, C _3-10 cycloalkyl, C _3-10 cycloalkenyl, 3 -10-membered heterocyclyl, C _6-14 aryl, 5-12-membered heteroaryl is unsubstituted or optionally substituted by one or more G ₇ ;

Each occurrence of G ₇ is independently selected from halogen, hydroxyl, amino, cyano, oxo, -C _1-12 alkyl -OG ₈ , -OG ₉ , Wherein, Cat is a cation, independently selected from sodium ion, potassium ion, triethylammonium ion, tripropylammonium ion, tributylammonium ion and tetrabutylammonium ion; r is an integer from 0 to 5; Solid support is a solid phase carrier;

G ₈ is independently selected from the following structures: H,

G ₉ is independently selected from

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, Y ₁ , Y ₂ , Y ₃ , Y ₄ , Y ₅ , and Y ₆ are each independently selected from -O-, -N(G ₁ )-, -C(G ₁ ) ₂ -.

In a further preferred embodiment, wherein Y ₁ , Y ₂ , Y ₃ , Y ₄ , Y ₅ , Y ₆ are each independently selected from -O-, -NH-, -CH(G ₁ )-.

In a further preferred embodiment, wherein Y ₁ , Y ₂ , Y ₃ , Y ₄ , Y ₅ , Y ₆ are each independently selected from -CH ₂ -.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, G ₁ Each occurrence is independently selected from hydrogen, halogen, hydroxy, amino, carboxyl, or C _1-10 optionally substituted by one or more of halogen, hydroxy, amino, carboxyl, C _1-6 alkyl Alkyl, -N(C _1-10 alkyl) ₂ .

In a further preferred embodiment, wherein each occurrence of _G1 is independently selected from hydrogen, halogen, hydroxy, amino, carboxyl, or may be optionally substituted by one or more of halogen, hydroxy, amino, carboxyl C _1-6 alkyl, -N(C _1-6 alkyl) ₂ .

In a further preferred embodiment, wherein each occurrence of G ₁ is independently selected from hydrogen, amino, carboxyl, C _1-4 alkyl, C _1-4 alkyl optionally substituted with one or more fluorine, -N(C _1-4 alkyl) ₂ .

In a further preferred embodiment, wherein each occurrence of G ₁ is independently selected from hydrogen, C _1-4 alkyl, C _1-4 alkyl optionally substituted with one or more fluorine.

In a further preferred embodiment, wherein each occurrence of _G1 is independently selected from hydrogen, methyl, ethyl, isopropyl, trifluoromethyl.

In a further preferred embodiment, wherein each occurrence of _G1 is independently selected from hydrogen, amino, carboxyl, methyl, ethyl, isopropyl, trifluoromethyl, N,N-diisopropylamino.

In a further preferred embodiment, wherein each occurrence of _G1 is independently selected from hydrogen.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, Y ₁ The branch unit composed of , Y ₂ , Y ₃ , Y ₄ , Y ₅ , Y ₆ and Z ₁ , Z ₂ , Z ₃ , Z ₄ is selected from the following structures:

In a further preferred embodiment, Y ₁ , Y ₂ , Y ₃ , Y ₄ , Y ₅ , Y ₆ together with Z ₁ , Z ₂ , Z ₃ , Z ₄ constitute The branch units are selected from

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, Z ₁ , Z ₂ , Z ₃ and Z ₄ are each independently selected from C.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, A ₁ , each occurrence of A ₂ is independently selected from hydrogen, or acetyl, propionyl, benzoyl, benzyl, benzyloxycarbonyl, which may be optionally substituted by one or more halogens.

In a further preferred embodiment, wherein each occurrence of A ₁ and A ₂ is independently selected from hydrogen, acetyl, trifluoroacetyl, difluoroacetyl, monofluoroacetyl, propionyl, benzoyl, Benzyl, benzyloxycarbonyl.

In a further preferred embodiment, wherein each occurrence of A ₁ and A ₂ is independently selected from hydrogen, acetyl, and benzoyl.

In a further preferred embodiment, wherein each occurrence of A ₁ and A ₂ is independently selected from hydrogen and acetyl.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, X _{1 , X 2 , _ _ _ _ _ _ _} ) ₂ C(G ₃ ) ₂ S) _m -, or C _2-4 alkenyl, C _2-4 alkynyl, C _3-6 cycloalkyl, 3- optionally substituted by one or more G ₃ 6-membered heterocycloalkyl, C _3-6 cycloalkenyl, C _6-10 aryl, 5-6-membered heteroaryl, the heteroatoms in the heterocycloalkyl and heteroaryl are independently selected from O, N or S, the number of heteroatoms is 1, 2 or 3.

In a further _{preferred embodiment} , _wherein each of _{X 1 ,} X _{2 ,} and ₂ CH ₂ O) _m -, -(CH ₂ CH ₂ S) _m -, or C _2-4 alkenyl, C _2-4 alkynyl, C _5-6 optionally substituted by one or more G ₃ Cycloalkyl, C _5-6 cycloalkenyl, 5-6 membered heterocycloalkyl, C _6-10 aryl, 5-6 membered heteroaryl, heteroatoms in the heterocycloalkyl and heteroaryl Independently selected from O or N, the number of heteroatoms is 1 or 2.

In a further _{preferred embodiment} , _wherein each of _{X 1 ,} X _{2 ,} and ₂ CH ₂ O) _m -, -(CH ₂ CH ₂ S) _m -, or optionally substituted with one or more G ₃

In a further _{preferred embodiment} , _wherein each of _{X 1 ,} X _{2 ,} and ₂ CH ₂ O) _m -, -(CH ₂ CH ₂ S) _m -,

In a further _{preferred embodiment} , _wherein each of _{X 1 ,} X _{2 ,} and ₂ CH ₂ O) _m -, -(CH ₂ CH ₂ S) _m -.

In a further preferred embodiment, wherein X ₁ , X ₂ , X ₃ are each independently selected from the group consisting of bonds, -(CH ₂ ) _n -, -(CH ₂ CH ₂ O) _m -.

In a further preferred embodiment, wherein _X1 , _X2 , _X3 are each independently selected from the group consisting of bonds, _-CH2- , -( _CH2 ) _2- , -( _CH2 ) _3- , -( _CH2 ) ₄ -, -(CH ₂ ) ₅ -, -(CH ₂ ) ₆ -, -CH ₂ CH ₂ O-, -(CH ₂ CH ₂ O) ₂ -, -(CH ₂ CH ₂ O) ₃ -.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, X ₁ , X ₂ , and X ₃ are each independently selected from the bond, -(CH ₂ ) _n -.

In a further preferred embodiment, wherein X ₁ , X ₂ , X ₃ are each independently selected from the group consisting of bonds, -(CH ₂ ) ₂ -, -(CH ₂ ) ₄ -, -(CH ₂ ) ₆ -.

In a further preferred embodiment, wherein X ₁ is independently selected from bond, X ₂ is independently selected from bond, -(CH ₂ ) ₂ -, -(CH ₂ ) ₄ -, and X ₃ is independently selected from -(CH ₂ ) ₂ -, -(CH ₂ ) ₄ -, -(CH ₂ ) ₆ -.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, G ₃ Each occurrence is independently selected from hydrogen, halogen, hydroxyl, amino, oxo, C _1-4 alkyl, C _1-4 alkoxy, C _1-4 haloalkyl, C _1-4 haloalkoxy.

In a further preferred embodiment, wherein each occurrence of G ₃ is independently selected from -H, -F, -Cl, -Br, -I, -OH, -NH ₂ , oxo, -CH ₃ , - CH ₂ CH ₃ , -CH ₂ CH ₂ CH ₃ , -C(CH ₃ ) ₃ , -OCH ₃ , -OCH ₂ CH ₃ , -OCH ₂ CH ₂ CH ₃ , -OC(CH ₃ ) ₃ , -O( CH ₂ F), -O(CHF ₂ ), -O(CF ₃ ), -CH ₂ F, -CHF ₂ , -CF ₃ , -CHFCH ₃ , -CH ₂ CHF ₂ .

In a further preferred embodiment, wherein each occurrence of G ₃ is independently selected from -H, -F, -Cl, -Br, -I, -OH, -NH ₂ , oxo, -CH ₃ , - CH ₂ CH ₃ .

In a further preferred embodiment, wherein each occurrence of _G3 is independently selected from hydrogen.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, X ₄ Independently selected from the group consisting of bonds, -(C(G ₄ ) ₂ ) _a -, -(C(G ₄ ) ₂ C(G ₄ ) ₂ O) _b -, -(C(G ₄ ) ₂ C(G ₄ ) ₂ S) _b -, or C _2-4 alkenyl, C _2-4 alkynyl, C 3-6 cycloalkyl, _{C 3-6 cycloalkenyl} , 3 optionally substituted by one or more G ₄ -6-membered heterocycloalkyl, C _6-10 aryl, 5-6-membered heteroaryl, the heteroatoms in the heterocycloalkyl or heteroaryl are independently selected from O, N or S, the number of heteroatoms 1, 2 or 3.

In a further preferred embodiment, wherein X ₄ is independently selected from the group consisting of bonds, -(CH ₂ ) _a -, -(CD ₂ ) _a -, -(CHD) _a -, -(CH ₂ CH ₂ O) _b - , -(CH ₂ CH ₂ S) _b -, or C _2-4 alkenyl, C _2-4 alkynyl, C _5-6 cycloalkyl, C _5- optionally substituted by one or more G _{4 6-} cycloalkenyl, 5-6-membered heterocycloalkyl, C _6-10 aryl, 5-6-membered heteroaryl, the heteroatoms in the heterocycloalkyl and heteroaryl are independently selected from O or N , the number of heteroatoms is 1 or 2.

In a further preferred embodiment, wherein X ₄ is independently selected from the group consisting of bonds, -(CH ₂ ) _a -, -(CD ₂ ) _a -, -(CHD) _a -, -(CH ₂ CH ₂ O) _b - , -(CH ₂ CH ₂ S) _b -, or optionally substituted by G ₄

In a further preferred embodiment, wherein X ₄ is independently selected from the group consisting of bonds, -(CH ₂ ) _a -, -(CD ₂ ) _a -, -(CHD) _a -, -(CH ₂ CH ₂ O) _b - ,-(CH ₂ CH ₂ S) _b -,

In a further preferred embodiment, wherein X ₄ is independently selected from the group consisting of bonds, -(CH ₂ ) _a -, -(CD ₂ ) _a -, -(CHD) _a -, -(CH ₂ CH ₂ O) _b - , -(CH ₂ CH ₂ S) _b -.

In a further preferred embodiment, wherein X ₄ is independently selected from the group consisting of bonds, -(CH ₂ ) _a -, -(CH ₂ CH ₂ O) _b -.

In a further preferred embodiment, wherein X ₄ is independently selected from bond, -(CH ₂ ) ₅ -, -(CH ₂ ) ₆ -, -(CH ₂ ) ₇ -, -(CH ₂ ) ₈ -, - (CH ₂ ) ₉ -, -(CH ₂ ) ₁₀ -, -(CH ₂ ) ₁₁ -, -(CH ₂ ) ₁₂ -, -(CH ₂ ) ₁₃ -, -(CH ₂ ) ₁₄ -, -CH ₂ CH ₂ O-, -(CH ₂ CH ₂ O) ₂ -, -(CH ₂ CH ₂ O) ₃ -.

In a further preferred embodiment, wherein _X4 is independently selected from the group consisting of bonds, -( _CH2 ) _8- , -( _CH2 ) _9- , -( _CH2 ) _10- , -( _CH2 ) _11- , - (CH ₂ ) ₁₂ -, -(CH ₂ ) ₁₃ -, -(CH ₂ ) ₁₄ -, -CH ₂ CH ₂ O-, -(CH ₂ CH ₂ O) ₂ -.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, X ₄ Independently selected from -(CH ₂ ) _a -.

In a further preferred embodiment, wherein X ₄ is independently selected from -(CH ₂ ) ₁₀ -, -(CH ₂ ) ₁₁ -, -(CH ₂ ) ₁₂ -.

In a preferred embodiment of the invention, the compound provided by the invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, G ₄ Each occurrence is independently selected from hydrogen, halogen, hydroxyl, amino, oxo, C _1-4 alkyl, C _1-4 alkoxy, C _1-4 haloalkyl, C _1-4 haloalkoxy.

In a further preferred embodiment, wherein each occurrence of G ₄ is independently selected from -H, -F, -Cl, -Br, -I, -OH, -NH ₂ , oxo, -CH ₃ , - CH ₂ CH ₃ , -CH ₂ CH ₂ CH ₃ , -C(CH ₃ ) ₃ , -OCH ₃ , -OCH ₂ CH ₃ , -OCH ₂ CH ₂ CH ₃ , -OC(CH ₃ ) ₃ , -O( CH ₂ F), -O(CHF ₂ ), -O(CF ₃ ), -CH ₂ F, -CHF ₂ , -CF ₃ , -CHFCH ₃ , -CH ₂ CHF ₂ .

