N-Acetylgalactosamine-Containing Targeting Ligands

Abstract

The present invention provides an N-acetylgalactosamine-containing targeting ligand, the ligand comprising a conjugate group of formula (X') and a nucleic acid molecule containing the targeting ligand. [C1] TIFF2025504082000406.tif24156

Description

[Technical field]

[0001] The present invention is in the field of medicine, and specifically relates to targeting ligands containing N-acetylgalactosamine and nucleic acid molecules containing said targeting ligands. [Background technology]

[0002] N-acetylgalactosamine (GalNAc) can specifically bind to the asialoglycoprotein receptor (ASGPR) on the surface of hepatocytes. Conjugation of GalNAc-containing targeting ligands to small interfering RNA (siRNA) allows for the delivery of siRNA targeted to the liver.

[0003] Although GalNAc-containing targeting ligands are disclosed in the prior art, there is a need in the art for targeting ligands that have better stability, targeting properties, and the like. Summary of the Invention [Means for solving the problem]

[0004] The present invention provides novel N-acetylgalactosamine-containing targeting ligands.

[0005] In one aspect, the present invention relates to an N-acetylgalactosamine-containing targeting ligand, said ligand comprising a conjugate group of formula (X'): [ka]

[0006] During the ceremony,

[0007] [ka] indicates the position of attachment to the biomolecule,

[0008] Q is independently H; [ka] and

[0009] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0010] L2 is a chemical bond or -CH2CH2C(O)-;

[0011] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0012] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0013] In the formula, a=0, 1, 2 or 3;

[0014] b=1, 2, 3, 4 or 5;

[0015] c=1, 2, 3, 4 or 5;

[0016] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0017] L is a chemical bond, -CHO- or -NHC(O)-;

[0018] L' is a chemical bond, -C(O)NH-, -NHC(O)- or -O(CH2CH2O) e - and

[0019] In the formula, e is 1, 2, 3, 4 or 5;

[0020] T is a chemical bond, -CH2-, -C(O)-, -M-, -CH2-M- or -C(O)-M-;

[0021] In the formula, M is [ka] and

[0022] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0023] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0024] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0025] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0026] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0027] In another aspect, the present invention relates to a targeting ligand containing N-acetylgalactosamine, wherein the conjugate group is shown in formula (I'): [ka]

[0028] During the ceremony,

[0029] [ka] indicates the position of attachment to the biomolecule,

[0030] Q is independently H; [ka] and

[0031] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0032] L2 is a chemical bond or -CH2CH2C(O)-;

[0033] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0034] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0035] In the formula, a=0, 1, 2 or 3;

[0036] b=1, 2, 3, 4 or 5;

[0037] c=1, 2, 3, 4 or 5;

[0038] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0039] L is -CH2O- or -NHC(O)-;

[0040] L' is a chemical bond, -C(O)NH- or -NHC(O)-;

[0041] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0042] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0043] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0044] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0045] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0046] In another aspect, the present invention relates to a targeting ligand containing N-acetylgalactosamine, wherein the conjugate group is shown in formula (X'):

[0047] Q is independently H; [ka] and

[0048] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0049] L2 is a chemical bond or -CH2CH2C(O)-;

[0050] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0051] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0052] In the formula, a=0, 1, 2 or 3;

[0053] b=1, 2, 3, 4 or 5;

[0054] c=1, 2, 3, 4 or 5;

[0055] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0056] L is a chemical bond, -CHO- or -NHC(O)-;

[0057] L' is a chemical bond, -C(O)NH-, -NHC(O)- or -O(CH2CH2O) e - and

[0058] In the formula, e is 1, 2, 3, 4 or 5;

[0059] T is a chemical bond, -CH2-, -M-, -CH2-M- or -C(O)-M-;

[0060] In the formula, M is

number

[0061] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0062] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0063] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0064] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0065] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0066] In another aspect, the present invention relates to a targeting ligand containing N-acetylgalactosamine, wherein the conjugate group is shown in formula (X'):

[0067] Q is independently H; [ka] and

[0068] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0069] L2 is a chemical bond or -CH2CH2C(O)-;

[0070] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0071] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0072] In the formula, a=0, 1, 2 or 3;

[0073] b=1, 2, 3, 4 or 5;

[0074] c=1, 2, 3, 4 or 5;

[0075] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0076] L is a chemical bond, -CHO- or -NHC(O)-;

[0077] L' is -O(CH2CH2O) e - and

[0078] In the formula, e is 1, 2, 3, 4 or 5;

[0079] T is a chemical bond, -CH2-, -C(O)-, -M-, -CH2-M- or -C(O)-M-;

[0080] In the formula, M is [ka] and

[0081] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0082] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0083] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0084] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0085] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0086] In another aspect, the present invention relates to a compound of formula (X) or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof,

[0087] [ka] During the ceremony,

[0088] Q is independently H; [ka] and

[0089] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0090] L2 is a chemical bond or -CH2CH2C(O)-;

[0091] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0092] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0093] In the formula, a=0, 1, 2 or 3;

[0094] b=1, 2, 3, 4 or 5;

[0095] c=1, 2, 3, 4 or 5;

[0096] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0097] L is a chemical bond, -CHO- or -NHC(O)-;

[0098] L' is a chemical bond, -C(O)NH-, -NHC(O)- or -O(CHCHO)e-;

[0099] In the formula, e is 1, 2, 3, 4 or 5;

[0100] T is a chemical bond, -CH2-, -C(O)-, -M-, -CH2-M- or -C(O)-M-;

[0101] In the formula, M is [ka] and

[0102] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0103] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0104] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0105] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0106] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0107] In another aspect, the present invention relates to a compound of formula (I) or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof,

[0108] [ka] During the ceremony,

[0109] Q is independently H; [ka] and

[0110] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0111] L2 is a chemical bond or -CH2CH2C(O)-;

[0112] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0113] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0114] In the formula, a=0, 1, 2 or 3;

[0115] b=1, 2, 3, 4 or 5;

[0116] c=1, 2, 3, 4 or 5;

[0117] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0118] L is -CH2O- or -NHC(O)-;

[0119] L' is a chemical bond, -C(O)NH- or -NHC(O)-;

[0120] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0121] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0122] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0123] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0124] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0125] In another aspect, the present invention relates to a compound of formula (X), or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof, wherein:

[0126] Q is independently H; [ka] and

[0127] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0128] L2 is a chemical bond or -CH2CH2C(O)-;

[0129] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0130] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0131] In the formula, a=0, 1, 2 or 3;

[0132] b=1, 2, 3, 4 or 5;

[0133] c=1, 2, 3, 4 or 5;

[0134] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0135] L is a chemical bond, -CHO- or -NHC(O)-;

[0136] L' is a chemical bond, -C(O)NH-, -NHC(O)- or -O(CHCHO)e-;

[0137] In the formula, e is 1, 2, 3, 4 or 5;

[0138] T is a chemical bond, -CH2-, -M-, -CH2-M- or -C(O)-M-;

[0139] In the formula, M is [ka] and

[0140] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0141] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0142] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0143] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0144] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0145] In another aspect, the present invention relates to a compound of formula (X), or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof, wherein:

[0146] Q is independently H; [ka] and

[0147] wherein L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O- or -NHC(O)-(CH2NHC(O))a-;

[0148] L2 is a chemical bond or -CH2CH2C(O)-;

[0149] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0150] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2)c-, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0151] In the formula, a=0, 1, 2 or 3;

[0152] b=1, 2, 3, 4 or 5;

[0153] c=1, 2, 3, 4 or 5;

[0154] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0155] L is a chemical bond, -CHO- or -NHC(O)-;

[0156] L' is -O(CH2CH2O) e - and

[0157] In the formula, e is 1, 2, 3, 4 or 5;

[0158] T is a chemical bond, -CH2-, -C(O)-, -M-, -CH2-M- or -C(O)-M-;

[0159] In the formula, M is [ka] and

[0160] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0161] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0162] wherein R is -OR', -CH2OR' or -CH2CH2OR', R' is H, a hydroxyl protecting group or a solid support, the hydroxyl protecting group being preferably -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytrityl.

[0163] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0164] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0165] In another aspect, the present invention relates to an intermediate compound, which is a compound of formula (V): [ka]

[0166] During the ceremony,

[0167] Pg is a hydroxyl protecting group selected, for example, from [ka] Q' is independently H; [ka] and

[0168] The other groups are as defined above.

[0169] In another aspect, the present invention relates to a process for preparing a compound of formula (I), the process comprising treating a compound of formula (V) under conditions to remove a hydroxyl protecting group, [ka]

[0170] Each group is as defined above.

[0171] In another aspect, the invention relates to a nucleic acid molecule, which comprises a nucleic acid and, attached thereto, a ligand as defined above.

[0172] In another aspect, the invention relates to a pharmaceutical composition, which comprises a nucleic acid molecule described herein and a pharma- ceutically acceptable carrier or excipient.

[0173] In another aspect, the invention relates to a kit, which comprises a nucleic acid molecule described herein.

[0174] In certain embodiments of the above ligands, compounds, intermediate compounds and nucleic acid molecules, the group definitions are preferably as follows:

[0175] Q In one embodiment, Q is H, and in another embodiment, Q is [ka] and in another embodiment, Q is [ka] and in another embodiment, Q is [ka] and in another embodiment, Q is [ka] and in another embodiment, Q is [ka] and in another embodiment, Q is [ka] and in another embodiment, Q is [ka] In another embodiment, Q is any combination of the above groups.

[0176] L1, L2, L3 and L4 In one embodiment, L1 is a chemical bond; in another embodiment, L1 is -CH2-; in another embodiment, L1 is -CH2CH2-; in another embodiment, L1 is -C(O)-; in another embodiment, L1 is -CH2O-; in another embodiment, L1 is -CH2O-; in another embodiment, L1 is -NHC(O)-(CH2NHC(O)) a -It is.

[0177] In one embodiment, L2 is a chemical bond, and in another embodiment, L2 is -CH2CH2C(O)-.

[0178] In one embodiment, L3 is a chemical bond, and in another embodiment, L3 is -(NHCH2CH2). b and in another embodiment, L3 is -C(O)CH2-.

[0179] In one embodiment, L4 is -(OCH2CH2) c In another embodiment, L4 is -(OCH2CH2CH2) c In another embodiment, L4 is -(OCH2CH2CH2CH2) c In another embodiment, L4 is -(OCH2CH2CH2CH2CH2) c In another embodiment, L4 is -NHC(O)-(CH2). d -It is.

[0180] L, L' and T In one embodiment, L is a chemical bond, in another embodiment, L is -CH2O-, and in another embodiment, L is -NHC(O)-.

[0181] In one embodiment, L' is a chemical bond, in another embodiment, L' is -C(O)NH-, and in another embodiment, L' is -O(CH2CH2O) e -It is.

[0182] In one embodiment, T is a chemical bond, in another embodiment, T is -C(O)-, in another embodiment, T is -M-, in another embodiment, T is -CH-M-, and in another embodiment, T is -C(O)-M-.

[0183] In one embodiment, M is [ka] and in another embodiment, M is [ka] and in another embodiment, M is [ka] and in another embodiment, M is [ka] It is.

[0184] R1, R2 and R3 In one embodiment, R1 and R2 together form -CH2CH2O- and R3 is H, in another embodiment, R1 and R2 together form -CH2CH(R)-O- and R3 is H, in another embodiment, R1 and R3 together form -C 1~2 alkylene- and R2 is H.

[0185] In one embodiment, R is -OR', in another embodiment, R is -CHOR', in another embodiment, R is -CHCHOR', in a more specific embodiment, R' is H, in another more specific embodiment, R' is a hydroxyl protecting group, in another more specific embodiment, R' is a solid phase support, in another more specific embodiment, R' is -C(O)CHCHC(O)OH, and in another more specific embodiment, R' is 4,4'-dimethoxytrityl. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0186] definition chemical definition Definitions of certain functional groups and chemical terms are explained in more detail below.

[0187] When numerical ranges are listed, each value and subrange within the range is intended to be included. For example, "C 1~6 Alkyl" is C1, C2, C3, C4, C5, C6, C 1~6 , C 1~5 , C 1~4 , C 1~3 , C 1~2 , C 2~6 , C 2~5 , C 2~4 , C 2~3 , C 3~6 , C 3~5 , C 3~4 , C 4~6 , C 4~5 and C 5~6 Contains alkyl.

[0188] "C 1~6 "Alkyl" refers to a straight or branched chain saturated hydrocarbon group having 1 to 6 carbon atoms. In some embodiments, preferably, 1~ 4 alkyl and C 1~2 It is an alkyl group. 1~6Examples of alkyl groups include methyl (C1), ethyl (C2), n-propyl (C3), isopropyl (C3), n-butyl (C4), tert-butyl (C4), sec-butyl (C4), isobutyl (C4), n-pentyl (C5), 3-pentyl (C5), pentyl (C5), neopentyl (C5), 3-methyl-2-butyl (C5), tert-pentyl (C5), and n-hexyl (C6). 1~6 The term "alkyl" further includes heteroalkyl, where one or more (e.g., 1, 2, 3, or 4) carbon atoms are replaced with a heteroatom (e.g., oxygen, sulfur, nitrogen, boron, silicon, phosphorus). Alkyl groups can be optionally substituted with one or more substituents, e.g., 1-5 substituents, 1-3 substituents, or 1 substituent. Common abbreviations for alkyl include Me(-CH3), Et(-CH2CH3), iPr(-CH(CH3)2), nPr(-CH2CH2CH3), n-Bu(-CH2CH2CH2CH3), or i-Bu(-CH2CH(CH3)2).

[0189] "Halo" or "halogen" refers to fluorine (F), chlorine (Cl), bromine (Br) and iodine (I).

[0190] Therefore, "C 1~6 Haloalkyl means the above C substituted with one or more halogen groups. 1~6 In some embodiments, preferably, C 1~4 Haloalkyl groups, more preferably C 1~2 Exemplary haloalkyl groups are -CF, -CHF, -CHFCHF, -CHCHF, -CFCF, -CC l3 Haloalkyl groups include, but are not limited to, -CH2Cl, -CHCl2, 2,2,2-trifluoro-1,1-dimethylethyl, and the like. Haloalkyl groups can be substituted at any available point of attachment, for example, with 1 to 5 substituents, 1 to 3 substituents, or 1 substituent.

[0191] "C 1~6 Alkylene" is C 1~6It refers to a divalent group formed by removing another hydrogen from an alkyl group, which may be substituted or unsubstituted. In some embodiments, preferably, C 1~4 Alkylene, C 2~4 Alkylene and C 1~2 Alkylene. Unsubstituted alkylene groups include, but are not limited to, methylene (-CH-), ethylene (-CHCH-), propylene (-CHCHCH-), butylene (-CHCHCHCHCH-), pentylene (-CHCHCHCHCHCH-), hexylene (-CHCHCHCHCHCHCH-), and the like. Exemplary substituted alkylene groups, such as alkylene groups substituted with one or more alkyl(methyl)s, include, but are not limited to, substituted methylene (-CH(CH3)-, -C(CH3)2-), substituted ethylene (-CH(CH3)CH2-, -CH2CH(CH3)-, -C(CH3)2CH2-, -CH2C(CH3)2-), substituted propylene (-CH(CH3)CH2CH2-, -CH2CH(CH3)CH2-, -CH2CH2CH(CH3)CH2-, -CH2CH2CH(CH3)-, -C(CH3)2CH2CH2-, -CH2C(CH3)2CH2-, -CH2CH2C(CH3)2-), and the like.

[0192] Other definitions The term "siRNA" herein refers to a class of double-stranded RNA molecules that can mediate the silencing of a target RNA (e.g., mRNA, e.g., a transcript of a gene encoding a protein) that is complementary thereto. siRNA is usually double-stranded, comprising an antisense strand that is complementary to the target RNA, and a sense strand that is complementary to the antisense strand. For convenience, such an mRNA is also referred to herein as the mRNA to be silenced. Such a gene is also referred to as the target gene. Usually, the RNA to be silenced is an endogenous gene or a pathogen gene. In addition, RNA other than mRNA (e.g., tRNA, etc.) and viral RNA can also be targeted.

[0193] "shRNA" refers to short hairpin RNA. shRNA contains two short inverted repeats. When cloned into an shRNA expression vector, the two short inverted repeats, separated by a stem-loop sequence in the middle, form a hairpin structure and are controlled by the pol III promoter. Then, 5 to 6 T's are bound as transcription terminators for RNA polymerase III.

[0194] The term "antisense strand" refers to the strand of an siRNA that contains a region that is completely, fully, or substantially complementary to a target sequence. The term "sense strand" refers to the strand of an siRNA that contains a region that is completely, fully, or substantially complementary to a region that is the antisense strand, as that term is defined herein.

[0195] The term "complementary region" refers to a region on the antisense strand that is completely, fully, or substantially complementary to the target mRNA sequence. If the complementary region is not completely complementary to the target sequence, mismatches may be located in the internal or terminal regions of the molecule. Usually, the most tolerable mismatches are in the terminal regions, e.g., within 5, 4, 3, 2, or 1 nucleotide of the 5' and / or 3' ends. The part of the antisense strand that is most sensitive to mismatches is called the "seed region". For example, in an siRNA that includes a 19nt strand, some mismatches at position 19 (5' to 3') are tolerated.

[0196] The term "complementary" refers to the ability of a first polynucleotide to hybridize to a second polynucleotide under certain conditions, such as stringent conditions. For example, stringent conditions may include 400 mM NaCl, 40 mM PIPES pH 6.4, 1 mM EDTA, at 50°C or 70°C for 12-16 hours. As long as it meets the above requirements for the ability to hybridize, a "complementary" sequence may also contain or be formed entirely from non-Watson-Crick base pairs and / or base pairs formed from non-natural and modified nucleotides. Such non-Watson-Crick base pairs include, but are not limited to, G:U wobble base pairs or Hoogstein base pairs.

[0197] A polynucleotide that is "at least partially complementary," "fully complementary," or "substantially complementary" to a messenger RNA (mRNA) refers to a polynucleotide that is substantially complementary to a continuous portion of the mRNA of interest. For example, a polynucleotide is complementary to at least a portion of a PCSK9 mRNA if the sequence is substantially complementary to an uninterrupted portion of the PCSK9 mRNA. As used herein, the terms "complementary," "fully complementary," "fully complementary," and "substantially complementary" can be used in reference to base pairing between the sense and antisense strands of an siRNA, or between the antisense strand of an siRNA agent and a target sequence.

[0198] "Sufficiently complementary" refers to the extent to which the sense strand needs to be complementary to the antisense strand to maintain the overall double-stranded nature of the molecule.In other words, complete complementarity is usually required, but in some cases, particularly in the antisense strand, one or more, for example, 6, 5, 4, 3, 2 or 1 mismatches (with respect to target mRNA) may be included, but the sense strand and the antisense strand can still maintain the overall double-stranded nature of the molecule.

[0199] A "nucleoside" is a compound composed of two substances, a purine or pyrimidine base and ribose or deoxyribose; a "nucleotide" is a compound composed of three substances, a purine or pyrimidine base, ribose or deoxyribose, and phosphate; and an "oligonucleotide" refers to a nucleic acid molecule (RNA or DNA) that is, for example, less than 100, 200, 300, or 400 nucleotides in length.

[0200] A "base" is a basic unit for synthesizing nucleosides, nucleotides, and nucleic acids, and its components contain nitrogen and are also called "nitrogenous bases." In this specification, unless otherwise specified, capital letters A, U, T, G, and C represent the base composition of nucleotides, and are adenine, uracil, thymine, guanine, and cytosine, respectively.

[0201] The "modification" of the nucleotide described herein includes, but is not limited to, methoxy modification, fluoro modification, phosphorothioate group binding, or protection with a common protecting group, etc. For example, the fluoro-modified nucleotide refers to a nucleotide in which the 2'-position hydroxyl group of the ribose group of the nucleotide is substituted with fluorine, and the methoxy-modified nucleotide refers to a nucleotide in which the 2'-position hydroxyl group of the ribose group is substituted with a methoxy group.

[0202] As used herein, "modified nucleotide" includes, but is not limited to, 2-O-methyl modified nucleotide, 2'-fluoro modified nucleotide, 2'-deoxy modified nucleotide, inosine ribonucleoside glycolate, abasic nucleotide, reverse base deoxyribonucleotide, nucleotide containing phosphorothioate group, vinyl phosphate modified nucleotide, locked nucleotide, 2'-amino modified nucleotide, 2'-alkyl modified nucleotide, morpholino nucleotide, phosphoramidate, non-natural base containing nucleotide, and terminal nucleotide connected to cholesteryl derivative or dodecyl dodecyl amide group, deoxyribonucleotide or protection by normal protecting group. For example, the above 2'-fluoro modified nucleotide refers to a nucleotide in which the hydroxyl group at the 2' position of the ribose group of the nucleotide is replaced by fluorine. The 2'-deoxy modified nucleotide refers to a nucleotide in which the 2'-hydroxyl group of the ribose group is replaced by a methoxy group.

[0203] The term "hydroxyl protecting group" refers to a group that can protect a hydroxyl group from chemical reactions and can be removed under certain conditions to restore the hydroxyl group. Mainly, it includes silane type protecting groups, acyl type protecting groups, and ether type protecting groups, but the following protecting groups are preferred:

[0204] Trimethylsilyl (TMS), triethylsilyl (TES), dimethylisopropylsilyl (DMIPS), diethylisopropylsilyl (DEIPS), tert-butyldimethylsilyl (TBDMS), tert-butyldiphenylsilyl (TBDPS), triisopropylsilyl (TIPS), acetyl (Ac), chloroacetyl, dichloroacetyl, trichloroacetyl, trifluoroacetyl (TFA), benzoyl, p-methoxybenzoyl, 9-fluorenylmethoxycarbonyl (Fmoc), allyloxycarbonyl Allyl (Alloc), 2,2,2-trichloroethoxycarbonyl (Troc), benzyloxycarbonyl (Cbz), tert-butoxycarbonyl (Boc), benzyl (Bn), p-methoxybenzyl (PMB), allyl, triphenylmethyl (Tr), bis-p-methoxytrityl (DMTr), methoxymethyl (MOM), phenoxymethyl (BOM), 2,2,2-trichloroethoxymethyl, 2-methoxyethoxymethyl (MEM), methylthiomethyl (MTM), p-methoxybenzyloxymethyl (PMBM).

[0205] As used herein, the term "pharmaceutically acceptable salts" refers to carboxylate salts, amino acid addition salts of the compounds of the invention, which are effective for their anticipated use within the scope of sound medical judgment for contact with patient tissues without undue toxicity, irritation, allergic response, and the like, commensurate with a reasonable benefit / risk ratio, and includes (where possible) zwitterionic forms of the compounds of the invention.

[0206] The compounds of the present invention may contain one or more asymmetric centers and therefore may exist in multiple stereoisomeric forms, such as enantiomeric and / or diastereomeric forms. For example, the compounds of the present invention may be in the form of individual enantiomers, diastereoisomers or geometric isomers (e.g., cis and trans isomers), or may be in the form of a mixture of stereoisomers, including racemic mixtures and mixtures enriched in one or more stereoisomers. Isomers can be separated from mixtures by methods known to those skilled in the art, including chiral high pressure liquid chromatography (HPLC) and the formation and crystallization of chiral salts, or preferred isomers can be separated by asymmetric synthesis.

[0207] The present invention further includes isotopically labeled compounds (isotopic variants) that are identical to those described in formula (I) except that one or more atoms are surrounded by atoms having essentially different atomic masses or mass numbers than the general formula (I). Illustrative examples of isotopes that can be incorporated into the compounds of the present invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, and chlorine, e.g., 2H, 3H, 4H, 5H, 6H, 7H, 8H, 9H, 10H, 11H, 12H, 13H, 14H, 15H, 16H, 17H, 18H, 19H, 20H, 21H, 22H, 23H, 24H, 25H, 26H, 27H, 28H, 29H, 30H, 31H, 32H, 33H, 34H, 35H, 36H, 37H, 38H, 39H, 40H, 41H, 42H, 43H, 44H, 45H, 46H, 47H, 48H, 49H, 50H, 51H, 52H, 53H, 54H, 55H, 56H, 57H, 58H, 59H, 60H, 61H, 62H, 63H, 64H, 65H, 66H, 67H, 68H, 69H, 70H, 71H, 72H, 73H, 74H, 75H, 76H, 77H, 78H, 79H, 80H, 81H, 82H, 83H, 84 3 H, 13 C. 11 C. 14 C. 15 N, 18 O. 17 O. 31 P, 32 P, 35 S, 18 F and 36 Compounds of the invention, prodrugs thereof, and pharma- ceutically acceptable salts of the compounds or prodrugs that contain the above isotopes and / or other isotopes of other atoms are within the scope of the invention. Certain isotopically labeled compounds of the invention, e.g., those that contain radioactive isotopes (e.g., 3 H and 14 Compounds incorporating C) may be used in drug and / or substrate tissue distribution assays. Tritium, i.e., 3H and carbon-14, i.e., 14 C isotopes are particularly preferred because they are easy to prepare and detect. Additionally, heavier isotopes such as deuterium, i.e. 2Substitution with H may be preferred in some cases due to the potential for greater metabolic stability resulting in therapeutic benefits such as increased in vivo half-life and reduced dosage requirements. Isotopically labeled compounds of formula (I) of the present invention and prodrugs thereof may generally be prepared by substituting a readily available isotopically labeled reagent for a non-isotopically labeled reagent in carrying out the processes disclosed in the following flow charts and / or examples and preparations.

[0208] The present invention particularly relates to the following technical solutions:

[0209] 1. An N-acetylgalactosamine-containing targeting ligand, said ligand comprising a conjugate group of formula (X'): [ka]

[0210] During the ceremony,

[0211] [ka] indicates the position of attachment to the biomolecule,

[0212] Q is independently H; [ka] and

[0213] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0214] L2 is a chemical bond or -CH2CH2C(O)-;

[0215] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0216] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0217] In the formula, a=0, 1, 2 or 3;

[0218] b=1, 2, 3, 4 or 5;

[0219] c=1, 2, 3, 4 or 5;

[0220] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0221] L is a chemical bond, -CHO- or -NHC(O)-;

[0222] L' is a chemical bond, -C(O)NH-, -NHC(O)- or -O(CH2CH2O) e - and

[0223] In the formula, e is 1, 2, 3, 4 or 5;

[0224] T is a chemical bond, -CH2-, -C(O)-, -M-, -CH2-M- or -C(O)-M-;

[0225] In the formula, M is [ka] and

[0226] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0227] Or, R1 and R3 are both -C 1~2alkylene- and R2 is H;

[0228] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0229] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0230] An N-acetylgalactosamine-containing targeting ligand, where n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0231] 2. The conjugated group is shown in formula (I'): [ka]

[0232] During the ceremony,

[0233] [ka] indicates the position of attachment to the biomolecule,

[0234] Q is independently H; [ka] and

[0235] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0236] L2 is a chemical bond or -CH2CH2C(O)-;

[0237] L3 is a chemical bond, -(NHCH2CH2)b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0238] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0239] In the formula, a=0, 1, 2 or 3;

[0240] b=1, 2, 3, 4 or 5;

[0241] c=1, 2, 3, 4 or 5;

[0242] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0243] L is -CH2O- or -NHC(O)-;

[0244] L' is a chemical bond, -C(O)NH- or -NHC(O)-;

[0245] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0246] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0247] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0248] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0249] The ligand according to technical solution 1, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0250] 3.Q is independently H or [ka] and

[0251] In the formula, L1 is -CH2O- or -NHC(O)-(CH2NHC(O)) a - and

[0252] L2 is -CH2CH2C(O)-,

[0253] L3 is -(NHCH2CH2) b -or-(NHCH2CH2CH2) b - and

[0254] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0255] In the formula, a=0, 1, 2 or 3;

[0256] b=1, 2, 3, 4 or 5;

[0257] c=1, 2, 3, 4 or 5;

[0258] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0259] L is -CHO-,

[0260] L' is a chemical bond;

[0261] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0262] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0263] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0264] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0265] The ligand according to technical solution 2, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0266] 4. The conjugated group is represented by formula (I'-1), formula (I'-2) or formula (I'-3), [ka]

[0267] During the ceremony,

[0268] [ka] indicates the position of attachment to the biomolecule,

[0269] Q is, [ka] and

[0270] In the formula, L1 is -CH2O- or -NHC(O)-;

[0271] L2 is -CH2CH2C(O)-,

[0272] L3 is -(NHCH2CH2) b -or-(NHCH2CH2CH2) b - and

[0273] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0274] Wherein b=1, 2, 3, 4 or 5;

[0275] c=1, 2, 3, 4 or 5;

[0276] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0277] L is -CHO-,

[0278] R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0279] The ligand according to technical solution 2, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0280] 5.Q is independent of H, [ka] and

[0281] In the formula, L1 is -CH2O-, -CH2O-CH2CH2O- or -NHC(O)-(CH2NHC(O)) a - and

[0282] L2 is a chemical bond,

[0283] L3 is -(NHCH2CH2) b -, -(NHCH2CH2CH2) b- or -C(O)CH2-,

[0284] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0285] In the formula, a=0, 1, 2 or 3;

[0286] b=1, 2, 3, 4 or 5;

[0287] c=1, 2, 3, 4 or 5;

[0288] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0289] L is -CH2O- or -NHC(O)-;

[0290] L' is a chemical bond or -C(O)NH-;

[0291] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0292] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0293] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0294] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0295] The ligand according to technical solution 2, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0296] 6. The conjugated group is represented by formula (II'-1) or formula (II'-2): [ka]

[0297] During the ceremony,

[0298] [ka] indicates the position of attachment to the biomolecule,

[0299] Q is independently [ka] and

[0300] In the formula, L1 is -CH2O- or -CH2O-CH2CH2O-;

[0301] L3 is -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0302] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0303] Wherein b=1, 2, 3, 4 or 5;

[0304] c=1, 2, 3, 4 or 5;

[0305] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0306] L is -NHC(O)-;

[0307] L' is a chemical bond or -C(O)NH-;

[0308] R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0309] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0310] The ligand according to technical solution 5, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0311] 7.Q is independently H, [ka] and

[0312] In the formula, L1 is -CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O- or -NHC(O)-(CH2NHC(O))a-;

[0313] L2 is a chemical bond,

[0314] L3 is -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0315] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0316] In the formula, a=0, 1, 2 or 3;

[0317] b=1, 2, 3, 4 or 5;

[0318] c=1, 2, 3, 4 or 5;

[0319] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0320] L is -CH2O- or -NHC(O)-;

[0321] L' is a chemical bond or -C(O)NH-;

[0322] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0323] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0324] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0325] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0326] The ligand according to technical solution 2, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0327] 8. The conjugated group is represented by the formula (II'-2): [ka]

[0328] During the ceremony,

[0329] [ka] indicates the position of attachment to the biomolecule,

[0330] Q is, [ka] and

[0331] L1 is -CH2O-,

[0332] L3 is -NHCH2CH2-,

[0333] L4 is -(OCH2CH2) c - and

[0334] c=1, 2 or 3;

[0335] L is -NHC(O)-;

[0336] R' is H,

[0337] The ligand according to technical solution 7, wherein n=6, 7, 8, 9 or 10, preferably 7, 8 or 9, more preferably 8.