In a further preferred embodiment, wherein each occurrence of G ₄ is independently selected from -H, -F, -Cl, -Br, -I, -OH, -NH ₂ , oxo, -CH ₃ , - CH ₂ CH ₃ .

In a further preferred embodiment, wherein each occurrence of G ₄ is independently selected from hydrogen.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein n is independent Ground is selected from an integer of 0-15, specifically 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15.

In a further preferred embodiment, n is independently selected from an integer from 0 to 6, specifically 0, 1, 2, 3, 4, 5, 6.

In a further preferred embodiment, wherein n is independently selected from 2, 4, and 6.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, m is independently The ground is selected from an integer from 0 to 6, Specifically, they are 0, 1, 2, 3, 4, 5, and 6.

In a further preferred embodiment, wherein m is independently selected from an integer from 0 to 3, specifically 0, 1, 2, 3.

In a further preferred embodiment, wherein m is independently selected from 0, 1, 2.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein a is independently Ground is selected from an integer of 0-15, specifically 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15.

In a further preferred embodiment, wherein a is independently selected from an integer from 6 to 12, specifically 6, 7, 8, 9, 10, 11, 12.

In a further preferred embodiment, wherein a is independently selected from 10, 11, 12.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, b is independently Ground is selected from an integer from 0 to 6, specifically 0, 1, 2, 3, 4, 5, and 6.

In a further preferred embodiment, wherein b is independently selected from an integer from 0 to 3, specifically 0, 1, 2, 3.

In a further preferred embodiment, wherein b is independently selected from 0, 1, 2.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, J ₁ , J ₂ , J ₃ , J ₄ are each independently selected from the group consisting of bonds, -O-, -S-, -NH-, -C(O)-, -NHC(O)-, -C(O)NH-, -C(O)O-, -OC(O)-, -N(G ₅ )C(O)-, -C(O)N(G ₅ )-, -N(G ₅ )-, -C( G ₅ ) ₂ -.

In a further preferred embodiment, wherein J ₁ , J ₂ , J ₃ , J ₄ are each independently selected from the group consisting of bonds, -O-, -NHC(O)-, -C(O)NH-.

In a further preferred embodiment, wherein J ₁ is independently selected from bond, J ₂ is independently selected from bond, -O-, and J ₃ is independently selected from -O-, -NHC(O)-, -C(O )NH-, J ₄ is independently selected from -NHC(O)-, -C(O)NH-.

In a further preferred embodiment, wherein J ₁ is independently selected from bond, J ₂ is independently selected from bond, -O-, J ₃ is independently selected from -O-, -NHC(O)-, and J ₄ is independently selected from bond, -O- Selected from -C(O)NH-.

In a preferred embodiment of the invention, the compound provided by the invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, G ₅ Each occurrence is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, carboxyl, -BH ₂ , -B(OH) ₂ , -N(C _1-6 alkyl) ₂ , C _1-6 alkyl, C _1-6 alkoxy, or C 1-6 alkyl, C _1-6 alkoxy, C _3-6 cycloalkyl optionally substituted by one or more _of deuterium, halogen, hydroxyl, and amino , C _3-6 cycloalkenyl, 3-6 membered heterocycloalkyl, C _6-10 aryl, 5-6 membered heteroaryl.

In a further preferred embodiment, wherein each occurrence of G ₅ is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, carboxyl, -N(C _1-6 alkyl) ₂ , C _1-6 alkyl, C _1-6 alkoxy, or C 1-6 alkyl, _{C 1-6} alkoxy, C _3-6 cycloalkyl, C _3- optionally substituted by one or more of deuterium and halogen _6- cycloalkenyl, 3-6 membered heterocycloalkyl, C _6-10 aryl, 5-6 membered heteroaryl, the heteroatoms in the heterocycloalkyl and heteroaryl are independently selected from O or N , the number of heteroatoms is 1 or 2.

In a further preferred embodiment, wherein each occurrence of _G5 is independently selected from -H, -F, -Cl, -Br, -I, _-NH2 , _-CD3 , _-CH2F , _-CHF2 , -CF ₃ , -CH ₂ Cl , -CH ₂ I , -N(CH ₃ )(CH ₃ ) , -CH ₃ , -CH ₂ CH ₃ , -CH ₂ CH ₂ CH ₃ , -C(CH ₃ ) ₃ .

In a further preferred embodiment, wherein each occurrence of G ₅ is independently selected from -H, -F, -NH ₂ , -CD ₃ , -CF ₃ , -CH ₃ , -CH ₂ CH ₃ , -CH ₂ CH ₂ CH ₃ , -C(CH ₃ ) ₃ .

In a further preferred embodiment, wherein each occurrence of G ₅ is independently selected from -H.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, J ₅ , J ₆ are each independently selected from the keys , -O-, -S-, -NH-, -C(O)-, -NHC(O)-, -C(O)NH-, -C(O)O-, -OC(O)-, -N(G ₆ )C(O)-, -C(O)N(G ₆ )-, -N(G ₆ )-, -C(G ₆ ) ₂ -.

In a further preferred embodiment, wherein J ₅ , J ₆ are each independently selected from the group consisting of bonds, -C(O)-, -NHC(O)-, -C(O)NH-, -N(G ₆ )C (O)-, -C(O)N(G ₆ )-.

In a further preferred embodiment, wherein J ₅ , J ₆ are each independently selected from the group consisting of bonds, -O-, -C(O)-, -NHC(O)-, -C(O)NH-, -OC( O)-.

In a further preferred embodiment, wherein _J5 is independently selected from -NHC(O)-, -C(O)NH-.

In a further preferred embodiment, wherein _J6 is independently selected from bond, -C(O)-.

In a preferred embodiment of the invention, the compound provided by the invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, G ₆ Each occurrence is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, carboxyl, -BH ₂ , -B(OH)(OH), -N(C _1-6 alkyl)(C _1-6 alkyl ), C _1-6 alkyl, C _1-6 alkoxy, or C 1-6 alkyl, C _1-6 alkoxy optionally substituted by one or more _of deuterium, halogen, hydroxyl, and amino base, C _3-6 cycloalkyl, C _3-6 cycloalkenyl, 3-6 membered heterocycloalkyl, C _6-10 aryl, 5-6 membered heteroaryl.

In a further preferred embodiment, wherein each occurrence of G ₆ is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, carboxyl, -N(C _1-6 alkyl)(C _1-6 alkyl), C _1-6 alkyl, C _1-6 alkoxy, _or C 1-6 alkyl, C _1-6 alkoxy, C _3-6 optionally substituted by one or more of deuterium and halogen Cycloalkyl, C _3-6 cycloalkenyl, 3-6 membered heterocycloalkyl, C _6-10 aryl, 5-6 membered heteroaryl, heteroatoms in the heterocycloalkyl and heteroaryl Independently selected from O or N, the number of heteroatoms is 1 or 2.

In a further preferred embodiment, wherein each occurrence of G ₆ is independently selected from -H, -F, -Cl, -Br, -I, -NH ₂ , -CD ₃ , -CH ₂ F, -CHF ₂ , -CF ₃ , -CH ₂ Cl , -CH ₂ I , -N(CH ₃ )(CH ₃ ) , -CH ₃ , -CH ₂ CH ₃ , -CH ₂ CH ₂ CH ₃ , -C(CH ₃ ) ₃ .

In a further preferred embodiment, wherein each occurrence of G ₆ is independently selected from -H, -F, -NH ₂ , -CD ₃ , -CF ₃ , -CH ₃ , -CH ₂ CH ₃ , -CH ₂ CH ₂ CH ₃ , -C(CH ₃ ) ₃ .

In a further preferred embodiment, wherein each occurrence of G ₆ is independently selected from -H.

In a preferred embodiment of the invention, the compound provided by the invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, X ₅ Each occurrence is independently selected from bond, C _1-6 alkyl, C _3-6 cycloalkyl, C _3-6 cycloalkenyl, 3-6 membered heterocycloalkyl, C _6-10 aryl, 5 -6-membered heteroaryl; the C _1-6 alkyl, C _3-6 cycloalkyl, C _3-6 cycloalkenyl, 3-6 membered heterocycloalkyl, C _6-10 aryl, 5- 6-membered heteroaryl groups are unsubstituted or optionally substituted with one or more _G7 .

_In a _further preferred embodiment, wherein _each occurrence of Heterocycloalkyl is unsubstituted or optionally substituted with one or more _G7 .

In a further preferred embodiment, wherein each occurrence _{of X} is independently selected from bond, or methyl, ethyl, n-propyl, isopropyl, optionally substituted by one or more _G

In a further preferred embodiment, wherein each occurrence of _X5 is independently selected from methyl substituted by one or two _G7 ,

In a further preferred embodiment, wherein each occurrence of X ₅ is independently selected from the group consisting of substituted with one or two G ₇

In a preferred embodiment of the invention, the compound provided by the invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, G ₇ Independently selected from hydroxyl, -C _1-6 alkyl-OG ₈ , -OG ₉ ,

In a further preferred embodiment, wherein G ₇ is independently selected from hydroxyl, -C _1-4 alkyl-OG ₈ , -OG ₉ ,

In a further preferred embodiment, wherein G ₇ is independently selected from hydroxyl, -C _1-3 alkyl-OG ₈ , -OG ₉ ,

In a further preferred embodiment, wherein G ₇ is independently selected from hydroxyl, -CH ₂ -OG ₈ , -OG ₉ ,

In a further preferred embodiment, wherein G ₇ is hydroxyl.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, G ₈ Independently selected from: H,

In a further preferred embodiment, wherein G ₈ is independently selected from H,

In a further preferred embodiment, wherein G ₈ is independently selected from H,

In a further preferred embodiment, wherein G ₈ is independently selected from H,

In a further preferred embodiment, wherein G ₈ is selected from H or

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, G ₉ independently selected from

In a further preferred embodiment, wherein G ₉ is independently selected from

In a further preferred embodiment, wherein G ₉ is independently selected from

In a further preferred embodiment, wherein G ₉ is independently selected from

In a further preferred embodiment, wherein G ₉ is selected from

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein Cat is independent Ground is selected from sodium ions, triethylammonium ions, and tetrabutylammonium ions; r is an integer from 1 to 2, specifically 1, 2; Solid support is a resin solid phase carrier.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein Cat is preferably Triethylammonium ion, sodium ion.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein r is 1.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, R is independent selected from

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, independently selected from

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, Independently selected from:

In a further preferred embodiment, Independently selected from:

In a further preferred embodiment, Independently selected from:

In a further aspect, the present invention provides a compound represented by formula (II), or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, It is a conjugate of a compound with liver-targeted delivery effect and an oligonucleotide, especially a novel GalNAc derivative-oligonucleotide conjugate. Conjugation of GalNAc derivatives can improve or enhance the pharmacokinetic and pharmacodynamic properties of the linked nucleic acid. Specifically, it has the following structure:

where each R is independently selected from:

Among them, Y ₁ , Y ₂ , Y ₃ , Y ₄ , Y ₅ , Y ₆ , Z ₁ , Z ₂ , Z ₃ , Z ₄ , X ₁ , X ₂ , X 3 , X _{4 ,} J ₁ , J ₂ , The definitions of J ₃ , J ₄ , J ₅ , J ₆ , A ₁ and A ₂ are the same as those of the compound of formula (I) above, and the definition of X ₅ ' is only the same as the definition of X ₅ in the compound of formula (I) above. Lack of an H, hydroxyl group, OG ₈ or OG ₉ to connect to E ₄ ;

E ₄ is independently selected from -O-, -S-;

E ₁ is -OH, and E ₂ is independently selected from =O, =S; alternatively, E ₁ is -O ^- , and E ₂ is independently selected from =O, =S; alternatively, E ₁ is =O, and E ₂ is independently selected from -CH ₃ , -O(C _1-6 alkyl), -NH(C _1-6 alkyl), -BH _{3 ,} -SH, -S ^- ; alternatively, E ₁ is =S , and E ₂ is independently selected from -CH ₃ , -SH, -S ^- ;

E ₃ is a functional oligonucleotide molecule, including but not limited to: small interfering RNA, microRNA, immune stimulators, alternative splice bodies, single-stranded RNA, double-stranded RNA, antisense nucleic acids, nucleic acid aptamers, One of stem-loop RNA, mRNA fragment, activating RNA or DNA.

In a preferred embodiment of the invention, the compound provided by the invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, E ₄ Selected from -O-.

In a preferred embodiment of the present invention, the compound provided by the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, E ₁ is -OH, and E ₂ is independently selected from =O, =S; alternatively, E ₁ is -O ^- , and E ₂ is independently selected from =O, =S; alternatively, E ₁ is =O, and E ₂ Independently selected from -CH ₃ , -O(C _1-2 alkyl), -BH _{3 ,} -SH, -S ^- ; alternatively, E ₁ is =S, and E ₂ is independently selected from -SH, -S ^- .

In further preferred embodiments, wherein E ₁ is -OH, and E ₂ is independently selected from =O; alternatively, E ₁ is -O ^- , and E ₂ is independently selected from =O; alternatively, E ₁ is = O, and E ₂ is independently selected from -CH ₃ , -OCH ₃ , -BH _{3 ,} -SH, -S ^- ; alternatively, E ₁ is =S, and E ₂ is independently selected from -SH, -S ^- .

In a further preferred embodiment, wherein E ₁ is -OH and E ₂ is selected from =O.

In a further preferred embodiment, _E3 is small interfering RNA, double-stranded RNA, single-stranded RNA or antisense nucleic acid.

In further preferred embodiments, relevant modifications can also be made to functional oligonucleotides, including but not limited to the following modifications: locked nucleic acid modification, ring-opening or non-locked nucleic acid modification, 2'-methoxyethyl modification, 2′-O-methyl modification, 2′-O-allyl modification, 2′-C-allyl modification, 2′-fluoro modification, 2′-deoxy modification, 2′-hydroxyl modification, thio Phosphate backbone modification, DNA modification, fluorescent probe modification, ligand modification.

In a preferred embodiment of the present invention, its E3 is a functional oligonucleotide, including but not limited to: small interfering RNA (siRNA), microRNA (microRNA), immune stimulatory (immune stimulatory), alternative splicing Splice variant, single-stranded RNA (ssRNA), double-stranded RNA (dsRNA), transfer RNA (tRNA), antisense nucleic acid (antisense), nucleic acid aptamer (Nucleic Acid Aptamer), stem-loop RNA (stem- One of loop RNA), mRNA fragment, small activating RNA (small activating RNA, saRNA) or DNA.

In a preferred embodiment of the present invention, the functional oligonucleotide is preferably small interfering RNA, single-stranded RNA, double-stranded RNA, or antisense nucleic acid.

In a preferred embodiment of the present invention, the 5' or 3' end of the functional oligonucleotide is connected to the GalNAc derivative of the present invention;

In a preferred embodiment of the present invention, each of the oligonucleotides binds 1-5 (specifically: 1, 2, 3, 4, 5) GalNAc derivatives of the present invention in one of the present invention. In preferred embodiments, it is preferred to carry out relevant modifications on functional oligonucleotides (for example: siRNA), including but not limited to: locked nucleic acid modification, ring-opening or non-locked nucleic acid modification, 2'-methoxyethyl modification, 2′-O-methyl modification, 2′-O-allyl modification, 2′-O-methoxyethyl, 2′-C-allyl modification, 2′-fluoro modification, 2′-deoxy Modification, 2′-hydroxyl modification, phosphorothioate backbone modification, DNA modification, fluorescent probe modification, ligand modification;

In a preferred embodiment of the present invention, the present invention also relates to an siRNA molecule that inhibits target gene expression, including a sense strand and an antisense strand that complement each other to form a double-stranded region, and the sense strand and/or the antisense strand Comprising or consisting of 15-25 nucleotides, the antisense strand is connected to at least 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or 25 consecutive nucleotides are completely or partially complementary, the length of the double-stranded region is 15-25bp, and at least one nucleotide in the siRNA molecule is modified;

In a preferred embodiment of the present invention, any strand of the siRNA molecule contains an overhang of 0-5 bases;

The target genes include but are not limited to: ApoB, ApoC, ANGPTL3, PCSK9, FVII, p53, HAV, HBV, HCV, HDV, HEV, AGT, Lp(a), XDH, HSD17B13, SCD1, PNPLA3, HMGCR, etc.

In a preferred embodiment of the present invention, the oligonucleotide in formula (II) that can be conjugated with the novel GalNAc derivative at the 5' or 3' end can be selected from the following sequences:

Preferably, the present invention provides a compound of formula (I) or formula (II) or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, wherein, The compound has the following structure:

The object of the present invention also includes providing the preparation of compounds represented by general formula (I), general formula (II) or their prodrugs, tautomers, stereoisomers, solvates, isotope derivatives or pharmaceutically acceptable compounds thereof. Salt method.

The compounds of general formula (I) and general formula (II) of the present invention can be prepared using the methods shown in the following schemes:

Another aspect of the present invention also provides a pharmaceutical composition, which contains the compound of the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable of salt.

Furthermore, the pharmaceutical composition of the present invention also contains pharmaceutically acceptable excipients.

The compound of the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or its pharmaceutical Administration of the above acceptable salts may be carried out in pure form or in the form of a suitable pharmaceutical composition by any acceptable mode of administration for drugs providing similar uses. The pharmaceutical composition of the present invention can be prepared by combining the compound of the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof with a suitable pharmaceutically acceptable salt. Prepared by combining auxiliary materials. The pharmaceutical composition of the present invention can be formulated into solid, semi-solid, liquid or gaseous preparations. Generally, the above-mentioned pharmaceutical compositions can be prepared by conventional preparation methods using conventional excipients in the field of preparation.

In another aspect, the present invention also provides the compound of the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof or the pharmaceutical composition of the present invention. Use in the preparation of medicines.

Further, the present invention also provides the compound of the present invention or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof or the pharmaceutical composition of the present invention. Use in the preparation of a medicament for the prevention and/or treatment of physiological conditions or diseases caused by the expression of specific genes in liver cells.

Further, the specific genes include but are not limited to: ApoB, ApoC, ANGPTL3, PCSK9, FVII, p53, HAV, HBV, HCV, HDV, HEV, AGT, Lp(a), XDH, HSD17B13, SCD1, PNPLA3, HMGCR wait.

Further, the invention provides uses, wherein the diseases are chronic liver diseases, hepatitis, liver fibrosis diseases, liver proliferative diseases and dyslipidemia.

In some embodiments, the dyslipidemia is hypercholesterolemia, hypertriglyceridemia, or atherosclerosis.

In some embodiments, the disease also includes other liver diseases, including malignant diseases characterized by cell proliferation, hematological diseases, diseases characterized by inflammation, or metabolic diseases. Among them, malignant diseases characterized by liver cell proliferation can be benign or malignant tumors, such as cancer, liver metastasis or hepatoblastoma. Hematologic disorders or disorders characterized by inflammation may be disorders involving coagulation factors, complement-mediated inflammation, or fibrosis. Metabolic diseases include dyslipidemia and disorders of glucose regulation.

In some embodiments, liver disease can also be treated by administering one or more oligonucleotides that have high sequence homology to genes involved in liver disease.

In yet another aspect, the invention provides a method for preventing and/or treating physiological conditions or diseases caused by the expression of specific genes in hepatocytes, comprising administering to an individual in need thereof a compound of the invention or a prodrug thereof. , tautomers, stereoisomers, solvates, isotope derivatives or pharmaceutically acceptable salts thereof or the pharmaceutical composition of the present invention; the diseases are chronic liver disease, hepatitis, liver fibrosis disease, and liver hyperplasia diseases and dyslipidemia.

In some embodiments, the dyslipidemia is hypercholesterolemia, hypertriglyceridemia, or atherosclerosis.

In some embodiments, the disease also includes other liver diseases, including malignant diseases characterized by cell proliferation, hematological diseases, diseases characterized by inflammation, or metabolic diseases. Among them, malignant diseases characterized by liver cell proliferation can be benign or malignant tumors, such as cancer, liver metastasis or hepatoblastoma. Hematologic disorders or disorders characterized by inflammation may be disorders involving coagulation factors, complement-mediated inflammation, or fibrosis. Metabolic diseases include dyslipidemia and disorders of glucose regulation.

In yet another aspect, the invention provides compounds of the invention or prodrugs, tautomers, stereoisomers thereof for preventing and/or treating physiological conditions or diseases caused by the expression of specific genes in hepatocytes. isotopes, solvates, isotope derivatives or pharmaceutically acceptable salts thereof or the pharmaceutical composition of the present invention;

The diseases described are chronic liver diseases, hepatitis, liver fibrosis diseases, liver proliferative diseases and dyslipidemia.

In some embodiments, the dyslipidemia is hypercholesterolemia, hypertriglyceridemia, or atherosclerosis.

In some embodiments, the disease also includes other liver diseases, including malignant diseases characterized by cell proliferation, hematological diseases, diseases characterized by inflammation, or metabolic diseases. Among them, malignant diseases characterized by liver cell proliferation can be benign or malignant tumors, such as cancer, liver metastasis or hepatoblastoma. Hematologic disorders or disorders characterized by inflammation may be disorders involving coagulation factors, complement-mediated inflammation, or fibrosis. Metabolic diseases include dyslipidemia and disorders of glucose regulation.

definition

It is the covalent attachment site of the group.

The terms "optionally," "any," "optionally" or "arbitrarily" mean that the subsequently described event or condition may, but need not, occur, and that the description includes circumstances in which the stated event or condition occurs and The event or condition described does not occur.

The term "oxo" means that two hydrogen atoms in the same substitution position are replaced by the same oxygen atom to form a double bond.

Unless otherwise specified, the term "alkyl" refers to a monovalent saturated aliphatic hydrocarbon group, a straight or branched chain group containing 1 to 20 carbon atoms, preferably 1 to 10 carbon atoms (i.e., C _{1 to 10} Alkyl), further preferably contains 1-8 carbon atoms (C _1-8 alkyl), more preferably contains 1-6 carbon atoms (i.e. C _1-6 alkyl), such as "C _1-6 alkyl" It means that the group is an alkyl group, and the number of carbon atoms on the carbon chain is between 1 and 6 (specifically 1, 2, 3, 4, 5 or 6). Examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, neopentyl, 1,1-dimethyl Propyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, n-heptyl , n-octyl, etc.

Unless otherwise specified, the term "alkenyl" refers to a straight-chain or branched unsaturated aliphatic hydrocarbon group composed of carbon atoms and hydrogen atoms and having at least one double bond. The alkenyl group may contain 2-20 carbon atoms, preferably 2-10 carbon atoms (i.e., C _2-10 alkenyl), further preferably 2-8 carbon atoms (C _2-8 alkenyl), and more preferably 2-8 carbon atoms (C 2-8 alkenyl). 2-6 carbon atoms (i.e. C _2-6 alkenyl), 2-5 carbon atoms (i.e. C _2-5 alkenyl), 2-4 carbon atoms (i.e. C _2-4 alkenyl), 2- 3 carbon atoms (i.e. C _2-3 alkenyl), 2 carbon atoms (i.e. C ₂ alkenyl), for example "C _2-6 alkenyl" means that the group is alkenyl, and the carbon chain The number of carbon atoms is between 2 and 6 (specifically 2, 3, 4, 5 or 6). Non-limiting examples of alkenyl groups include, but are not limited to, vinyl, 1-propenyl, 2-propenyl, 1-butenyl, isobutenyl, 1,3-butadienyl, and the like.

Unless otherwise specified, the term "alkynyl" refers to a straight-chain or branched unsaturated aliphatic hydrocarbon group composed of carbon atoms and hydrogen atoms and having at least one triple bond. The alkynyl group may contain 2-20 carbon atoms, preferably 2-10 carbon atoms (i.e., C _2-10 alkynyl group), further preferably 2-8 carbon atoms (C _2-8 alkynyl group), and more preferably 2-8 carbon atoms (C 2-8 alkynyl group). 2-6 carbon atoms (i.e. C _2-6 alkynyl), 2-5 carbon atoms (i.e. C _2-5 alkynyl), 2-4 carbon atoms (i.e. C _2-4 alkynyl), 2- 3 carbon atoms (i.e. C _2-3 alkynyl), 2 carbon atoms (i.e. C ₂ alkynyl), for example "C _2-6 alkynyl" means that the group is an alkynyl group, and the carbon chain The number of carbon atoms is between 2 and 6 (specifically 2, 3, 4, 5 or 6). Non-limiting examples of alkynyl groups include, but are not limited to, ethynyl, 1-propynyl, 2-propynyl, 1-butynyl, and the like.

Unless otherwise specified, the term "alkoxy" refers to -O-alkyl, which is as defined above, that is, containing 1-20 carbon atoms, preferably 1-10 carbon atoms, preferably 1-8 carbon atoms, more preferably 1-6 carbon atoms (specifically 1, 2, 3, 4, 5 or 6). Representative examples include, but are not limited to, methoxy, ethoxy, propoxy, isopropoxy, butoxy, 1-methylpropoxy, 2-methylpropoxy, tert-butoxy, pentoxy Oxygen, 1-methylbutoxy, 2-methylbutoxy, 3-methylbutoxy, 1,1-dimethylpropoxy, 1,2-dimethylpropoxy, 2 , 2-dimethylpropoxy, 1-ethylpropoxy, etc.