[0338] 9. The conjugated group is represented by the formula (II'-2): [ka]

[0339] During the ceremony,

[0340] [ka] indicates the position of attachment to the biomolecule,

[0341] Q is independently [ka] and

[0342] In the formula, L1 is -CH2- or -C(O)-;

[0343] L3 is -(NHCH2CH2) b - and

[0344] L4 is -(OCH2CH2) c- and

[0345] Wherein b=1, 2, 3, 4 or 5;

[0346] c=1, 2, 3, 4 or 5;

[0347] L is -CH2O- or -NHC(O)-;

[0348] R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0349] The ligand according to technical solution 7, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0350] 10. The conjugated group is represented by the formula (II'-3): [ka]

[0351] During the ceremony,

[0352] [ka] indicates the position of attachment to the biomolecule,

[0353] Q1 is, [ka] wherein L1 is -CH2-;

[0354] Q2 is, [ka] wherein L1 is -C(O)-;

[0355] L3 is -NHCH2CH2-,

[0356] L4 is -(OCH2CH2) c - and

[0357] c=1, 2 or 3;

[0358] L is -NHC(O)-;

[0359] R' is H,

[0360] The ligand according to technical solution 9, wherein n=6, 7, 8, 9 or 10, preferably 7, 8 or 9, more preferably 8.

[0361] 11.Q is independently H, [ka] and

[0362] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0363] L2 is a chemical bond or -CH2CH2C(O)-;

[0364] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0365] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0366] In the formula, a=0, 1, 2 or 3;

[0367] b=1, 2, 3, 4 or 5;

[0368] c=1, 2, 3, 4 or 5;

[0369] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0370] L is a chemical bond, -CHO- or -NHC(O)-;

[0371] L' is a chemical bond, -C(O)NH-, -NHC(O)- or -O(CHCHO)e-;

[0372] In the formula, e is 1, 2, 3, 4 or 5;

[0373] T is a chemical bond, -CH2-, -M-, -CH2-M- or -C(O)-M-;

[0374] In the formula, M is [ka] and

[0375] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0376] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0377] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0378] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0379] The ligand according to technical solution 1, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0380] 12. T is -M-, -CH2-M- or -C(O)-M-, where M is [ka] The ligand according to technical solution 11,

[0381] 13. Q is independently H or [ka] and

[0382] In the formula, L1 is -CH2O- or -NHC(O)-(CH2NHC(O)) a - and

[0383] L2 is -CH2CH2C(O)-,

[0384] L3 is -(NHCH2CH2) b -or-(NHCH2CH2CH2) b - and

[0385] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0386] In the formula, a=0, 1, 2 or 3;

[0387] b=1, 2, 3, 4 or 5;

[0388] c=1, 2, 3, 4 or 5;

[0389] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0390] L is a chemical bond or -CH2O-;

[0391] L' is a chemical bond or -O(CH2CH2O) e - and

[0392] In the formula, e is 1, 2, 3, 4 or 5;

[0393] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0394] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0395] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0396] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0397] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0398] The ligand according to technical solution 11 or 12, wherein T is as defined in technical solution 11 or 12.

[0399] 14. The conjugated group is represented by formula (III'-1), formula (III'-2) or formula (III'-3), [ka]

[0400] During the ceremony,

[0401] Q is, [ka] and

[0402] In the formula, L1 is -CH2O- or -NHC(O)-;

[0403] L2 is -CH2CH2C(O)-,

[0404] L3 is -(NHCH2CH2) b -or-(NHCH2CH2CH2) b - and

[0405] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0406] Wherein b=1, 2, 3, 4 or 5;

[0407] c=1, 2, 3, 4 or 5;

[0408] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0409] L is a chemical bond or -CH2O-;

[0410] wherein R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0411] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0412] The ligand according to technical solution 13, wherein T is as defined in technical solution 11 or 12.

[0413] 15.Q is independently H, [ka] and

[0414] In the formula, L1 is -CH2-, -CH2O-, or -C(O)-;

[0415] L2 is a chemical bond,

[0416] L3 is -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0417] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0418] Wherein b=1, 2, 3, 4 or 5;

[0419] c=1, 2, 3, 4 or 5;

[0420] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0421] L is a chemical bond or -NHC(O)-;

[0422] L' is a chemical bond;

[0423] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0424] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0425] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0426] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0427] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0428] The ligand according to technical solution 11 or 12, wherein T is as defined in technical solution 11 or 12.

[0429] 16. The conjugated group is represented by formula (IV'-1) or formula (IV'-2): [ka]

[0430] During the ceremony,

[0431] Q is independently [ka] and

[0432] In the formula, L1 is -CH2-, -CH2O-, or -C(O)-;

[0433] L3 is -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0434] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0435] Wherein b=1, 2, 3, 4 or 5;

[0436] c=1, 2, 3, 4 or 5;

[0437] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0438] L is a chemical bond or -NHC(O)-;

[0439] L' is a chemical bond;

[0440] wherein R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0441] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0442] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0443] The ligand according to technical solution 11 or 12, wherein T is as defined in technical solution 11 or 12.

[0444] 17. The conjugated group is represented by the formula (II'-3): [ka]

[0445] During the ceremony,

[0446] [ka] indicates the position of attachment to the biomolecule,

[0447] Q1 is, [ka] wherein L1 is -CH2-;

[0448] Q2 is, [ka] where L1 is a chemical bond;

[0449] L3 is -NHCH2CH2-,

[0450] L4 is -(OCH2CH2)c-,

[0451] c=1, 2 or 3;

[0452] L is -NHC(O)-;

[0453] R' is H,

[0454] The ligand according to technical solution 1, wherein n=6, 7, 8, 9 or 10, preferably 7, 8 or 9, more preferably 8.

[0455] 18. Q is independently H, [ka] and

[0456] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0457] L2 is a chemical bond or -CH2CH2C(O)-;

[0458] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0459] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0460] In the formula, a=0, 1, 2 or 3;

[0461] b=1, 2, 3, 4 or 5;

[0462] c=1, 2, 3, 4 or 5;

[0463] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0464] L is a chemical bond, -CHO- or -NHC(O)-;

[0465] L' is -O(CH2CH2O) e - and

[0466] In the formula, e is 1, 2, 3, 4 or 5;

[0467] T is a chemical bond, -CH2-, -C(O)-, -M-, -CH2-M- or -C(O)-M-;

[0468] In the formula, M is [ka] and

[0469] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0470] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0471] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0472] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0473] The ligand according to technical solution 1, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0474] 19. The ligand according to technical solution 1, wherein the conjugated group is selected from the following: [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5]

[0475] 20. The ligand according to technical solution 1, wherein the conjugated group is selected from the following: [Table 2-1] [Table 2-2] [Table 2-3] [Table 2-4] [Table 2-5] [Table 2-6]

[0476] 21. A ligand according to any one of technical solutions 1 to 20, which targets ASGPR.

[0477] 22. A nucleic acid molecule, comprising a nucleic acid and a ligand according to any one of technical solutions 1 to 21 connected thereto.

[0478] 23. The nucleic acid molecule according to technical solution 22, wherein said nucleic acid is selected from DNA, RNA and DNA / RNA hybrids.

[0479] 24. The nucleic acid molecule according to technical solution 23, wherein the nucleic acid molecule is single-stranded or double-stranded.

[0480] 25. The nucleic acid molecule according to technical solution 23, wherein the nucleic acid molecule is selected from small interfering RNA (siRNA) and short hairpin RNA (shRNA).

[0481] 26. The nucleic acid molecule according to technical solution 23, wherein the nucleic acid molecule is a double-stranded RNA molecule, comprising a sense strand and an antisense strand, and the conjugated group is connected to the 3'-end or 5'-end, preferably the 3'-end, of the sense strand of the double-stranded RNA molecule.

[0482] 27. The nucleic acid molecule according to technical solution 26, wherein the sense strand and the antisense strand of the double-stranded RNA molecule each have 10-30, preferably 15-25, more preferably 20-25 nucleotides.

[0483] 28. The nucleic acid molecule according to technical solution 26 or 27, wherein one or more nucleotides of the sense strand and / or the antisense strand of said double-stranded RNA molecule comprise a chemical modification.

[0484] 29. The nucleic acid molecule according to technical solution 26 or 27, wherein one or more nucleotides in the sense strand and / or the antisense strand of said double-stranded RNA molecule are replaced with a nucleotide analogue.

[0485] 30. A nucleic acid molecule according to any one of technical solutions 22 to 29, which specifically binds to the asialoglycoprotein receptor (ASGPR) on the surface of liver cells.

[0486] 31. A pharmaceutical composition, comprising a nucleic acid molecule according to any one of technical solutions 22-30 and a pharma- ceutically acceptable carrier or excipient.

[0487] 32. A kit, comprising a nucleic acid molecule according to any one of technical solutions 22 to 30.

[0488] 33. A compound of formula (X) or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof, [ka]

[0489] During the ceremony,

[0490] Q is independently H; [ka] and

[0491] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0492] L2 is a chemical bond or -CH2CH2C(O)-;

[0493] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0494] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0495] In the formula, a=0, 1, 2 or 3;

[0496] b=1, 2, 3, 4 or 5;

[0497] c=1, 2, 3, 4 or 5;

[0498] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0499] L is a chemical bond, -CHO- or -NHC(O)-;

[0500] L' is a chemical bond, -C(O)NH-, -NHC(O)- or -O(CHCHO)e-;

[0501] In the formula, e is 1, 2, 3, 4 or 5;

[0502] T is a chemical bond, -CH2-, -C(O)-, -M-, -CH2-M- or -C(O)-M-;

[0503] In the formula, M is [ka] and

[0504] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0505] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0506] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0507] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0508] A compound of formula (X), or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0509] 34. A compound having the formula (I): [ka]

[0510] During the ceremony,

[0511] Q is independently H; [ka] and

[0512] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0513] L2 is a chemical bond or -CH2CH2C(O)-;

[0514] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0515] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0516] In the formula, a=0, 1, 2 or 3;

[0517] b=1, 2, 3, 4 or 5;

[0518] c=1, 2, 3, 4 or 5;

[0519] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0520] L is -CH2O- or -NHC(O)-;

[0521] L' is a chemical bond, -C(O)NH- or -NHC(O)-;

[0522] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0523] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0524] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0525] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0526] The compound according to technical solution 33, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0527] 35. Q is independently H or [ka] and

[0528] In the formula, L1 is -CH2O- or -NHC(O)-(CH2NHC(O)) a - and

[0529] L2 is -CH2CH2C(O)-,

[0530] L3 is -(NHCH2CH2) b -or-(NHCH2CH2CH2) b - and

[0531] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0532] In the formula, a=0, 1, 2 or 3;

[0533] b=1, 2, 3, 4 or 5;

[0534] c=1, 2, 3, 4 or 5;

[0535] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0536] L is -CHO-,

[0537] L' is a chemical bond;

[0538] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0539] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0540] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0541] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0542] The compound according to technical solution 34, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0543] 36. Having the formula (I-1), (I-2) or (I-3), [ka]

[0544] During the ceremony,

[0545] Q is, [ka] and

[0546] In the formula, L1 is -CH2O- or -NHC(O)-;

[0547] L2 is -CH2CH2C(O)-,

[0548] L3 is -(NHCH2CH2) b -or-(NHCH2CH2CH2) b - and

[0549] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0550] Wherein b=1, 2, 3, 4 or 5;

[0551] c=1, 2, 3, 4 or 5;

[0552] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0553] L is -CH2O-,

[0554] R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0555] The compound according to technical solution 34, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0556] 37. Q is independently H, [ka] and

[0557] In the formula, L1 is -CH2O-, -CH2O-CH2CH2O- or -NHC(O)-(CH2NHC(O)) a - and

[0558] L2 is a chemical bond,

[0559] L3 is -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0560] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0561] In the formula, a=0, 1, 2 or 3;

[0562] b=1, 2, 3, 4 or 5;

[0563] c=1, 2, 3, 4 or 5;

[0564] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0565] L is -CH2O- or -NHC(O)-;

[0566] L' is a chemical bond or -C(O)NH-;

[0567] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0568] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0569] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0570] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0571] The compound according to technical solution 34, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0572] 38. Having the formula (II-1) or (II-2), [ka]

[0573] During the ceremony,

[0574] Q is independently [ka] and

[0575] In the formula, L1 is -CH2O- or -CH2O-CH2CH2O-;

[0576] L3 is -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0577] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0578] Wherein b=1, 2, 3, 4 or 5;

[0579] c=1, 2, 3, 4 or 5;

[0580] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0581] L is -NHC(O)-;

[0582] L' is a chemical bond or -C(O)NH-;

[0583] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0584] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0585] The compound according to technical solution 37, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0586] 39. Q is independently H, [ka] and

[0587] In the formula, L1 is -CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0588] L2 is a chemical bond,

[0589] L3 is -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0590] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0591] In the formula, a=0, 1, 2 or 3;

[0592] b=1, 2, 3, 4 or 5;

[0593] c=1, 2, 3, 4 or 5;

[0594] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0595] L is -CH2O- or -NHC(O)-;

[0596] L' is a chemical bond or -C(O)NH-;

[0597] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0598] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0599] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0600] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0601] The compound according to technical solution 37, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0602] 40. Having the formula (II-2): [ka]

[0603] During the ceremony,

[0604] Q is, [ka] and

[0605] L1 is -CH2O-,

[0606] L3 is -NHCH2CH2-,

[0607] L4 is -(OCH2CH2) c - and

[0608] c=1, 2 or 3;

[0609] L is -NHC(O)-;

[0610] R' is H,

[0611] The compound according to technical solution 39, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof, wherein n=10-30, preferably 15-25, more preferably 8.

[0612] 41. Having the formula (II-2): [ka]

[0613] During the ceremony,

[0614] Q is independently [ka] and

[0615] In the formula, L1 is -CH2- or -C(O)-;

[0616] L3 is -(NHCH2CH2) b - and

[0617] L4 is -(OCH2CH2) c - and

[0618] Wherein b=1, 2, 3, 4 or 5;

[0619] c=1, 2, 3, 4 or 5;

[0620] L is -CH2O- or -NHC(O)-;

[0621] R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0622] The compound according to technical solution 39, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0623] 42. Having the formula (II-3), [ka]

[0624] During the ceremony,

[0625] Q1 is, [ka] wherein L1 is -CH2-;

[0626] Q2 is, [ka] wherein L1 is -C(O)-;

[0627] L3 is -NHCH2CH2-,

[0628] L 4 is -(OCH2CH2) c - and

[0629] c=1, 2 or 3;

[0630] L is -NHC(O)-;

[0631] R' is H,

[0632] The compound according to technical solution 41, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof, wherein n=10-30, preferably 15-25, more preferably 8.

[0633] 43. Q is independently H, [ka] and

[0634] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0635] L2 is a chemical bond or -CH2CH2C(O)-;

[0636] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0637] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0638] In the formula, a=0, 1, 2 or 3;

[0639] b=1, 2, 3, 4 or 5;

[0640] c=1, 2, 3, 4 or 5;

[0641] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0642] L is a chemical bond, -CHO- or -NHC(O)-;

[0643] L' is a chemical bond, -C(O)NH-, -NHC(O)- or -O(CHCHO)e-;

[0644] In the formula, e is 1, 2, 3, 4 or 5;

[0645] T is a chemical bond, -CH2-, -M-, -CH2-M- or -C(O)-M-;

[0646] In the formula, M is [ka] and

[0647] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0648] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0649] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0650] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0651] The compound according to technical solution 33, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0652] 44. T is -M-, -CH2-M- or -C(O)-M-, where M is [ka] or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0653] 45.Q is independently H or [ka] and

[0654] In the formula, L1 is -CH2O- or -NHC(O)-(CH2NHC(O)) a - and

[0655] L2 is -CH2CH2C(O)-,

[0656] L3 is -(NHCH2CH2) b -or-(NHCH2CH2CH2) b - and

[0657] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0658] In the formula, a=0, 1, 2 or 3;

[0659] b=1, 2, 3, 4 or 5;

[0660] c=1, 2, 3, 4 or 5;

[0661] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0662] L is a chemical bond or -CH2O-;

[0663] L' is a chemical bond or -O(CH2CH2O) e - and

[0664] In the formula, e is 1, 2, 3, 4 or 5;

[0665] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0666] or R1 and R3 together form -C1-2 alkylene- and R2 is H;

[0667] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0668] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0669] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0670] T is as defined in technical solution 43 or 44, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0671] 46. ​​Having the formula (III-1), (III-2) or (III-3), [ka]

[0672] During the ceremony,

[0673] Q is, [ka] and

[0674] In the formula, L1 is -CH2O- or -NHC(O)-;

[0675] L2 is -CH2CH2C(O)-,

[0676] L3 is -(NHCH2CH2) b -or-(NHCH2CH2CH2) b - and

[0677] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0678] Wherein b=1, 2, 3, 4 or 5;

[0679] c=1, 2, 3, 4 or 5;

[0680] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0681] L is a chemical bond or -CH2O-;

[0682] R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0683] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0684] T is as defined in technical solution 43 or 44, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0685] 47.Q is independently H, [ka] and

[0686] In the formula, L1 is -CH2-, -CH2O-, or -C(O)-;

[0687] L2 is a chemical bond,

[0688] L3 is -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0689] L4 is -(OCH2CH2) c- or -NHC(O)-(CH d - and

[0690] Wherein b=1, 2, 3, 4 or 5;

[0691] c=1, 2, 3, 4 or 5;

[0692] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0693] L is a chemical bond or -NHC(O)-;

[0694] L' is a chemical bond;

[0695] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0696] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0697] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0698] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0699] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0700] T is as defined in technical solution 43 or 44, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0701] 48. Having the formula (IV-1) or (IV-2), [ka]

[0702] During the ceremony,

[0703] Q is independently [ka] and

[0704] In the formula, L1 is -CH2-, -CH2O-, or -C(O)-;

[0705] L3 is -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0706] L4 is -(OCH2CH2) c - or -NHC(O)-(CH d - and

[0707] Wherein b=1, 2, 3, 4 or 5;

[0708] c=1, 2, 3, 4 or 5;

[0709] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0710] L is a chemical bond or -NHC(O)-;

[0711] L' is a chemical bond;

[0712] R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0713] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0714] n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0715] T is as defined in technical solution 43 or 44, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0716] 49. Having the formula (II-3): [ka]

[0717] During the ceremony,

[0718] Q1 is, [ka] wherein L1 is -CH2-;

[0719] Q2 is, [ka] where L1 is a chemical bond;

[0720] L3 is -NHCH2CH2-,

[0721] L4 is -(OCH2CH2) c - and

[0722] c=1, 2 or 3;

[0723] L is -NHC(O)-;

[0724] R' is H,

[0725] The compound according to technical solution 33, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof, wherein n=10-30, preferably 15-25, more preferably 8.

[0726] 50.Q is independently H, [ka] and

[0727] In the formula, L1 is a chemical bond, -CH2-, -CH2CH2-, -C(O)-, -CH2O-, -CH2O-CH2CH2O-, or -NHC(O)-(CH2NHC(O)) a - and

[0728] L2 is a chemical bond or -CH2CH2C(O)-;

[0729] L3 is a chemical bond, -(NHCH2CH2) b -, -(NHCH2CH2CH2) b - or -C(O)CH2-,

[0730] L4 is -(OCH2CH2) c -, -(OCH2CH2CH2) c -, -(OCH2CH2CH2CH2) c -, -(OCH2CH2CH2CH2CH2) c - or -NHC(O)-(CH d - and

[0731] In the formula, a=0, 1, 2 or 3;

[0732] b=1, 2, 3, 4 or 5;

[0733] c=1, 2, 3, 4 or 5;

[0734] d=1, 2, 3, 4, 5, 6, 7 or 8;

[0735] L is a chemical bond, -CHO- or -NHC(O)-;

[0736] L' is -O(CH2CH2O) e - and

[0737] In the formula, e is 1, 2, 3, 4 or 5;

[0738] T is a chemical bond, -CH2-, -C(O)-, -M-, -CH2-M- or -C(O)-M-;

[0739] In the formula, M is [ka] and

[0740] R1 and R2 together form -CH2CH2O- or -CH2CH(R)-O-, and R3 is H;

[0741] Or, R1 and R3 are both -C 1~2 alkylene- and R2 is H;

[0742] wherein R is -OR', -CHOR' or -CHCHOR', R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CHCHC(O)OH or 4,4'-dimethoxytrityl;

[0743] m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;

[0744] The compound according to technical solution 33, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0745] 51. The compound is selected from: [Table 3-1] [Table 3-2] [Table 3-3] [Table 3-4] [Table 3-5]

[0746] wherein R' is -C(O)CH2CH2C(O)OH or 4,4'-dimethoxytrityl, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof.

[0747] 52. The compound is a compound according to technical solution 33, or a pharma- ceutically acceptable salt, stereoisomer or isotopic variant thereof, selected from the following: [Table 4-1] [Table 4-2] [Table 4-3] [Table 4-4] [Table 4-5] [Table 4-6]

[0748] 53. An intermediate compound, which is a compound of formula (V): [ka]

[0749] During the ceremony,

[0750] Pg is a hydroxyl protecting group selected, for example, from [ka]

[0751] Q' is independently H; [ka] and

[0752] The intermediate compound, wherein the other groups are as defined in any one of technical solutions 33-50.

[0753] 54. An intermediate compound selected from the following: [Table 5-1] [Table 5-2] [Table 5-3] [Table 5-4] [Table 5-5]

[0754] 55. An intermediate compound selected from the following: [Table 6-1] [Table 6-2] [Table 6-3] [Table 6-4] [Table 6-5] [Table 6-6]

[0755] 56. A process for preparing a compound of formula (I), comprising treating a compound of formula (V) under conditions to remove a hydroxyl protecting group; [ka]

[0756] The method for preparing a compound of formula (I), wherein the other groups are as defined in any one of technical solutions 33-50.

[0757] (Example) The following examples are intended to illustrate the invention and are not intended to limit the scope of the invention.

[0758] CbzCl: benzyl chloroformate

[0759] CbzOSu: N-benzyloxycarbonyloxysuccinimide

[0760] DCI: dicyclohexylcarbodiimide

[0761] DIEA / DIPEA: Diisopropylethylamine

[0762] DMAP: Dimethylaminopyridine

[0763] EDCl: 1-Ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride

[0764] HATU: 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylureonium hexafluorophosphate

[0765] HBTU: O-benzotriazole-tetramethylureonium hexafluorophosphate

[0766] HOBt: 1-hydroxybenzotriazole

[0767] TEBA: Benzyltriethylammonium bromide

[0768] TEBAC: Triethylbenzylammonium chloride

[0769] TMSOTf: Trimethylsilyl triflate

[0770] Sequence information used in the examples: [Table 7-1] [Table 7-2] [Table 7-3] [Table 7-4]

[0771] In this specification, the meanings of each abbreviation are as follows:

[0772] The distribution of A, U, G and C indicates the natural adenine, uracil, guanine and cytosine ribonucleotides.

[0773] m indicates that the adjacent nucleotide to its left is a 2'-OCH3 modified nucleotide, for example, Am, Um, Gm, and Cm indicate 2'-OCH3 modified A, U, G, and C.

[0774] f indicates that the adjacent nucleotide to its left is a 2-F modified nucleotide, for example, Af, Uf, Gf, and Cf indicate 2'-F modified A, U, G, and C, respectively.

[0775] "s" or "s-" means that two adjacent nucleotides and / or delivery vectors are joined via a phosphorothioate.

[0776] L96 represents a GalNAc delivery vector known in the art, of the following structure: [ka] indicates the position of attachment to the siRNA via a phosphate or thiophosphate group; see, e.g., PCT Publication Nos. WO2009073809 and WO2009082607. [ka]

[0777] NAG37 represents a GalNAc delivery vector known in the art and of the following structure: [ka] indicates the position of attachment to the siRNA via a phosphate or thiophosphate group, see, e.g., PCT Publication No. WO2018044350. [ka]

[0778] IB stands for reverse base deoxyribonucleotide, and can include the following three structures depending on the position / connection method in siRNA. IB is well known in the art, see, for example, F. Czauderna, Nucleic Acids Res., 2003, 31(11), 2705-16 and PCT publication numbers WO2016011123 and WO2019051402. [ka]

[0779] The structures of GL1 to GL32 are as defined above.

[0780] Example 1 Preparation of Compound E1 [ka]

[0781] 1. Preparation of Compound 3 [ka] Compound 1 (100 g, 734 mmol) is added to water (176.28 mL) in which sodium hydroxide (176.28 g, 4.40 mol) is dissolved at 25 ° C., then TEBAC (83.65 g, 367 mmol) and compound 2 (564.83 g, 4.407 mol) are added, and after three nitrogen purges, the reaction solution is stirred at 25 ° C. for 18 h. Thin layer chromatography (dichloromethane / methanol = 10 / 1) shows the complete consumption of compound 1, and thin layer chromatography (petroleum ether / ethyl acetate = 5 / 1) shows the formation of a new product spot. The reaction solution is diluted with 1000 mL of ice water, extracted with ethyl acetate (500 mL × 3), the organic phase is dried over anhydrous sodium sulfate, and concentrated in vacuum to obtain the crude product, which is purified by column chromatography (petroleum ether / ethyl acetate = 1 / 0 to 10 / 1) to obtain compound 3 (about 56 g) as a colorless oily liquid. 1 H NMR (400MHz, CD3OD) δ3.59(t, J=6.4Hz, 8H), 3.36(s, 8H), 2.43(t, J=6.4Hz, 8H), 1.47(s, 36H).

[0782] 2. Preparation of Compound 4 [ka] Dissolve compound 3 (56 g, 87.372 mmol) in DCM (400 mL) at 25 °C, slowly add TFA (100 mL) dropwise, and stir the reaction solution at 25 °C for 18 h. Thin layer chromatography (petroleum ether / ethyl acetate = 5 / 1) shows complete consumption of the reactants and the formation of one new spot. Concentrate the reaction solution to dryness to obtain the crude pink oily liquid compound (about 20 g). 1 H NMR (400MHz, CD3OD) δ3.63(t, J=6.0Hz, 8H), 3.36(s, 8H), 2.51(t, J=6.4Hz, 8H).

[0783] 3 Preparation of compound 5 [ka] Compound 4 (4.2 g, 9.896 mmol) is dissolved in DMF (21 mL) at 25 ° C., TEA (1.376 mL, 9.896 mmol) and compound 4A (3.46 g, 9.896 mmol) are added in sequence, and the mixture is heated to 83 ° C. The reaction solution is stirred under nitrogen atmosphere at 83 ° C. for 24 hours. Thin layer chromatography (dichloromethane / methanol = 8 / 1) shows the complete consumption of compound 4 and the formation of a new spot. LCMS shows the MS value of compound 5. The reaction solution is concentrated in vacuum to obtain the crude product, which is purified by MPLC (dichloromethane / methanol = 1 / 0 to 10 / 1) to obtain compound 5 (about 6.6 g) as a yellow oily liquid. 1 H NMR (400MHz, CDCl3) δ7.32-7.37(m, 5H), 5.14(s, 2H), 3.60-3.70(m, 8H), 3.35(s, 8H), 2.53-2.65(m, 8H). LCMS:m / z=515.0(M+H) + .

[0784] 4 Preparation of compound 6 [ka] Compound 5 (6.6 g, 12.827 mmol) is dissolved in DCM (200 mL) at 25 °C, then compound 5A (11.81 g, 64.137 mmol) and EDCI (12.30 g, 64.137 mmol) are added sequentially, and the reaction solution is heated at 25 °C for 6 h. Thin layer chromatography (dichloromethane / methanol = 8 / 1) shows the complete consumption of compound 1, and thin layer chromatography (petroleum ether / ethyl acetate = 5 / 1) shows the formation of a new product spot. LCMS shows the MS value of compound 6. The reaction solution is concentrated in vacuum to obtain the crude product, which is purified by MPLC (petroleum ether / ethyl acetate = 1 / 0 to 5 / 1) to obtain compound 6 (about 4.9 g) as a colorless oily liquid. 1 H NMR (400MHz, CDCl3) δ7.32-7.37(m, 5H), 5.14(s, 2H), 3.74(t, J=6.0Hz, 6H), 3.68 (t, J=6.4Hz, 2H), 3.36-3.45(m, 8H), 2.87(t, J=6.0Hz, 6H), 2.62(t, J=6.4Hz, 2H). LCMS:m / z=1013.2(M+H) + .

[0785] 5. Preparation of Compound 7 [ka] Compound 6 (3.0 g, 2.96 mmol) and compound 6A (6.81 g, 11.8 mmol, prepared according to the method of intermediate 3-4 in Example 23) are dissolved in DCM (50.0 mL), DIEA (3.91 mL, 23.7 mmol) is added, nitrogen is purged three times, and the reaction solution is reacted at 20° C. for 16 hours. Thin layer chromatography (first development, dichloromethane:methanol=10 / 1, second development, petroleum ether:ethyl acetate=3:1) shows the disappearance of raw materials and the formation of a new spot. The reaction solution is diluted with 150 ml of dichloromethane, washed with aqueous citric acid solution (50.0 ml x 3), saturated aqueous sodium bicarbonate solution (50.0 ml x 3), and saturated saline solution (50.0 ml x 3). The organic phase is dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (dichloromethane:methanol = 100:1 to 8:1) to obtain a white solid product, compound 7 (about 400 mg). 1 H NMR (400MHz, CD3OD) δ7.29-7.39(m, 5H), 5.31-5.36(m, 3H), 5.16(s, 2H), 5.05- 5.09(m, 3H), 4.61-4.68(m, 3H), 4.06-4.19(m, 10H), 4.00-4.05(m, 3H), 3.90-3 .96(m, 3H), 3.58-3.76(m, 33H), 3.51-3.57(m, 7H), 3.35-3.41(m, 8H), 2.58-2. 65(m, 2H), 2.42(t, J=6.0Hz, 6H), 2.14(s, 9H), 2.03(s, 9H), 1.92-1.96(m, 18H)

[0786] 6 Preparation of compound 8 [ka] Compound 7 (5.00 g, 2.63 mmol) is dissolved in MeOH (60.0 mL) at 20 °C, 10% mass fraction Pd / C (1.50 g) is added, and the mixture is purged with H2 three times and stirred at 20 °C and 15 psi for 14 h. Thin layer chromatography (dichloromethane:methanol = 8 / 1) shows the disappearance of the raw material and the formation of a new spot. The reaction solution is filtered through Celite, and the filtrate is concentrated under reduced pressure to obtain compound 8 (approximately 4.40 g) as a white solid. 1H NMR (400MHz, CD3OD) δ5.34(d, J=2.8Hz, 3H), 5.04-5.11(m, 3H), 4.62-4.67(m, 3H), 4.08-4.20(m, 9H), 4.00-4.08(m, 4H), 3.91-3.98(m, 3H), 3. 59-3.77(m, 33H), 3.53-3.59(m, 7H), 3.37-3.40(m, 8H), 2.49-2.53(m, 2 H), 2.43(t, J=6.0Hz, 6H), 2.15(s, 9H), 2.03(s, 9H), 1.93-1.97(m, 18H)

[0787] 7 Preparation of Compound 10 [ka] Compound 8 (1.50 g, 0.831 mmol), HATU (0.470 g, 1.24 mmol) and DIEA (0.412 mL, 2.492 mmol) are dissolved in anhydrous DMF (15.0 mL) at 20 ° C., nitrogen replacement is performed three times, and the mixture is stirred at 20 ° C. for 0.5 hours. Compound 9 (0.130 g, 1.246 mmol) is added and the mixture is stirred at 20 ° C. for 2 hours. LC / MS shows that all raw materials have disappeared and a main peak has been generated. Thin layer chromatography (dichloromethane: methanol = 8 / 1) shows the disappearance of raw materials and the generation of a new spot. The reaction solution is diluted with dichloromethane (100 mL), washed with saturated sodium bicarbonate aqueous solution (20.0 mL × 3) and saturated saline (20.0 mL × 3) in that order, and the organic phase is dried over anhydrous sodium sulfate, and then concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography (dichloromethane:methanol = 10 / 1 to 8 / 1) to obtain a brown solid product, compound 10 (about 1.00 g). 1H NMR (400MHz, CD3OD) δ5.31-5.35(m, 3H), 5.04-5.10(m, 3H), 4.63-4.67(m, 3H), 4.08 -4.21(m, 9H), 4.01-4.08(m, 4H), 3.91-3.98(m, 3H), 3.83-3.90(m, 2H), 3.69-3.77(m , 4H), 3.59-3.69(m, 27H), 3.53-3.58(m, 6H), 3.36-3.41(m, 15H), 2.62-2.68(m, 2H), 2.40-2.46(m, 6H), 2.15(s, 9H), 2.03(s, 9H), 1.80-1.98(m, 20H), 1.35-1.53(m, 2H). LCMS:m / z=945.5(M / 2+H) + .

[0788] 8. Preparation of E1 [ka] Compound 10 (1.00 g, 0.529 mmol) is dissolved in acetonitrile and centrifuged to remove water in the compound, then dissolved in DCM (10.0 mL) under nitrogen atmosphere, DCI (0.03 g, 0.265 mmol) is added, compound 11 (0.32 g, 1.06 mmol) is added dropwise to the above solution, and stirred at 20 ° C for 1 h. LCMS shows disappearance of raw materials and 40.0% product. The reaction solution is cooled to 0 ° C, 6 ml of saturated aqueous sodium bicarbonate solution and 6 ml of saturated saline solution are added dropwise, diluted with 150 ml of dichloromethane, separated, and washed with 50 ml of a 1:1 mixed aqueous solution of saturated aqueous sodium bicarbonate and saturated saline. The organic phase is dried over anhydrous sodium sulfate, filtered, and centrifuged to obtain the crude product. The crude product is diluted with 15 ml of dichloromethane, slowly dripped into 60 ml of methyl tert-butyl ether, filtered, the filter cake is washed with methyl tert-butyl ether three times, the filter cake is dissolved in acetonitrile, and then centrifuged to remove the methyl tert-butyl ether in the compound. A yellow solid E1 (615 mg, 0.252 mmol, 47.6%) is obtained.

[0789] LCMS(ESI):m / z=1103.3((M+59*2) / 2+H)+ ,

[0790] 1 H NMR (400MHz, DMSO-d6) δ7.86-7.93 (m, 3H), 7.78-7.83 (m, 3H), 5.19-5.23 (m, 3H), 4.97 (dd, J=3.2, 11.2Hz, 3H ), 4.51-4.57(m, 3H), 3.99-4.06(m, 10H), 3.84-3.92(m, 3H), 3.75-3.82(m, 3H), 3.62-3.75(m, 4H), 3.45-3.6 2(m, 34H), 3.37-3.41(m, 6H), 3.17-3.23(m, 13H), 2.76(t, J=6.0Hz, 2H), 2.53-2.57(m, 2H), 2.28(t, J=6.4Hz , 6H), 2.12(s, 9H), 2.00(s, 9H), 1.9-1.89(s, 9H), 1.71-1.82(m, 11H), 1.38-1.60(m, 2H), 1.09-1.17(m, 12H).

[0791] 31 P NMR (400MHz, DMSO-d6) δ-145.22.

[0792] Example 2 Preparation of Compound E1

change

[0793] 1 Preparation of compound 2

change

[0794] 2. Preparation of Compound 3 [ka] Compound 2 (4.10 g, 2.33 mmol) is dissolved in MeOH (100 mL) at room temperature, and 10% mass fraction of wet Pd / C (0.50 g, 4.70 mmol) is added to the solution, hydrogen is replaced three times, and the reaction mixture is stirred under hydrogen atmosphere (14.696 psi) at 25° C. for 18 hours. The MS response of the product is detected by liquid chromatography / mass spectrometry, and thin layer chromatography (dichloromethane / methanol=10 / 1, phosphomolybdic acid coloring) shows complete consumption of raw materials and the formation of a new spot. The reaction solution is filtered, and the filtrate is concentrated under reduced pressure to obtain a white solid 3 (about 3.87 g). 1 H NMR (400MHz, CD3OD) δ5.30-5.36(m, 3H), 5.07-5.09(m, 3H), 4.60-4.66(m, 3 H), 4.06-4.22(m, 9H), 4.01-4.06(m, 3H), 3.92-3.96(m, 3H), 3.69-3.75(m, 3 H), 3.61-3.67(m, 14H), 3.48-3.60(m, 7H), 3.32-3.45(m, 16H), 2.49-2.51(m , 2H), 2.43-2.47(m, 6H), 2.15(s, 9H), 2.03(s, 9H), 1.95(d, J=5.20Hz, 18H). LCMS:m / z=837.9(M / 2+H) + .

[0795] 3 Preparation of compound 5 [ka] At 25 °C, compound 3 (1.54 g, 0.920 mmol) is dissolved in DMF (15.0 mL), HATU (0.52 g, 1.38 mmol) and DIEA (0.456 mL, 2.76 mmol) are added in sequence, the reaction solution is stirred at 25 °C for 30 min, then compound 4 is added, and the reaction solution is stirred at 25 °C for 2.5 h. LCMS shows the complete consumption of compound 3, the presence of MS value of compound 5. The reaction solution is diluted with 50 mL of DCM, washed with saturated NaHCO3 solution (20 mL × 3), then the organic phase is washed with saturated saline solution (20 mL × 3), followed by drying with anhydrous Na2SO4, and the organic phase is centrifuged to obtain the crude product. The obtained crude product is purified by column chromatography (dichloromethane / methanol = 1 / 0 to 5 / 1) to obtain compound 5 (about 1.26 g) as a white solid. 1 H NMR (400MHz, CD3OD) δ5.32-5.36(m, 3H), 5.06(dd, J=3.2, 11.2Hz, 3H), 4.61-4.66(m , 3H), 4.06-4.20(m, 9H), 4.01-4.05(m, 3H), 3.91-3.97(m, 3H), 3.70-3.76(m, 3H), 3. 60-3.67(m, 14H), 3.50-3.57(m, 7H), 3.33-3.40(m, 18H), 2.66(t, J=6.0Hz, 2H), 2.4 4(t, J=6.0Hz, 6H), 2.15(s, 9H), 2.03(s, 9H), 1.92-1.98(m, 18H), 1.27-1.50(m, 4H). LCMS:m / z=879.5(M / 2+H) + .

[0796] 4. Preparation of E2 [ka] Compound 5 (1.26 g, 0.717 mmol) is dissolved in DCM (10.0 mL) at room temperature, compound 6 (0.43 g, 1.43 mmol) and DCI (0.08 g, 0.717 mmol) are added to the solution, nitrogen replacement is performed three times, and the reaction mixture is stirred at 25 ° C for 1 hour. The MS response of the product is detected by liquid chromatography / mass spectrometry, the reaction solution is cooled to 0 ° C in an ice salt bath, 6 mL of saturated NaHCO3 aqueous solution and 6 mL of saturated saline are added to the reaction solution, 50 mL of DCM is added to extract, the organic phase is washed with a 1:1 mixed solution of saturated NaHCO3 and saturated saline (20 mL × 3), the organic phase is then dried with Na2SO4, and the organic phase is centrifuged to obtain the crude product. Dissolve the crude product in 15 mL of DCM, slowly add the crude product solution dropwise to MTBE in a dry ice bath, and filter to obtain a white solid E2 (640 mg, 45.60% yield, 77.02% purity).

[0797] 1 H NMR (400MHz, DMSO-d6) δ7.79-7.91(m, 6H), 5.20-5.23(m, 3H), 4.96-4.98(m, 3H) , 4.51-4.56(m, 3H), 4.03(s, 10H), 3.65-3.92(m, 11H), 3.46-3.61(m, 21H), 3.34- 3.42(m, 8H), 3.19-3.22(m, 15H), 2.74-2.76(m, 2H), 2.52-2.54(m, 2H), 2.27-2.2 9(m, 6H), 2.10(s, 9H), 2.00(s, 9H), 1.89(s, 9H), 1.77(s, 9H), 1.12-1.16(m, 12H)

[0798] 31 P NMR (400MHz, DMSO-d6)δ-145.243

[0799] LCMS: m / z=1037.4 (M / 2+CH3COOH) + .

[0800] Example 3 Preparation of compound E3 [ka]

[0801] 1. Preparation of Compound 3 [ka] Compound 2 (0.17 g, 1.43 mmol) is dissolved in DMF (10.0 mL) at room temperature, and compound 1 (1.20 g, 0.717 mmol, prepared according to the method of compound 3 in Example 2), DIEA (0.178 mL, 1.08 mmol), and HATU (0.41 g, 1.08 mmol) are added to the solution, and nitrogen substitution is performed three times. The reaction mixture is stirred at 25 ° C for 1 hour. The MS response of the product is detected by liquid chromatography / mass spectrometry. Thin layer chromatography (dichloromethane / methanol = 8 / 1) shows the complete disappearance of the raw material and the formation of a new spot. The reaction solution is concentrated under reduced pressure, and the obtained crude product is purified by column chromatography (dichloromethane / methanol = 10 / 1 to 8 / 1) to obtain a white solid 3 (0.96 g). LCMS: m / z = 887.4 (M / 2 + H) + .

[0802] 2. Preparation of E3 [ka] Compound 3 (0.96 g, 0.542 mmol) is dissolved in DCM (10.0 mL) at room temperature, compound 4 (0.33 g, 1.08 mmol) and DCI (0.03 g, 0.271 mmol) are added to the solution, nitrogen replacement is performed three times, and the reaction mixture is stirred at 25 ° C for 1 hour. The MS response of the product is detected by liquid chromatography / mass spectrometry. The reaction solution is cooled to 0 ° C in an ice-salt bath, 6 mL of saturated NaHCO3 aqueous solution and 6 mL of saturated saline are added to the reaction solution, and 50 mL of DCM is added sequentially to extract, the organic phase is washed with a 1:1 mixed solution of saturated NaHCO3 and saturated saline (20 mL × 3), the organic phase is dried with Na2SO4, and the organic phase is centrifuged to obtain the crude product. Dissolve the crude product in 15 mL of DCM, slowly add the crude product solution dropwise to MTBE in a dry ice bath, and filter to obtain a white solid E3 (520 mg, 48.67% yield, 85.29% purity).

[0803] LCMS(ESI):m / z=1045.3(M / 2+CH3COOH) + ,

[0804] 1 H NMR (400MHz, DMSO-d6) δ7.75-7.92(m, 6H), 5.19-5.22(m, 3H), 4.96-4.99(m, 3H), 4. 52-4.57(m, 3H), 4.01-4.04(m, 10H), 3.73-3.92(m, 11H), 3.46-3.61(m, 21H), 3.37- 3.41(m, 8H), 3.19-3.22(m, 15H), 2.75-2.78(m, 3H), 2.27-2.32(m, 6H), 2.06-2.13( m, 11H), 1.99-2.02 (m, 9H), 1.86-1.90 (m, 9H), 1.76-1.79 (m, 9H), 1.12-1.15 (m, 12H)

[0805] 31 P NMR(400MHz, DMSO-d6)δ-147.960

[0806] Example 4 Preparation of Compound E4

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[0807] 1 Preparation of compound 3

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[0808] 2. Preparation of E4 [ka] Compound 3 (1.00 g, 0.525 mmol) is dissolved in acetonitrile, then centrifuged to remove water in the compound, then dissolved in DCM (10.0 mL) under nitrogen atmosphere, compound 4 (0.32 g, 1.05 mmol) and DCI (0.03 g, 0.262 mmol) are slowly added dropwise, and reacted at 20 ° C for 1 hour. LCMS shows disappearance of raw materials, the reaction solution is cooled to 0 ° C, 6 ml of saturated sodium bicarbonate aqueous solution and 6 ml of saturated saline are added dropwise, diluted with 100 ml of dichloromethane, separated, and washed with 50 ml of a 1:1 mixed aqueous solution of saturated sodium bicarbonate aqueous solution and saturated saline. The organic phase is dried over anhydrous sodium sulfate, filtered, and centrifuged to obtain the crude product. The crude product is diluted with 15 ml of dichloromethane, slowly dropped into 60 ml of methyl tert-butyl ether, filtered, the filter cake is washed with methyl tert-butyl ether three times, the filter cake is dissolved in acetonitrile, and then centrifuged to remove the methyl tert-butyl ether in the compound. A yellow solid product E4 (712 mg, 0.339 mmol, 64.4%) is obtained.

[0809] LCMS(ESI):m / z=1111.5((M+59*2) / 2+H) + ,

[0810] 1 H NMR (400MHz, DMSO-d6) δ7.86-7.94(m, 3H), 7.76-7.84(m, 3H), 5.19-5.26(m, 3H), 4.92-5.00 (m, 3H), 4.51-4.57(m, 3H), 3.98-4.08(m, 10H), 3.82-3.93(m, 5H), 3.73-3.81(m, 5H), 3.45-3 .63(m, 36H), 3.38-3.41(m, 6H), 3.15-3.26(m, 15H), 2.75-2.83(m, 2H), 2.52-2.54(m, 2H), 2. 28(t, J=6.4Hz, 6H), 2.10(s, 9H), 2.00(s, 9H), 1.89(s, 9H), 1.77(s, 9H), 1.07-1.17(m, 12H).

[0811] 31P NMR (400MHz, DMSO-d6)δ -147.928, -148.145

[0812] Example 5 Preparation of Compound E5 [ka]

[0813] 1. Preparation of Compound 2 [ka] At 25°C, compound 1 (10g, 22.349mmol, prepared according to the method of intermediate 1-3 in Example 25) is dissolved in dichloromethane (300mL), compound 2A (4.67g, 26.819mmol), HOBt (3.62g, 26.819mmol), EDCI (5.14g, 26.819mmol) and DIEA (11.082mL, 67.048mmol) are added in sequence, and the reaction solution is stirred at 25°C for 18 hours. Thin layer chromatography (dichloromethane / methanol=10 / 1) shows the complete consumption of compound 1 and the formation of a new spot. LCMS shows the MS value of compound 2. The reaction solution is diluted with 300 mL of dichloromethane, washed with saturated aqueous citric acid solution (30 mL×3), the organic phase is washed with saturated sodium bicarbonate solution (50 mL×3), and then the organic phase is dried over anhydrous sodium sulfate, and the organic phase is centrifuged to obtain compound 2 (about 11.5 g) as a white solid. 1 H NMR (400MHz, CD3OD) δ5.33-5.37(m, 1H), 5.07(dd, J=3.2, 11.2Hz, 1H), 4.54 -4.62(m, 1H), 4.11-4.26(m, 3H), 4.00-4.06(m, 1H), 3.85-3.93(m, 1H), 3.48 -3.60(m, 1H), 3.18-3.24(m, 2H), 3.05-3.11(m, 2H), 2.20(t, J=7.2Hz, 2H), 2 .16(s, 3H), 2.04(s, 3H), 1.92-1.98(m, 6H), 1.55-1.75(m, 6H), 1.45(s, 9H). LCMS:m / z=604.1(M+H) + .

[0814] 2. Preparation of Compound 3 [ka] Dissolve compound 2 (10 g, 16.565 mmol) in dichloromethane (100 mL) at 25 °C, slowly add TFA (10 mL) dropwise, and stir the reaction solution at 25 °C for 18 h. Thin layer chromatography (dichloromethane / methanol = 10 / 1) shows complete consumption of the reactants and the formation of a new spot. Concentrate the reaction solution to dryness to obtain crude compound 3 (about 20 g) as a yellow oily liquid. 1 H NMR (400MHz, CD3OD) δ5.32-5.36(m, 1H), 5.04(dd, J=3.2, 11.2Hz, 1H), 4.50-4 .55(m, 1H), 3.97-4.16(m, 4H), 3.85-3.92(m, 1H), 3.48-3.57(m, 1H), 3.26-3. 30(m, 2H), 2.90-2.98(m, 2H), 2.24(t, J=7.2Hz, 2H), 2.14(s, 3H), 2.02(s, 3H) , 1.92-1.96(m, 6H), 1.80-1.88(m, 2H), 1.63-1.73(m, 2H), 1.57-1.62(m, 2H).

[0815] 3 Preparation of compound 5 [ka] Compound 4 (4.37 g, 4.315 mmol, prepared according to the method of compound 6 in Example 1) is dissolved in dichloromethane (150 mL) at 25 ° C., compound 3 (8.7 g, 17.277 mmol) and DIEA (6 mL, 36.302 mmol) are added in sequence, and the reaction solution is stirred at 25 ° C. for 18 hours. Thin layer chromatography (dichloromethane / methanol = 10 / 1) shows the complete consumption of compound 4, and thin layer chromatography (petroleum ether / ethyl acetate = 5 / 1) shows the formation of a new product spot. LCMS shows the MS value of compound 5. The reaction solution is diluted with 100 mL of dichloromethane, washed with saturated aqueous citric acid solution (30 mL x 3), the organic phase is washed with saturated sodium bicarbonate solution (30 mL x 3), then the organic phase is dried over anhydrous sodium sulfate, and the organic phase is centrifuged to obtain the crude product. The crude product was purified by MPLC (petroleum ether / ethyl acetate = 1 / 0 to 10 / 1) to obtain compound 5 (approximately 7.1 g) as a white solid. 1 H NMR (400MHz, CD3OD) δ7.30-7.40(m, 5H), 5.31-5.36(m, 3H), 5.15(s, 2H), 5.06(dd, J=3.2, 11.2Hz, 3H), 4.52 -4.58(m, 3H), 4.10-4.32(m, 12H), 3.83-3.90(m, 3H), 3.67-3.73(m, 1H), 3.62-3.66(m, 2H), 3.56-3.61(m, 6 H), 3.48-3.55(m, 3H), 3.33-3.36(m, 5H), 3.17-3.24(m, 13H), 2.60(t, J=6.0Hz, 2H), 2.39(t, J=6.0Hz, 6H), 2.20(t, J=7.2Hz, 6H), 2.14(s, 9H), 2.02(s, 9H), 1.91-1.96(m, 18H), 1.62-1.73(m, 13H), 1.52-1.62(m, 6H). LCMS:m / z=987.6(M / 2+H) + .

[0816] 4 Preparation of compound 6 [ka] Compound 5 (7.1 g, 3.602 mmol) is dissolved in methanol (100 mL) at 25 °C, 10% mass fraction Pd / C (0.5 g, 4.698 mmol) is added, and the reaction solution is stirred under hydrogen (14.696 psi) atmosphere at 25 °C for 18 h. Thin layer chromatography (dichloromethane / methanol = 8 / 1) shows complete consumption of compound 5 and the formation of a new spot. The reaction solution is filtered, and the filtrate is centrifuged to obtain compound 6 (approximately 6.7 g).

[0817] 5 Preparation of compound 8 [ka] Compound 6 (1.50 g, 0.797 mmol), HATU (0.450 g, 1.196 mmol), DIEA (0.395 mL, 2.392 mmol) are added to DMF (15.0 mL), nitrogen replacement is performed three times, and after reacting at 20 ° C for 30 minutes, compound 7 (0.120 g, 1.19 mmol) is added. The reaction solution is reacted at 20 ° C for 2 hours. Thin layer chromatography (dichloromethane / methanol = 8 / 1) shows the disappearance of raw materials and the formation of a new spot. The reaction solution is diluted with dichloromethane (100 mL), washed with saturated sodium bicarbonate aqueous solution (30.0 mL × 3) and saturated saline (30.0 mL × 3) in sequence, and the organic phase is dried over anhydrous sodium sulfate, and then concentrated under reduced pressure to obtain the crude product. The crude product is purified by column chromatography (dichloromethane:methanol = 100 / 1 to 8 / 1) to obtain compound 8 (about 1.00 g) as a yellow solid. 1H NMR (400MHz, CD3OD) δ5.31-5.36(m, 3H), 5.06(dd, J=3.2, 11.2Hz, 3H), 4.50-4.58(m, 3H) , 4.07-4.23(m, 10H), 3.98-4.06(m, 4H), 3.84-3.92(m, 5H), 3.60-3.65(m, 8H), 3.50-3.56 (m, 3H), 3.33-3.36(m, 8H), 3.18-3.26(m, 13H), 2.60-2.68(m, 2H), 2.41(t, J=6.0Hz, 6H), 2.21(t, J=7.2Hz, 6H), 2.14(s, 9H), 2.03(s, 9H), 1.90-1.98(m, 20H), 1.56-1.74(m, 20H). LCMS:m / z=983.1(M / 2+H) + .

[0818] 6. Preparation of E5 [ka] At 25°C, compound 8 (1g, 0.509mmol) is dissolved in dichloromethane (10mL), DCI (0.03g, 0.255mmol) and compound 9 (0.31g, 1.018mmol) are added in sequence, and the reaction solution is stirred under nitrogen atmosphere at 25°C for 2 hours. LCMS shows complete consumption of compound 8, presence of MS value of E5. The reaction solution is cooled to 0°C in an ice-salt bath, 6mL of saturated sodium bicarbonate solution and 6mL of saturated saline solution are added to the reaction solution, then 50mL of dichloromethane is added, and the organic phase is washed with a 1:1 mixed solution of saturated sodium bicarbonate and saturated saline (20mL x 3), followed by drying the organic phase with anhydrous sodium sulfate, and the organic phase is centrifuged to obtain the crude product. The crude product is dissolved in 15 mL of dichloromethane and the crude product solution is slowly added dropwise to methyl tert-butyl ether in a dry ice bath to precipitate a white solid which is filtered to give a white solid E5 (722 mg, 0.222 mmol, 43.61%).

[0819] 1H NMR (400MHz, DMSO-d6) δ7.80-7.85 (m, 6H), 7.70-7.76 (m, 3H), 5.19-5.23 (m, 3H), 4.93-5.00 (m, 3H), 4.45-4.51 ( m, 3H), 3.99-4.05 (m, 10H), 3.83-3.90 (m, 3H), 3.64-3.74 (m, 6H), 3.56-3.62 (m, 2H), 3.47-3.55 (m, 9H), 3.36-3. 44(m, 4H), 3.21(s, 8H), 2.98-3.07(m, 13H), 2.76(t, J=6.0Hz, 2H), 2.53-2.58(m, 2H), 2.26(t, J=6.0Hz, 6H), 2.1 0(s, 9H), 2.04(t, J=7.2Hz, 6H), 2.00(s, 9H), 1.89(s, 9H), 1.77(s, 9H), 1.38-1.55(m, 22H), 1.06-1.17(m, 12H).

[0820] 31 P NMR(400MHz, DMSO-d6)δ-145.243

[0821] LCMS: m / z=1041.7((M-83) / 2+H) + .

[0822] Example 6 Preparation of Compound 6

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[0823] 1 Preparation of compound 3

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[0824] 2. Preparation of E6 [ka] At 25°C, compound 3 (800mg, 0.404mmol) is dissolved in dichloromethane (10mL), DCI (23.86mg, 0.202mmol) and compound 4 (0.31g, 1.018mmol) are added in sequence, and the reaction solution is stirred under nitrogen atmosphere at 25°C for 2 hours. LCMS shows complete consumption of compound 3, presence of MS value of E5. The reaction solution is cooled to 0°C in an ice-salt bath, 6mL of saturated sodium bicarbonate solution and 6mL of saturated saline solution are added to the reaction solution, then 50mL of dichloromethane is added, and the organic phase is washed with a 1:1 mixed solution of saturated sodium bicarbonate and saturated saline (20mL x 3), followed by drying the organic phase with anhydrous sodium sulfate, and the organic phase is centrifuged to obtain the crude product. The crude product is dissolved in 15 mL of dichloromethane, and the crude product solution is slowly added dropwise to methyl tert-butyl ether in a dry ice bath to precipitate a white solid, which is filtered to give a white solid E6 (595 mg, 0.117 mmol, 29.07%).

[0825] 1 HNMR (400MHz, DMSO-d6) δ7.79-7.86(m, 6H), 7.70-7.77(m, 3H), 5.21(d, J=3.2Hz, 3H), 4.96(dd, J=3.2, 11.2Hz , 3H), 4.48(d, J=8.0Hz, 3H), 4.02(s, 9H), 3.80-3.92(m, 5H), 3.66-3.79(m, 6H), 3.47-3.65(m, 13H), 3.36-3.45 (m, 5H), 3.21(s, 8H), 2.96-3.07(m, 13H), 2.74-2.81(m, 2H), 2.54-2.56(m, 2H), 2.26(t, J=6.0Hz, 6H), 2.10(s, 9H), 2.04(t, J=7.2Hz, 6H), 2.00(s, 9H), 1.89(s, 9H), 1.77(s, 9H), 1.41-1.53(m, 18H), 1.14(d, J=6.4Hz, 12H).

[0826] 31 P NMR (400MHz, DMSO-d6)δ-147.942

[0827] LCMS: m / z=1049.4 (M / 2+H) + .

[0828] Example 7 Preparation of Compound E7 [ka]

[0829] 1. Preparation of Compound 3 [ka] Compound 1 (2.00 g, 1.87 mmol, prepared according to the method of intermediate 3-3 in Example 24) is dissolved in DCM (20.0 mL) at room temperature, DIEA (0.135 mL, 0.814 mmol) and compound 2 (0.550 g, 0.814 mmol) are added to the solution in sequence, and nitrogen purge is performed three times. The reaction mixture is stirred at 25 ° C for 16 hours. The MS response of the product is detected by liquid chromatography / mass spectrometry, and thin layer chromatography (dichloromethane / methanol = 5 / 1) shows the disappearance of the raw material and the formation of a new spot. The reaction solution is concentrated under reduced pressure, and the obtained crude product is purified by column chromatography (dichloromethane / methanol = 5 / 1) to obtain a white solid 3 (about 780 mg). 1 H NMR (400MHz, CD3OD) δ7.28-7.42(m, 5H), 5.30-5.34(m, 4H), 5.04-5.14(m, 6H ), 4.63-4.67(m, 4H), 4.36-4.44(m, 2H), 4.00-4.20(m, 23H), 3.91-3.95(m, 4H) ), 3.69-3.77(m, 9H), 3.52-3.67(m, 32H), 3.34-3.43(m, 9H), 2.29-2.31(m, 4H ), 2.14(s, 12H), 2.03(s, 12H), 1.92-1.96(m, 24H).LCMS:m / z=1221.6(M / 2+H) + .

[0830] 2. Preparation of Compound 4 [ka] Compound 3 (1.10 g, 0.451 mmol) is dissolved in MeOH (10.0 mL) at room temperature, 10% mass fraction of wet Pd / C (0.050 g, 0.451 mmol) is added to the solution, hydrogen is replaced three times, and the reaction mixture is stirred under hydrogen atmosphere (14.696 psi) at 25 ° C for 18 hours. The MS response of the product is detected by liquid chromatography / mass spectrometry, and thin layer chromatography (dichloromethane / methanol = 10 / 1, phosphomolybdic acid coloring) shows complete consumption of raw materials and the formation of a new spot. The reaction solution is filtered, and the filtrate is concentrated under reduced pressure to obtain a white solid 4 (about 840 mg). 1 H NMR (400MHz, CD3OD) δ5.32-5.34(m4H), 5.06-5.10(m, 4H), 4.63-4.65(m, 4H), 4.38-4.40(m, 2H), 3.99-4.20(m, 20H), 3.90-3.97(m, 4H), 3.69-3.76(m, 6H), 3.50-3.68(m, 36H), 3.35-3.44(m, 11H), 2.28-2.38(m, 4H), 2.15(s, 12H), 2.03(s, 12H), 1.90-1.94(m, 24H). LCMS:m / z=1154.7(M / 2+H) + .

[0831] 3 Preparation of compound 6 [ka] Compound 5 (232 mg, 0.364 mmol, prepared according to the method of compound Int-6 in Example 13) is dissolved in DCM (10.0 mL) at room temperature, and HBTU (207 mg, 0.546 mmol), DIEA (0.181 mL, 1.09 mmol) and compound 4 (840 mg, 0.364 mmol) are added to the solution in sequence, and nitrogen substitution is performed three times. The reaction mixture is stirred at 25 ° C for 1 hour. The disappearance of the raw material is detected by liquid chromatography / mass spectrometry, and thin layer chromatography (dichloromethane / methanol = 5 / 1) shows the disappearance of the raw material and the formation of a new spot. The reaction solution is concentrated under reduced pressure, and the obtained crude product is purified by column chromatography (dichloromethane / methanol = 8 / 1 to 5 / 1) to obtain a white solid 6 (about 620 mg). 1H NMR (400MHz, CD3OD) δ7.41-7.43(m, 2H), 7.23-7.34(m, 7H), 6.83-6.90(m, 4H), 5.31-5.35(m, 4H) , 5.01-5.12(m, 4H), 4.63-4.65(m, 4H), 4.41-4.45(m, 2H), 4.31-4.33(m, 1H), 3.99-4.22(m, 22H), 3.87-3.97(m, 6H), 3.58-3.81(m, 45H), 3.34-3.43(m, 10H), 2.19-2.40(m, 10H), 2.14(s, 12H), 2. 02(s, 12H), 1.92-1.96(mz, 24H), 1.48-1.63(m, 4H), 1.28-1.38(m, 8H).LCMS:m / z=1460.0(M / 2+H) + .

[0832] 4. Preparation of E7 [ka] Compound 6 (300 mg, 0.103 mmol) is dissolved in DCM (10.0 mL) at room temperature, DIEA (0.102 mL, 0.618 mmol) and compound 7 (10.3 mg, 0.103 mmol) and DMAP (12.6 mg, 0.103 mmol) are added to the solution in sequence, and nitrogen substitution is performed three times. The reaction mixture is stirred at 25 ° C for 2 hours. The disappearance of the raw material is detected by liquid chromatography / mass spectrometry. The reaction solution is concentrated under reduced pressure, and the obtained crude product is prepared and separated by preparative high performance liquid chromatography (preparative HPLC, column: Waters Xbridge BEH C18 100 * 30 mm * 10 um, mobile phase: water-ACN, B%: 17% ~ 57%, 5 min) to obtain a white solid E7 (53.0 mg, yield 17.08%, purity 78.94%). 1H NMR (400MHz, CD3OD) δ7.41-7.45(m, 2H), 7.17-7.34(m, 7H), 6.85-6.89(m, 4H), 5.32-5.36(m, 4H), 5.03-5.13 (m, 4H), 4.63-4.67(m, 4H), 4.38-4.47(m, 2H), 4.32-4.34(m, 1H), 4.01-4.26(m, 22H), 3.88-4.00(m, 6H), 3.7 7-3.81(m, 7H), 3.49-3.76(m, 45H), 3.33-3.47(m, 10H), 2.56-2.62(m, 2H), 2.45-2.55(m, 3H), 2.21-2.38(m, 7H), 2.14(s, 12H), 2.05-2.11(m, 2H), 2.02(s, 12H), 1.92-1.96(m, 24H), 1.47-1.68(m, 4H), 1.28-1.34(m, 8H) MS: m / z=3022.36(M+H) + .