Unless otherwise specified, the term "halogen" or "halogenated" refers to F, Cl, Br, I. The term "haloalkyl" means an alkyl group as defined above in which one, two or more hydrogen atoms or all of the hydrogen atoms are replaced by halogen. Representative examples of haloalkyl groups include CCl ₃ , CF ₃ , CHCl ₂ , CH ₂ Cl, CH ₂ Br, CH ₂ I, CH ₂ F, CH ₂ CF ₃ , CF ₂ CF ₃ , and the like.

Unless otherwise specified, the term "cycloalkyl" refers to a monocyclic saturated aliphatic hydrocarbon group with a specific number of carbon atoms, preferably containing 3-12 carbon atoms (i.e., C _3-12 cycloalkyl), more preferably containing 3-10 carbon atoms (C _3-10 cycloalkyl), more preferably 3-7 carbon atoms (C _3-7 cycloalkyl), 4-6 carbon atoms (C _4-6 cycloalkyl), 5-6 carbon atoms (C _5-6 cycloalkyl). Examples include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, methylcyclopropyl, 2-ethyl-cyclopentyl, dimethylcyclobutyl, and the like.

Unless otherwise specified, "cycloalkenyl" means composed of the subgroups monocyclic hydrocarbon ring, bicyclic hydrocarbon ring and spiro-hydrocarbon ring, however, the system is unsaturated, that is, there is at least one CC double bond but no aromatic system. Preferably it contains 3-12 carbon atoms (i.e. C _3-12 cycloalkenyl), more preferably 3-10 carbon atoms (C _3-10 cycloalkenyl), further preferably 3-6 carbon atoms (C _{3 -6} cycloalkenyl), 4-6 carbon atoms (C _4-6 cycloalkenyl), 5-6 carbon atoms (C _5-6 cycloalkenyl).

Unless otherwise specified, the term "heterocyclyl" or "heterocycle" refers to a saturated or partially unsaturated monocyclic or polycyclic cyclic non-aromatic substituent having ring carbon atoms and 1 to 4 ring heteroatoms, including 3-20 ring atoms, of which 1, 2, 3 or more ring atoms are selected from N, O or S, and the remaining ring atoms are C. Preferably contains 3 to 12 ring atoms (3-12 membered heterocyclyl), further One step preferably contains 3-10 ring atoms (3-10-membered heterocyclyl), or 3-8 ring atoms (3-8-membered heterocyclyl), or 3-6 ring atoms (3-6-membered heterocyclyl) base), or 4 to 6 ring atoms (4-6 membered heterocyclyl group), or 5 to 6 ring atoms (5-6 membered heterocyclyl group). The number of heteroatoms is preferably 1 to 4, more preferably 1 to 3 (ie, 1, 2 or 3). Examples of monocyclic heterocyclyl groups include pyrrolidinyl, imidazolidinyl, tetrahydrofuryl, dihydropyrrolyl, piperidinyl, piperazinyl, pyranyl, and the like. Polycyclic heterocyclyl groups include spirocyclic, fused cyclic and bridged cyclic heterocyclyl groups. "Heterocyclyl" may be a monocyclic ("monocyclic heterocyclyl") or a fused ("fused heterocyclyl" or "heterofused cyclyl"), bridged ("heterobridged cyclyl" or "bridged heterocyclyl") or spiro-fused ("heterospiryl" or "spiroheterocyclyl") ring system, such as a bicyclic system ("bicyclic heterocyclyl"), and can Is saturated or may be partially unsaturated. Heterocyclyl bicyclic systems may include one or more heteroatoms in one or both rings. "Heterocyclyl" also includes ring systems in which the heterocyclyl ring as defined above is fused with one or more carbocyclyl groups, wherein the point of attachment is on the carbocyclyl or heterocyclyl ring, or "Heterocyclyl" also includes ring systems in which the heterocyclyl ring, as defined above, is fused with one or more aryl or heteroaryl groups, or a cycloalkyl ring, as defined above, is fused with one or more aryl or heteroaryl groups. A fused ring system of a heteroaryl group, wherein the point of attachment is on the heterocyclyl ring or cycloalkyl ring, and in such cases the membership of the heterocyclyl ring system is the post-fused ring system Number of atoms. In certain embodiments, each instance of heterocyclyl is independently optionally substituted, e.g., unsubstituted (an "unsubstituted heterocyclyl") or substituted with one or more substituents (a "substituted heterocyclyl"). Exemplary 3-membered heterocyclyl groups containing 1 heteroatom include, but are not limited to, aziridinyl, oxiranyl, and thiorenyl. Exemplary 4-membered heterocyclyl groups containing 1 heteroatom include, but are not limited to, azetidinyl, oxetanyl, and thietanyl. Exemplary 5-membered heterocyclyl groups containing 1 heteroatom include, but are not limited to, tetrahydrofuryl, dihydrofuryl, tetrahydrophenylthio, dihydrophenylthio, pyrrolidinyl, dihydropyrrolyl, and pyrrole. Base-2,5-dione. Exemplary 5-membered heterocyclyl groups containing 2 heteroatoms include, but are not limited to, dioxolanyl, oxathiolanyl, dithiolyl, and oxazolidin-2-one . Exemplary 5-membered heterocyclyl groups containing 3 heteroatoms include, but are not limited to, triazolinyl, oxadiazolinyl, and thiadiazolinyl. Exemplary 6-membered heterocyclyl groups containing 1 heteroatom include, but are not limited to, piperidinyl, tetrahydropyranyl, dihydropyridyl, and thianyl. Exemplary 6-membered heterocyclyl groups containing 2 heteroatoms include, but are not limited to, piperazinyl, morpholinyl, dithianyl, and dioxanyl. Exemplary 6-membered heterocyclyl groups containing 3 heteroatoms include, but are not limited to, triazacyclohexyl, oxadiazinyl, thiadiazinyl, oxothiazinyl, and dioxo Dioxazinanyl. Exemplary 7-membered heterocyclyl groups containing 1 heteroatom include, but are not limited to, azepanyl, oxpanyl, and thiepanyl. Exemplary 8-membered heterocyclyl groups containing 1 heteroatom include, but are not limited to, azacyclooctyl, oxacyclooctanyl, and thiocyclyl. Exemplary 5-membered heterocyclyl groups fused to a _C6 aryl ring (also referred to herein as a 5,6-bicyclic heterocycle) include, but are not limited to, indolyl, isodihydro Indolyl, dihydrobenzofuranyl, dihydrobenzothienyl, benzoxazolinonyl, etc. Exemplary 6-membered heterocyclyl groups fused to an aryl ring (also referred to herein as a 6,6-bicyclic heterocycle) include, but are not limited to, tetrahydroquinolyl, tetrahydroisoquinoline Key et al.

Unless otherwise specified, "heterocycloalkyl" refers to a monocyclic, saturated "heterocyclyl" or "heterocycle" as defined above. The ring atoms are as defined above, that is, containing 3 to 20 ring atoms ("3 -20-membered heterocycloalkyl"), the number of heteroatoms is 1 to 4 (1, 2, 3 or 4), preferably 1 to 3 (1, 2 or 3), wherein heteroatoms The atoms are each independently selected from N, O, or S. It preferably contains 3 to 12 ring atoms ("3-12 membered heterocycloalkyl"), more preferably contains 3 to 10 ring atoms ("3-10 membered heterocycloalkyl"), and even more preferably contains 3 to 8 ring atoms. ring atoms ("3-8-membered heterocycloalkyl"), more preferably 4-7 ring atoms ("4-7-membered heterocycloalkyl"), still more preferably 5-10 ring atoms ("5-10 membered heterocycloalkyl"), further preferably contains 5-6 ring atoms ("5-6 membered heterocycloalkyl"). In certain embodiments, each instance of heterocycloalkyl is independently optionally substituted, e.g., unsubstituted (an "unsubstituted heterocycloalkyl") or substituted with one or more substituents Substituted (a "substituted heterocycloalkyl"). The above "heterocyclyl" or "heterocycle" section has given some exemplary "heterocycloalkyl", which also includes, but is not limited to, aziridinyl, oxirinyl, thiirane base, azetidinyl, oxetanyl, thietanyl, tetrahydrofuranyl, oxetanyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl , oxathiacyclohexyl, oxazolidinyl, dioxanyl, dithiocyclohexyl, thiazolidinyl, pyrrolidinyl, pyrazolidinyl, imidazolidinyl, etc.

Unless otherwise specified, the term "aryl" or "aryl" means an aryl group containing 6 to 16 carbon atoms, or 6 to 14 carbon atoms, or 6 to 12 carbon atoms. carbon atoms, or monocyclic, bicyclic and tricyclic aromatic carbocyclic ring systems of 6 to 10 carbon atoms, preferably 6 to 10 carbon atoms. The term "aryl" can be used interchangeably with the term "aromatic ring". Examples of aryl groups may include, but are not limited to, phenyl, naphthyl, anthracenyl, phenanthrenyl, or pyrenyl, and the like.

Unless otherwise specified, the term "heteroaryl" or "heteroaryl ring" means a 5-14-membered structure, or preferably a 5-10-membered structure, or preferably a 5-8-membered structure, more preferably a 5-6-membered structure. An aromatic monocyclic or polycyclic ring system in which 1, 2, 3 or more ring atoms are heteroatoms and the remaining atoms are carbon, the heteroatoms are independently selected from O, N or S, the number of heteroatoms Preferably 1, 2 or 3. Examples of heteroaryl groups include, but are not limited to, furyl, thienyl, oxazolyl, thiazolyl, isoxazolyl, oxadiazolyl, thiadiazolyl, pyrrolyl, pyrazolyl, imidazolyl, triazolyl , tetrazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, thiodiazolyl, triazine, phthalazinyl, quinolyl, isoquinolinyl, pyridinyl, purinyl, indyl Indolyl, isoindolyl, indazolyl, benzofuranyl, benzothienyl, benzopyridinyl, benzopyrimidinyl, benzopyrazinyl, benzimidazolyl, benzophthalazinyl, pyrrole Para[2,3-b]pyridyl, imidazo[1,2-a]pyridyl, pyrazolo[1,5-a]pyridyl, pyrazolo[1,5-a]pyrimidinyl, imidazole [1,2-b]pyridazinyl, [1,2,4]triazolo[4,3-b]pyridazinyl, [1,2,4]triazolo[1,5-a] Pyrimidinyl, [1,2,4]triazolo[1,5-a]pyridyl, etc.

The term "Solid support" or "solid support" refers to inorganic particles, polymers or other solid materials as a matrix, with active groups that can be linked to target compounds through surface modification, thereby being used for the synthesis of oligonucleotides.

Unless otherwise specified, the term "pharmaceutically acceptable salt" or "pharmaceutically acceptable salt" means salts that are suitable within the scope of reasonable medical judgment for contact with mammalian, especially human, tissue without undue toxicity, irritation, or allergic reaction. Medically acceptable salts of amines, carboxylic acids, and other types of compounds are well known in the art and are commensurate with a reasonable benefit/risk ratio. The salts may be prepared in situ during the final isolation and purification of the compounds of the invention, or separately by reacting the free base or free acid with a suitable reagent.

Unless otherwise specified, the term "isotopic derivative" means that the compounds of the invention may exist in isotopically traced or enriched form, containing one or more atoms having an atomic weight or mass number different from that found in nature The atomic weight or mass number of the largest number of atoms. Isotopes may be radioactive or non-radioactive isotopes. The isotopes commonly used as isotope labels are: hydrogen isotopes, ² H and ³ H; carbon isotopes: ¹³ C and ¹⁴ C; chlorine isotopes: ³⁵ Cl and ³⁷ Cl; fluorine isotopes: ¹⁸ F; iodine isotopes: ¹²³ I and ¹²⁵ I ; Nitrogen isotopes: ¹³ N and ¹⁵ N; Oxygen isotopes: ¹⁵ O, ¹⁷ O and ¹⁸ O and sulfur isotope ³⁵ S. These isotopically labeled compounds can be used to study the distribution of pharmaceutical molecules in tissues. Especially ² H and ¹³ C are more widely used because they are easy to label and detect. Substitution of certain heavy isotopes, such as deuterium ( ^2H ), can enhance metabolic stability, extend half-life, thereby reducing dosage and providing therapeutic advantages. Isotopically labeled compounds generally start from labeled starting materials and are synthesized using known synthetic techniques as for non-isotopically labeled compounds.