[0833] Example 8 Preparation of Compound E8 [ka]

[0834] 1. Preparation of Compound 1 [ka] Compound 1A (5.00 g, 17.8 mmol) is added to DCM (200 mL) at 25 ° C, then compound pentafluorophenol (8.18 g, 44.4 mmol) and EDCI (10.2 g, 53.3 mmol) are added successively, and the reaction solution is stirred at 25 ° C for 14 h. Thin layer chromatography (petroleum ether / ethyl acetate = 5 / 1) shows the formation of a new spot. The reaction solution is concentrated under reduced pressure, and the obtained crude product is purified by column chromatography (petroleum ether / ethyl acetate = 1 / 0 to 5 / 1) to obtain compound 1 (about 9.00 g) as a white solid. 1H NMR (400MHz, CDCl3) δ7.34-7.38(m, 5H), 5.39-5.41(m, 1H), 5.19(s, 2H), 4 .87-4.92(m, 1H), 2.86-2.97(m, 2H), 2.55-2.58(m, 1H), 2.28-2.34(m, 1H).

[0835] 2. Preparation of Compound 3 [ka] Compound 1 (800 mg, 1.30 mmol) was dissolved in DCM (20.0 mL), DIEA (421 mg, 3.26 mmol) and compound 2 (2076 mg, 3.26 mmol, prepared according to the method of intermediate 5 in Example 24) were added, and the reaction solution was stirred at 25 ° C for 18 hours, LC / MS detected the MS response that the product should have, and the spot plate (DCM: MeOH = 10: 1) showed the complete reaction of the raw material, and the reaction solution was directly concentrated through a normal phase silica gel column, eluted with MeOH: DCM = 0: 1 to 1: 10, and a yellow solid compound 3 (about 1.3 g) was obtained. LCMS: m / z = 1290.4 (M + H) + .

[0836] 3 Preparation of compound 4 [ka] Compound 3 (1.30 g, 1.01 mmol) was dissolved in MeOH (30 mL), and 10% mass fraction Pd / C (0.20 g, 1.88 mmol) was added. Hydrogen was replaced three times, and the reaction solution was reacted under H2 (15 psi) atmosphere at 25 °C for 18 hours. LC / MS detected the MS response that the product should have, and the reaction solution was filtered to remove Pd / C, washed with methanol, and the filtrate was combined and concentrated to obtain compound 4 (about 1.08 g) as a yellow oil. LCMS: m / z = 1156.9 (M + H) + .

[0837] 4 Preparation of compound 6 [ka] Compound 4 (1080 mg, 0.934 mmol) is dissolved in DCM (20 mL), compound 5 (302.2 mg, 0.449 mmol, prepared according to the method of compound 2 in Example 7) and DIEA (0.17 mL, 0.934 mmol) are added, and the reaction solution is stirred at 25 °C for 18 h. LC / MS detects the MS response that the product should have, and TLC (DCM:MeOH = 10:1) shows the formation of a new spot, and the reaction solution is concentrated directly onto a column (DCM:MeOH = 1:0 to DCM:MeOH = 10:1) to obtain compound 6 (about 820 mg). LCMS: m / z = 1309.5 (M / 2 + H) + .

[0838] 5 Preparation of compound 7 [ka] Compound 6 (800 mg, 0.306 mmol) was dissolved in MeOH (10 mL), and 10% mass fraction Pd / C (100 mg, 0.940 mmol) was added. Hydrogen was replaced three times, and the reaction solution was reacted under H2 (15 psi) atmosphere at 25°C for 18 hours. LC / MS detected the MS response that the product should have, and the reaction solution was filtered to remove Pd / C, washed with methanol, and the filtrate was combined and concentrated to obtain compound 7 (about 1.08 g) as a white solid. LCMS: m / z = 1242.4 (M / 2 + H) + .

[0839] 6 Preparation of compound 9 [ka] Compound 7 (700 mg, 0.282 mmol) is dissolved in DCM (20 mL), HBTU reagent (160 mg, 0.423 mmol), DIEA (0.140 mL, 0.846 mmol) and compound 8 (180 mg, 0.282 mmol, prepared according to the method of compound Int-6 in Example 13) are added in sequence, and the reaction solution is stirred at 25 ° C for 18 hours. LC / MS detects the MS response that the product should have. TLC (DCM: MeOH = 10: 1) shows the complete reaction of the raw materials, the formation of a new spot. The reaction solution is washed with 20 ml of saturated sodium bicarbonate aqueous solution, extracted with dichloromethane (20 ml × 2), the organic phase is combined and concentrated, and passed through a normal phase column (DCM: MeOH = 1: 0 to 10: 1) to obtain compound 9 (about 750 mg) as a white solid. LCMS: m / z=1399.1 [(M-DMTr) / 2+H] + .

[0840] 7. Preparation of E8 [ka] Compound 9 (500mg, 0.161mmol) is dissolved in DCM (5mL), compound 10 (96.9mg, 0.968mmol), DIEA (0.160mL, 0.968mmol) and DMAP (19.7mg, 0.161mmol) are added, and the reaction solution is stirred at 25℃ for 18h. LCMS detects the MS response that the product should have. The reaction solution is directly centrifuged, and 5ml of acetonitrile is added to dissolve it, and then the mixture is subjected to preparative HPLC (chromatography column: Waters Xbridge BEH C18 100*30mm*10um, mobile phase: water-acetonitrile], B%: 20%~50%, 11min, flow rate: 50ml / min), to obtain compound E8 as a white solid. LCMS (ESI): m / z=1449.0[(M-DMTr) / 2+H]+, 1H NMR (400MHz, CD3OD) δ7.43(d, J=8.0Hz, 2H), 7.30-7.32(m, 6H), 7.20-7.25(m, 1H), 6.85-6.89(m, 4H), 5.33(d, J=3.2Hz, 4H), 5 .06(dd, J=11.2, 3.2Hz, 4H), 4.66(d, J=8.8Hz, 4H), 4.33-4.43(m, 4H), 4.09-4.17(m, 12H), 4.01-4.04(m, 8H), 3.89-3.94(m, 4H) ), 3.79(d, J=2.0Hz, 6H), 3.71-3.76(m, 9H), 3.60-3.66(m, 48H), 3.52-3.58(m, 8H), 3.37-3.40(m, 8H), 2.54-2.60(m, 4H), 2.2 6-2.31(m, 8H), 2.14(s, 12H), 2.02(s, 12H), 1.96-2.00(m, 2H), 1.94(d, J=4.0Hz, 24H), 1.53-1.61(m, 4H), 1.29-1.38(m, 10H).

[0841] Example 9 Preparation of Compound E9 [ka]

[0842] 1. Preparation of compound 1C [ka] Compound 1A (10 g, 82.6 mmol), compound 1B (42.3 g, 330 mmol) and NaOH (5 M, 1.65 mL) are added to DMSO (300 mL) in sequence at room temperature and reacted at 24 ° C for 12 hours. Thin layer chromatography (dichloromethane / methanol = 10 / 1) shows the formation of a new spot. The reaction solution is diluted with water (1000 mL), extracted with dichloromethane (500 mL × 3), the organic phase is dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain compound 1C (45 g) as a colorless oil. 1 H NMR (400MHz, CDCl3) δ3.62-3.70(m, 6H)3.25-3.49(m, 6H), 2.41-2.52(m, 6H), 1.45(s, 27H)

[0843] 2. Preparation of Compound 1 [ka] Compound 1C (30.0 g, 59.3 mmol), CbzOSu (17.8 g, 71.2 mmol) and DIEA (11.5 g, 89 mmol) are added in DCM (300 mL) successively under room temperature conditions. The mixture is reacted at 25 °C for 14 h. Thin layer chromatography (petroleum ether / ethyl acetate = 3 / 1) shows the formation of a new spot. The reaction solution is diluted with water (500 mL), extracted with dichloromethane (200 mL x 3), the organic phase is concentrated under reduced pressure, and the crude product is purified by column chromatography (petroleum ether / ethyl acetate = 30 / 1 to 2 / 1) to obtain compound 1 (about 19.0 g) as a colorless oil. 1 H NMR (400MHz, CDCl3) δ7.29-7.39(m, 5H), 5.30(s, 1H), 5.04(s, 2H), 3.57-3.72(m, 12H), 2.44(t, J=6.4Hz, 6H), 1.44(s, 27H)

[0844] 3 Preparation of compound 2 [ka] Compound 1 (8.00 g, 21.083 mmol) is dissolved in DCM (30 mL) at room temperature, TFA (10 mL, 130.591 mmol) is added, and the mixture is stirred at 25 °C for 15 h. Liquid chromatography / mass spectrometry indicates complete reaction of the raw materials and formation of the product. The reaction solution is directly concentrated under reduced pressure to obtain the crude product of colorless oil 2 (approximately 7.27 g). 1 H NMR (400MHz, CD3OD) δ7.26-7.38 (m, 5H), 5.03 (s, 2H), 3.59-3.70 (m, 12H), 2.51 (t, J=6.0Hz, 6H). LCMS:m / z=472.1(M+H) + .

[0845] 4 Preparation of Compound 4 [ka] Compound 2 (1.00 g, 2.121 mmol) is dissolved in DMF (30 mL) at room temperature, EDCI (2.44 g, 12.726 mmol) and compound 3 (2.34 g, 12.726 mmol) are added, and the mixture is stirred at 25 °C for 2 h. Thin layer chromatography (petroleum ether: ethyl acetate = 5:1) shows the disappearance of the raw material and the formation of a new spot. The reaction solution is concentrated under reduced pressure to obtain the crude product, which is purified by column chromatography (petroleum ether: ethyl acetate = 20 / 3 to 5 / 1). A white solid 4 (about 1.24 g) is obtained. 1 H NMR (400MHz, CD3OD) δ7.26-7.38(m, 5H), 4.98(s, 2H), 3.80(t, J=5.6Hz, 6H), 3.72(s, 6H), 2.93(t, J=5.2Hz, 6H)

[0846] 5 Preparation of Compound 5 [ka] Compound 4 (1.24 g, 1.279 mmol) is dissolved in DMF (40 mL) at room temperature, and compound A2 (1.84 g, 3.837 mmol, prepared according to the method of intermediate 3-4 in Example 23) and DIEA (2.114 mL, 12.789 mmol) are added, and the mixture is stirred at 25 ° C for 15 hours. Thin layer chromatography (dichloromethane:methanol = 10:1, phosphomolybdic acid coloring) shows the disappearance of raw materials and the formation of a new spot. The reaction solution is concentrated under reduced pressure, and the crude product is purified by column chromatography (dichloromethane:methanol = 25 / 2 to 10 / 1). A white solid 5 (about 1.60 g) is obtained. 1H NMR (400MHz, CD3OD) δ7.27-7.41(m, 5H), 5.34(d, J=3.6Hz, 3H), 5.07(dd, J=3.2, 11. 2Hz, 3H), 5.04(s, 2H), 4.64(d, J=8.4Hz, 3H), 4.06-4.19(m, 9H), 4.00-4.05(m, 3H), 3 .90-3.96(m, 3H), 3.70-3.75(m, 3H), 3.59-3.70(m, 30H), 3.51-3.56(m, 6H), 3.34-3. 39(m, 6H), 2.43(t, J=6.0Hz, 6H), 2.14(s, 9H), 2.03(s, 9H), 1.95(s, 9H), 1.93(s, 9H)

[0847] 6 Preparation of compound 6 [ka] Compound 5 (1.6 g, 0.864 mmol) is dissolved in MeOH (20 mL) at room temperature, 10% mass fraction Pd / C (0.3 g, 2.819 mmol) is added, the mixture is purged with H2 three times, and stirred at 25 °C for 15 h. Thin layer chromatography (dichloromethane / methanol = 10 / 1) shows the disappearance of raw materials and the formation of a new spot. Liquid chromatography / mass spectrometry shows the complete reaction of the raw materials and the formation of the product. The reaction solution is filtered through Celite, and the filtrate is concentrated under reduced pressure to obtain a colorless transparent solid 6 (approximately 1.40 g). 1 H NMR (400MHz, CD3OD) δ5.34(d, J=2.8Hz, 3H), 5.07(dd, J=3.2, 11.2Hz, 3H), 4.65(d, J=8.4Hz, 3H), 4.06-4.20(m, 9H), 4.01-4.06(m, 3H), 3.90-3.9 8(m, 3H), 3.72-3.76(m, 3H), 3.52-3.70(m, 30H), 3.36-3.41(m, 12H), 2. 46(t, J=6.0Hz, 6H), 2.15(s, 9H), 2.03(s, 9H), 1.95(s, 9H), 1.94(s, 9H). LCMS: m / z=860.1 (M / 2+H) + .

[0848] 7 Preparation of compound 7 [ka] Compound 7A (20 g, 98.888 mmol) is added to DCM (500 mL) at 25° C., then compound 7B (45.51 g, 247.3219 mmol) and EDCI (56.87 g, 296.663 mmol) are added sequentially, and the reaction solution is stirred at 25° C. for 18 hours. Thin layer chromatography (petroleum ether / ethyl acetate=1 / 1) shows complete consumption of the reactants, and thin layer chromatography (petroleum ether / ethyl acetate=5 / 1) shows the formation of a new product spot. The reaction solution is concentrated under reduced pressure to obtain the crude product, which is purified by MPLC to obtain compound 7 (about 48.7 g) as a white solid. 1 H NMR (400MHz, CDCl3) δ2.68 (t, J=7.6Hz, 4H), 1.72-1.87 (m, 4H), 1.34-1.51 (m, 8H).

[0849] 8 Preparation of compound 8 [ka] Compound 6 (1.00 g, 0.582 mmol) is dissolved in DMF (20 mL) at room temperature, 7 (0.93 g, 1.745 mmol) and DIEA (0.481 mL, 2.909 mmol) are added, and the mixture is reacted at 25 °C for 15 h. Thin layer chromatography (dichloromethane:methanol = 10:1) shows the disappearance of the raw material and the formation of a new spot. The reaction solution is concentrated under reduced pressure, and the crude product is purified by column chromatography (dichloromethane:methanol = 25 / 2 to 10 / 1) to obtain a colorless oily liquid 8 (about 809 mg). 1H NMR (400MHz, CD3OD) δ5.34(d, J=2.8Hz, 3H), 5.07(dd, J=3.2, 11.2Hz, 3H), 4.65(d, J=8.4H) z, 3H), 4.06-4.18(m, 9H), 4.01-4.05(m, 3H), 3.90-3.98(m, 3H), 3.61-3.76(m, 35H), 3.54- 3.58(m, 6H), 3.36-3.41(m, 6H), 2.73(t, J=7.2Hz, 2H), 2.45(t, J6.0Hz, 6H), 2.14(s, 9H), 2.03(s, 9H), 1.90-1.97(m, 18H), 1.70-1.84(m, 2H), 1.52-1.65(m, 2H), 1.30-1.35(m, 8H). 1 H NMR:RW0008-15-P1A(400MHz, CD3OD)

[0850] 9 Preparation of Compound 10 [ka] Compound 8 (350 mg, 0.169 mmol) was dissolved in DCM (2.00 mL) at room temperature, and compound 9 (76.1 mg, 0.169 mmol) and DIEA (0.112 mL, 0.677 mmol) were added. After reacting for 12 hours, thin layer chromatography (dichloromethane / methanol = 10 / 1) showed the disappearance of the raw material and the formation of a new spot. The reaction solution was diluted with water (10 mL), extracted with dichloromethane (10 mL x 3), the organic phase was dried over sodium sulfate, and the filtrate was concentrated under reduced pressure to obtain colorless oil 10 (about 388 mg). 1H NMR (400MHz, METHANOL-d4) δ7.43(d, J=8.4Hz, 2H), 7.20-7.35(m, 7H), 6.82-6.92(m, 4H), 5.34(d, J=2.8Hz, 3H), 5.0 7(dd, J=3.6, 11.2Hz, 3H), 4.65(d, J=8.4Hz, 3H), 4.06-4.17(m, 9H), 3.99-4.05(m, 3H), 3.90-3.96(m, 4H), 3.79(d, J =2.8Hz, 6H), 3.58-3.75(m, 39H), 3.53-3.57(m, 7H), 3.38(t, J=5.6Hz, 8H), 3.22-3.24(m, 1H), 2.45(t, J=6.0Hz, 6H) , 2.15-2.20(m, 2H), 2.13-2.15(m, 9H), 2.02-2.04(m, 9H), 1.91-1.96(m, 18H), 1.49-1.61(m, 4H), 1.27-1.35(m, 8H)

[0851] 10. Preparation of E9 [ka] Compound 10 (388 mg, 0.166 mmol) is dissolved in DCM (4 mL) at room temperature, DIEA (0.165 mL, 0.997 mmol), DMAP (5.08 mg, 0.042 mmol) and compound 11 (99.79 mg, 0.997 mmol) are added, and the mixture is reacted at 25° C. for 3 hours. LCMS shows the complete reaction of the raw materials and the formation of the product. Dichloromethane (30 mL) is added to dilute, and then the organic phase is washed three times with saturated NaCl solution (10 mL×3), followed by drying over anhydrous sodium sulfate. The reaction solution was concentrated under reduced pressure, and the obtained crude product was prepared and separated by preparative high performance liquid chromatography (preparative HPLC, column: Welch Xtimate C18 150*25mm*5um, mobile phase: water-ACN, B%: 20%-50%, 5 min) to obtain a white solid E9 (78.0 mg, yield 19.28%, purity 76.01%). LCMS: m / z=1216.1 (M / 2+H) + , 1H NMR (400MHz, CD3OD) δ7.43(d, J=8.0Hz, 2H), 7.22-7.35(m, 7H), 6.80-6.92(m, 4H), 5.34(d, J=3.2Hz, 3H), 5.08(dd, J=3. 2, 11.2Hz, 3H), 4.65(d, J=8.4Hz, 3H), 4.05-4.18(m, 10H), 4.00-4.04(m, 3H), 3.90-3.97(m, 4H), 3.79(d, J=2.4Hz, 6H), 3 .58-3.75(m, 39H), 3.55(t, J=5.2Hz, 7H), 3.32-3.42(m, 8H), 3.20-3.24(m, 1H), 2.56(dd, J=5.6, 18.8Hz, 4H), 2.44(t, J =6.0Hz, 6H), 2.16-2.21(m, 2H), 2.14(s, 9H), 2.02(s, 9H), 1.90-1.98(m, 18H), 1.48-1.61(m, 4H), 1.31(d, J=13.2Hz, 8H)

[0852] Example 10 Preparation of Compound E10 [ka]

[0853] 1. Preparation of Compound 1A [ka] Compound 1C (20 g, 73.8 mmol) is dissolved in anhydrous toluene (200 mL), tetrabutylammonium hydrogen sulfate (3.75 g, 11.0 mmol), tert-butyl bromoacetate (85.9 g, 442.8 mmol) and 30% NaOH (160 mL) are added in sequence, and the reaction solution is stirred at 45 ° C for 18 h. TCL plate (PE:EA = 1:1, UV 254 nm) shows complete consumption of the raw material and the formation of a new spot. The reaction solution is cooled to room temperature, left to stand and separated, the lower alkaline aqueous phase is extracted with methyl tert-butyl ether (200 ml × 2), the organic phase is combined and concentrated, and passed through a normal phase column (silica gel, PE:EA = 1:0 to PE:EA = 3:1) to obtain compound 1A (about 28.5 g).

[0854] 2. Preparation of Compound 1B [ka] LiAlH4 (1.67 g, 44.0 mmol) is added to THF (100 mL) with stirring, and after nitrogen replacement, it is cooled to 0 ° C in an ice bath, and compound 1A (10.0 g, 20.0 mmol) is dissolved in 20 ml of THF, and slowly dropped into the above solution at 0 ° C using a syringe, and the reaction solution is stirred at 0 ° C for 2 hours. LC / MS detects the complete consumption of the raw material and the presence of MS response that the product should have. 2 ml of water, 2 ml of 15% NaOH aqueous solution, and 6 ml of water are added to the reaction solution with stirring, and after sufficient stirring, anhydrous sodium sulfate powder is added, and after stirring for 15 minutes, it is filtered and the organic phase is concentrated to obtain colorless oily compound 1B (about 6.60 g). LCMS: m / z = 360.3 (M + H) + .

[0855] 3 Preparation of compound 1 [ka] Compound 1B (6.60 g, 18.4 mmol) is dissolved in anhydrous toluene (100 mL), tetrabutylammonium hydrogen sulfate (1.25 g, 3.67 mmol), tert-butyl bromoacetate (21.5 g, 110 mmol) and 30% NaOH (80 mL) are added in sequence, and the reaction solution is stirred at 45 ° C for 18 hours. LC / MS detects the MS response that the product should have, and the TCL plate (PE: EA = 3: 1, UV 254 nm) shows the complete consumption of the raw material and the generation of a new spot. The reaction solution is cooled to room temperature, left to stand and separated, the lower alkaline aqueous phase is extracted with methyl tert-butyl ether (50 ml × 2), the organic phase is combined and concentrated, and passed through a normal phase column (silica, PE: EA = 1: 0 to PE: EA = 3: 1) to obtain compound 1 (about 9.6 g). LCMS: m / z = 589.9 (M + H) + .

[0856] 4. Preparation of Compound 2 [ka] Compound 1 (9.60 g, 16.4 mmol) is dissolved in EtOH (50 mL), 10% mass fraction Pd / C (1 g, 9.40 mmol) is added, hydrogen replacement is performed three times, and the reaction solution is reacted under H2 (15 psi) atmosphere at 45 ° C for 18 hours. LC / MS detects the MS response that the product should have, and the reaction solution is filtered to remove palladium carbon and concentrated to obtain compound 2 (about 6.60 g). 1 H NMR (400MHz, CD3OD) δ4.04(s, 4H), 3.68-3.70(m, 4H), 3.62-3.64(m, 4H), 3.50-3.54(m, 2H), 3.40-3.44(m, 2H), 3.06-3.12(m, 1H), 1.48(s, 18H). LCMS:m / z=408.7(M+H) + .

[0857] 5. Preparation of Compound 3 [ka] Compound 2 (6.60 g, 16.4 mmol) is dissolved in DCM (100 mL), DIEA (5.41 mL, 32.7 mmol) is added first, then benzyl chloroformate (2.53 mL, 18.0 mmol) is added dropwise with stirring, and the reaction solution is stirred at 25 °C for 18 h. LC / MS detects the MS response that the product should have. TLC (PE:EA = 3:1, PMA) shows the complete reaction of the raw materials, the generation of a new pot, and the reaction solution is passed through a normal phase column (PE:EA = 1:0 to 3:1) and directly concentrated to obtain colorless oily compound 3 (about 7.5 g). 1 H NMR (400MHz, CD3OD) δ7.29-7.35(m, 5H), 5.08(s, 2H), 4.02(s, 4H), 3.87-3.92(m, 1H), 3.63-3.67(m, 8H), 3.55-3.56(m, 4H), 1.47(s, 18H). LCMS:m / z=542.3(M+H) + .

[0858] 6 Preparation of compound 4 [ka] Compound 3 (3.7 g, 6.88 mmol) is dissolved in DCM (18 mL), TFA (6 mL) is added, and the reaction solution is stirred for 18 h at 25° C. LC / MS detects the MS response that the product should have, and the reaction solution is directly concentrated to give compound 4 (approximately 4.3 g) as a yellow oil. 1 H NMR (400MHz, CD3OD) δ7.28-7.35(m, 5H), 5.08(s, 2H), 4.12(s, 4H), 3.87-3.93(m, 1H), 3.63-3.68(m, 8H), 3.55-3.56(m, 4H). LCMS:m / z=430.1(M+H) + .

[0859] 7 Preparation of compound 6 [ka] Compound 4 (1 g, 2.33 mmol) is dissolved in DCM (30 mL), DIEA (4.06 mL, 23.3 mmol) and compound 5 (1.96 g, 6.99 mmol) are added, the reaction solution is stirred at 25 °C for 1 h, and LC / MS detects the MS response that the product should have. TLC (PE:EA = 3:1) shows the complete reaction of the raw materials, the formation of a new spot. The reaction solution is passed through a normal phase column (PE:EA = 1:0 to PE:EA = 3:1) and concentrated directly to give compound 6 (about 900 mg) as a white solid. LCMS: m / z = 762.6 (M + H) + .

[0860] 8 Preparation of compound 8 [ka] Compound 6 (1132 mg, 1.16 mmol) is dissolved in DCM (10 mL), DIEA (0.26 mL, 1.58 mmol) and compound 7 (400 mg, 0.53 mmol, prepared according to the method of compound 6 in Example 12) are added, and the reaction solution is stirred at 25 °C for 18 h. TLC (DCM:MeOH = 10:1) shows the complete reaction of the raw materials and the formation of a new spot, and LCMS detects the MS response that the product should have. The reaction solution is directly concentrated through a normal phase column (DCM:MeOH = 1:0 to 10:1) to give compound 8 (about 1100 mg) as a colorless oil. LCMS: m / z = 1177.4 (M / 2 + H) + .

[0861] 9 Preparation of Compound 9 [ka] Compound 8 (1100 mg, 0.467 mmol) was dissolved in MeOH (20 mL), and 10% mass fraction Pd / C (100 mg, 0.940 mmol) was added. Hydrogen was replaced three times, and the reaction solution was reacted under H2 (15 psi) atmosphere at 25°C for 18 hours. LC / MS detected the MS response that the product should have, and the reaction solution was filtered to remove Pd / C, washed with methanol, and the filtrate was combined and concentrated to obtain compound 9 (about 960 mg) as a white solid. LCMS: m / z = 1110.3 (M / 2 + H) + .

[0862] 10 Preparation of compound 11 [ka] Compound 9 (960 mg, 0.282 mmol) is dissolved in DMF (20 mL), HBTU reagent (246 mg, 0.649 mmol), DIEA (0.214 mL, 1.30 mmol) and compound 10 (277 mg, 0.433 mmol, prepared according to the method of compound Int-6 in Example 13) are added in sequence, and the reaction solution is stirred at 25 ° C for 18 hours. LC / MS detects the MS response that the product should have. TLC (DCM: MeOH = 10: 1) shows the complete reaction of the raw materials, the formation of a new spot. The reaction solution is washed with 20 ml of saturated sodium bicarbonate aqueous solution, extracted with dichloromethane (20 ml × 2), the organic phase is combined and concentrated, and passed through a normal phase column (DCM: MeOH = 1: 0 to 10: 1) to obtain compound 11 (about 1060 mg) as a white solid. LCMS: m / z=1266.9[(M-DMTr) / 2+H] + .

[0863] 11. Preparation of E10 [ka] Compound 11 (500 mg, 0.161 mmol) is dissolved in DCM (5 mL), tetrahydrofuran-2,5-dione (96.86 mg, 0.968 mmol), DIEA (0.160 mL, 0.968 mmol) and DMAP (19.7 mg, 0.161 mmol) are added and the reaction solution is stirred for 18 hours at 25° C. LCMS detects the MS response that the product should have. The reaction solution was directly centrifuged and dissolved by adding 5 ml of acetonitrile. The mixture was then subjected to preparative HPLC (chromatography column: Waters Xbridge BEH C18 100*30mm*10um, mobile phase: water-acetonitrile), B%: 20%-50%, 11 min, flow rate: 50 ml / min. A white solid E10 (56 mg, 0.018 mmol, 10.9%) was obtained. LCMS (ESI): m / z=1317.0 [(M-DMTr) / 2+H]+, 1H NMR (400MHz, CD3OD) δ7.43 (d, J=8.0Hz, 2H), 7.27-7.32 (m, 6H), 7.18-7.24 (m, 1H), 6.85-6.89 (m, 4H), 5.34 (d, J=3.6Hz, 4H), 5.07-5.12 (m, 4H), 4.62-4.67 (m, 4H), 4.42-4.48 (m, 2H), 4.20-4.24 (m, 2H), 3.98-4.17 (m, 22H), 3.90-3.94 (m, 4H), 3.79 (d, J=2.0Hz, 6H), 3.69-3. 74(m,12H),3.62-3.63(m,14H),3.52-3.55(m,8H),3.35-3.40(m,8H),3.20(s,2H),2.48-2.60(m,4H),2.28-2.34(m,5H),2.20-2.28(m , 2H), 2.14(s, 12H), 2.06-2.11(m, 2H), 2.02(s, 12H), 1.97-1.99(m, 2H), 1.94(d, J=5.2Hz, 24H), 1.53-1.59(m, 4H), 1.28-1.38(m, 12H).

[0864] Example 11 Preparation of Compound E11

change

[0865] 1 Preparation of compound 2

change

[0866] 2. Preparation of Compound 3 [ka] Compound 2 (6.80 g, 11.3 mmol) is dissolved in DCM (50.0 mL) at room temperature, HCl / EtOAc (20.0 mL) is added to the solution, and the reaction mixture is stirred at 25 °C for 1 h. Thin layer chromatography (dichloromethane / methanol = 10 / 1, phosphomolybdic acid coloring) shows complete consumption of the raw material and the formation of a new spot. The reaction solution is concentrated under reduced pressure to give the crude product of white solid 3 (about 6.90 g), which is used directly in the next reaction step without purification.

[0867] 3 Preparation of compound 5 [ka] Compound 3 (6.80 g, 13.5 mmol) is dissolved in DCM (20.0 mL) at room temperature, DIEA (2.23 mL, 13.5 mmol) and compound 4 (3.61 g, 6.08 mmol) are added to the solution in sequence, and nitrogen substitution is performed three times. The reaction mixture is stirred at 25 °C for 18 h. Thin layer chromatography (dichloromethane / methanol = 10 / 1) shows complete consumption of the raw material and the formation of a new spot. The reaction solution is concentrated under reduced pressure, and the obtained crude product is purified by column chromatography (dichloromethane / methanol = 12 / 1 to 10 / 1) to obtain a white solid 5 (about 6.00 g). 1 H NMR (400MHz, CD3OD) δ5.3-5.34(m, 2H), 5.04-5.08(m, 2H), 4.54-4.58(m, 2H), 3.97-4.21(m, 9H), 3.81-3.92(m, 2H), 3.51-3.53(m, 2H), 3.14-3.28 (m, 9H), 2.25-2.32(m, 2H), 2.19-2.23(m, 4H), 2.14(s, 6H), 2.03(s, 6H), 1.94(d, J=9.2Hz, 12H), 1.62-1.75(m, 9H), 1.56-1.62(m, 4H), 1.44(s, 9H)

[0868] 4 Preparation of compound 6 [ka] Compound 5 (1.0 g, 0.821 mmol) is dissolved in DCM (10.0 mL) at room temperature, HCl / EtOAc (1.00 mL) is added to the solution, and the reaction mixture is stirred at 25 °C for 16 h. Thin layer chromatography (dichloromethane / methanol = 10 / 1, phosphomolybdic acid coloring) shows complete consumption of the raw material and the formation of a new spot. The reaction solution is concentrated under reduced pressure to obtain the crude product of white solid 6 (about 1.57 g). The crude product is used directly in the next reaction step without purification.

[0869] 5 Preparation of compound 8 [ka] Compound (31.57 g, 0.772 mmol) is dissolved in DCM (20.0 mL) at room temperature, DIEA (0.255 mL, 1.544 mmol) and compound 7 (0.21 g, 0.309 mmol) are added to the solution in sequence, and nitrogen substitution is performed three times. The reaction mixture is stirred at 25 °C for 16 hours. The MS response of the product is detected by liquid chromatography / mass spectrometry, and thin layer chromatography (dichloromethane / methanol = 5 / 1) shows the disappearance of the raw material and the formation of a new spot. The reaction solution is concentrated under reduced pressure, and the obtained crude product is purified by column chromatography (dichloromethane / methanol = 5 / 1) to obtain a white solid 8 (about 780 mg). 1 H NMR (400MHz, CD3OD) δ7.25-7.42(m, 5H), 5.30-5.33(m, 4H), 5.04-5.06(m, 4H), 4. 54-4.56(m, 4H), 4.30-4.40(m, 2H), 3.96-4.19(m, 21H), 3.83-3.90(m, 4H), 3.67-3 .76(m, 4H), 3.50-3.52(m, 4H), 3.11-3.27(m, 18H), 2.25-2.33(m, 4H), 2.18-2.20 (m, 8H), 2.14(s, 12H), 2.02(s, 12H), 1.94(d, J=9.20Hz, 24H), 1.56-1.70(m, 24H). LCMS:m / z=1272.5(M / 2+H) + .

[0870] 6 Preparation of compound 9 [ka] Compound 8 (380 mg, 0.150 mmol) is dissolved in MeOH (10.0 mL) at room temperature, and 10% mass fraction of wet Pd / C (100 mg, 0.940 mmol) is added to the solution in sequence, hydrogen substitution is performed three times, and the reaction mixture is stirred under hydrogen atmosphere (14.696 psi) at 25 °C for 18 h. The MS response of the product is detected by liquid chromatography / mass spectrometry, and thin layer chromatography (dichloromethane / methanol = 10 / 1, phosphomolybdic acid coloring) shows complete consumption of the raw material and the generation of a new spot. The reaction solution is filtered, and the filtrate is concentrated under reduced pressure to obtain a white solid 9 (about 330 mg). 1 H NMR (400MHz, CD3OD) δ5.31-5.35(m, 4H), 5.04-5.06(m, 4H), 4.53-4.56(m, 4H), 4.34-4.36(m, 2H), 3.99-4.16(m, 21H), 3.85-3.90(m, 4H), 3.51- 3.56(m, 4H), 3.18-3.22(m, 18H), 2.28-2.33(m, 4H), 2.19-2.22(m, 8H), 2.14(s, 12H), 2.03(s, 12H), 1.94(d, J=8.8Hz, 24H), 1.58-1.71(m, 24H). LCMS: m / z=1204.9 (M / 2+H) + .

[0871] 7 Preparation of compound 11 [ka] Compound 9 (170 mg, 0.071 mmol) is dissolved in DCM (10.0 mL) at room temperature, and compound 10 (45.2 mg, 0.071 mmol, prepared according to the method of compound Int-6 in Example 13), DIEA (0.035 mL, 0.212 mmol), HBTU (40.2 mg, 0.106 mmol), DMAP (8.63 mg, 0.071 mmol) are added to the solution in sequence, and nitrogen substitution is performed three times. The reaction mixture is stirred at 25 ° C for 1 h. The disappearance of raw materials is detected by liquid chromatography / mass spectrometry. Thin layer chromatography (dichloromethane / methanol = 5 / 1) shows the disappearance of raw materials and the formation of a new spot. The reaction solution is concentrated under reduced pressure, and the obtained crude product is purified by column chromatography (dichloromethane / methanol = 8 / 1 to 5 / 1) to obtain a white solid 11 (about 340 mg).

[0872] 8. Preparation of E11 [ka] Compound 11 (340 mg, 0.112 mmol) is dissolved in DCM (10.0 mL) at room temperature, DIEA (14.5 mg, 0.112 mmol), compound 12 (11.3 mg, 0.112 mmol) and DMAP (6.84 mg, 0.056 mmol) are added to the solution in sequence, and nitrogen substitution is performed three times. The reaction mixture is stirred at 25 ° C for 1 hour. The disappearance of the raw material is detected by liquid chromatography / mass spectrometry. The reaction solution is concentrated under reduced pressure, and the obtained crude product is prepared and separated by preparative high performance liquid chromatography (preparative HPLC, column: Waters Xbridge BEH C18 100 * 30 mm * 10 um, mobile phase: water-ACN, B%: 19% ~ 49%, 5 min) to obtain a white solid E11 (72.3 mg, yield 20.59%, purity 80.47%). 1H NMR (400MHz, CD3OD) δ7.41-7.43 (m, 2H), 7.29-7.31 (m, J=8.8Hz, 6H), 7.17-7.26 (m, 1H), 6.83-6.90 (m, 4H), 5.31-5.34 (m, 4H), 5.02-5. 08(m, 4H), 4.50-4.56(m, 4H), 4.32-4.40(m, 3H), 4.20-4.25(m, 1H), 4.09-4.19(m, 10H), 4.05-4.09(m, 7H), 3.95-4.04(m, 6H), 3.83-3. 90 (m, 5H), 3.76-3.80 (m, 6H), 3.58-3.75 (m, 8H), 3.48-3.52 (m, 4H), 3.18-3.23 (m, 15H), 2.56-2.62 (m, 2H), 2.48-2.54 (m, 2H), 2.24-2. 35(m, 8H), 2.18-2.20(m, 8H), 2.14(s, 12H), 2.09-2.11(m, 2H), 2.02(s, 12H), 1.90-1.98(m, 28H), 1.55-1.72(m, 28H), 1.28-1.32(m, 8H)

[0873] Example 12 Preparation of Compound E12

change

[0874] 1 Preparation of compound 14

change

[0875] 2. Preparation of Compound 6 [ka] Compound 14 (5.00 g, 4.49 mmol) was dissolved in methanol (100 mL) at room temperature, and 10% mass fraction of wet Pd / C (1.00 g, 9.40 mmol) was added to the solution. Hydrogen was replaced three times, and the reaction mixture was stirred under hydrogen atmosphere (14.696 psi) at 25 °C for 18 h. Thin layer chromatography (dichloromethane / methanol = 10 / 1, phosphomolybdic acid coloring) showed complete consumption of the raw material and the formation of a new spot. The reaction solution was filtered, and the filtrate was concentrated under reduced pressure to give compound 6 (about 4.2 g) as a white solid. 1 H NMR (400MHz, CH3OD) δ5.32-5.34(m, 2H), 5.07-5.11(m, 2H), 4.631-4.63(m, 2H), 4.01-4.17(m, 9H), 3.92-3.96(m, 2H), 3.68-3.75 (m, 2H), 3.61-3.66(m, 4H), 3.54-3.56(m, 4H), 3.32-3.43(m, 6H), 2.25-2.34(m, 2H), 2.15(s, 6H), 2.03(s, 6H), 1.92-1.96(m, 12H)

[0876] 3 Preparation of compound 2 [ka] Compound 1 (10.0 g, 20.0 mmol) was dissolved in ethanol (100 mL), and 10% mass fraction Pd / C (1.00 g, 9.39 mmol) was added. Hydrogen was replaced three times, and the reaction solution was reacted under H2 (15 psi) atmosphere at 45 °C for 18 hours. Liquid chromatography / mass spectrometry detected the MS response that the product should have, and the reaction solution was filtered through Celite to remove Pd / C, washed three times with ethanol (10.0 ml x 3), and the filtrates were combined and concentrated to obtain yellow oily product 2. LCMS: m / z = 320.3 (M + H) + .

[0877] 4. Preparation of Compound 3 [ka] Compound 2 (1.00 g, 3.17 mmol) is dissolved in tetrahydrofuran (10.0 mL) and H2O (10.0 mL) at 25 °C, and sodium bicarbonate (798 mg, 9.51 mmol), CbzCl (1.18 g, 4.75 mmol) are added to it. The reaction solution is stirred at 25 °C for 1 hour. Liquid chromatography / mass spectrometry shows the complete reaction of the raw materials, the appearance of the MS value of the target product. Thin layer chromatography (petroleum ether: ethyl acetate = 3: 1) shows the complete reaction of the raw materials, the formation of a new spot. 150.0 ml of ethyl acetate is added to the reaction solution, and the mixture solution is washed with saturated saline three times (15.0 ml × 3). The organic phase is dried with anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product is separated and purified by column (petroleum ether: ethyl acetate = 3: 1) to obtain colorless oily compound 3 (about 770 mg). 1 H NMR (400MHz, CD3OD) δ7.25-7.39(m, 5H), 5.08(s, 2H), 4.00(s, 4H), 3.88-3.93(m, 1H), 3.62(d, J=5.6Hz, 4H), 1.47(s, 18H). LCMS:m / z=454.3(M+H) + .

[0878] 5. Preparation of Compound 4 [ka] Compound 3 (770 mg, 1.71 mmol) is dissolved in anhydrous dichloromethane (6.00 mL) at 25°C, and TFA (3.00 mL, 39.1 mmol) is added to it. The reaction solution is stirred at 25°C for 12 hours. Liquid chromatography / mass spectrometry indicates complete reaction of the raw materials, and the MS value of the product is generated. Thin layer chromatography (petroleum ether: ethyl acetate = 3:1) indicates complete reaction of the raw materials. The reaction solution is directly concentrated under reduced pressure to obtain the product 4 (about 770 mg) as a colorless oil. 1 H NMR (400MHz, CD3OD) δ7.24-7.39(m, 5H), 5.08(s, 2H), 4.10(s, 4H), 3.90-3.95(m, 1H), 3.57-3.68(m, 4H). LCMS:m / z=342.1(M+H) + .

[0879] 6 Preparation of compound 5 [ka] Compound 4 (770 mg, 2.25 mmol) is dissolved in anhydrous dichloromethane (6.00 mL) at 25 °C, and pentafluorophenol (913 mg, 4.96 mmol) and EDCI (432 mg, 2.25 mmol) are added. The mixture is stirred at 25 °C for 1 h. Liquid chromatography / mass spectrometry indicates the formation of product MS. Thin layer chromatography (petroleum ether: ethyl acetate = 3:1) indicates the formation of a new spot. The reaction solution is concentrated under reduced pressure to obtain the crude product, which is purified by column chromatography (petroleum ether: ethyl acetate = 3:1) to obtain compound 5 (about 500 mg) as a colorless oil. 1 H NMR (400MHz, CD3OD) δ7.20-7.44(m, 5H), 5.08(s, 2H), 4.58(s, 4H), 3.95-4.05(m, 1H), 3.71-3.81(m, 4H). LCMS:m / z=674.0(M+H) + .

[0880] 7 Preparation of compound 7 [ka] Compound 5 (500 mg, 0.742 mmol) and compound 6 (2.72 g, 1.86 mmol) are dissolved in DMF (6.00 mL) at 25 °C, and DIEA (0.123 mL, 0.742 mmol) is added to it. The mixture solution is stirred at 25 °C for 12 hours. Liquid chromatography / mass spectrometry shows the complete reaction of the raw materials and the formation of the product. Thin layer chromatography (petroleum ether: ethyl acetate = 3: 1, T1) shows the complete reaction of the raw materials. The reaction solution is concentrated under reduced pressure to obtain the crude product. The crude product is purified by column chromatography (dichloromethane: methanol = 10: 1, T2) to obtain compound 7 (about 860 mg) as a yellow solid. 1H NMR (400MHz, CD3OD) δ7.25-7.45(m, 5H), 5.34(d, J=3.2Hz, 4H), 5.07-5.11(m, 4H), 4.60-4.71( m, 4H), 4.35-4.44(m, 2H), 4.13-4.16(m, 6H), 4.10(d, J=3.6Hz, 2H), 4.08(d, J=3.6Hz, 2H), 4.0 0-4.06(m, 9H), 3.88-3.96(m, 4H), 3.67-3.75(m, 15H), 3.61-3.62(m, 8H), 3.47-3.56(m, 9H), 3 .34-3.40(m, 8H), 2.25-2.35(m, 4H), 2.14(s, 12H), 2.01-2.04(m, 13H), 1.94(d, J=7.6Hz, 25H). LCMS:m / z=1133.3(M / 2+H) + .

[0881] 8 Preparation of compound 8 [ka] Dissolve compound 7 (350 mg, 0.155 mmol) in ethanol (6.00 mL) at 25 °C, add 10% mass fraction Pd / C (500 mg, 4.70 mmol), and stir the mixture at 25 °C for 12 h. Liquid chromatography / mass spectrometry indicates complete reaction of the raw materials. Filter the reaction solution through Celite, and concentrate the filtrate under reduced pressure to obtain compound 8 (approximately 200 mg) as a yellow oil. 1 H NMR (400MHz, CD3OD) δ5.34(s, 4H), 5.07-5.11(m, 4H), 4.65(dd, J=9.6, 11.2H z, 4H), 4.36-4.45(m, 2H), 4.12-4.20(m, 8H), 4.00-4.12(m, 13H), 3.91-3.95( m, 5H), 3.69-3.74(m, 5H), 3.62-3.63(m, 12H), 3.53-3.56(m, 10H), 3.36-3.4 2(m, 8H), 2.29-2.40(m, 4H), 2.15(s, 12H), 2.03(s, 12H), 1.90-1.97(m, 24H). LCMS:m / z=1066.3(M / 2+H) + .

[0882] 9 Preparation of Compound 10 [ka] Compound 8 (450 mg, 0.211 mmol) is dissolved in DMF (6.00 mL) at 25° C., HBTU (104 mg, 0.274 mmol), DIEA (0.105 mL, 0.633 mmol) are added, and finally, compound 9 (135 mg, 0.211 mmol, synthesized according to the method of compound Int-6 in Example 13) is added to it, and the mixture is stirred at 25° C. for 12 hours. Thin layer chromatography (dichloromethane:methanol=10:1) shows the formation of a new spot. The mixture is washed three times (5 ml×3) with dichloromethane (100 ml), saturated sodium bicarbonate (5 ml×3) and brine, respectively. The organic phase is dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product is purified by column chromatography (dichloromethane:methanol=10:1) to obtain compound 10 (about 350 mg) as a yellow oil. LCMS: m / z=1371.8 (M / 2+H) + .

[0883] 10. Preparation of E12 [ka] Compound 10 (350 mg, 0.127 mmol) is dissolved in anhydrous dichloromethane (5.00 mL) at 25 degrees Celsius, compound 11 (76.5 mg, 0.765 mmol) is added thereto, DIEA (0.126 mL, 0.765 mmol) and DMAP (3.89 mg, 0.032 mmol) are added to the above mixture solution in sequence. The reaction solution is stirred at 25 degrees Celsius for 3 hours. Liquid chromatography / mass spectrometry shows the complete reaction of the raw materials and the formation of the product. 200 ml of dichloromethane is added to the mixture solution, washed with saturated saline three times (15.0 ml x 3), the organic phase is dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product is separated by high performance liquid chromatography (chromatography column: Waters Xbridge BEH C18 100*30mm*10um, mobile phase: water-ACN, gradient: 17%-57% / 11min, flow rate: 50ml / min) to obtain a white solid E12 (35mg, 0.012mmol, 9.65%).

[0884] LCMS(ESI):m / z=1422.0(M / 2+H) + ,

[0885] 1H NMR (400MHz, CD3OD) δ7.42-7.44(m, 2H), 7.19-7.35(m, 7H), 6.81-6.92(m, 4H), 5.32-5.34(m, 4H), 5.06-5.12(m, 4H), 4.59-4.68(m, 4) H), 4.40-4.42(m, 2H), 4.30-4.34(m, 1H), 4.18-4.28(m, 2H), 4.11-4.18(m, 8H), 3.99-4.11(m, 14H), 3.82-3.99(m, 7H), 3.78-3.79(m, 6H), 3.67-3.75(m, 8H), 3.65-3.66(m, 1H), 3.60-3.62(m, 10H), 3.52-3.54(m, 8H), 3.47-3.49(m, 1H), 3.37-3.38(m, 7H), 3.11-3.18( m, 2H), 2.50-2.61(m, 4H), 2.21-2.41(m, 10H), 2.14(s, 12H), 2.02(s, 12H), 1.93-1.94(m, 24H), 1.49-1.66(m, 4H), 1.29-1.31(m, 8H).

[0886] Example 13 Preparation of compound E13 [ka]

[0887] 1. Preparation of Compound 1-2 [ka] Compound 1-1 (38.0 g, 231 mmol), TsNHBoc (75.4 g, 278 mmol), K2CO3 (6.40 g, 46.3 mmol) and TEBA (5.27 g, 23.1 mmol) are added to a three-neck flask under room temperature conditions. The mixture is reacted at 95°C for 2 hours, then cooled to 25°C. TLC (PE / EA=2 / 1, UV 254 nm) indicates the completion of the reaction. The reaction solution is diluted with water (500 mL), extracted with dichloromethane (200 mL x 3), the organic phase is concentrated under reduced pressure, and the crude product is separated and purified by column (PE / EA=10 / 1 to 3 / 1) to obtain colorless oily compound 1-2 (40.0 g). 1H NMR (400MHz, CDCl3) δ7.72(d, J=8.4Hz, 2H), 7.28-7.39(m, 7H), 4.72-4.86(m, 2H) , 4.42-4.56(m, 2H), 3.52-3.61m, 2H), 3.14-3.39(m, 2H), 2.43(s, 3H), 1.47(s, 9H)

[0888] 2. Preparation of Compounds 1-4 [ka] Compound 1-2 (79.0 g, 181 mmol), compound 1-3 (4.21 g, 30.5 mmol), and TEBA (3.47 g, 15.2 mmol) were reacted at 95 °C for 3 hours, and TLC (PE / EA = 2 / 1, UV 254 nm) showed the completion of the reaction, and the reaction solution was diluted with water (500 mL), extracted with dichloromethane (200 mL × 3), and the organic phase was concentrated under reduced pressure, and the crude product was separated and purified by column (PE / EA = 10 / 1 to 3 / 1) to obtain colorless oily compound 1-4 (about 40.0 g). 1 H NMR (400MHz, CDCl3) δ7.64-7.77(m, 2H), 7.27-7.38(m, 13H), 5.05-5.16(m, 1H), 4.50-4.59(m, 4H), 4.03-4.11 (m, 1H), 3.62-3.76(m, 2H), 3.44-3.58(m, 3H), 3.25-3.33(m, 2H), 3.15-3.20(m, 1H), 2.42(s, 3H), 1.46(s, 9H)

[0889] 3 Preparation of compounds 1-5 [ka] Under ice bath conditions, compound 1-4 (77.0 g, 128 mmol) and Et3N (28.6 mL, 205 mmol) are added successively to dichloromethane (800 mL), and methanesulfonyl chloride (24.2 g, 212 mmol) is slowly added dropwise. The reaction is carried out at 25° C. for 2 hours, and LCMS shows the completion of the reaction. The reaction solution is washed with water (800 mL), and the organic phase is concentrated under reduced pressure to obtain colorless oily compound 1-5 (about 90.0 g). LCMS (ESI): m / z=700.1 (M+Na).+ , 1 H NMR (400MHz, CDCl3) δ7.68(d, J=8.4Hz, 2H), 7.28-7.39(m, 12H), 5.10-5.20(m, 1H), 4.93-5.02(m, 1H), 4.52-4.61(m, 4 H), 3.72-3.88(m, 2H), 3.52-3.67(m, 4H), 3.27-3.36(m, 1H), 3.15-3.22(m, 1H), 3.04(s, 3H), 2.43(s, 3H), 1.43(s, 9H). LCMS:m / z=700.1(M+Na) + .

[0890] 4. Preparation of Compounds 1-6 [ka] Compound 1-5 (90.0 g, 133 mmol) and potassium carbonate (91.8 g, 664 mmol) are added to MeOH (900 mL) at room temperature. The mixture is reacted at 66 °C for 2 hours, and TLC (PE / EA = 2 / 1, UV 254 nm) shows the formation of a new spot. The organic phase is concentrated under reduced pressure, the reaction solution is diluted with water (200 mL), extracted with dichloromethane (200 mL x 3), the organic phase is concentrated under reduced pressure, and the crude product is separated and purified by column (PE / EA = 30 / 1 to 2 / 1) to obtain compound 1-6 (about 64.0 g) as a white solid. 1 H NMR (400MHz, CDCl3) δ7.62(d, J=8.4Hz, 2H), 7.27-7.39(m, 12H), 4.48-4.62( m, 4H), 3.93-4.08(m, 2H), 3.55-3.69(m, 4H), 2.84-3.10(m, 4H), 2.44(s, 3H)

[0891] 5 Preparation of Compounds 1-7 [ka] Compound 1-6 (69.0 g, 143 mmol) and magnesium scraps (54.7 g, 2.28 mol) are added to MeOH (400 mL) at room temperature and reacted at 66 °C for 1 hour. TLC (DCM / MeOH = 10 / 1, UV 254 nm) shows complete reaction of the raw materials and the formation of a new spot. The reaction solution is diluted with water (3000 mL) and saturated aqueous ammonium chloride solution (3000 mL), extracted with dichloromethane (1000 mL x 3), the organic phase is washed with saturated sodium bicarbonate (300 mL x 3), and the organic phase is concentrated under reduced pressure to obtain compound 1-7 (about 32.0 g) as a colorless oil. 1 H NMR (400MHz, CDCl3) δ7.27-7.40(m, 10H), 4.57(s, 4H), 3.90-4.00(m, 2H), 3.59-3.69(m, 4H), 2.77-3.04(m, 4H)

[0892] 6 Preparation of Compounds 1-8 [ka] Compound 1-7 (8.00 g, 24.4 mmol) was added to HCl (100 mL, 12 M) at room temperature and reacted at 50° C. for 2 hours. The reaction solution was concentrated under reduced pressure to obtain colorless oily compound 1-8 (approximately 3.60 g, HCl salt). 1 H NMR (400MHz, CDCl3) δ4.12-4.22(m, 2H), 3.78(d, J=4.4Hz, 4H), 3.17-3.30(m, 4H)

[0893] 7 Preparation of compound Int-1 [ka] Compound 1-8 (8.29 g, 24.5 mmol) is added to pyridine (50 mL, 618 mmol) at room temperature, followed by DMTrCl (4.77 g, 12.3 mmol). The reaction is allowed to react at 25° C. for 18 hours, and TLC (DCM / MeOH=10 / 1, UV254 nm) shows the formation of the product. The reaction solution is concentrated under reduced pressure, diluted with saturated ammonium chloride (200 mL), extracted with DCM (100 mL×3), and the organic phase is concentrated under reduced pressure. The crude product is subjected to column chromatography (DCM / MeOH=99 / 1 to 10 / 1) and separated to obtain Int-1 (approximately 2.6 g, 5.78 mmol, 23.6%) as a yellow solid. 1 H NMR (400MHz, CDCl3) δ7.38-7.50(m, 2H), 7.25-7.36(m, 6H), 7.18-7.25(m, 1H), 6.87(d, J=8.8Hz, 4H), 4.26-4.40(m, 1H) ), 3.94-4.05(m, 1H), 3.85(d, J=4.4Hz, 2H) 3.73-3.82(m, 6H), 3.34-3.41(m, 1H), 3.23-3.30(m, 3H), 3.05-3.23(m, 2H)

[0894] 8 Preparation of Compound 3 [ka] Compound 1 (1.00 g, 3.05 mmol) is dissolved in anhydrous N,N-dimethylformamide (10.0 mL) at 25 °C, and HOBt (0.54 g, 3.97 mmol), EDCI (0.76 g, 3.97 mmol), and DIEA (1.51 mL, 9.16 mmol) are added in sequence. The mixture is stirred at 25 °C for 2 hours. Liquid chromatography / mass spectrometry indicates complete reaction of the raw materials and the formation of the product. Thin layer chromatography plate (dichloromethane:methanol = 10:1) indicates complete reaction of the raw materials and the formation of the product. Dichloromethane (200 ml) is added to the mixture, and it is washed three times with saturated aqueous citric acid solution (20 ml x 3), saturated sodium bicarbonate solution (20 ml x 3), and saturated saline solution (20 ml x 3). The organic phase is dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (dichloromethane:methanol=10:1) to give compound 3 (about 1.00 g) as a yellow oil. 1 H NMR (400MHz, CD3OD) δ7.21-7.41(m, 10H), 4.47-4.60(m, 4H), 3.88-4.01(m, 2H), 3.75(dd, J=3.2, 13.2 Hz, 1H), 3.63-3.65(m, 3H), 3.40-3.63(m, 7H), 2.27-2.34(m, 4H), 1.50-1.60(m, 4H), 1.28-1.34(m, 8H)

[0895] LCMS: m / z=526.8 (M+H) + .

[0896] 9 Preparation of compound 4 [ka] Compound 3 (380 mg, 0.20 mmol) is dissolved in anhydrous methanol (10.0 mL) at 25° C., and 10% mass fraction wet Pd / C (500 mg, 4.70 mmol) is added thereto. The reaction solution is stirred at 50° C. for 12 h. Liquid chromatography / mass spectrometry indicates the formation of the product. The reaction solution is filtered through Celite, and the filtrate is concentrated under reduced pressure to give compound 4 (approximately 600 mg) as a yellow oil. 1H NMR (400MHz, CD3OD) δ3.79-3.89(m, 2H), 3.73(dd, J=3.6, 13.2Hz, 1H), 3.64-3.68(m, 5H), 3.57-3.62(m, 2H), 3.48-3.56(m, 2H), 3.38-3.47(m, 1H), 1.58-1.62(m, 5H), 1.34(d, J=2.8Hz, 11H), 1.30-1.37(m, 1H). LCMS:m / z=346.5(M+H) + .

[0897] 10 Preparation of Compound 5 [ka] Compound 4 (600 mg, 1.74 mmol) is dissolved in pyridine (3.00 mL) at 25 °C, and DMTrCl (883 mg, 2.61 mmol) is added. The reaction solution is stirred at 25 °C for 1 h. Liquid chromatography / mass spectrometry indicates the formation of the product. Thin layer chromatography (dichloromethane:methanol = 10:1) indicates the complete reaction of the raw materials and the formation of the product. Dichloromethane (200 ml) is added to the mixture, and the mixture is washed three times with saturated sodium bicarbonate solution (20 ml x 3) and brine (20 ml x 3), respectively. The organic phase is dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product is purified by column chromatography (petroleum ether:ethyl acetate = 3:1) to obtain compound 5 (about 475 mg) as a colorless liquid. 1 H NMR (400MHz, CD3OD) δ7.41-7.43(m, 2H), 7.16-7.34(m, 7H), 6.83-6.87(m, 4H), 3.88-4.04(m, 2H), 3.77-4.09(m, 7H), 3.61-3.68(m, 4H), 3.4 3-3.58(m, 3H), 3.33-3.38(m, 1H), 3.14-3.20(m, 1H), 3.05(dd, J=8.4, 13.2Hz, 1H), 2.20-2.42(m, 4H), 1.51-1.60(m, 4H), 1.25-1.39(m, 8H). LCMS: m / z=648.3 (M+H) + .

[0898] 11 Preparation of compound Int-6 [ka] Compound 5 (475 mg, 0.73 mmol) is dissolved in tetrahydrofuran (8.00 mL) and H2O (2.00 mL) at 25°C, and lithium hydroxide monohydrate (36.9 mg, 0.89 mmol) is added to it. The reaction solution is stirred at 25°C for 12 hours. Thin layer chromatography (dichloromethane:methanol=10:1) shows the complete reaction of the raw materials and the formation of the product. The reaction solution is concentrated under reduced pressure and then directly freeze-dried in a freeze dryer to obtain compound 6 (about 430 mg) as a white solid.

[0899] 12 Preparation of compound 7 [ka] At 25°C, compound Int-6 (100mg, 0.156mmol) is dissolved in anhydrous N,N-dimethylformamide (3.00mL), and HBTU reagent (89.0mg, 0.234mmol), DIEA (0.078mL, 0.47mmol) and intermediate C (254mg, 0.156mmol, synthesized according to the method of intermediate 2-2 in Example 26) are added in sequence. The reaction solution is stirred at 25°C for 12 hours. Thin layer chromatography (dichloromethane:methanol=10:1) shows the formation of a new spot. Dichloromethane (200ml) is added to the mixture, and the mixture is washed three times with saturated sodium bicarbonate solution (20ml x 3) and three times with brine (20ml x 3). The organic phase is dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (dichloromethane:methanol=10:1) to give compound 7 (about 140 mg) as a yellow oil. 1H NMR (400MHz, CD3OD) δ7.43(d, J=8.4Hz, 2H), 7.18-7.34(m, 7H), 6.81-6.91(m, 4H), 5.34(d, J=2.8Hz, 3H), 5.09-5 .11(m, 3H), 4.62-4.66(m, 3H), 4.27-4.35(m, 2H), 4.07-4.17(m, 9H), 4.00-4.07(m, 4H), 3.89-3.98(m, 5H), 3.79 (d, J=2.4Hz, 6H), 3.69-3.75(m, 5H), 3.60-3.64(m, 9H), 3.52-3.56(m, 10H), 3.43-3.50(m, 4H), 3.39-3.40(m, 1H) ), 2.25-2.40(m, 8H), 2.13-2.15(m, 9H), 2.02(s, 9H), 1.94-1.96(m, 18H), 1.48-1.66(m, 4H), 1.21-1.36(m, 11H)

[0900] 13. Preparation of E13 [ka] Compound 7 (400 mg, 0.187 mmol) is dissolved in anhydrous dichloromethane (5.00 mL), DIEA (0.18 mL, 1.12 mmol), DMAP (5.71 mg, 0.047 mmol) are added in sequence, and finally compound 8 (112 mg, 1.12 mmol) is added. The mixture is stirred at 25°C for 3 hours. Thin layer chromatography (dichloromethane:methanol = 10:1) shows the formation of a new spot. Dichloromethane (200 ml) is added to the mixture, and the mixture is washed three times with saturated saline (15.0 ml x 3). The organic phase is dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product is separated by high performance liquid chromatography (chromatography column: Welch Xtimate C18 150*25mm*5um, mobile phase: water-ACN, gradient: 27%-57% / 11min, flow rate: 25ml / min) to obtain compound E13 (74.8mg, 0.033mmol, yield 17.9%) as a white solid. LCMS (ESI): m / z=1149.5(M / 2+H) + , 1H NMR (400MHz, CD3OD) δ7.42-7.44(m, 2H), 7.18-7.34(m, 7H), 6.83-6.90(m, 4H), 5.33-5.34(m, 3H), 5.08-5.11(m, 3H), 4.61-4.68(m, 3 H), 4.27-4.36(m, 2H), 4.19-4.27(m, 1H), 4.10-4.19(m, 7H), 4.08(s, 2H), 4.00-4.07(m, 4H), 3.90-3.94(m, 4H), 3.83-3.89(m, 1H), 3 .79(d, J=2.4Hz, 6H), 3.70-3.72(m, 4H), 3.53-3.54(m, 15H), 3.44-3.48(m, 3H), 3.35-3.40(m, 4H), 3.10-3.23(m, 2H), 2.53-2.63(m, 4H), 2.22-2.40(m, 8H), 2.11-2.16(m, 9H), 2.05-2.10(m, 2H), 2.02(s, 9H), 1.90-1.97(m, 19H), 1.48-1.65(m, 4H), 1.29-1.32(m, 8H)

[0901] Example 14 Preparation of Compound E14

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[0902] 1 Preparation of compound 3

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[0903] 2. Preparation of compound E14 [ka] Compound 3 (612 mg, 0.254 mmol) is dissolved in DCM (5 mL) at room temperature, DIEA (0.252 mL, 1.524 mmol), DMAP (7.76 mg, 0.063 mmol) and 4 (152.49 mg, 1.524 mmol) are added and reacted at 25 °C for 3 h. Liquid chromatography / mass spectrometry indicates complete reaction of the raw materials and formation of the product. Dichloromethane (30 mL) is added to dilute the reaction solution, and the organic phase is washed three times with saturated NaCl solution (30 mL × 3) and dried over anhydrous sodium sulfate. The reaction solution was concentrated under reduced pressure, and the obtained crude product was subjected to preparative high performance liquid chromatography (preparative HPLC, column: Waters Xbridge BEH C18 100×30 mm×10 um, mobile phase: water-ACN, B%: 17%-57%, 12 min) to obtain a white solid E14 (160.1 mg, yield 25.1%, purity 73.34%).