Unless otherwise specified, the terms "solvate" and "solvate" mean the physical association of a compound of the invention with one or more solvent molecules, whether organic or inorganic. This physical association includes hydrogen bonding. In certain circumstances, such as when one or more solvent molecules are incorporated into the crystal lattice of a crystalline solid, solvates will be able to be separated. The solvent molecules in a solvate may exist in regular and/or disordered arrangements. Solvates may contain stoichiometric or non-stoichiometric amounts of solvent molecules. "Solvate" encompasses both solution phase and isolable solvates. Exemplary solvates include, but are not limited to, hydrates, ethanolates, methoxides, and isopropoxides. Solvation methods are well known in the art.

Unless otherwise specified, the term "stereoisomer" refers to compounds that have the same chemical structure but different arrangements of atoms or groups in space. Stereoisomers include enantiomers, diastereomers, conformational isomers (rotamers), geometric isomers (cis/trans) isomers, atropisomers, etc. Any resulting mixture of stereoisomers may be separated into pure or substantially pure geometric isomers, enantiomers, and diastereomers based on differences in the physicochemical properties of the components, for example, by chromatography. method and/or fractional crystallization method.

Unless otherwise specified, the term "tautomers" refers to structural isomers with different energies that are interconvertible through a low energy barrier. If tautomerism is possible (eg in solution), a chemical equilibrium of tautomers can be achieved. For example, proton tautomers (also known as proton transfer tautomers) include interconversions by proton migration, such as keto-enol isomerization and imine-enamine isomerization. Valence tautomers involve interconversions through the reorganization of some of the bonding electrons.

Unless otherwise indicated, the structural formulas described in the present invention include all isomeric forms (such as enantiomers, diastereomers, and geometric isomers (or conformational isomers)): for example, those containing asymmetric centers R, S configuration, double bond (Z), (E) isomers, and (Z), (E) conformational isomers. Therefore, individual stereochemical isomers or mixtures of enantiomers, diastereomers, or geometric isomers (or conformational isomers) of the compounds of the present invention are within the scope of the present invention.

Unless otherwise specified, the term "prodrug" refers to a drug that is converted in the body to the parent drug. Prodrugs are often useful in that they improve some defined, undesirable physical or biological property. Physical properties are often related to solubility (too high or insufficient lipid or water solubility) or stability, while problematic biological properties include too rapid metabolism or poor bioavailability, which may themselves be related to physicochemical properties. For example, they are bioavailable via oral administration, whereas the parent body is not. Prodrugs also have increased solubility in pharmaceutical compositions compared to the parent drug. An example of a prodrug, but not limited thereto, may be any compound of the invention administered as an ester ("prodrug") to facilitate delivery across cell membranes, where water solubility is detrimental to mobility, but once inside Intracellular water solubility is beneficial and is subsequently metabolically hydrolyzed to carboxylic acids, the active entities. Another example of a prodrug may be a short peptide (polyamino acid) bound to an acid group, where the peptide is metabolized to reveal the active moiety.

Functional oligonucleotides described in the present invention refer to oligonucleotides that can utilize RNA activation (RNA activation) by generating stable and specific hybridization with target sequences. Principles such as RNAa), RNA interference (RNAi), antisense nucleic acid technology, and exon skipping technology can up-regulate or down-regulate the expression of target genes, or lead to alternative splicing of mRNA. In some aspects, functional oligonucleotides can also be nucleic acid structures that produce stable and specific binding to target proteins. In addition, those skilled in the art will easily understand that polynucleotides (such as mRNA itself or fragments thereof) are also suitable for conjugation with the conjugation molecules provided by the present disclosure to form conjugates to achieve targeted delivery, such as liver targeting. delivery, thereby regulating the expression of proteins transcribed from the mRNA.

"Target sequence" refers to the target mRNA. In the context of the present invention, "target mRNA" refers to the mRNA corresponding to a gene that is abnormally expressed in liver cells. It can be either the mRNA corresponding to an overexpressed gene or the mRNA corresponding to an underexpressed gene. In some embodiments of the present disclosure, corresponding to the above-mentioned abnormally expressed genes, the target mRNA can be ApoB, ApoC, ANGPTL3, PCSK9, FVII, p53, HAV, HBV, HCV, HDV, HEV, AGT, Lp(a ), XDH, HSD17B13, SCD1, PNPLA3, HMGCR and other genes corresponding mRNA. In some embodiments, the target mRNA can be the mRNA transcribed from the corresponding PCSK9 gene, or the mRNA corresponding to the ANGPTL3 gene, or the mRNA corresponding to the XDH gene, or the mRNA corresponding to the APOC3 gene.

The complete complementarity in the present invention means that the bases of nucleotides follow the classic Watson-Crick base pairing principle (for example: A-U, G-C, A-T); the partial complementarity means that some bases are not Satisfy the Watson-Crick base pairing principle, for example: there is non-Watson-Crick base pairing mode (for example: G-U, A-A).

The "conjugate" or "conjugated molecule" used in the present invention refers to a compound formed by covalent linkage between various chemical moieties.

The abbreviations used in the examples and elsewhere herein are:
CPG glass-based solid carrier
CPG- _NH2 glass solid phase carrier with amino-modified surface
CBz benzyloxycarbonyl
DMTr dimethoxytrityl
DIPEA N,N'-Diisopropylethylamine
DCM dichloromethane
DMF N,N-dimethylformamide
EDCI.HCl 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride
HATU 2-(7-benzotriazole oxide)-N,N,N',N'-tetramethylurea hexafluorophosphate
MeOH methanol
Pd/C palladium carbon
HOBT 1-Hydroxybenzotriazole

DMAP 4-dimethylaminopyridine

TBTU 2-(1H-benzotrisazoL-1-yl)-1,1,3,3-tetramethylurea tetrafluoroborate

Fmoc 9-fluorenylmethoxycarbonyl

The beneficial effects of the present invention are:

The present invention designs a class of compounds with liver-targeted delivery effects and oligonucleotide conjugates thereof. This compound with liver-targeted delivery effects is a new type of GalNAc derivative, which is used for the treatment of diseases occurring in the liver or related to the liver. Liver-related diseases provide a new direction. Tests have shown that compared with the existing technology (ligand L96), the oligonucleotide conjugate of the novel GalNAc derivative molecule of the present invention has stronger binding ability to ASGPR, is more easily absorbed by liver cells, and inhibits target mRNA higher activity. In addition, the present invention studies a specific synthesis method, which has simple process, convenient operation, and is conducive to large-scale industrial production and application.

Description of the drawings

Figure 1: In vivo imaging of mice administered subcutaneous injection of PC01 and PC02 at 2h and 4h (Figure 1-1 is PC012h; Figure 1-2 is PC022h; Figure 1-3 is PC014h; Figure 1-4 is PC024h);

Figure 2: Average fluorescence density of PC01 and PC02 in Huh7 cells;

Figure 3: Average fluorescence density of PC01 and PC02 in mouse liver primary cells;

Figure 4: The number of positive cells of PC01 and PC02 in mouse liver primary cells; where AF647 refers to the name of the detection channel of the flow cytometer;

Figure 5: Effect diagram of in vitro uptake efficiency of GalNAc ligand-coupled siRNA;

Figure 6: The results of using human primary hepatocytes to evaluate the in vitro inhibition of PCSK9 mRNA activity by siRNA of the test compound;

Figure 7: In vivo drug efficacy experimental results in PC15, PC17, and PC18 mice;

Figure 8: In vivo drug efficacy experimental results in PC15 and PC19 mice.

Detailed ways

The present invention will be further described below in conjunction with specific embodiments. It should be understood that these examples are only used to illustrate the invention and are not intended to limit the scope of the invention. Experimental methods without specifying specific conditions in the following examples usually follow conventional conditions or conditions recommended by the manufacturer. Unless otherwise defined, all professional and scientific terms used herein have the same meaning as commonly understood by professionals in the art. In addition, any methods and materials similar or equivalent to those described can be applied to the method of the present invention. The preferred implementation methods and materials shown in this article are for demonstration purposes only.

The following are preparation examples of exemplary compounds of the present invention.

Example 1:

Among them, the synthetic route of compound 6 is:

Synthesis of compound 2:

After adding 1.46g of compound 1, 15mL of ethanol and 146.0mg of Pd/C into the three-necked flask, the mixture was stirred at room temperature under H ₂ environment overnight. After the reaction, the filtrate was collected by filtration and concentrated to obtain crude product 2, which was directly used in the next reaction.

Synthesis of compound 4:

Add 209.4 mg of compound 3 and 15 mL of DMF into the three-neck flask. After stirring and dissolving, add 575.1 mg EDCI.HCl and 405.5 mg HOBT, and stir at room temperature for 30 minutes under nitrogen protection. Then add the crude compound 2 obtained in the previous step, 5 mL dry DMF and 0.75 mL DIPEA, and stir at room temperature overnight. After the reaction, 100 mL of DCM was added, washed with saturated ammonium chloride solution (2x50 mL), water (2x50 mL) and saturated brine (1x100 mL). The organic phase was dried over anhydrous sodium sulfate and filtered. The collected organic phase was concentrated under vacuum and purified by silica gel column chromatography (DCM/MeOH 20:1) to obtain 272.3 mg of compound 4 in a yield of 32%. ESI-MS: m/z 1703.6[M+H] ⁺ .

Synthesis of compound 5:

After adding 272.3 mg of compound 4, 0.5 mL of tetrahydrofuran and 30 mg of Pd/C into the three-neck flask, the mixture was stirred at room temperature overnight in a H ₂ environment. After the reaction, the filtrate was collected by filtration and concentrated to obtain crude product 5, which was directly used in the next reaction.

Synthesis of compound 6:

①Synthesis of compound 6-b:

Compound 6-a (10g, 283.00mmol, 1eq.) was dissolved in anhydrous tetrahydrofuran (1.5L), and cooled in an ice bath (5°C). Borane dimethyl sulfide (991.00 mmol 3eq.) was added for 20 minutes. After the addition of borane dimethyl sulfide was completed, the reaction mixture was heated to 60°C overnight, during which time a white precipitate formed. Then methanol (150 ml) was slowly added to the reaction mixture to quench excess borane dimethyl sulfide, and then co-evaporated with methanol three times (100 ml each time) to remove the borate. The product could be put into the next step without further purification. reaction, the yield was 81%. ESI-MS: m/z 340.1[M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ ) δ: 7.89 (d, J = 7.4Hz, 2H), 7.65 (d, J = 7.3Hz, 2H), 7.55–7.13 (m, 4H), 5.07–4.95 ( m,1H),4.95–4.87(m,1H),4.80–4.70(m,1H),4.70–4.63(m,1H),4.36–4.2(m,4H),3.92–3.80(m,1H), 3.80–3.68(m,1H),3.57–3.46(m,2H),3.35–3.02(m,2H),2.07–1.91(m,1H),1.91–1.74(m,1H).

②Synthesis of compound 6-c:

Dissolve the crude product 6-b (7.64g, 225.00mmol, 1eq.) in anhydrous pyridine (1.0L), and add 4,4'-dimethoxytriacyl chloride (DMTrCl, 247mmol, 1.1eq.) in small amounts in batches. ), adding each batch for 10 minutes at room temperature. The reaction mixture was stirred at room temperature for 4 h, concentrated under reduced pressure to half the volume, and diluted with ethyl acetate (500 mL). The organic phase was separated and washed with saturated _aqueous NaHCO solution (2 × 500 mL) and saturated brine (500 mL). Separate the organic layer, dry with Na ₂ SO ₄ , filter, and evaporate to dryness. After alkalizing the column with a very small amount of pyridine, purify the crude product through column chromatography using EA:PE=30% eluent. The crude product is directly applied to the column. May cause product decomposition, yield 63%.

¹ H NMR (600MHz, DMSO-d ₆ ) δ: 7.89 (dd, J = 7.8, 4.0 Hz, 2H), 7.65 (t, J = 6.8 Hz, 2H), 7.42 (t, J = 7.5 Hz, 2H) ,7.37–7.31(m,2H),4.95(d,J＝3.9Hz,1H),4.69(t,J＝5.6Hz,1H),4.28(dddd,J＝31.9,21.0,11.6,6.3Hz,4H ),3.82(d,J＝10.8Hz,1H),3.48(h,J＝5.4Hz,1H),3.41–3.21(m,5H),1.99(dp,J＝16.3,5.7Hz,1H),1.83 (ddd,J=12.9,8.0,4.1Hz,1H).

③Synthesis of compound 6-d:

The product 6-c (8.79g, 137mmol, 1eq.) obtained in the above step was dissolved in anhydrous DMF (500mL), and piperidine (548mmol 4eq.) was added. The reaction mixture was stirred at room temperature for 3 h. The reaction mixture was diluted with chloroform (500 mL) and the organic phase was washed with saturated _aqueous NaHCO (2 x 500 mL) and brine (500 mL). The organic layer was separated, dried over Na ₂ SO ₄ , filtered, and evaporated under reduced pressure to obtain an oil, which was purified by column chromatography (methanol: dichloromethane 5% elution column must be alkalized first), and the yield was 71%. .

¹ H NMR (600MHz, DMSO-d ₆ ) δ: 7.39 (dd, J＝8.4, 1.3Hz, 2H), 7.35–7.17 (m, 7H), 6.91–6.80 (m, 4H), 4.55 (s, 1H) ),4.11(tt,J＝5.0,2.3Hz,1H),3.73(s,6H),3.42–3.39(m,1H),2.90(dd,J＝8.7,6.2Hz,1H),2.85(dd, J＝11.1,4.8Hz,1H),2.75(dd,J＝8.7,5.8Hz,1H),2.61(ddd,J＝11.1,2.8,1.1Hz,1H),1.72–1.63(m,1H),1.39 (ddd,J=12.9,8.4,5.9Hz,1H).

④Synthesis of compound 6-e:

Monomethyl sebacate (39.3 mmol, 1.1 eq.) was dissolved in DMF (200 mL). Add HBTU (42.80mmol 1.2eq.) and N,N-diisopropylethylamine (107.10mmol 3eq.), and stir at room temperature for 5 minutes. A solution of the product in step 4 above (35.7mmol 1eq.) dissolved in anhydrous DMF (50mL) was added, and the reaction mixture was stirred at room temperature. After 18 hours, the reaction mixture was Pour into ice water and extract with dichloromethane. The organic _layer was washed with saturated aqueous sodium bicarbonate solution, dried with brine ( _Na2SO4 ) and filtered. After alkalizing the column, use EA:PE=50% eluent to pass through the column, and the yield is 83%.

¹ H NMR (600MHz, Chloroform-d) δ: 7.39–7.33 (m, 1H), 7.32–7.22 (m, 3H), 7.25–7.15 (m, 1H), 6.81 (ddd, J＝16.1, 9.0, 2.6 Hz,2H),4.12(p,J＝7.1Hz,1H),3.77(d,J＝6.5Hz,4H),3.66(d,J＝1.3Hz,2H),3.49–3.37(m,1H), 3.16–3.06(m,1H),2.65(s,1H),2.33–2.06(m,3H),2.04(s,1H),1.67–1.56(m,1H),1.36–1.16(m,10H), 0.89(s,1H).

⑤Synthesis of compound 6:

Compound 6-e (3.12 g, 4.94 mmol, 1 eq.) was dissolved in a mixture of methanol (15 mL), THF (9.7 mL) and water (9.7 mL). Add LiOH (34.76mmol 7eq.), stir at room temperature for 1 hour, and remove the solvent under reduced pressure. Dichloromethane (10 ml) was added to the reaction to obtain a slurry. After alkalinizing the column, the slurry was loaded onto the column and eluted with 5% dichloromethane-methanol. The yield was 85%.

¹ H NMR (400MHz, DMSO-d ₆ )δ:7.35-7.25(m,4H),7.22-7.17(m,5H),6.89–6.68(m,4H),4.43–4.33(m,1H),4.13 (dd,J＝9.1,4.8Hz,1H),3.73(s,6H),3.56–3.16(m,4H),3.01–2.95(m,1H),2.20(t,J＝7.5Hz,1H), 2.04-1.80(m,5H),1.43(dt,J＝25.5,7.0Hz,4H),1.23(dd,J＝8.6,4.1Hz,12H).

Synthesis of compound 7:

Add 151.6 mg of compound 6 and 0.2 mL of DMF to the three-neck flask. After stirring and dissolving, add 46.0 mg EDCI.HCl and 32.4 mg HOBT, and stir at room temperature for 30 minutes under nitrogen protection. Then add the crude compound 5 obtained in the previous step, 0.3 mL dry DMF and 105.1 μL DIPEA, and stir at room temperature overnight. After the reaction, add 10 mL of DCM, wash with saturated ammonium chloride solution (2x10 mL), water (2x10 mL) and saturated brine (1x10 mL). The organic phase is dried over anhydrous sodium sulfate and filtered. The collected organic phase was concentrated under vacuum and purified by silica gel column chromatography to obtain 109.6 mg of compound 7, with a yield of 32%. ESI-MS: m/z 1089.7[M-2H] ^2- .

Synthesis of compound 8:

Add 109.6 mg of compound 7 and 0.5 mL of DMF to the three-neck flask. After stirring to dissolve, add 10.1 mg succinic anhydride, 38.2 μL triethylamine and 0.6 mg DMAP, and stir at room temperature overnight under nitrogen protection. After the reaction, the collected organic phase was concentrated under vacuum and purified by silica gel column chromatography to obtain 83.6 mg of compound 8, with a yield of 82%. ESI-MS: m/z 1140.5[M-2H] ^2- .

Synthesis of compound 9:

Add 78.9 mg compound 8, 15.7 mg HOBT, 12 μL DIPEA and 1 mL DMF to the three-neck flask. After the reaction was stirred at room temperature for 10 min, 230 mg of solid phase carrier CPG-NH ₂ (amine content: 75 μmol/g) was added, and stirred at room temperature for 20 h. After the reaction was completed, it was filtered, and the solid was washed with DMF, acetonitrile, and DCM respectively, and dried to obtain 232.2 mg of solid. Afterwards, it was capped with acetic anhydride/pyridine/N-methylimidazole, washed again with DMF, acetonitrile, and DCM respectively, and dried to obtain 220.8 mg of compound 9 with a loading capacity of 14.9 μmol/g.

Synthesis of conjugate 9:

Conjugate 9 was prepared from compound 9 with reference to the method of Example 7 below.

Examples 2 to 6:

Referring to the synthetic route and operation of Example 1, using conventional synthesis methods in this field, Example 2-6 (compounds 16-18, 24-26, 33, 34-36, 37-39 and conjugates 18, 26, 33, 36, 39).

Example 7: Synthesis and Purification of Oligonucleotides (Conjugates)

Select a certain amount of solid-phase carrier (or other general-purpose solid-phase carrier) connected to GalNAc of the specific structure of the present invention, and prepare all oligonucleotides on the LK-192X synthesizer. According to the sequence requirements, the phosphoramidite monomers corresponding to the nucleosides were diluted with 1:40 (g/mL) anhydrous acetonitrile solvent, and the coupling time was 3 minutes, for a total of two couplings. Use 3% TCA for deprotection, use 0.3M benzylthiotetrazole acetonitrile solution for activation, and cap and oxidize (or thio) by CAPA/CAPB and 50mM _I2 solution (or PADS reagent) respectively. After trityl-offsynthesis, transfer the solid phase carrier to a 2 mL centrifuge tube, add 1.2 mL ammonia water and heat it in a 65°C oven for 3 hours to remove the protecting group. Then cool to room temperature, concentrate under vacuum for 30 minutes, filter the solution through a 0.22um filter membrane into a sampling bottle, and use a semi-preparative reverse-phase purifier for single-strand purification with an elution gradient of 7% to 30% (ACN: 100mM TEAA) , time 10min; flow rate: 5mL/min. After purification and preparation, concentrate in vacuum and spin to dryness at room temperature. Finally, the samples were dissolved in water and each solution was desalted on a GE Hi-Trap desalting column. salt to elute the final oligonucleotide product. All identities and purity were confirmed using ESI-MS and IEX HPLC respectively. Use a microplate reader to determine the concentration using UV light. Mix equal molar amounts of the sense strand and antisense strand into a new experimental tube, heat at 95°C for 5 minutes, and slowly anneal to room temperature. Finally, use a vacuum concentrator to spin dry at room temperature to obtain the final solution. product.

Oligonucleotide conjugates of related GalNAc derivative molecules prepared according to the above method and their related oligonucleotide sequences:

Note: The relevant sequence numbered XD targets the XDH gene; the relevant sequence numbered PC targets the PCSK9 gene; and the relevant sequence numbered AN targets the ANGPTL3 gene.

The following are the effect tests and data of the compounds of the present invention:

Test Example 1: Evaluation experiment of liver targeting effect of GalNAc ligand-conjugated siRNA with different structures in C57BL/6 mice

A total of 12 C57BL/6 female mice were used in the experiment. They were divided into 2 groups. The group information is as follows: Group 1 PC01 and Group 2 PC02. There were 6 mice in each group. They were all administered by subcutaneous injection. The drugs PC01 and PC02 were subcutaneously injected respectively. PC02 (recorded as 0h at the time of administration), the dosage was 5 mg/kg. In vivo imaging was performed on all mice in Group 1 PC01 and Group 2 PC02 at 2h, 4h, 24h and 48h; at 48h, mice in Group 1 PC01 and Group 2 PC02 were euthanized.

As shown in Figure 1, the 2h and 4h experimental results showed that after subcutaneous injection of the test drug in Group 1 PC01 and Group 2 PC02, the fluorescence signal was mainly detected in the liver, and gradually weakened as time went by. After 24 hours, the in vivo fluorescence signal decreased to an unobservable level. After 48 hours, the hearts, livers, spleens, lungs and kidneys of the euthanized mice were separated and in vitro images were taken. Signals of similar levels were detected in the isolated livers of PC01 in group 1 and PC02 in group 2.

As shown in Table 1, through the total fluorescence intensity of the in vivo fluorescence signals of PC01 and PC02 at 2h, 4h and 24h, the data were compared after obtaining the average fluorescence intensity. It was found that the average fluorescence intensity of PC02 was higher than that of the corresponding phase at 2h, 4h and 24h. The corresponding average fluorescence intensity of PC01 shows that PC02 has a relatively stronger enrichment effect in the liver than PC01.

Table 1 Fluorescence intensity values related to in vivo imaging of mice in each group at 2h, 4h and 24h

Test Example 2: Comparison of the in vitro uptake efficiency of GalNAc ligand-coupled siRNA with different structures

Huh7 cells were cultured in a 37°C, 5% _CO2 saturated humidity incubator containing 10% FBS/DMEM medium (100 μg/mL streptomycin, 100 U/mL penicillin). Huh7 cells cultured to 75% confluence were digested using trypsin (Gibico) and resuspended in 2% BSA/PBS buffer to prepare a cell suspension (1×10 ⁶ cells/mL). Take 100 μL of each cell suspension and add it to two different centrifuge tubes, and then add PC01 and PC02 coupled with fluorescent groups to the two centrifuge tubes (so that the final concentrations are both 20 nM). Incubate at room temperature in the dark for 1 hour, wash the cells twice with 2% BSA/PBS buffer, and use a flow cytometer to detect the average fluorescence density of the cells at an excitation wavelength of 647 nm.

The results in Figure 2 show that the average fluorescence density of Huh7 cells treated with PC02 was 213, which was significantly higher than the average fluorescence density of 147 cells treated with PC01 (P=0.0031). This shows that compared with PC01, PC02 obtained by the present invention is more easily absorbed by liver cells.

To further verify this conclusion, the uptake efficiency of PC01 and PC02 was tested in primary mouse liver cells. Resuspend fresh mouse liver primary cells in 2% BSA/PBS to make a cell suspension (1×10 ⁶ cells/mL). Take 100 μL of cell suspension into two different centrifuge tubes and add PC01 and PC02 coupled with fluorescent groups (final concentration 20nM). After incubation at room temperature in the dark for 1 hour, the cells were washed twice with PBS buffer, and then a flow cytometer was used to detect the average fluorescence density of the cells and the number of positive cells at an excitation wavelength of 647 nm.

The results in Figure 3 and Figure 4 show that: consistent with the results in Huh7, the average fluorescence density of primary mouse liver cells treated with PC02 was 287, while the average fluorescence density of primary mouse liver cells treated with PC01 was 173. In comparison, MFI is about doubled. It can be seen that compared with PC01, the efficiency of uptake of PC02 by liver cells is significantly higher.

In order to further prove that the molecules designed in the present invention are more easily taken up by liver cells, after the molecules designed in the present invention are coupled to different sequences targeting the same target or sequences targeting different targets, the following process is followed to evaluate the response of Huh7 cells to these molecules. uptake efficiency.

Prepare Huh7 cells in advance. The Huh7 cells cultured in a 10cm culture dish were digested and counted, and plated according to the number of 250,000 cells per well. Follow-up treatment will be carried out after 24 hours of adhesion to the wall. Wash Huh7 cells twice with PBS, then use DMEM containing 2% FBS to dilute different GalNac-coupled siRNA, adjust the concentration of siRNA to 200 nM, add it to the culture dish, and culture for 24 hours. Then aspirate the culture supernatant, wash the cells twice with PBS, count the cells after digestion, centrifuge and aspirate the supernatant, wash them twice more with PBS, and then directly put it on the machine for flow cytometry detection, use the histogram to read the fluorescence signal, and record The mean fluorescence intensity (MFI) of different samples was used for subsequent analysis. Read at least 10,000 signals per sample.