[0904] 1 H NMR (400MHz, CD3OD) δ7.43(d, J=8.0Hz, 2H), 7.31(d, J=8.8Hz, 5H), 7.17-7.27(m, 2H), 6.85-6.90(m, 4H), 5.33(d, J=3.2Hz, 3H), 5.06(dd, J=3.6, 11.2Hz, 3H), 4.56(d, J=8.4Hz, 3H), 4.05-4.20(m, 10H), 3.94-4.03(m, 5H), 3.80-3.94(m, 5H), 3.79(d , J=2.4Hz, 6H), 3.57-3.72(m, 15H), 3.40-3.57(m, 4H), 3.18-3.26(m, 15H), 2.48-2.61(m, 4H), 2.42(t, J=6.0Hz, 6H), 2.16- 2.25(m, 8H), 2.14(s, 9H), 2.02(s, 9H), 1.92-1.96(m, 18H), 1.62-1.71(m, 12H), 1.51-1.62(m, 10H), 1.30(d, J=13.2Hz, 8H)

[0905] LCMS: m / z=1283.5 ((M+59) / 2) + .

[0906] Example 15 Preparation of Compound E15 [ka]

[0907] 1 Preparation of compound 2_3 [ka] Compound 2_1 (11.6 g, 58.59 mmol) and compound 2_2 (4.5 g, 29.29 mmol) are dissolved in MeOH (100 mL). The reaction solution is reacted at 20 °C for 2 hours, and then sodium cyanoborohydride (7.37 g, 117.180 mmol) is added. The reaction solution is reacted at 20 °C for 16 hours. TLC (DCM / MeOH = 10 / 1) shows the formation of a new spot. 60 mL of DCM and 40 mL of water are added to the reaction solution, the reaction solution is separated, and the aqueous phase is extracted with DCM (55 mL * 1). The combined organic phase is washed with saturated brine (50 mL × 1), dried over anhydrous sodium sulfate, filtered, and centrifuged to obtain the crude product. The crude product is purified by column chromatography (DCM / MeOH = 1 / 0 ~ 10 / 1) to obtain compound 2_3 (about 2.6 g) as a white solid. 1 H NMR (400MHz, MeOD-d4) δ4.11(brd, J=13.6Hz, 2H), 3.86-3.93(m, 1H), 3.42-3.77(m, 5H), 2.92(brd, J=11.2H) z, 1H), 2.69-2.86(m, 3H), 2.24-2.19-2.51(m, 2H), 2.00-2.12(m, 1H), 1.79-1.97(m, 2H), 1.39-1.52(m, 10H)

[0908] 2 Preparation of compound 2 [ka] Compound 2_3 (6.5 g, 21.6 mmol) is dissolved in DCM (50 mL) and HCl / EtOAc (20 mL) is added. The reaction solution is reacted at 20 degrees Celsius for 1 hour. TLC (DCM / MeOH=10 / 1) shows the formation of a new spot. The reaction solution is centrifuged to obtain the crude product compound 2 (about 5.1 g) as a white solid. 1H NMR (400MHz, MeOD-d4) δ4.14-4.22(m, 1H), 3.90-4.04(m, 2H), 3.53-3.73(m, 7H), 2.98-3.24(m, 4H), 2.42-2.54(m, 2H), 2.03-2.17(m, 2H).

[0909] 3 Preparation of compound 3 [ka] Compound 1 (1500mg, 0.896mmol), HATU (511mg, 1.34mmol) and DIEA (0.741mL, 4.48mmol) are dissolved in DMF (15.0mL). The reaction solution is reacted at 20 degrees Celsius for 30 hours. Then, compound 2 (269mg, 1.34mmol) is added and the reaction is carried out at 20°C for 2 hours. LCMS shows the formation of MS value of product. TLC (DCM / MeOH=5 / 1) shows the formation of a new spot. 15mL of DCM and 20mL of saturated aqueous sodium bicarbonate solution are added to the reaction solution, the reaction solution is separated, and the aqueous phase is extracted with DCM (15.0mLx1). The combined organic phase is washed with saturated brine (25mLx1), dried over anhydrous sodium sulfate, filtered and centrifuged to obtain the crude product. The crude product was purified by column chromatography (DCM / MeOH=1 / 05 / 1) to give compound 3 (about 850 mg) as a yellow solid product. 1H NMR (400MHz, DMSO-d6) δ7.79-7.95(m, 6H), 5.21(d, J=3.2Hz, 3H), 4.98(dd, J=3.2, 11.2Hz, 3H), 4.65(s, 1H), 4.55(d, J=8.4Hz, 3H), 4.37(d, J=12.0Hz, 1H), 4.03(s, 9H), 3.72-3.95(m, 8H), 3.45-3.66(m, 18H), 3 .37-3.42(m, 8H), 3.08-3.27(m, 16H), 2.93-3.06(m, 1H), 2.76-2.89(m, 1H), 2.62-2.75(m, 1H), 2.53-2. 59(m, 3H), 2.29(t, J=6.4Hz, 6H), 2.12(s, 9H), 2.00(s, 9H), 1.89(s, 9H), 1.77(s, 9H), 1.23-1.30(m, 6H). LCMS:m / z=929.1(M / 2+H) + .

[0910] 4. Preparation of E15 [ka] Compound 3 (850 mg, 0.458 mmol) is dissolved in acetonitrile, then centrifuged to remove water in the compound, then dissolved in DCM (10.0 mL) under nitrogen atmosphere, DCI (27.0 mg, 0.229 mmol) is added, compound 4 (276 mg, 0.916 mmol) is slowly added dropwise, and the reaction solution is reacted at 20 ° C for 1 hour. LCMS shows the disappearance of raw materials, the reaction solution is cooled to 0 ° C, 6 ml of saturated aqueous sodium bicarbonate solution and 6 ml of saturated saline solution are added dropwise, diluted with 100 ml of dichloromethane, separated, and washed with 50 ml of a 1:1 mixed aqueous solution of saturated aqueous sodium bicarbonate solution and saturated saline solution. The organic phase is dried over anhydrous sodium sulfate, filtered, and centrifuged to obtain the crude product. The crude product is diluted with 15 ml of dichloromethane, slowly dripped into 60 ml of methyl tert-butyl ether, filtered, the filter cake is washed with methyl tert-butyl ether three times, the filter cake is dissolved in acetonitrile, and then centrifuged to remove the methyl tert-butyl ether in the compound. The product E15 (555 mg, 0.269 mmol, 58.9%) is obtained as a yellow solid.

[0911] LCMS(ESI):m / z=1087.0((M+59*2) / 2+H) + ,

[0912] 1 H NMR (400MHz, DMSO-d6) δ7.85-7.90 (m, 3H), 7.75-7.84 (m, 3H), 5.20-5.24 (m, 3H), 4.98 (dd, J=3.6, 11.2Hz, 3H), 4.51 -4.58(m, 3H), 3.98-4.07(m, 10H), 3.83-3.91(m, 4H), 3.69-3.82(m, 6H), 3.47-3.62(m, 23H), 3.35-3.42(m, 8H), 3.15 -3.26 (m, 15H), 2.95-3.02 (m, 1H), 2.73-2.84 (m, 3H), 2.63-2.65 (m, 1H), 2.36-2.42 (m, 2H), 2.29 (t, J=6.4Hz, 6H), 2 .10-2.13 (m, 1H), 2.10 (s, 9H), 2.00 (s, 9H), 1.89 (s, 9H), 1.69-1.79 (m, 11H), 1.24-1.39 (m, 2H), 1.03-1.17 (m, 12H).

[0913] 31 P NMR(400MHz, DMSO-d6)δ-147.834

[0914] Example 16 Preparation of Compound E16

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[0915] 1 Preparation of compound 3

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[0916] 2. Preparation of E16 [ka] Compound 3 (850 mg, 0.428 mmol) is dissolved in acetonitrile, centrifuged to remove water, then dissolved in DCM (10.0 mL), DCI (25.2 mg, 0.214 mmol) is added, compound 4 (257 mg, 0.855 mmol) is added under nitrogen atmosphere, nitrogen replacement is performed three times, and the reaction is carried out at 20 ° C for 1 hour. LCMS shows the disappearance of the raw material, the reaction solution is cooled to 0 ° C, 6 ml of saturated aqueous sodium bicarbonate and 6 ml of saturated saline are added dropwise, diluted with 100 ml of dichloromethane, separated, and washed with 50 ml of a 1:1 mixed aqueous solution of saturated aqueous sodium bicarbonate and saturated saline. The organic phase is dried over anhydrous sodium sulfate, filtered, and centrifuged to obtain the crude product. The crude product is diluted with 15 ml of dichloromethane, slowly dripped into 60 ml of methyl tert-butyl ether, filtered, the filter cake is washed with methyl tert-butyl ether three times, the filter cake is dissolved in acetonitrile, and then centrifuged to remove the methyl tert-butyl ether in the compound. The product E16 (591 mg, 0.270 mmol, 63.17%) is obtained as a yellow solid.

[0917] LCMS(ESI): m / z=2246.3(M+59+H) + , LCMS:m / z=1053.7((M-83) / 2+H) + .

[0918] 1H NMR (400MHz, DMSO-d6) δ7.87-7.93 (m, 3H), 7.78-7.84 (m, 3H), 5.19-5.24 (m, 3H), 4.93-5.01 (m, 3H), 4.53-4.58 (m , 3H), 3.96-4.08(m, 10H), 3.83-3.92(m, 4H), 3.69-3.82(m, 6H), 3.44-3.64(m, 37H), 3.37-3.41(m, 6H), 3.14-3.2 6(m, 15H), 2.92-3.00(m, 1H), 2.72-2.84(m, 3H), 2.63-2.68(m, 1H), 2.35-2.44(m, 2H), 2.28(t, J=6.0Hz, 6H), 2.1 6-2.22 (m, 1H), 2.10 (s, 9H), 2.00 (s, 9H), 1.89 (s, 9H), 1.69-1.80 (m, 11H), 1.20-1.40 (m, 2H), 1.00-1.17 (m, 12H).

[0919] 31 P NMR (400MHz, DMSO-d6) δ-147.815.

[0920] Example 17 Preparation of Compound E17

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[0921] 1 Preparation of compound 3

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[0922] 2. Preparation of E17 [ka] Compound 3 (890 mg, 0.431 mmol) is dissolved in acetonitrile, then centrifuged to remove water in the compound, then dissolved in DCM (10.0 mL) under nitrogen atmosphere, DCI (25.4 mg, 0.216 mmol) is added, compound 4 (260 mg, 0.863 mmol) is slowly added dropwise, and the reaction solution is reacted at 20 ° C for 1 hour. LCMS shows the disappearance of raw materials, the reaction solution is cooled to 0 ° C, 6 ml of saturated sodium bicarbonate aqueous solution and 6 ml of saturated saline are added dropwise, diluted with 100 ml of dichloromethane, separated, and washed with 50 ml of a 1:1 mixed aqueous solution of saturated sodium bicarbonate aqueous solution and saturated saline. The organic phase is dried over anhydrous sodium sulfate, filtered, and centrifuged to obtain the crude product. The crude product is diluted with 15 ml of dichloromethane, slowly dripped into 60 ml of methyl tert-butyl ether, filtered, the filter cake is washed with methyl tert-butyl ether three times, the filter cake is dissolved in acetonitrile, and then centrifuged to remove the methyl tert-butyl ether in the compound. The product E17 (708 mg, 0.313 mmol, 72.51%) is obtained as a yellow solid.

[0923] LCMS(ESI):m / z=2246.3((M+59*2) / 2+H) + ,

[0924] 1H NMR (400MHz, DMSO-d6) δ7.78-7.86 (m, 6H) 7.71-7.76 (m, 3H), 5.21 (d, J=3.2Hz, 3H), 4.96 (dd, J=3.2, 11.2Hz, 3H), 4.48 (d, J=8.4Hz, 3H), 3.99-4.06(m, 10H), 3.82-3.91(m, 4H), 3.65-3.79(m, 6H), 3.37-3.58(m, 18H), 3.21(s, 8H), 3.00-3.09(m, 13 H), 2.86-2.99(m, 2H), 2.73-2.86(m, 3H), 2.62-2.69(m, 1H), 2.54-2.59(m, 2H), 2.26(t, J=6.0Hz, 6H), 2.15-2.20(m, 1H), 2.10 (s, 9H), 2.04 (t, J=6.8Hz, 6H), 1.99 (s, 9H), 1.89 (s, 9H), 1.68-1.78 (m, 11H), 1.41-1.55 (m, 20H), 1.03-1.17 (m, 12H).

[0925] 31 P NMR(400MHz, DMSO-d6)δ-147.834

[0926] Example 18 Preparation of Compound E18

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[0927] 1 Preparation of compound 3

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[0928] LCMS: m / z=316.1 (M+H) + .

[0929] 2. Preparation of Compound 4 [ka] Compound 3 (1.10 g, 3.49 mmol) is dissolved in pyridine (20.0 mL) at 25 °C, and a solution of DMTrCl (1.18 g, 3.488 mmol) in DCM (10.0 mL) is added. The mixture is stirred at 25 °C for 2 h. LCMS shows the formation of the product. DCM (200 ml) is added to the mixture, and the mixture is washed three times with saturated sodium bicarbonate (20.0 ml x 3) and saturated saline (20 ml x 3), respectively. The organic phase is dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product is purified by TLC (DCM / MeOH = 10 / 1 to 5 / 1) to obtain compound 4 (560 mg, 0.928 mmol, 26.6% yield) as a yellow oil, which is characterized by LCMS.

[0930] LCMS: m / z=640.2 (M+H) + .

[0931] 3. Preparation of compound Int-5 [ka] Compound 4 (560 mg, 0.906 mmol) was added to THF (3.00 mL) and HO (1.50 mL), lithium hydroxide (57.05 mg, 1.360 mmol) was added, and the mixture was reacted at 20°C for 12 hours. TLC (dichloromethane:methanol = 8:1) showed the disappearance of the raw material and the formation of a new spot. The reaction solution was centrifuged, and then 5 mL of water was added and the mixture was lyophilized to obtain the white solid lithium salt product compound Int-5 (630 mg, 1.033 mmol, 113.99%).

[0932] 4 Preparation of compound 7 [ka] Compound 6 (prepared according to the method of compound 4 in Example 7, 190 mg, 0.08 mmol) is dissolved in DCM (5.00 mL) at 25 ° C., Int-5 (50.2 mg, 0.08 mmol), HBTU (39.0 mg, 0.10 mmol) and DIEA (0.04 mL, 0.25 mmol) are added successively. The mixture is stirred at 25 ° C. for 12 hours. LCMS shows the formation of the product. Dichloromethane (60.0 ml) is added to the reaction solution, followed by washing with saturated sodium bicarbonate solution (10.0 ml × 3) and saturated saline (10 ml × 3) three times, the organic phase is dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain crude compound 7 (270 mg, crude product). Use directly in the next step.

[0933] LCMS: m / z=1445.2 (M / 2+H) + .

[0934] 5. Preparation of Compound E18 [ka] Compound 7 (270 mg, crude product) and compound 8 (56.0 mg, 0.56 mmol) are added to DCM (5.00 mL) at 25 °C, followed by DIEA (0.093 mL, 0.56 mmol) and DMAP (2.85 mg, 0.023 mmol). The mixture is stirred at 25 °C for 12 hours, and LCMS shows the disappearance of the raw material. Dichloromethane (100 ml) is added to the reaction solution, and the solution is washed three times with saturated sodium bicarbonate solution (15.0 ml x 3) and saturated saline (15.0 ml x 3). The organic phase is dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain crude product, which is separated by preparative HPLC (chromatography column: Waters Xbridge BEH C18 150*25mm*5um, mobile phase: TEAA-ACN, gradient: 20%-53% / 15min, flow rate: 15ml / min) to obtain product E18 (74mg, 0.025mmol, 26.49%) as white solid. 1 H NMR (400MHz, CD3OD) δ7.38(d, J=7.2Hz, 2H), 7.17-7.32(m, 7H), 6.78-6.94(m, 4H), 5.34(d, J=2.80Hz, 4H), 5.05-5.11(m, 4H), 4.65(dd, J=3.2, 8.8Hz, 4H), 4.08-4.45(m, 14H), 3.88-4.08(m, 13H), 3.75-3.83(m, 7H), 3.66-3.74(m, 8H), 3.58-3.66(m, 24H) ), 3.48-3.58(m, 9H), 3.32-3.48(m, 11H), 3.11-3.21(m, 24H), 3.07(d, J=7.2Hz, 1H), 2.51-2.59(m, 2H), 2.44-2.51(m, 2H), 2. 19-2.40(m, 9H), 2.14(s, 12H), 2.05-2.12(m, 3H), 2.02(s, 13H), 1.95(s, 12H), 1.92(s, 13H), 1.61(s, 3H), 1.16-1.38(m, 44H)

[0935] Example 19 Preparation of compound E19 [ka]

[0936] 1. Preparation of Compound 3 [ka] Compound 2 (630 mg, 3.15 mmol) and compound 1 (1.42 g, 3.15 mmol) are dissolved in MeOH (20.0 mL) at 25 °C, Et3N (0.874 mL, 6.29 mmol) is added, and the reaction solution is stirred at 25 °C for 12 h. NaBH3CN (593 mg, 9.44 mmol) is added, and the reaction solution is stirred at 25 °C for 24 h. LCMS shows the MS value of compound 3. Thin layer chromatography (DCM / MeOH = 10 / 1) shows complete consumption of the reactants, and the formation of a new spot. The reaction solution is diluted with 100 mL of dichloromethane, washed with saturated NaHCO3 solution (10.0 mL x 3), then saturated NaCl solution, and the organic phase is dried over Na2SO4 and centrifuged to obtain the crude product. The crude product was purified by MPLC (DCM / MeOH = 1 / 0 to 10 / 1) to obtain compound 3 (970 mg, 48.65% yield) as a yellow oily liquid. 1 H NMR (400MHz, CD3OD) δ7.38-7.47(m, 2H), 7.16-7.34(m, 7H), 6.85(dd, J=8.8, 1.6Hz, 4H), 3.93-4.01(m, 1H), 3.77(s, 6H), 3.64-3.71(m, 2H), 3.64(s, 3H), 3.60-3.60(m, 1H), 3.52-3.59(m, 1H), 3.38(dd, J=9.2, 6.0Hz, 1H), 3.16-3.23(m, 1H), 2.35-2.53( m, 3H), 2.14-2.32(m, 5H), 1.53-1.64(m, 2H), 1.40-1.49(m, 2H), 1.26-1.36(m, 10H).

[0937] 2. Preparation of Compound 4 [ka] Compound 3 (570 mg, 0.899 mmol) was dissolved in a mixed solvent of THF (4.00 mL) and HO (2.00 mL) at 25 °C, LiOH (25.9 mg, 1.08 mmol) was added, and the reaction solution was stirred at 40 °C for 18 h. Thin layer chromatography (DCM / MeOH = 10 / 1) showed complete consumption of the reactants and the formation of a new spot. The reaction solution was centrifuged, then dissolved in 1.00 mL of acetonitrile and 4.00 mL of water, and lyophilized to obtain a white solid product, which was identified as compound 4 (510 mg, 90.63% yield) by LCMS.

[0938] 3 Preparation of compound 6 [ka] Compound 4 (54.4 mg, 0.087 mmol) is dissolved in DCM (10.0 mL) at 25 ° C, HBTU (41.2 mg, 0.109 mmol) and DIEA (0.086 mL, 0.522 mmol) are added in sequence, the reaction solution is stirred at 25 ° C for 0.5 hours, then compound 5 (prepared according to the method of compound 4 in Example 7, 200 mg, 0.087 mmol) is added, and the reaction solution is stirred at 25 ° C for 17.5 hours. The reaction solution is diluted with DCM (20.0 mL), washed with saturated NaHCO3 aqueous solution (5.00 mL × 3), the organic phase is washed with saturated saline (5.00 mL × 3), then the organic phase is dried with Na2SO4, and the organic phase is centrifuged to obtain crude compound 6 (270 mg). The crude product is used directly in the next step without purification.

[0939] 4. Preparation of Compound E19 [ka] Dissolve compound 6 (270 mg, 0.093 mmol) in DCM (10.0 mL) at 25 °C, add DIEA (0.092 mL, 0.557 mmol), DMAP (4.54 mg, 0.037 mmol) and compound 7 (55.7 mg, 0.557 mmol), and stir the reaction solution at 25 °C for 18 h. LCMS shows the MS value of the product. Centrifuge the reaction solution to obtain the crude product, which is separated on a reverse phase column (chromatography column: Waters Xbridge BEH C18 100*25 mm*5 um, mobile phase: TEAA-ACN, B%: 20%-60%, 18 min, flow rate: 15 ml / min) to obtain white solid compound E19 (64.0 mg, yield 22.92%, purity 95.34%).

[0940] 1 H NMR (400MHz, CD3OD) δ7.41-7.47(m, 2H), 7.17-7.35(m, 7H), 6.83-6.90(m, 4H), 5.32-5.36(m, 4H), 5.05-5.11(m, 4H), 4.64-4. 68(m, 4H), 4.28-4.45(m, 4H), 4.00-4.21(m, 21H), 3.86-3.99(m, 6H), 3.79(s, 6H), 3.67-3.77(m, 8H), 3.59-3.67(m, 24H), 3.55 (q, J=5.2Hz, 8H), 3.34-3.50(m, 10H), 3.08(q, J=7.2Hz, 17H), 2.56-2.63(m, 2H), 2.42-2.51(m, 4H), 2.21-2.36(m, 10H), 2.12 -2.16(m, 12H), 2.06-2.12(m, 2H), 2.00-2.05(m, 13H), 1.98(s, 1H), 1.58-1.66(m, 2H), 1.41-1.49(m, 2H), 1.21-1.35(m, 35H).

[0941] Example 20 Preparation of compound E20 [ka]

[0942] 1. Preparation of Compound 3 [ka] Compound 1 (1.60 g, 7.20 mmol) is dissolved in DCM (60 mL) at 25 ° C., then compound 2 (3.31 g, 18.0 mmol) and EDCI (4.14 g, 21.6 mmol) are added successively, and the reaction solution is stirred at 25 ° C. for 18 h. LCMS shows the MS value of the product, and thin layer chromatography (PE / EA = 5 / 1) shows the formation of a new spot. The reaction solution is concentrated under reduced pressure to obtain the crude product, which is purified by MPLC (PE / EA = 1 / 0 to 5 / 1) to obtain compound 3 (1.70 g, 42.59% yield) as a yellow oil. 1 H NMR (400MHz, CDCl3) δ4.54-4.57(m, 4H), 3.83-3.87(m, 4H), 3.74-3.79(m, 4H).

[0943] 2. Preparation of Compound 6 [ka] At 25°C, compound 4 (prepared according to the method of intermediate 3-3 of Example 24, 2.30g, 2.15mmol) is dissolved in DCM (20.0mL), compound 5 (0.690g, 1.03mmol) and DIEA (0.508mL, 3.08mmol) are added successively, and the reaction solution is stirred at 25°C for 10 hours. LCMS shows the reaction produces the target compound, and thin layer chromatography (DCM / MeOH=8 / 1) shows the complete consumption of the reactants and the production of a new spot. The reaction solution is directly centrifuged, and when the eluent is DCM / MeOH, the product is eluted to obtain compound 6 (1.93g, 77.09% yield) as a white solid. 1 H NMR (400MHz, CD3OD) δ7.29-7.40(m, 5H), 5.31-5.36(m, 4H), 5.05-5.13(m , 6H), 4.62-4.68(m, 4H), 4.36-4.43(m, 2H), 4.00-4.20(m, 22H), 3.89-3.9 6(m, 4H), 3.67-3.76(m, 9H), 3.49-3.66(m, 34H), 3.34-3.42(m, 8H), 2.27- 2.35(m, 4H), 2.12-2.16(m, 12H), 2.01-2.04(m, 12H), 1.90-1.96(m, 24H).

[0944] 3 Preparation of compound 7 [ka] Compound 6 (930 mg, 0.381 mmol) is dissolved in THF (10.0 mL) at 25° C., TFA (47.8 mg, 0.419 mmol) and Pd / C 10% (100 mg, 0.940 mmol) are added successively, the reaction solution is stirred and reacted under hydrogen atmosphere (14.696 psi) at 25° C. for 18 hours. Thin layer chromatography (DCM / MeOH=8 / 1) shows complete consumption of the reactants and the formation of a new spot. The reaction solution is filtered, and the filtrate is centrifuged to obtain compound 7 (830 mg, 94.44% yield) as a white solid. 1 H NMR (400MHz, CD3OD) δ5.33-5.36(m, 4H), 5.09(d, J=3.6Hz, 2H), 5.06(d, J=2.4Hz, 2H), 4.60-4.67(m, 4H), 4.36-4.43(m, 2H), 4.00-4.19(m, 22H), 3.92-3.98(m, 4H), 3.69-3.77(m, 9H), 3.55-3.67(m, 34H), 3.35-3.45(m, 8H), 2.34-2.43(m, 4H), 2.15(s, 12H), 2.03(s, 12H), 1.93-1.97(m, 24H).

[0945] 4. Preparation of Compound 8 [ka] Compound 3 is dissolved in DCM (10.0 mL) at 25 °C, compound 7 (400 mg, 0.173 mmol) and TEA (0.144 mL, 1.04 mmol) are added successively, and the reaction solution is stirred at 25 °C for 18 h. LCMS shows the MS value of the product. TLC (DCM / MeOH = 10 / 1) shows the complete reaction of the raw materials, and the formation of a new spot. The reaction solution is concentrated under reduced pressure to obtain the crude product, which is purified by MPLC (DCM / MeOH = 1 / 0 to 5 / 1) to obtain compound 8 (340 mg, 78.09% yield) as a white solid. 1H NMR (400MHz, CD3OD) δ5.32-5.36(m, 4H), 5.06-5.12(m, 4H), 4.64-4.69(m, 4H), 4.38 -4.47(m, 3H), 4.03-4.18(m, 22H), 3.90-3.96(m, 6H), 3.68-3.75(m, 16H), 3.61-3.6 6(m, 24H), 3.53-3.58(m, 8H), 3.35-3.43(m, 8H), 2.27-2.34(m, 4H), 2.13-2.17(m, 1 2H), 2.08-2.12(m, 2H), 2.02-2.04(m, 12H), 1.97-2.01(m, 2H), 1.93-1.96(m, 23H).

[0946] 5. Preparation of Compound 10 [ka] At 25 ° C, compound 8 (310 mg, 0.123 mmol) is dissolved in DCM (10 mL), then HBTU (140 mg, 0.370 mmol) and DIEA (0.122 mL, 0.741 mmol) are added in sequence, the reaction solution is stirred at 25 ° C for 0.5 h, and finally, compound 9 (166 mg, 0.370 mmol) is added, and the reaction solution is stirred at 25 ° C for 17.5 h. LCMS shows the MS value of the product, complete consumption of the raw material. The reaction solution is centrifuged to obtain the crude product. The crude product is prepared and separated by reverse phase column (chromatography column: 01-Waters Xbridge BEH C18 19 * 150 mm, mobile phase: TEAA-ACN, B%: 20% ~ 58%, flow rate: 15 ml / min), and a white solid compound 10 (170 mg, yield 46.80%) is obtained. 1H NMR (400MHz, CD3OD) δ7.41-7.45(m, 2H), 7.25-7.34(m, 7H), 6.87(t, J=8.0Hz, 4H), 5.34(d, J=3.2Hz, 4H), 5.08(ddd, J=11.2, 3 .2, 1.6Hz, 4H), 4.65(dd, J=8.4, 3.2Hz, 4H), 4.58(brs, 4H), 4.39-4.44(m, 3H), 4.00-4.19(m, 24H), 3.93(dt, J=11.2, 4.4Hz, 4H) ), 3.79(s, 6H), 3.69-3.76(m, 11H), 3.58-3.67(m, 34H), 3.51-3.57(m, 9H), 3.34-3.41(m, 8H), 3.15(q, J=7.2Hz, 11H), 2.27-2 .34(m, 4H), 2.14(s, 12H), 2.06-2.12(m, 2H), 2.02(s, 12H), 1.96-2.00(m, 2H), 1.94(d, J=6.8Hz, 24H), 1.29(t, J=7.2Hz, 15H).

[0947] 6 Preparation of compound E20 [ka] Compound 10 (170 mg, 0.058 mmol) is dissolved in DCM (6.00 mL) at 25 °C, then DIEA (0.057 mL, 0.347 mmol), DMAP (2.82 mg, 0.023 mmol) and compound 11 (34.7 mg, 0.347 mmol) are added sequentially, and the reaction solution is stirred at 25 °C for 18 h. LCMS shows the MS value of the product. The reaction solution is centrifuged to obtain the crude product, which is separated by reverse phase column (chromatography column: C18, mobile phase: H2O (0.1% TEA)-ACN, B%: 30%-70%, 20 min, flow rate: 20 ml / min) to obtain white solid compound E20 (96 mg, yield 54.61%, purity 90.93%). 1H NMR (400MHz, CD3OD) δ7.44 (brd, J=8.0Hz, 2H), 7.19-7.34 (m, 7H), 6.88 (brdd, J=8.0, 6.4Hz, 4H), 5.34 (d, J=3.2Hz, 4H), 5.05-5.11(m, 4H), 4.65(dd, J=8.4, 2.8Hz, 4H), 4.38-4.45(m, 3H), 4.20-4.31(m, 3H), 4.01-4.19(m, 24H), 3.9 0-3.95(m, 4H), 3.79(s, 6H), 3.45-3.77(m, 53H), 3.34-3.42(m, 9H), 3.18(q, J=7.2Hz, 5H), 2.57-2.63(m, 2H), 2.48 -2.54(m, 2H), 2.27-2.34(m, 4H), 2.08-2.20(m, 13H), 2.02(s, 13H), 1.94(d, J=5.6Hz, 23H), 1.30(t, J=7.2Hz, 8H).

[0948] Example 21 Preparation of Compound E21

change

[0949] 1 Preparation of compound 6

change

[0950] 2. Preparation of intermediate 3-2 [ka] To a solution of compound 6 (37.0 g, 15.5 mmol, 1.00 equiv) in DME (740 mL) is added TFA (1.77 g, 15.54 mmol, 1.15 mL, 1.00 equiv) and Pd / C (3.70 g, 10% purity). The suspension is degassed and purged with hydrogen three times. The mixture is stirred at 15 °C and H2 (15 Psi) for 16 h. LCMS shows complete reaction. Filter through Celite and concentrate the filtrate in vacuo to remove the solvent to give intermediate 3-2 (77.2 g, crude, TFA salt) as a white solid.