As shown in Figure 5, compound ligands are used for different targets and siRNA sequences (the related sequence numbered XD targets the XDH gene; the related sequence numbered PC targets the PCSK9 gene; and the related sequence numbered AN targets the ANGPTL3 gene). The 26 oligonucleotide conjugates (XD05 and XD11, PC07 and PC13, and AN05 and AN11) all had the highest uptake efficiency. The above experimental results show that the present invention obtains the targeting effect of the compound with liver-targeted delivery function. Does not depend on the target and siRNA sequence delivered.

Test Example 3: Using primary human hepatocytes (PHH) to evaluate the in vitro inhibition of PCSK9 mRNA activity by siRNA of the test compound

On day 0, first dilute siRNA to different concentrations with PBS, then add a certain volume of diluted siRNA to a 96-well cell culture plate; resuscitate the frozen PHH, adjust the PHH to an appropriate density, and then seed the PHH into in 96-well plates. Also set up control wells without compound. The tested compounds were tested at 8 concentration points and 3 replicate wells. 48 hours after plating, aspirate the supernatant and collect the cells. Extract the total RNA from the cells, follow the instructions of the reverse transcription kit, add random primers and reverse-transcribe it into cDNA, and then detect the cDNA in the sample by qPCR. At the same time, GAPDH primers and probes specifically detect GAPDH cDNA. The expression level of target gene mRNA in each sample was calculated by the ΔΔCt relative quantification method. The relative expression level of the target gene is represented by 2 ^-ΔΔCT , and the calculation formula is as follows:
ΔCT = average Ct value of the target gene - average Ct value of the internal reference gene;
ΔΔCT=ΔCT (drug-added group) group-ΔCT (no compound control group);
Relative mRNA expression = 2 ^-ΔΔCT

Figure 6 and the results in Table 2 show that PC02 containing ligand 9 inhibits PCSK9 mRNA activity in primary human hepatocytes (PHH) with an IC ₅₀ of 5.65 nM, which is better than the IC ₅₀ value of PC01 containing ligand L96 in the control group of 16.57 nM. PC02 is more active than PC01.

Table 2 Results of inhibiting PCSK9mRNA activity in primary human hepatocytes (PHH)

In addition, multiple studies have shown (1. Mariano Severgnini et al, A rapid two-step method for isolation of functional primary mouse hepatocytes: cell characterization and asialoglycoprotein receptor based assay development, Cytotechnology, 2012; 2. Nair et al, Multivalent N- acetylgalactosamine-conjugated siRNA localizes in hepatocytes and elicits robust RNAi-mediated gene silencing, 2014): GalNAc structures with different affinities (measured using Kd values), when coupled to the same small nucleic acid sequence, the coupler with higher affinity (i.e. Kd The smaller the coupling value), the higher the biological activity of the target gene in effective reduction in cell experiments (i.e., the smaller the IC ₅₀ ). Therefore, those skilled in the art can reasonably expect that the compound of the present invention conjugated to siRNA of other targets and other sequences can also have the effect of reducing the biological activity of the target gene.

Test Example 5: Effectiveness Test in Mouse

A total of 32 6-8 week old humanized PCSK9 male and female mice (Shanghai Southern Model Biotechnology Co., Ltd.) were used in the experiment. They were randomly divided into groups according to body weight, with 8 mice in each group, 4 males and 4 males, and a single dose of 3 mg/kg. Subcutaneous injection was administered in 4 groups. The blank is the negative control group injected with normal saline, PC15 is the positive control group, and the two groups PC17 and PC18 are the experimental groups. Blood was collected on days -3 (the third day before administration), 7, 14, 21, 28, and 35 (fasting for 4 hours before blood collection, the first dose was on day 0), and 0.1 ml of blood was collected by bleeding from the back of the eyeball. Human PCSK9 ELISA Kit (proteintech) was used to detect the expression level of PCSK9 protein in the serum of the blood separated serum, and comparisons were made between groups.

The mouse drug efficacy results show (Figure 7) that PCSK9 protein levels of PC15 (L96), PC17 (ligand 18) and PC18 (ligand 26) all dropped to the lowest on the 4th day, and then began to rebound. On the 28th day, PC15 (L96) basically returned to the pre-drug level, while the PCSK9 protein levels in the PC17 (ligand 18) and PC18 (ligand 26) groups were still lower than the pre-drug level.

Another group of experiments used 6-8 weeks old humanized PCSK9 male and female mice, a total of 24, randomly divided into groups according to body weight, 8 in each group, 4 males and 4 in each group, and administered with a single subcutaneous injection of 3 mg/kg. The blank is the negative control group injected with normal saline. PC15 is the positive control group, and PC19 is the experimental group. Blood was collected on days -3 (the 3rd day before administration), 7, 14, 21, and 28 (fasting for 4 hours before blood collection, the first dose was on day 0), and 0.1ml of blood was collected by bleeding from the back of the eye. Separation of serum using Human PCSK9 ELISA Kit (proteintech) was used to detect the expression level of PCSK9 protein in serum and make comparisons between groups.

The drug efficacy results in mice showed (Figure 8) that the PCSK9 protein levels of both PC15 (L96) and PC19 (ligand 39) dropped to the lowest on the 7th day, then began to increase, and returned to the pre-administration level on the 28th day.

Based on the above experimental results, it is shown that: from the perspective of experimental effect indicators such as inhibition level and rebound speed, when other conditions remain unchanged, the in vivo efficacy of the oligonucleotide conjugated with the compound with liver-targeted delivery effect of the present invention is Experiments were significantly better with L96-conjugated oligonucleotides.

Claims (21)