[0951] 1 H NMR:ET53345-86-P1A(400MHz, DMSO) δ8.9(m, 1H), 8.40-8.20(m, 6H), 7.83-7.81(m, 6H), 5.32(d, J=3.2Hz, 1H), 5.2 1(d, J=2.8Hz, 4H), 4.96(dd, J=3.2Hz, 2H), 4.55(d, J=8.4Hz, 4H), 4.35(m, 1H), 4.20(m, 1H), 4.02(s, 12H), 3.86-3.77(m, 10H), 3.61-3.49(m, 42H), 3.42-3.19 (m, 10H), 2.15-2.10(m, 18H), 1.99(s, 12H), 1.89(s, 14H), 1.77-1.74(m, 21H).

[0952] 3 Preparation of compound 3 [ka] Dissolve intermediate 3-2 (500 mg, 0.222 mmol) in DCM (10.0 mL), add HBTU (127 mg, 0.334 mmol), DIEA (0.110 mL, 0.667 mmol) and compound 2 (142 mg, 0.222 mmol) in sequence, and stir the reaction solution at 25 °C for 18 h. LCMS detects the MS response of the product. Thin layer chromatography (DCM / MeOH = 10 / 1) shows the complete reaction of the raw material, the formation of a new spot. Wash the reaction solution with saturated aqueous sodium bicarbonate solution (20.0 mL), extract with dichloromethane (10.0 mL x 2), combine the organic phase, concentrate, and pass through a normal phase column (DCM / MeOH = 1:0 to 10:1) to obtain a white solid 3 (545 mg, 85.56% yield).

[0953] 4. Preparation of Compound E21 [ka] Compound 3 (545 mg, 0.190 mmol) is dissolved in DCM (20.0 mL), compound 4 (114 mg, 1.14 mmol), DIEA (0.189 mL, 1.14 mmol) and DMAP (23.3 mg, 0.190 mmol) are added, and the reaction solution is stirred at 25 °C for 18 h. LCMS detects the MS response of the product. The reaction solution is directly centrifuged and dried, and dissolved by adding acetonitrile (5.00 mL), and passed through preparative HPLC (chromatography column: Waters Xbridge BEH C18 100 * 30 mm * 10 um, mobile phase: water-acetonitrile], B%: 17% ~ 57%, flow rate: 50 ml / min) to obtain white solid compound E21 (50.0 mg, yield 7.47%, purity 84.29%). LCMS (ESI): m / z = 2962.3 (M + H) + , 1 H NMR (400MHz, CD3OD) δ7.43(d, J=8.0Hz, 2H), 7.27-7.32(m, 6H), 7.18-7.25(m, 1H), 6.85-6.88(m, 4H), 5.32-5.34(m, 4H), 5.06-5 .09(m, 4H), 4.64-4.67(m, 4H), 4.32-4.39(m, 2H), 4.02-4.21(m, 20H), 3.83-3.99(m, 6H), 3.79(d, J=2.0Hz, 6H), 3.69-3.74(m, 5) H), 3.58-3.67(m, 28H), 3.51-3.57(m, 6H), 3.35-3.41(m, 8H), 3.14-3.23(m, 2H), 2.51-2.60(m, 4H), 2.30-2.40(m, 8H), 2.20-2. 23(m, 2H), 2.14(s, 12H), 2.06-2.09(m, 2H), 2.02-2.03(m, 12H), 1.94(d, J=5.2Hz, 24H), 1.54-1.59(m, 4H), 1.29-1.38(m, 12H).

[0954] Example 22 Preparation of Intermediate 2-3 [ka]

[0955] 1. Preparation of Compound 8 [ka] To a solution of compound 7 (100 g, 256 mmol) in DCM (300 mL), add TMSOTf (68.5 g, 308 mmol, 55.7 mL) dropwise over 0.5 h at 10-15 °C. The mixture is then stirred at 25 °C for 12 h. TLC (ethyl acetate) indicates complete consumption of compound 7 (R f =0.3) and the formation of compound 8 (R f =0.4). Cool the compound to 0 °C and slowly pour into aqueous NaHCO3 (100 g dissolved in 1.00 L water) at 0-5 °C, separate the organic phase using DCM (200 mL x 3), extract the aqueous phase, dry over Na2SO4, and concentrate in vacuum to obtain compound 8 (approximately 83.0 g) as a yellow oil. 1 H NMR: (400MHz, DMSO-d6). δ6.03(d, J=7.2Hz, 2H), 5.24-5.22(m, 1H), 4.89-4.86(m, 1H), 4.26-4.24(m, 1H), 4. 13-4.05(m, 2H), 4.04-3.94(m, 1H), 2.06(s, 3H), 1.99(d, J=3.2Hz, 6H), 1.98(s, 3H).

[0956] 2. Preparation of compound 2A [ka] Both reactions proceed in parallel.

[0957] To a solution of compound 2A-1 (450 g, 4.28 mol, 428 mL) in 2-methyl-THF (1.80 L), compound 2A-2 (365 g, 2.14 mol, 304 mL) in THF (450 mL) is added dropwise at 0° C. After addition, the compound is stirred under nitrogen atmosphere at 25° C. for 12 hours. TLC (ethyl acetate, compound 2A-2 R f =0.9, compound 2A R f=0.3) indicates complete consumption of compound 2A-2 and formation of compound 2A. The two reactions are combined. The reaction mixture is added with HCl / EtOAc (1M, 900 mL), stirred for 30 min, the white solid is filtered through Celite, and the filtrate is concentrated in vacuo to give compound 2A (approximately 990 g) as a colorless oil. 1 H NMR: (400MHz, CDCl3). δ7.26-7.21(m, 5H), 5.56(s, 1H), 5.00(s, 2H), 3.62-3.59(m, 2H), 3.45-3.23(m, 4H), 3.29(s, 2H).

[0958] 3 Preparation of compound 9 [ka] To a mixture of compound 8 (83.0 g, 252 mmol) and compound 2A (42.2 g, 176 mmol) in DCM (500 mL) at 0-10 °C, add TMSOTf (28.0 g, 126 mmol, 22.8 mL) dropwise and stir the mixture at 20 °C for 12 h. TLC (petroleum ether / ethyl acetate = 0 / 1) indicates complete consumption of compound 8, compound 9 (R f =0.3). LCMS (compound 9: RT = 0.744 min) shows complete consumption of compound 8. The reaction mixture is poured into saturated aqueous NaHCO3 (200 mL) at 0-5 °C, washed with brine (200 mL), dried over Na2SO4, filtered and concentrated in vacuo. The residue is purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 0 / 1). The crude product is triturated with petroleum ether / ethyl acetate = 1 / 1 (100 mL) at 25 °C for 30 min to give compound 9 (approximately 59.0 g) as a white solid. 1H NMR: (400MHz, DMSO-d6). δ7.82(d, J=9.2Hz, 1H), 7.40-7.28(m, 5H), 5.23(d, J=3.2Hz, 1H), 5.26-4.98(m, 3H), 4.56(d, J=8.4Hz, 1H), 4.05(s, 3H), 3.93-3.86(m , 1H), 3.81-3.76(m, 1H), 3.63-3.48(m, 3H), 3.43-3.40(m, 2H), 3.35-3.15(m, 2H), 2.11(s, 3H), 2.01(s, 3H), 1.91(s, 3H), 1.79(s, 3H).

[0959] 4. Preparation of intermediate 2-3 (TFA salt) [ka] To Pd / C (9.50 g, content 10%) in THF (475 mL) is added compound 9 (95.0 g, 167 mmol) and TFA (19.0 g, 167 mmol, 12.4 mL) under an argon atmosphere. The reaction mixture is flushed with argon three times and with hydrogen three times, then the mixture is stirred under an atmosphere of H2 (45 psi) at 25 °C for 3 h. TLC (dichloromethane / methanol = 10 / 1) indicates complete consumption of compound 9 (R f =0.5) and intermediate 2-3 (TFA salt) (R f =0.2). The reaction mixture was concentrated under reduced pressure to give intermediate 2-3 (TFA salt) (55.53 g, yield: 57.9%, purity: 95.7%) as an off-white solid. 1 H NMR: (400MHz, CDCl3). δ8.00-7.97(m, 5H), 5.29(d, J=3.2Hz, 1H), 5.07(d, J=3.6Hz, 1H), 4.63(d, J=8.4Hz, 1H), 4.10(s, 3H), 3.97-3.95 (m, 1H), 3.89-3.88(m, 1H), 3.70-3.63(m, 5H), 3.04(s, 2H), 2.17(s, 3H), 2.07(s, 3H), 1.96(s, 3H), 1.85(s, 3H).

[0960] Example 23 Preparation of Intermediate 3-4 [ka]

[0961] 1. Preparation of Compound 2 [ka] To compound 1 (300 g, 2.01 mol) in DCM (1.80 L) at 15° C., benzyl (2,5-dioxopyrrolidin-1-yl) carbonate was slowly added and TEA (203 g, 2.01 mol, 280 mL) was added dropwise. After the addition, the mixture was stirred at 25° C. for 16 h. TLC (dichloromethane:methanol=10:1) showed that reactant 1 (R f =0.32) and also shows a significant new spot (R f =0.52). The reaction mixture (1.00 L x 2) is washed with saturated sodium bicarbonate solution, and the organic phase is washed with brine (1.00 L), dried over anhydrous Na2SO4, and concentrated in vacuo. Without purification, compound 2 (approximately 385 g) is a yellow oil.

[0962] 2. Preparation of compound 2A [ka] To a solution of compound 4 (350 g, 1.62 mol, HCl) and Ac2O (994 g, 9.74 mol, 912 mL) in pyridine (1.75 L) at 0-15 °C, DMAP (19.8 g, 162 mmol) is added in one portion and TEA (164 g, 1.62 mol, 226 mL) is also added dropwise. The mixture is stirred at 25 °C for 16 h. LCMS (product: RT = 0.687 min) indicates complete consumption of starting reactant. EtOAc (1.40 L) is added to the mixture at 25 °C and stirred for 30 min, after which the mixture is filtered and the filter cake is washed with EtOAc (300 mL). The filter cake is triturated with water (1.45 L) for 30 min at 25 °C. The mixture is filtered and the filter cake is washed with water (175 mL x 3) and the filter cake is collected to give compound 2A (about 580 g) as a white solid.

[0963] 3. Preparation of compound 2B [ka] The three reactions proceed in parallel.

[0964] To a solution of compound 2A (200 g, 514 mmol) in DCM (800 mL) at 10-15 °C, add TMSOTf (137 g, 616 mmol, 111 mL) dropwise over 0.5 h. Then, stir the mixture at 25 °C for 3 h. TLC (dichloromethane:methanol = 20:1) indicates complete consumption of compound 2A (R f =0.54) and new spots (R f =0.24). The three reactions are combined. The mixture is cooled to 0-15 °C and slowly poured into NaHCO3 solution (300 g dissolved in 3.00 L water) at 0-5 °C, the organic phase is separated, the aqueous phase is extracted with DCM (1.00 L × 3), the organic layers are combined, dried over Na2SO4, filtered and concentrated in vacuo. Compound 2B (approximately 507 g) is obtained as a yellow oil, which is used in the next step without purification.

[0965] 4. Preparation of Compound 3 [ka] To a mixture of compound 2B (250 g, 759 mmol) and compound 2 (151 g, 531 mmol) in DCM (1.00 L), TMSOTf (84.4 g, 380 mmol, 69.0 mL) is added dropwise, and the mixture is stirred at 20° C. for 12 h. TLC (dichloromethane:methanol=20:1) indicates complete consumption of compound 2 (R f =0.33) and new spots (R f =0.03). Cool the combined reactions to 0-5 °C, then pour the reactions into NaHCO3 (aq, 100 g in 1 L water), stir at 5-10 °C for 10 min, and separate the phases. Extract the aqueous phase with DCM (500 mL x 2), dry the combined organic phase over Na2SO4, filter, and concentrate the filtrate in vacuo. Without purification, compound 3 (approximately 360 g) is a yellow oil. 1H NMR: (400MHz, DMSO). δ7.79-7.37(m, 1H), 7.35-7.26(m, 5H), 5.21-5.20(m, 1H), 5.00-4.95(m, 3H), 4.55-4.53(m3-3.8, 1H), 4.06(m, 3H), 3.61-3.59(m, 1H) , 3.59-3.57(m, 1H), 3.48-3.40(m, 6H), 3.39-3.31(m, 2H), 3.14-3.13(m, 2H), 2.09(s, 3H), 1.99(s, 3H), 1.88(s, 3H), 1.76-1.74(m, 3H)

[0966] 5 Preparation of intermediate 3-4 (TFA salt) [ka] The three reactions proceed in parallel.

[0967] Compound 3 (180 g, 293 mmol) and TFA (33.5 g, 293 mmol, 21.8 mL) are added to a mixture of Pd / C (18.0 g, 16.3 mmol, content 10%) in THF (1.80 L) under argon atmosphere. The suspension is evacuated and ventilated with hydrogen three times. The mixture is stirred under H2 (50 Psi) at 30 °C for 2 h. LCMS (product: RT = 0.697 min) shows consumption of compound 3 and detects a product peak. The three reactions are combined. The mixture is filtered through Celite and the filtrate is concentrated under reduced pressure to remove the solvent. Intermediate 3-4 (TFA salt) (393 g, 660 mmol, 74.8% yield, 99.6% purity, TFA) is obtained as a yellow solid without purification.

[0968] 1 H NMR: (400MHz, DMSO-d6)

[0969] δ=7.92(d, J=9.1Hz, 4H), 5.27-5.17(m, 1H), 5.03-4.91(m, 1H), 4.60-4.50(m, 1H), 4.09-3.97(m, 4H), 3.85(s , 2H), 3.65-3.46(m, 10H), 3.04-2.92(m, 2H), 2.10(s, 3H), 2.00(s, 3H), 1.94-1.86(m, 3H), 1.82-1.71(m, 4H).

[0970] Example 24 Preparation of Intermediate 3 and Intermediate 5 [ka]

[0971] 1. Preparation of Compound 5 [ka] To a solution of compound 4B (10.0 g, 35.5 mmol, 1.00 equiv) and compound 3-4 (46.3 g, 78.2 mmol, 2.20 equiv, TFA) in DCM (1.00 L) at 25° C., add DIEA (30.3 g, 234 mmol, 40.8 mL, 6.60 equiv) in one portion. Stir at 25° C. for 30 min. Add HBTU (30.3 g, 234 mmol, 40.8 mL, 6.60 equiv) to the mixture. Stir at 25° C. for 16 h. LCMS (product: RT=0.681 min) indicates completion of the reaction and the mixture is concentrated in vacuo. Add 0.50N HCl (200mLx2) to the mixture at 20 °C, extract with DCM (3x500mL), combine the organic layers, wash with saturated NaHCO3 (3x800mL) until pH = 8, wash with brine (3x500mL), dry over Na2SO4, concentrate in vacuum and purify. Purify the residue by column chromatography (SiO2, DCM:MeOH = 50:1 to 15:1). Concentrate the residue in vacuum at 40 °C and purify by preparative MPLC (column: 800g Agela C18, mobile phase: [water-ACN], 15-45% for 25 min, 45% for 10 min). Dry under vacuum to obtain yellow solid compound 5 (approximately 180g + 75.0g + 87.0g + 40.0g + 38.0g).

[0972] 417.0 g of compound 3-4 is converted to compound 5 in 9 batches.

[0973] 2. Preparation of intermediate 3-3 [ka] To Pd / C (3.00 g, content 10%) in THF (300 mL) is added compound 5 (73.0 g, 61.7 mmol, 1.00 equiv.) and TFA (7.04 g, 61.7 mmol, 4.57 mL, 1.00 equiv.) under argon atmosphere. The suspension is degassed and purged with hydrogen three times. H2 (20 Psi) is added and stirred at 20 °C for 16 h. TLC (dichloromethane:methanol = 8:1, R f = 0.0) indicates completion of the reaction. The mixture is filtered through Celite, and the filtrate is concentrated under reduced pressure to remove the solvent to give compound 3-3 (approximately 33.4 g + 129 g + 75.0 g) as a white solid.

[0974] 1 H NMR: (400MHz, DMSO)

[0975] δ8.53(t, J=5.2Hz, 1H), 8.18(d, J=2.4Hz, 3H), 8.03(t, J=5.2Hz, 1H), 7.84(dd, J=3.6H z, 2H), 5.22(d, J=3.2Hz, 2H), 4.96(dd, J=3.2Hz, 2H), 4.55(d, J=8.4Hz, 2H), 4.02(t, J= 8.8Hz, 6H), 3.77-3.59(m, 5H), 3.58-3.45(m, 21H), 3.40-3.20(m, 4H), 2.18(t, J=7.6Hz) , 2H), 2.17(d, J=8.0Hz, 6H), 2.10(s, 6H), 1.99(s, 6H), 1.90-1.80(m, 8H), 1.77(s, 6H).

[0976] 3 Preparation of compound 2 [ka] To a solution of compound 1 (90.0 g, 465 mmol, 1.00 equiv) in DCM (540 mL) at 0° C., TEA (47.1 g, 465 mmol, 64.8 mL, 1.00 equiv) and Cbz-OSu (110 g, 442 mmol, 0.95 equiv) are added. The mixture is stirred at 20° C. for 16 h. LCMS (product: RT=0.625 min) indicates completion of the reaction. The reaction mixture is washed with saturated sodium bicarbonate solution (450 mL×2), the organic phase is washed with brine (500 mL), dried over anhydrous NaSO4, and concentrated to give compound 2 (approx. 136 g) as a yellow oil.

[0977] 4. Preparation of Compound 3 [ka] To a solution of compound 2 (51.0 g, 155 mmol, 1.00 equiv) in DCE (300 mL) at 20 °C, compound 2A (84.9 g, 218 mmol, 1.40 equiv) and Sc(OTf)3 (12.0 g, 62.3 mmol, 0.40 equiv) are added. Stir at 85 °C for 3 h. LCMS (product: RT = 0.783 min) indicates completion of the reaction. Cool the reaction mixture to 15 °C and extract with DCM (300 mL) by adding saturated NaHCO3 (300 mL) at 15-20 °C. The combined organic layers are washed with 1 M HCl (50.0 mL) and brine (300 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure at 45 °C to give compound 3 (84.0 g) as a brown oil.

[0978] 5 Preparation of intermediate 5 [ka] To Pd / C (7.00 g, content 10%) in THF (300 mL) is added compound 3 (70.0 g, 109 mmol, 1.00 equiv.) and TFA (12.5 g, 109 mmol, 8.14 mL, 1.00 equiv.) under argon atmosphere. The suspension is degassed and ventilated with hydrogen three times. The mixture is stirred under H2 (40 Psi) at 30 °C for 2 h. TLC (dichloromethane:methanol = 8:1) shows the reaction is complete. The mixture is filtered through Celite and the filtrate is concentrated under reduced pressure to remove the solvent to give compound 5 (66.0 g, 103 mmol, yield 94.29%, TFA) as a yellow oil.

[0979] 1 H NMR: (400MHz, DMSO)

[0980] δ7.86-7.84(m, 4H), 5.22(d, J=3.6Hz, 1H), 4.97(dd, J=3.2Hz, 1H), 4.54(d, J=8.4Hz, 1H), 4.03(s, 3H), 3.7 7-3.60(m, 3H), 3.60-3.51(m, 15H), 3.00-2.97(m, 2H), 2.10(s, 3H), 2.00(s, 3H), 1.89(s, 3H), 1.81(s, 3H).

[0981] Example 25 Preparation of Intermediate 1-3 [ka] To a mixture of DCM (100 mL), ACN (100 mL) and H2O (300 mL), NaIO4 (224 g, 1.05 mol) and RuCl3 (966 mg, 4.66 mmol) are added in one portion at -5 to 0 °C. Compound 1 is dissolved in a mixture of DCM (100 mL) and ACN (100 mL), and then the solution is added dropwise to the above reaction mixture at -5 to 0 °C. The reaction mixture is stirred at 15 °C for 12 h. TLC (petroleum ether / ethyl acetate = 1 / 1, product R f=0.20) indicates the formation of a new spot. The reaction mixture is adjusted to pH=8 using NaHCO3 (50.0 g) and the mixture is extracted using DCM (500 mL x 4). The aqueous phase is adjusted to pH=3 using citric acid (100 g). The mixture is then extracted using DCM (500 mL x 4). The organic phase is separated, dried over Na2SO4, filtered and concentrated under reduced pressure to form a residue. Purify the residue by recrystallization from EtOAc (1.00 L) at 15 °C. The crude product is washed with EtOAc (1.00 L). Intermediate 1-3 (55.3 g, 52.2% yield, 98.4% purity) is obtained as an off-white solid.

[0982] 1 HNMR: (400MHz DMSO-d6)

[0983] 11.9(s, 1H), 7.80(d, J=9.2Hz, 9H), 5.20(d, J=3.6Hz1H), 4.97-4.93(m, 1H), 4.48(d, J=8.8Hz, 1H), 4.02(s, 3H), 3.90-3.83(m, 1H), 3.73-3.69(m, 1H), 3.44-3.39(m, 1H), 2.19(t, J=6.8Hz, 2H), 2.10(s, 3H), 1.99(s, 3H), 1.88(s, 3H), 1.76(s, 3H), 1.49-1.48(m, 4H).

[0984] Example 26 Preparation of Intermediate 2-2 [ka]

[0985] 1. Preparation of Compound 3 [ka] The four reactions proceed in parallel.

[0986] 4-Methylmorpholine (434 g, 4.30 mol, 472 mL) is added to a solution of compound 1 (250 g, 741 mmol) in THF (1.75 L) and then cooled to 0° C. Isobutyl chloroformate (243 g, 1.78 mol, 233 mL) is added to the reaction mixture within 10 min while maintaining the reaction temperature below 4.0° C. After the addition, the mixture is stirred for 40 min or more and compound 2 (526 g, 1.78 mol) is added sequentially to the reaction mixture within 10 min while maintaining the reaction temperature below 4.0° C. After the addition, the ice bath is removed and then the reaction is heated to room temperature and maintained for 2 h. TLC (petroleum ether / ethyl acetate=1 / 1, compound 1 R f =0.43) indicates complete consumption of compound 1, formation of a new spot. Combine the four reactions. Pour the reaction solution into a stirring cold (0°C) 0.50 M HCl (aq) (12.0 L) solution and stir for approximately 10 min. Then add EtOAc (4.00 L x 3), stir for a period of time, separate the layers, and wash the organic phase with brine (10.0 L), dry over Na2SO4, and concentrate in vacuo to give a thick, colorless oil. Add hexane (1.20 L) to the stirring oil. White smoke appears in the solution and then dissipates with further stirring. Add seed crystals (1.20 g, 0.10 wt%) and white crystals slowly form. Within 20 min, the suspension becomes thick enough to prevent stirring, at which point additional hexane (6.00 L) is added and stirred for 12 h. Filter the suspension, wash with hexane (1.20 L), and dry to give compound 3 (approximately 1.20 kg) as a white solid.

[0987] 1 H NMR: (400MHz DMSO)

[0988] δ8.10-8.08(m, 1H), 7.62-7.60(m, 1H), 7.38-7.31(m, 5H), 5.11-4.94(m, 2H), 4.12-4.09(m, 1H), 3.92-3.87(m, 1H), 2.25-2.19(m, 4H), 1.90-1.88(m, 2H), 1.73-1.67(m, 2H), 1.40-1.39(m, 27H).

[0989] 2. Preparation of Compound 4 [ka] The five reactions proceed in parallel.

[0990] A solution of compound 3 (240 g, 415 mmol) in HCOOH (2.40 L) is stirred at 45° C. for 2 h. LCMS (compound 4 = 0.570 min) indicates complete consumption of compound 3, with the main peak of the required m / z appearing. The five reactions are combined. Dilute and concentrate using toluene and ACN (1.50 L each). Formic acid is azeotropically removed using 1:1 ACN to toluene (500 mL) and three times ACN (500 mL each). Compound 4 is dried under high vacuum. The residue is then stirred with DCM (500 mL) and the organic layer is removed, which is then azeotropically dried twice with ACN (400 mL), dried under vacuum, and then azeotroped nine times with toluene (400 mL) to give compound 4 (approximately 800 g) as a white solid.

[0991] 1 H NMR: (400MHz DMSO)

[0992] δ12.4-12.2(m, 2H), 8.12-8.10(m, 1H), 7.62-7.60(m, 1H), 7.38-7.34(m, 5H), 5.07-5.02(m, 2H) , 4.22-4.17(m, 1H), 3.99-3.96(m, 1H), 2.31-2.21(m, 4H), 2.00-1.93(m, 2H), 1.76-1.75(m, 2H).

[0993] 3 Preparation of compound 6 [ka] To a stirred solution of compound 4 (80.0 g, 194 mmol) and compound 2-3 (384 g, 700 mmol, TFA) in DMF (2.00 L) was added HOBT (103 g, 760 mmol), EDCI (146 g, 760 mmol), and DIEA (113 g, 877 mmol, 153 mL) in sequence. The reaction was stirred at 20° C. for 2 h. TLC (dichloromethane / methanol=5 / 1, compound 6 R f =0.43) indicating complete consumption of compound 4, formation of a new spot. The reaction mixture is slowly poured into stirred cold 0.5 M aqueous HCl (230 mL) and after stirring for 10 min a white solid forms which is filtered and the aqueous phase is extracted twice with DCM (1.50 L). The combined organic phase is washed with 5% NaHCO3(aq) (200 mL) before it is dried (Na2S) and concentrated by evaporation under pressure. The residue is purified by column chromatography (SiO2, dichloromethane / methanol = 5 / 1, compound 6 R f =0.43) to give compound 6 (approximately 180 g) as a yellow solid.

[0994] 1 H NMR: (400MHz DMSO)

[0995] δ7.95-7.91(m, 3H), 7.82-7.80(m, 4H), 7.39-7.31(m, 6H), 5.21-5.01 (m, 3H), 5.00-4.96(m, 5H), 4.56-4.53(m, 3H), 4.02(s, 1H), 3.88(s, 9 H), 3.85(s, 4H), 3.76(s, 3H), 3.50-349(m, 9H), 3.39-3.36(m, 6H), 3. 19-3.15(m, 6H), 2.15-2.05(m, 12H), 1.99(s, 9H), 1.89-1.77(m, 21H).

[0996] 4. Preparation of Intermediate 2-2 [ka] In a dry hydrogenation bottle under argon atmosphere, Pd(OH)2 / C (4.94 g, 3.52 mmol, content 10%) was added, MeOH (350 mL) was added, followed by compound 6 (47.4 g, 28.5 mmol), TFA (3.26 g, 28.5 mmol, 2.11 mL) and stirred under H2 (50 psi) at 2 °C for 3 h. The reaction was monitored by TLC (dichloromethane / methanol = 5 / 1, intermediate 2-2 R f =0.25), indicating consumption of compound 6. The reaction mixture is filtered and the filtrate is concentrated in vacuo to give intermediate 2-2 (TFA salt) (46.3 g, 94.6% yield, 95.7% purity, TFA) as a white solid.

[0997] 1 H NMR: (400MHz DMSO)

[0998] δ8.56-8.50(m, 1H), 8.14-8.05(m, 4H), 7.99(s, 1H), 7.88-7.82(m, 4H), 5.2 2-5.21(m, 3H), 4.99-4.96(m, 3H), 4.55-4.53(m, 3H), 4.09(s, 1H), 4.03(s, 9) H), 3.87-3.79(m, 6H), 3.59-3.52(m, 1H), 3.51-347(m, 9H), 3.39-3.37(m, 5 H), 3.17-3.16(m, 6H), 2.22-2.10(m, 12H), 1.99(s, 9H), 1.93-1.73(m, 21H).

[0999] Example 27 Preparation of Intermediate 1-2 [ka]

[1000] 1. Preparation of Compound 3 [ka] To a solution of compound 1 (500 g, 4.13 mol) in DMSO (800 mL), add NaOH (5 M, 82.6 mL), followed by compound 2 (2.12 kg, 16.5 mol) at 25° C. Stir the mixture at 40° C. for 12 h. Reflux the mixture at 80° C. for 3.5 h. TLC (petroleum ether / ethyl acetate=3 / 1, compound 3 R f =0.49) indicates the completion of the reaction. MTBE (1.00 L) is added to the mixture, which is then washed with water (1.00 L × 2) and brine (300 mL). The organic phase is dried over Na2SO4, filtered, and concentrated under reduced pressure to give the crude product. The residue is purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 5 / 1, compound 3 R f =0.49) to obtain compound 3 (approximately 690 g) as a yellow oil.

[1001] 1 H NMR: (400MHz, DMSO-d6)

[1002] δ3.53-3.56(m, 6H), 3.71(s, 6H), 2.39(t, J=6.0Hz, 6H), 1.40(s, 27H), 1.29(s, 2H).

[1003] 2. Preparation of Compound 4 [ka] The two reactions proceed in parallel.

[1004] To a solution of compound 3 (365 g, 722 mmol) in DCM (1.25 L) at 25° C., add a solution of NaHCO3 (121 g, 1.44 mol, 56.2 mL) in water (1.25 L), then add CbzCl (185 g, 1.08 mol, 154 mL) dropwise and stir for 12 h. TLC (petroleum ether / ethyl acetate=5 / 1, compound 4 R f=0.46), indicating complete consumption of compound 3. Combine the two reactions. Extract the reaction mixture with DCM (1.00 L × 2), and wash the combined organic layers with brine (200 mL), dry over Na2SO4, and concentrate in vacuo to give a residue. Purify the residue by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 5 / 1, compound 4 R f =0.46) to give compound 4 (approximately 735 g) as a yellow oil.

[1005] 1 H NMR: (400MHz, DMSO-d6)

[1006] δ7.31-7.35(m, 5H), 4.97(s, 2H), 3.49-.3.45(m, 12H), 2.37-2.40(m, 6H), 1.39(s, 27H).

[1007] 3 Preparation of compound 5 [ka] Compound 4 (735 g, 1.15 mol) is added with HCOOH (2.20 L) at 25° C. The solution is then stirred at 25° C. for 16 h. TLC (petroleum ether / ethyl acetate=5 / 1, compound 5 R f = 0.1) indicates completion of the reaction. The reaction is concentrated in vacuo to form a residue. The residue is diluted with toluene (200 ml) and acetonitrile (200 mL) and concentrated in vacuo to give compound 5 (about 545 g, crude product) as a yellow oil. 1 H NMR: (400MHz, DMSO-d6) δ12.2(s, 3H), 7.30-7.39(m, 5H), 4.98(s, 2H), 3.54-3.57(m, 6H), 3.49(s, 6H), 2.40-2.50(m, 6H).

[1008] 4 Preparation of compound 7 [ka] To a solution of compound 5 (520 g, 1.10 mol) in DCM (600 mL) was added DIPEA (570 g, 4.41 mol), HBTU (1.38 kg, 3.64 mol) and compound 6 (634 g, 3.64 mol). The mixture was stirred for 12 h at 25° C. TLC (dichloromethane / methanol=10 / 1, compound 7 R f =0.22) indicates the completion of the reaction. The reaction is washed with saturated NaHCO3 solution (500 mL × 3) and brine (200 mL), dried over Na2SO4, filtered, and concentrated in vacuo to give a residue. The residue is purified by column chromatography (SiO2, dichloromethane / methanol = 1 / 0 to 5 / 1, dichloromethane / methanol = 10 / 1, compound 7 R f =0.22) to give compound 7 (approximately 361 g) as a yellow oil.