A compound represented by formula (I) or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, which has the following structure:
where each R is independently selected from:
Y ₁ , Y ₂ , Y ₃ , Y ₄ , Y ₅ , Y ₆ are each independently selected from -O-, -S-, -N(G ₁ )-, -C(G ₁ ) ₂ -; G ₁ is each The second occurrence is independently selected from hydrogen, halogen, hydroxyl, amino, carboxyl, or C _1-10 alkyl optionally substituted by one or more of halogen, hydroxyl, amino, carboxyl, C _1-6 alkyl Base, C _1-10 alkoxy group, -N(C _1-10 alkyl) ₂ ; Z ₁ , Z ₂ , Z ₃ and Z ₄ are each independently selected from C, N ⁺ and Si; Each occurrence of A ₁ and A ₂ is independently selected from hydrogen, or acetyl, propionyl, benzoyl, benzyl, which may be optionally substituted by one or more of halogen, hydroxyl, amino, and carboxyl. benzyloxycarbonyl; _{X 1 , X 2 , and _ _ _ _} G ₃ ) ₂ C(G ₃ ) ₂ S) _m -, or C _2-6 alkenyl, C _2-6 alkynyl, C _3-10 cycloalkyl optionally substituted by one or more G ₃ , C _3-10 cycloalkenyl, 3-10 membered heterocyclyl, C _6-14 aryl, 5-12 membered heteroaryl; Each occurrence of G ₃ is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, oxo, C _1-6 alkyl, C _1-6 alkoxy, or by deuterium, halogen, hydroxyl, amino One or more optionally substituted C _1-6 alkyl, C _1-6 alkoxy; X ₄ is independently selected from bonds, -(C(G ₄ ) ₂ ) _a -, -(C(G ₄ ) ₂ C(G ₃ ) ₂ O) _b -, -(C(G ₄ ) ₂ C(G ₃ ) ₂ S) _b -, or C _2-6 alkenyl, C _2-6 alkynyl, C _3-10 cycloalkyl, C _3-10 cycloalkenyl optionally substituted by one or more G ₄ , 3-10 membered heterocyclyl, C _6-14 aryl, 5-12 membered heteroaryl; Each occurrence of G ₄ is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, oxo, C _1-6 alkyl, C _1-6 alkoxy, or by deuterium, halogen, hydroxyl, amino One or more optionally substituted C _1-6 alkyl, C _1-6 alkoxy; n and a are independently selected from the integers from 0 to 24 each time they appear, and m and b are independently selected from the integers from 0 to 12 each time they appear; J ₁ , J ₂ , J ₃ , and J ₄ are each independently selected from the group consisting of bonds, -O-, -S-, -NH-, -C(O)-, -C(S)-, -S(O)- , -S(O) ₂ -, -NHC(O)-, -C(O)NH-, -C(O)O-, -OC(O)-, -SS-, -N(G ₅ )C(O)-, -C(O)N(G ₅ )- , -N(G ₅ )-, -C(G ₅ ) ₂ -, -Si(G ₅ ) ₂ -, -P(O)(OH)-, -P(O)O-, Each occurrence of G ₅ is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, carboxyl, -BH ₂ , -B(OH) ₂ , -N(C _1-6 alkyl) ₂ , C _1-12 alkyl group, C _1-12 alkoxy group, _or C 1-12 alkyl group, C _1-12 alkoxy group, C _3-10 ring optionally substituted by one or more of deuterium, halogen, hydroxyl, and amino group Alkyl, C _3-10 cycloalkenyl, 3-10 membered heterocyclyl, C _6-14 aryl, 5-12 membered heteroaryl; Each occurrence of J ₅ and J ₆ is independently selected from the group consisting of bonds, -O-, -S-, -NH-, -C(O)-, -C(S)-, -S(O)-, - S(O) ₂ -, -NHC(O)-, -C(O)NH-, -C(O)O-, -OC(O)-, -SS-, -N(G ₆ )C(O )-, -C(O)N(G ₆ )-, -N(G ₆ )-, -C(G ₆ ) ₂ -, -Si(G ₆ ) ₂ -, -P(O)(OH)- ,-P(O)O-, Each occurrence of G ₆ is independently selected from hydrogen, deuterium, halogen, hydroxyl, amino, carboxyl, -BH ₂ , -B(OH) ₂ , -N(C _1-6 alkyl) ₂ , C _1-12 alkyl group, C _1-12 alkoxy group, _or C 1-12 alkyl group, C _1-12 alkoxy group, C _3-10 ring optionally substituted by one or more of deuterium, halogen, hydroxyl, and amino group Alkyl, C _3-10 cycloalkenyl, 3-10 membered heterocyclyl, C _6-14 aryl, 5-12 membered heteroaryl; _{Each occurrence of _ _} , C _6-14 aryl, 5-12 membered heteroaryl; the C _1-6 alkyl, C _1-6 alkoxy, C _3-10 cycloalkyl, C _3-10 cycloalkenyl, 3 -10-membered heterocyclyl, C _6-14 aryl, 5-12-membered heteroaryl is unsubstituted or optionally substituted by one or more G ₇ ; Each occurrence of G ₇ is independently selected from halogen, hydroxyl, amino, cyano, oxo, -C _1-12 alkyl -OG ₈ , -OG ₉ , Wherein, Cat is a cation, independently selected from sodium ion, potassium ion, triethylammonium ion, tripropylammonium ion, tributylammonium ion and tetrabutylammonium ion; r is an integer from 0 to 5; Solid support is a solid phase carrier; G ₈ is independently selected from the following structures: H, G ₉ is independently selected from The compound according to claim 1 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: wherein, Y ₁ , Y ₂ , Y ₃ , Y ₄ , Y ₅ , Y ₆ are each independently selected from -O-, -N(G ₁ )-, -C(G ₁ ) ₂ -, further selected from -O-, -NH-, - CH(G ₁ )-; Each occurrence of G ₁ is independently selected from hydrogen, halogen, hydroxyl, amino, carboxyl, or may be replaced by one or more of halogen, hydroxyl, amino, carboxyl, C _1-6 alkyl Optionally substituted C _1-10 alkyl, -N(C _1-10 alkyl) ₂ , preferably, each occurrence of G ₁ is independently selected from hydrogen, methyl, ethyl, isopropyl, tris Fluoromethyl. The compound according to any one of claims 1 to 2 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: Y The branch unit composed of ₁ , Y ₂ , Y ₃ , Y ₄ , Y ₅ , Y ₆ and Z ₁ , Z ₂ , Z ₃ , Z ₄ is selected from the following structure: Preferably, the branch unit composed of Y ₁ , Y ₂ , Y ₃ , Y ₄ , Y ₅ , Y ₆ and Z ₁ , Z ₂ , Z ₃ , Z ₄ is The compound according to any one of claims 1 to 3 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: Z ₁ , Z ₂ , Z ₃ and Z ₄ are each independently selected from C. The compound according to any one of claims 1 to 4 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: A _1. Each occurrence of A ₂ is independently selected from hydrogen, or acetyl, propionyl, benzoyl, benzyl, benzyloxycarbonyl which may be optionally substituted by one or more halogens; preferably, A ₁ , each occurrence of A ₂ is independently selected from hydrogen and acetyl. The compound according to any one of claims 1 to 5 or its prodrug, tautomer, stereoisomer, solvate, Isotope _derivatives or pharmaceutically _acceptable salts thereof, characterized in that _: X ₁ , X _{2 , and 2} C(G ₃ ) ₂ O) _m -, -(C(G ₃ ) ₂ C(G ₃ ) ₂ S) _m -, or C _2-4 alkenyl optionally substituted by one or more G ₃ , C _2-4 alkynyl, C _3-6 cycloalkyl, 3-6 membered heterocycloalkyl, C _3-6 cycloalkenyl, C _6-10 aryl, 5-6 membered heteroaryl, the above The heteroatoms in the heterocycloalkyl and heteroaryl groups are independently selected from O, N or S, and the number of heteroatoms is 1, 2 or 3; preferably, X ₁ , X ₂ and X ₃ are each independently selected Self bond, -(CH ₂ ) _n -, -(CD ₂ ) _n -, -(CHD) _n -, -(CH ₂ CH ₂ O) _m -, -(CH ₂ CH ₂ S) _m -; preferably , X ₁ , X ₂ and X ₃ are each independently selected from the bond, -(CH ₂ ) _n -; preferably, X ₁ , X ₂ and X ₃ are each independently selected from the bond, -(CH ₂ ) ₂ -, -(CH ₂ ) ₄ -, -(CH ₂ ) ₆ -; preferably, X ₁ is independently selected from bonds, and X ₂ is independently selected from bonds, -(CH ₂ ) ₂ -, -(CH ₂ ) ₄ - , X ₃ is independently selected from -(CH ₂ ) ₂ -, -(CH ₂ ) ₄ -, -(CH ₂ ) ₆ -; preferably, G ₃ is independently selected from hydrogen, halogen, hydroxyl, Amino, oxo, C _1-4 alkyl, C _1-4 alkoxy, C _1-4 haloalkyl, C _1-4 haloalkoxy; each occurrence of G ₃ is independently selected from -H, - F, -Cl, -Br, -I, -OH, -NH ₂ , oxo, -CH ₃ , -CH ₂ CH ₃ ; preferably, G ₃ is independently selected from hydrogen. The compound according to claim 6 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: n is independently selected from 0 An integer of -15, m is independently selected from an integer of 0-6. The compound according to any one of claims 1 to 7 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: X ₄ is independently selected from the group consisting of bonds, -(C(G ₄ ) ₂ ) _a -, -(C(G ₄ ) ₂ C(G ₄ ) ₂ O) _b -, -(C(G ₄ ) ₂ C(G ₄ ) ₂ S) _b -, or C _2-4 alkenyl, C _2-4 alkynyl, C _3-6 cycloalkyl, C _3-6 cycloalkenyl optionally substituted by one or more G ₄ , 3-6 membered heterocycloalkyl, C _6-10 aryl, 5-6 membered heteroaryl, the heteroatoms in the heterocycloalkyl and heteroaryl are independently selected from O, N or S, the heteroatoms The number is 1, 2 or 3; preferably, X ₄ is independently selected from the group consisting of bonds, -(CH ₂ ) _a -, -(CD ₂ ) _a -, -(CHD) _a -, -(CH ₂ CH ₂ O) _b -, -(CH ₂ CH ₂ S) _b -; preferably, X ₄ is independently selected from the group consisting of bonds, -(CH ₂ ) _a -, -(CH ₂ CH ₂ O) _b -; preferably, X ₄ is independently selected from -(CH ₂ ) _a -; preferably, X ₄ is independently selected from bonds, -(CH ₂ ) ₈ -, -(CH ₂ ) ₉ -, -(CH ₂ ) ₁₀ -, - (CH ₂ ) ₁₁ -, -(CH ₂ ) ₁₂ -, -(CH ₂ ) ₁₃ -, -(CH ₂ ) ₁₄ -, -CH ₂ CH ₂ O-, -(CH ₂ CH ₂ O) ₂ -; Preferably, X ₄ is independently selected from -(CH ₂ ) ₁₀ -, -(CH ₂ ) ₁₁ -, -(CH ₂ ) ₁₂ -; preferably, G ₄ is independently selected from hydrogen, halogen, Hydroxy, amino, oxo, C _1-4 alkyl, C _1-4 alkoxy, C _1-4 haloalkyl, C _1-4 haloalkoxy; preferably, G ₄ is selected independently for each occurrence From -H, -F, -Cl, -Br, -I, -OH, -NH ₂ , oxo, -CH ₃ , -CH ₂ CH ₃ ; preferably, each occurrence of G ₄ is independently selected from hydrogen. The compound according to claim 8 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: a is independently selected from 0 An integer of -15, b is independently selected from an integer of 0-6. The compound according to any one of claims 1 to 9 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: J ₁ , _J2 , _J3 , _J4 are each independently selected from the group consisting of bonds, -O-, -S-, -NH-, -C(O)-, -NHC(O)-, -C(O)NH- , -C(O)O-, -OC(O)-, -N(G ₅ )C(O)-, -C(O)N(G ₅ )-, -N(G ₅ )-, -C (G ₅ ) ₂ -; Preferably, J ₁ , J ₂ , J ₃ , and J ₄ are each independently selected from bonds, -O-, -NHC(O)-, -C(O)NH-; Preferably, J ₁ is independently selected from bond, J ₂ is independently selected from bond, -O-, J ₃ is independently selected from -O-, -NHC(O)-, -C(O)NH-, and J ₄ is independently selected from From -NHC(O)-, -C(O)NH-; preferably, J ₁ is independently selected from bond, J ₂ is independently selected from bond, -O-, and J ₃ is independently selected from -O-, - NHC(O)-, J ₄ is independently selected from -C(O)NH-; preferably, G ₅ is independently selected from -H, -F, -Cl, -Br, -I, -NH at each occurrence _2. -CD ₃ , -CH ₂ F , -CHF ₂ , -CF ₃ , -CH ₂ Cl , -CH ₂ I , -N(CH ₃ )(CH ₃ ), -CH ₃ , -CH ₂ CH ₃ , -CH ₂ CH ₂ CH ₃ , -C(CH ₃ ) ₃ , Preferably, each occurrence of G ₅ is independently selected from -H. The compound according to any one of claims 1 to 10 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: J ₅ , J ₆ are each independently selected from the group consisting of bonds, -C(O)-, -NHC(O)-, -C(O)NH-, -N(G ₆ )C(O)-, -C(O) N(G ₆ )-; preferably, J ₅ and J ₆ are each independently selected from bond, -O-, -C(O)-, -NHC(O)-, -C(O)NH-, -OC (O)-; Preferably, J ₅ is independently selected from -NHC(O)-, -C(O)NH-; Preferably, J ₆ is independently selected from bond, -C(O)-; Preferably, Each occurrence of G ₆ is independently selected from -H, -F, -Cl, -Br, -I, -NH ₂ , -CD ₃ , -CH ₂ F, -CHF ₂ , -CF ₃ , -CH ₂ Cl , -CH ₂ I, -N(CH ₃ )(CH ₃ ), -CH ₃ , -CH ₂ CH ₃ , -CH ₂ CH ₂ CH ₃ , -C(CH ₃ ) ₃ , Preferably, each occurrence of G ₆ is independently selected from -H, -F, -NH ₂ , -CD ₃ , -CF ₃ , -CH ₃ , -CH ₂ CH ₃ , -CH ₂ CH ₂ CH ₃ , -C(CH ₃ ) ₃ ; preferably, G ₆ is independently selected from -H at each occurrence. The compound according to any one of claims 1 to 11 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: X Each occurrence of ₅ is independently selected from bond, C _1-6 alkyl, C _3-6 cycloalkyl, C _3-6 cycloalkenyl, 3-6 membered heterocycloalkyl, C _6-10 aryl, 5-6 membered heteroaryl; the C _1-6 alkyl, C _3-6 cycloalkyl, C _3-6 cycloalkenyl, 3-6 membered heterocycloalkyl, C _6-10 aryl, 5 -6-membered heteroaryl is unsubstituted or optionally substituted by one or more _G7 ; preferably, _X5 is independently selected for each occurrence of bond, _C1-4 alkyl, 5-6 membered heteroaryl Cycloalkyl, the C _1-4 alkyl, 5-6 membered heterocycloalkyl is unsubstituted or optionally substituted by one or more G ₇ ; preferably, X ₅ is independently selected each time it appears. Methyl, ethyl, n-propyl, isopropyl, or optionally substituted by one or more G ₇ Preferably, each occurrence of _X5 is independently selected from methyl substituted by one or two _G7 , Preferably, each occurrence of X ₅ is independently selected from the group consisting of substituted by one or two G ₇ The compound according to claim 12 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: G ₇ is independently selected from hydroxyl , -C _1-6 alkyl-OG ₈ , -OG ₉ , Preferably, G ₇ is independently selected from hydroxyl, -C _1-4 alkyl -OG ₈ , -OG ₉ , More preferably, G ₇ is independently selected from hydroxyl, -C _1-3 alkyl-OG ₈ , -OG ₉ , More preferably, G ₇ is independently selected from hydroxyl, -CH ₂ -OG ₈ , -OG ₉ , The compound according to claim 13 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: G ₈ is independently selected from H , Preferably, G ₈ is independently selected from H, More preferably, G ₈ is H or G ₉ is independently selected from Preferably, G ₉ is independently selected from Preferably, G ₉ is independently selected from The compound according to any one of claims 1 to 14 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: Cat Independently selected from sodium ions, triethylammonium ions, and tetrabutylammonium ions; r is an integer from 1 to 2, specifically 1, 2; preferably, r is 1; Solid support is a resin solid phase carrier. The compound according to any one of claims 1 to 15 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: R independently selected from
A compound represented by formula (II) or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, which has the following structure:
where each R is independently selected from:
Among them, Y ₁ , Y ₂ , Y ₃ , Y ₄ , Y ₅ , Y ₆ , Z ₁ , Z ₂ , Z ₃ , Z ₄ , X ₁ , X ₂ , X 3 , X _{4 ,} J ₁ , J ₂ , The definitions of J ₃ , J ₄ , J ₅ , J ₆ , A ₁ and A ₂ are the same as those of the compound of formula (I) above, and the definition of X ₅ ' is only the same as the definition of X ₅ in the compound of formula (I) above. Lack of an H, hydroxyl group, OG ₈ or OG ₉ to connect to E ₄ ; E ₄ is independently selected from -O-, -S-; E ₁ is -OH, and E ₂ is independently selected from =O, =S; alternatively, E ₁ is -O ^- , and E ₂ is independently selected from =O, =S; alternatively, E ₁ is =O, and E ₂ is independently selected from -CH ₃ , -O(C _1-6 alkyl), -NH(C _1-6 alkyl), -BH _{3 ,} -SH, -S ^- ; alternatively, E ₁ is =S , and E ₂ is independently selected from -CH ₃ , -SH, -S ^- ; E ₃ is a functional oligonucleotide molecule, including but not limited to: small interfering RNA, microRNA, immune stimulators, alternative splice bodies, single-stranded RNA, double-stranded RNA, antisense nucleic acids, nucleic acid aptamers, One of stem-loop RNA, mRNA fragment, activating RNA or DNA. The compound according to claim 1 or 17 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, characterized in that: the compound is selected from :






The compound according to claim 18 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof, the siRNA includes a sense of complementary double-stranded region strand and antisense strand, the sense strand and/or the antisense strand includes or consists of 15-25 nucleotides, and the antisense strand is consistent with at least 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or 25 consecutive complementary nucleotides, the length of the double-stranded region is 15-25bp, the At least one nucleotide in the siRNA molecule is modified; preferably, any strand of the siRNA molecule contains an overhang of 0-5 bases; the target genes include but are not limited to: ApoB, ApoC, ANGPTL3, PCSK9, SCD1, FVII, p53, HAV, HBV, HCV, HDV, HEV, AGT, Lp(a), XDH, HSD17B13, PNPLA3, HMGCR, etc. A pharmaceutical composition, characterized by: comprising the compound according to any one of claims 1 to 19 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or its pharmaceutical Acceptable salts, and pharmaceutically acceptable excipients. The compound according to any one of claims 1 to 19 or its prodrug, tautomer, stereoisomer, solvate, isotope derivative or pharmaceutically acceptable salt thereof or claim 20 The use of a pharmaceutical composition in the preparation of medicaments for the treatment, prevention and/or treatment of physiological conditions or diseases caused by the expression of specific genes in liver cells; preferably, the diseases include: chronic liver disease, hepatitis, liver fibrosis chemical diseases, liver proliferative diseases and dyslipidemia.