[1009] 1 H NMR: (400MHz, DMSO-d6)

[1010] δ7.80-7.83(m, 3H), 7.29-7.35(m, 5H), 6.76-6.77(m, 3H), 4.97(s, 2H), 3.52-3.55(m, 6H), 3.47( s, 6H), 2.99-3.04(m, 6H), 2.87-2.92(m, 6H), 2.25-2.28(m, 6H), 1.46-1.50(m, 6H), 1.36(s, 27H).

[1011] 5 Preparation of compound 8 [ka] To a solution of compound 7 (360 g, 382.9 mmol) in DCM (1.00 L) at 25° C., add TFA (1.66 kg, 14.6 mol, 1.08 L). Stir the reaction for 12 h at 25° C. TLC (dichloromethane:methanol=10:1, compound 8 R f = 0.0) indicates completion of the reaction. The solution is concentrated in vacuo to give a residue. The residue is diluted with toluene (500 mL) and concentrated in vacuo to give compound 8 (approximately 245 g) as a yellow oil.

[1012] 6 Preparation of compound 9 [ka] To a solution of compound 1-3 (302 g, 675 mmol) in DCM (720 mL), DIPEA (339 g, 2.63 mol) and HBTU (256 g, 675 mmol) are added and the mixture is stirred at 25 °C for 30 min. Then, compound 8 (120 g, 188 mmol) is added to the mixture and the mixture is stirred at 25 °C for 12 h. LCMS (compound 9 RT = 1.737 min) indicates the completion of the reaction. The solution is washed with saturated aqueous NaHCO3 (400 mL × 3) and brine (200 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure to give the residue. The residue is purified by column chromatography (SiO2, dichloromethane / methanol = 1 / 0 to 5 / 1, compound 9 R f =0.35) to give the crude product, which is purified by reverse HPLC (MeCN / H2O) to give compound 9 (approximately 200 g) as a yellow oil.

[1013] 1 H NMR: (400MHz, DMSO-d6)

[1014] δ7.81-7.84(m, 6H), 7.72-7.75(m, 3H), 7.33-7.35(m, 5H), 5.20-5.21(m, 3H), 4.94-4.97 (m, 5H), 4.46-4.49(m, 3H), 4.02(s, 9H), 3.83-.3.90(m, 3H), 3.67-3.72(m, 3H), 3.52-3. 55(m, 6H), 3.47(s, 5H), 3.34-3.42(m, 3H), 2.99-3.05(m, 12H), 2.27(t, J=6.4Hz, 6H), 2. 10(s, 9H), 2.03(t, J=7.2H), 1.99(s, 9H), 1.88(s, 9H), 1.77(s, 9H), 1.72-1.53(m, 14H).

[1015] 7 Preparation of intermediate 1-2 [ka] To a solution of compound 9 (109 g, 56.5 mmol) in THF (654 mL) under argon at 15° C., add TFA (6.45 g, 56.5 mmol, 4.19 mL) and Pd / C (21.8 g, 10% content). The mixture is stirred under H2 (15 Psi) at 15° C. for 12 h. LCMS (Intermediate 1-2 RT=0.696 min) indicates completion of the reaction. The mixture is filtered and the filtrate is concentrated in vacuo to give Intermediate 1-2 (84.2 g, 73.3% yield, 94.1% purity, TFA) as an off-white solid.

[1016] 1 H NMR: (400MHz, DMSO-d6)

[1017] δ7.76-7.91(m, 9H), 5.21(d, J=3.6Hz, 3H), 4.94-4.98(m, 3H), 4.48(d, J=8.4Hz, 3 H), 3.99-4.04(m, 9H), 3.83-3.90(m, 3H), 3.67-3.72(m, 3H), 3.57-3.64(m, 9H), 3. 58-3.65(m, 9H), 3.44(s, 5H), 3.01-3.07(m, 12H), 2.34(t, J=6.0Hz, 6H), 2.10(s, 9 H), 2.02-2.06(m, 6H), 1.99(s, 9H), 1.88(s, 9H), 1.77(s, 9H), 1.46-1.53(m, 17H).

[1018] Example 28 Preparation of siRNA

[1019] The siRNAs of the present invention are prepared using solid-phase phosphoramidite methods well known in the art. Particular methods can be found, for example, in PCT Publication Nos. WO2016081444 and WO2019105419, and are briefly described below.

[1020] 1. Attachment of ligands to CPG supports Below, compound E7 is used as an example to illustrate the attachment of ligands to CPG carriers, however, other GalNAc ligands of the invention can be attached to CPG carriers using similar methods.

[1021] Compound E7 (53 mg, 0.018 mmol) and HBTU (13.3 mg, 0.035 mmol) are mixed, then acetonitrile (5 mL) is added and shaken to dissolve, then DIEA (9.0 mg, 0.07 mmol) and DMAP (2.1 mg, 0.018 mmol) are added and shaken to dissolve until transparent. Blank carrier resin (550 mg, CPG pore size 1000 Å) is weighed and added to the reaction solution, the temperature is controlled at 20 °C, and reacted on a shaker overnight. Samples are taken to monitor, and thin layer chromatography (TLC) is performed, with DCM / methanol = 4 / 1 as the developer and phosphomolybdic acid as the coloring agent, which shows the completion of the reaction. Filter using a sand funnel, wash the filter cake with anhydrous acetonitrile (20 mL x 5), take out the filter cake, and filter using an oil pump under reduced pressure for 6 hours to obtain 530 mg of off-white solid.

[1022] Put 530 mg of the above condensate into a 50 mL round-bottom bottle, add CapC (DMAP / acetonitrile), CapB (N-methylimidazole / pyridine / acetonitrile), and CapA (acetic anhydride / acetonitrile) in sequence, filter, wash the filter cake with acetonitrile (20 mL × 4), take out the filter cake, and filter under reduced pressure by oil pump for 8 hours to obtain 200 mg of off-white solid to be used for solid-phase synthesis.

[1023] 2. Synthesis of the sense strand (SS strand) and antisense strand (AS strand) Using the solid-phase phosphoramidite synthesis method, a blank CPG solid support or a CPG solid support connected to a GalNAc ligand according to step 1 above is used as the starting cycle, and nucleoside monomers or GalNAc ligands are connected one by one from the 3'-5' direction according to the order of the base sequence. It includes four steps of reactions: each connection of nucleoside monomers, deprotection, coupling, capping, oxidation or thiolation. The synthesis conditions for a synthesis scale of 5 umol of oligonucleic acid are as follows.

[1024] The nucleoside monomer was provided in 0.05 mol / L acetonitrile solution, and the reaction conditions for each step were the same, i.e., the temperature was 25 degrees, the deprotection was performed using 3% trichloroacetic acid-dichloromethane solution, deprotected three times, the activating agent used in the coupling reaction was 0.25 mol / L ETT-acetonitrile solution, coupled twice, 10% acetic anhydride-acetonitrile and pyridine / N-methylimidazole / acetonitrile (10:14:76, v / v / v) were used for capping, capped twice, oxidation was performed using 0.05 mol / L iodine in tetrahydrofuran / pyridine / water (70 / 20 / 10, v / v / v) solution, oxidized twice, and thiolation was performed using 0.2 mol / L PADS acetonitrile / 3-methylpyridine (1 / 1, v / v) to thiolate twice.

[1025] 3. Oligonucleotide Purification and Annealing 3.1 Aminolysis The synthesized solid phase carrier (sense strand or antisense strand) is added to a 5 mL centrifuge tube, 3% diethylamine / ammonia (v / v) is added, and the mixture is reacted in a thermostatic water bath at 35 degrees (or 55 degrees) for 16 hours (or 8 hours), filtered, and the solid phase carrier is washed three times with ethanol / water (1 mL each time), and the crude product is purified after centrifugation and concentration of the filtrate.

[1026] 3.2 Purification Methods of purification and desalting are well known to those skilled in the art, for example, a strong anionic packing material can be used to pack the column, a sodium chloride-sodium hydroxide system can be used for elution and purification, the product can be collected and transferred to a tube, a gel-packed purification column can be used for desalting, and the elution system is pure water.

[1027] 3.3 Annealing Mix the sense strand (SS strand) and the antisense strand (AS strand) in a molar ratio (SS strand / AS strand = 1 / 1.05), heat the water bath to 70-95 degrees, hold for 3-5 minutes, allow to cool naturally to room temperature, and freeze-dry the system to obtain the product.

[1028] Example 29: Examination of the long-term efficacy of the compounds of the present invention in a C57BL / 6 mouse model C57BL / 6 mice (male, 18-21g, 6-8 weeks old) were randomly divided into groups, and the dosage was calculated according to the body weight of each animal. A single dose was administered by subcutaneous injection. The siRNA conjugate was administered as a 1mg / mL solution (solvent: 0.9% sodium chloride aqueous solution). Specifically, before the experiment, the siRNA conjugate was dissolved in 0.9% sodium chloride aqueous solution and adjusted to the required concentration and volume. The administration volume of saline (control group) and siRNA conjugate was 5mL / kg.

[1029] Ten milligrams of liver will be harvested before dosing (recorded as day 0) and at days 7, 14, 21, 28, 35, 42, 56, and 70 after dosing, placed in RNAlater™ solution, and stored frozen at −80° C. for use in detection of hepatic mTTR mRNA.

[1030] Detection of hepatic mTTR mRNA: Cellular RNA was extracted using a nucleic acid extraction device (Hangzhou Aosheng, Auto-pure96) according to the operation protocol of the High Throughput Tissue RNA Extraction Kit (Fanzhi Medical, FG0412), for reverse transcription refer to the PrimeScript™ II 1st Strand cDNA Synthesis Kit (Takara, 6210B), for fluorescent quantitative PCR reaction (ABI, QuantStudio3) detection refer to the TaqMan™ Fast Advanced Master Mix (ABI, 4444965) 20μL system, for primers refer to Table 1. [Table 8]

[1031] Data Statistics and Analysis

[1032] 2 -ΔΔCt Calculate the value and convert it to a percentage to get the remaining inhibition,

[1033] △△Ct = [(Ct gene of interest in experimental group - Ct internal standard for experimental group) - (Ct gene of interest in control group - Ct internal standard for control group)].

[1034] The gene of interest is mTTR and the internal standard is mGAPDH. [Table 9] [Table 10]

[1035] Here, " / " means not tested.

[1036] Example 30: Examination of the long-term efficacy of the compounds of the present invention in a C57BL / 6 mouse model The long-term efficacy of the test compound (see Table 4) in C57BL / 6 mouse model is verified by the experimental procedure in Example 29. The dosage is calculated according to the animal's weight, and administered by subcutaneous injection once, the siRNA conjugate is administered as a 1mg / mL solution (solvent is 0.9% sodium chloride aqueous solution), and the dosage is 1mpk, and 10mg of liver is taken at each time point, put into RNAlater™ solution, and frozen and stored at -80℃, for use in detecting liver mTTR mRNA, and the experimental results are shown in Table 5. [Table 11] [Table 12]

[1037] Example 31: Examination of the long-term efficacy of the compounds of the present invention in a C57BL / 6 mouse model For validation of long-term efficacy of test compounds (see Table 6) in the C57BL / 6 mouse model, refer to the experimental procedures in Example 29.

[1038] The dosage was calculated according to the animal's weight and administered as a single dose by subcutaneous injection. The siRNA conjugate was administered as a 1 mg / mL solution (solvent: 0.9% sodium chloride aqueous solution), with a dosage of 1 mpk. Blood was collected from the orbit at each time point, and serum mTTR protein was detected using an ELISA kit (Abcam, ab282297). The experimental results are shown in Table 7. [Table 13] [Table 14]

[1039] Example 32, Activity detection in HDI mouse model (APOC3) 1.HDI Animal Modeling In vivo transfection modeling is performed on 6-8 week-old female Balb / c mice by a dual gene stable transfection system using the high pressure injection method of the tail vein, and the TransIT®-QR Delivery Solution (total volume is 10% of the animal's body weight, Mirusbio-MIR 5240) of Piggy-Bac transposon plasmid containing target gene cDNA sequence at different mass ratios and helper plasmid is injected into the mice within 5-7 seconds through a 27-gauge needle, and after injection, the animals are returned to their cages and observed for 30 minutes. The modeling day is taken as day 0, and the SEAP expression level is detected at each time point after modeling (day 7-day 35).

[1040] 2. Detection of SEAP Expression The kit (Phospha-Light™ SEAP Reporter Gene Detection System, Invitrogen, T1016) standard started at 15 mU / mL and was diluted two-fold for a total of seven concentration points.

[1041] Mix CSPD substrate and reaction buffer diluent in a ratio of 1:20 to form a reaction solution;

[1042] Dilute the 5x buffer dilution to 1x buffer dilution with RNase-free, DNase-distilled water.

[1043] Mix serum and 1x buffer diluent in a centrifuge tube to form sample diluent, incubate sample diluent at 65°C for 30 min, then cool to room temperature.

[1044] Add 50 μL of sample dilution to a 96-well plate, then add 50 μL of assay buffer to each well and incubate at room temperature for 5 minutes.

[1045] Add 50 μL of reaction solution to each well, incubate at room temperature for 20 min, and read the SEAP chemiluminescence value on a microplate reader (Tecan, Infinite 200).

[1046] 3. Liveness Detection Select Piggy-Bac helper plasmid and Piggy-Bac transposon recombinant plasmid with a mass ratio of 1:1, and model with a total amount of plasmid of 50ug. On the 15th day after modeling, each mouse is subcutaneously administered a single dose according to Table 10, and 200μl saline containing 3mg / kg (mpk) Apoc3 RNAi reagent or 200μl saline without Apoc3 RNAi reagent is used as control. The experimental screening results of HDI model are shown in Table 11. [Table 15] [Table 16]

[1047] Example 33, Activity detection in HDI mouse model (ANGPTL3) Referring to the experimental procedure of Example 32, in this experiment, for the construction of HDI model and drug administration detection, 6-8 week old female Balb / c mice are selected, and Piggy-Bac helper plasmid and Piggy-Bac transposon recombination plasmid with a mass ratio of 1:1 are selected, and modeled with a total amount of plasmid of 100ug. On the 14th day after modeling, each mouse is subcutaneously administered once according to Table 14, and 200μl of saline containing 1mg / kg (mpk) ANGPTL3 RNAi reagent or 200μl of saline without Apoc3 RNAi reagent is used as control. The experimental screening results of HDI model are shown in Table 15. [Table 17] [Table 18] Note: Serum SEAP was normalized to pre-treatment, with pre-treatment set at 100%.

[1048] Example 34, In vivo activity detection (PCSK9) Experimental animals: 40 humanized PCSK9 Tg mice, male, 11–12 weeks old.

[1049] The baseline levels of PCSK9 protein in the animals are measured by ELISA on D-12 (ie, 12 days before administration, the same below) and D-5 before administration, and the baseline level is defined as 100%.

[1050] On DO, the compound to be tested is administered subcutaneously in a single dose of 3 mg / kg, see Table 19 for details. [Table 19]

[1051] After administration, whole blood was collected on D7, D14, D21, and D28, and allowed to stand at room temperature. Serum was then collected by centrifugation and measured by ELISA. The results are shown in Table 20. [Table 20]

[1052] The above is a more detailed description of the present invention combined with certain preferred embodiments, and it cannot be concluded that the specific implementation of the present invention is limited to these descriptions. Those skilled in the art to which the present invention belongs can make some simple deductions or substitutions without departing from the concept of the present invention, which should be considered as belonging to the protection scope of the present invention.

Claims

1. An N-acetylgalactosamine-containing targeting ligand, said ligand comprising a conjugate group of formula (X'): 【Chemistry 1】 During the ceremony, 【Chemistry 2】 indicates the position of attachment to the biomolecule, Q is independently H; 【Chemistry 3】 and In the ceremony, L １ is a chemical bond, -CH ２ --, --CH ２ CH ２ -, -C(O)-, -CH ２ O-, -CH ２ O-CH ２ CH ２ O- or -NHC(O)-(CH ２ NHC (O) ａ - and L ２ is a chemical bond or -CH ２ CH ２ C(O)—, L ３ is a chemical bond, -(NHCH ２ CH ２ ) ｂ -, -(NHCH ２ CH ２ CH ２ ) ｂ -or-C(O)CH ２ - and L ４ is -(OCH ２ CH ２ ), -(OCH ｃ CH ２ CH ２ CH ２ ), -(OCH ｃ CH ２ CH ２ CH ２ CH ２ ), -(OCH ｃ CH ２ CH ２ CH ２ CH ２ CH ２ ), -(OCH ｃ CH ２ ), - or -NHC(O)-(CH ｄ ), and In the formula, a=0, 1, 2 or 3; b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is a chemical bond, -CH ２ -O- or -NHC(O)-, L' is a chemical bond, -C(O)NH-, -NHC(O)- or -O(CH ２ CH ２ O) e-; In the formula, e is 1, 2, 3, 4 or 5; T is a chemical bond, -CH ２ -, -C(O)-, -M-, -CH ２ -M- or -C(O)-M-, In the formula, M is 【Chemistry 4】 and R １ and R ２ Both are -CH ２ CH ２ O- or -CH ２ CH(R)-O-, and R ３ is H, Or, R １ and R ３ Both are -C １～２ alkylene-, and R ２ is H, In the formula, R is -OR', -CH ２ OR' or -CH ２ CH ２ OR', where R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group is preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; An N-acetylgalactosamine-containing targeting ligand, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

2. The conjugated group is shown in formula (I'): 【Chemistry 5】 During the ceremony, 【Chemistry 6】 indicates the position of attachment to the biomolecule, Q is independently H; 【Chemistry 7】 and In the ceremony, L １ is a chemical bond, -CH ２ --, --CH ２ CH ２ -, -C(O)-, -CH ２ O-, -CH ２ O-CH ２ CHO- or -NHC(O)-(CH ２ NHC (O) ａ - and L ２ is a chemical bond or -CH ２ CHC(O)-, L ３ is a chemical bond, -(NHCH ２ CH ２ ) ｂ -, -(NHCH ２ CH ２ CH ２ ) ｂ -or-C(O)CH ２ - and L ４ is -(OCH ２ CH ２ ) ｃ -, -(OCH ２ CH ２ CH ２ ) ｃ -, -(OCH ２ CH ２ CH ２ CH ２ ) ｃ -, -(OCH ２ CH ２ CH ２ CH ２ CH ２ ) ｃ - or -NHC(O)-(CH ２ ) ｄ -, and In the formula, a=0, 1, 2 or 3; b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is -CH ２ -O- or -NHC(O)-, L' is a chemical bond, -C(O)NH- or -NHC(O)-; R １ and R ２ Both are -CH ２ CH ２ O- or -CH ２ CH(R)-O-, and R ３ is H, Or, R １ and R ３ Both are -C １～２ alkylene-, and R ２ is H, In the formula, R is -OR', -CH ２ OR' or -CH ２ CH ２ OR', where R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group is preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; The ligand of claim 1, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

3. Q is independently H or 【Chemistry 8】 and In the ceremony, L １ is -CH ２ O- or -NHC(O)-(CH ２ NHC (O) ａ - and L ２ is -CH ２ CH ２ C(O)—, L ３ is -(NHCH ２ CH ２ ) ｂ - or - (NHCH ２ CH ２ CH ２ ) ｂ - and L ４ is -(OCH ２ CH ２ ) ｃ - or -NHC(O)-(CH ２ ) ｄ - and In the formula, a=0, 1, 2 or 3; b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is -CH ２ O-, L' is a chemical bond; R １ and R ２ Both are -CH ２ CH ２ O- or -CH ２ CH(R)-O-, and R ３ is H, Or, R １ and R ３ Both are -C １～２ alkylene-, and R ２ is H, In the formula, R is -OR', -CH ２ OR' or -CH ２ CH ２ OR', where R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group is preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; The ligand of claim 2, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

4. The conjugated group is represented by formula (I'-1), formula (I'-2) or formula (I'-3): 【Chemistry 9】 During the ceremony, 【Chemistry 10】 indicates the position of attachment to the biomolecule, Q is, 【Chemistry 11】 and In the ceremony, L １ is -CH ２ -O- or -NHC(O)-, L ２ is -CH ２ CH ２ C(O)—, L ３ is -(NHCH ２ CH ２ ) ｂ - or - (NHCH ２ CH ２ CH ２ ) ｂ - and L ４ is -(OCH ２ CH ２ ) ｃ - or -NHC(O)-(CH ２ ) ｄ - and wherein b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is -CH ２ O-, R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; The ligand of claim 2, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

5. Q is independently H; 【Chemistry 12】 and In the ceremony, L １ is -CH ２ O-, -CH ２ O-CH ２ CH ２ O- or -NHC(O)-(CH ２ NHC (O) ａ - and L ２ is a chemical bond, L ３ is -(NHCH ２ CH ２ ) ｂ -, -(NHCH ２ CH ２ CH ２ ) ｂ -or-C(O)CH ２ - and L ４ is -(OCH ２ CH ２ ) ｃ - or -NHC(O)-(CH ２ ) ｄ - and In the formula, a=0, 1, 2 or 3; b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is -CH ２ -O- or -NHC(O)-, L' is a chemical bond or -C(O)NH-; R １ and R ２ Both are -CH ２ CH ２ O- or -CH ２ CH(R)-O-, and R ３ is H, Or, R １ and R ３ Both are -C １～２ alkylene-, and R ２ is H, In the formula, R is -OR', -CH ２ OR' or -CH ２ CH ２ OR', where R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group is preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; The ligand of claim 2, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

6. The conjugated group is represented by formula (II'-1) or formula (II'-2): 【Chemistry 13】 During the ceremony, 【Chemistry 14】 indicates the position of attachment to the biomolecule, Q is independently 【Chemistry 15】 and In the ceremony, L １ is -CH ２ O- or -CH ２ O-CH ２ CH ２ O-, L ３ is -(NHCH ２ CH ２ ) ｂ -, -(NHCH ２ CH ２ CH ２ ) ｂ -or-C(O)CH ２ - and L ４ is -(OCH ２ CH ２ ) ｃ - or -NHC(O)-(CH ２ ) ｄ - and wherein b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is -NHC(O)-, L' is a chemical bond or -C(O)NH-; R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; The ligand of claim 5, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

7. Q is independently H; 【Chemistry 16】 and In the ceremony, L １ is -CH2-, -C(O)-, -CH ２ O-, -CH ２ O-CH ２ CH ２ O- or -NHC(O)-(CH ２ NHC (O) ａ - and L ２ is a chemical bond, L ３ is -(NHCH ２ CH ２ ) ｂ -, -(NHCH ２ CH ２ CH ２ ) ｂ -or-C(O)CH ２ - and L ４ is -(OCH ２ CH ２ ) ｃ - or -NHC(O)-(CH ２ ) ｄ - and In the formula, a=0, 1, 2 or 3; b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is -CH ２ -O- or -NHC(O)-, L' is a chemical bond or -C(O)NH-; R １ and R ２ Both are -CH ２ CH ２ O- or -CH ２ CH(R)-O-, and R ３ is H, Or, R １ and R ３ Both are -C １～２ alkylene-, and R ２ is H, In the formula, R is -OR', -CH ２ OR' or -CH ２ CH ２ OR', where R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group is preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; The ligand of claim 2, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

8. The conjugated group is represented by formula (II'-2): 【Chemistry 17】 During the ceremony, 【Chemistry 18】 indicates the position of attachment to the biomolecule, Q is, 【Chemistry 19】 and L １ is -CH ２ O-, L ３ is -NHCH ２ CH ２ - and L ４ is -(OCH ２ CH ２ ) ｃ - and c=1, 2 or 3; L is -NHC(O)-, R′ is H; 8. The ligand of claim 7, wherein n=6, 7, 8, 9 or 10, preferably 7, 8 or 9, more preferably 8.

9. The conjugated group is represented by formula (II'-2): 【Chemistry 20】 During the ceremony, 【Chemistry 21】 indicates the position of attachment to the biomolecule, Q is independently 【Chemical 22】 and In the ceremony, L １ is -CH ２ - or -C(O)-, L ３ is -(NHCH ２ CH ２ ) ｂ - and L ４ is -(OCH ２ CH2) ｃ - and wherein b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; L is -CH ２ -O- or -NHC(O)-, R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group being preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; The ligand of claim 7, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

10. The conjugated group is represented by formula (II'-3): 【Chemistry 23】 During the ceremony, 【Chemistry 24】 indicates the position of attachment to the biomolecule, Q １ teeth, 【Chemistry 25】 Wherein L １ is -CH ２ - and Q ２ teeth, 【Chemistry 26】 Wherein L １ is —C(O)—, L ３ is -NHCH ２ CH ２ - and L ４ is -(OCH ２ CH ２ ) ｃ - and c=1, 2 or 3; L is -NHC(O)-, R′ is H; 10. The ligand of claim 9, wherein n=6, 7, 8, 9 or 10, preferably 7, 8 or 9, more preferably 8.

11. Q is independently H; 【Chemistry 27】 and In the ceremony, L １ is a chemical bond, -CH ２ --, --CH ２ CH ２ -, -C(O)-, -CH ２ O-, -CH ２ O-CH ２ CH ２ O- or -NHC(O)-(CH ２ NHC (O) ａ - and L ２ is a chemical bond or -CH ２ CH ２ C(O)—, L ３ is a chemical bond, -(NHCH ２ CH ２ ) ｂ -, -(NHCH ２ CH ２ CH ２ ) ｂ -or-C(O)CH ２ - and L ４ is -(OCH ２ CH ２ ) ｃ -, -(OCH ２ CH ２ CH ２ ) ｃ -, -(OCH ２ CH ２ CH ２ CH ２ ) ｃ -, -(OCH ２ CH ２ CH ２ CH ２ CH ２ ) ｃ -, or -NHC(O)-(CH ２ ) ｄ -, and In the formula, a=0, 1, 2 or 3; b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is a chemical bond, -CH ２ -O- or -NHC(O)-, L' is a chemical bond, -C(O)NH-, -NHC(O)-, or -O(CH ２ CH ２ O) e-; In the formula, e is 1, 2, 3, 4 or 5; T is a chemical bond, -CH ２ -, -M-, -CH ２ -M- or -C(O)-M-, In the formula, M is 【Chemistry 28】 and R １ and R ２ Both are -CH ２ CH ２ O- or -CH ２ CH(R)-O-, and R ３ is H, Or, R １ and R ３ Both are -C １～２ alkylene-, and R ２ is H, In the formula, R is -OR', -CH ２ OR' or -CH ２ CH ２ OR', where R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group is preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; The ligand of claim 1, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

12. T is -M-, -CH ２ -M- or -C(O)-M-, where M is 【Chemical Formula 29】 The ligand of claim 11 ,

13. Q is independently H or 【Chemistry 30】 and In the ceremony, L １ is -CH ２ O- or -NHC(O)-(CH ２ NHC (O) ａ - and L ２ is -CH ２ CH ２ C(O)—, L ３ is -(NHCH ２ CH ２ ) ｂ - or - (NHCH ２ CH ２ CH ２ ) ｂ - and L ４ is -(OCH ２ CH ２ ) ｃ - or -NHC(O)-(CH ２ ) ｄ - and In the formula, a=0, 1, 2 or 3; b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is a chemical bond or -CH ２ O-, L' is a chemical bond or -O(CH ２ CH ２ O) ｅ - and In the formula, e is 1, 2, 3, 4 or 5; R １ and R ２ Both are -CH ２ CH ２ O- or -CH ２ CH(R)-O-, and R ３ is H, Or, R １ and R ３ Both are -C １～２ alkylene-, and R ２ is H, In the formula, R is -OR', -CH ２ OR' or -CH ２ CH ２ OR', where R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group is preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; 13. The ligand of claim 11 or 12, wherein T is as defined in claim 11 or 12.

14. The conjugated group is represented by formula (III'-1), formula (III'-2) or formula (III'-3): 【Chemistry 31】 During the ceremony, Q is, 【Chemistry 32】 and In the ceremony, L １ is -CH ２ -O- or -NHC(O)-, L ２ is -CH ２ CH ２ C(O)—, L ３ is -(NHCH ２ CH ２ ) ｂ - or - (NHCH ２ CH ２ CH ２ ) ｂ - and L ４ is -(OCH ２ CH ２ ) ｃ - or -NHC(O)-(CH ２ ) ｄ - and wherein b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is a chemical bond or -CH ２ O-, wherein R′ is H, a hydroxyl protecting group or a solid support, and the hydroxyl protecting group is preferably —C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; 14. The ligand of claim 13, wherein T is as defined in claim 11 or 12.

15. Q is independently H; 【Chemical 33】 and In the ceremony, L １ is -CH ２ --, --CH ２ -O- or -C(O)-; L ２ is a chemical bond, L ３ is -(NHCH ２ CH ２ ) ｂ -, -(NHCH ２ CH ２ CH ２ ) ｂ -or-C(O)CH ２ - and L ４ is -(OCH ２ CH ２ ) ｃ - or -NHC(O)-(CH ２ ) ｄ - and wherein b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is a chemical bond or -NHC(O)-; L' is a chemical bond; R １ and R ２ Both are -CH ２ CH ２ O- or -CH ２ CH(R)-O-, and R ３ is H, Or, R １ and R ３ Both are -C １～２ alkylene-, and R ２ is H, In the formula, R is -OR', -CH ２ OR' or -CH ２ CH ２ OR', where R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group is preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; 13. The ligand of claim 11 or 12, wherein T is as defined in claim 11 or 12.

16. The conjugated group is represented by formula (IV'-1) or formula (IV'-2): 【Chemical Formula 34】 During the ceremony, Q is independently 【Chemistry 35】 and In the ceremony, L １ is -CH ２ --, --CH ２ -O- or -C(O)-, L ３ is -(NHCH ２ CH ２ ) ｂ -, -(NHCH ２ CH ２ CH ２ ) ｂ -or-C(O)CH ２ - and L ４ is -(OCH ２ CH ２ ) ｃ - or -NHC(O)-(CH ２ ) ｄ - and wherein b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is a chemical bond or -NHC(O)-; L' is a chemical bond; wherein R′ is H, a hydroxyl protecting group or a solid support, and the hydroxyl protecting group is preferably —C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; 13. The ligand of claim 11 or 12, wherein T is as defined in claim 11 or 12.

17. The conjugated group is represented by formula (II'-3): 【Chemical Formula 36】 During the ceremony, 【Chemical 37】 indicates the position of attachment to the biomolecule, Q １ teeth, 【Chemical 38】 Wherein L １ is -CH ２ - and Q ２ teeth, 【Chemical 39】 Wherein L １ is a chemical bond, L ３ is -NHCH ２ CH ２ - and L ４ is -(OCH ２ CH ２ ) ｃ - and c=1, 2 or 3; L is -NHC(O)-, R′ is H; 2. The ligand of claim 1, wherein n=6, 7, 8, 9 or 10, preferably 7, 8 or 9, more preferably 8.

18. Q is independently H; 【Chemistry 40】 and In the ceremony, L １ is a chemical bond, -CH ２ --, --CH ２ CH ２ -, -C(O)-, -CH ２ O-, -CH ２ O-CH ２ CH ２ O- or -NHC(O)-(CH ２ NHC (O) ａ - and L ２ is a chemical bond or -CH ２ CH ２ C(O)—, L ３ is a chemical bond, -(NHCH ２ CH ２ ) ｂ -, -(NHCH ２ CH ２ CH ２ ) ｂ -or-C(O)CH ２ - and L ４ is -(OCH ２ CH ２ ), -(OCH ２ CH ２ CH ２ ), ｃ -(OCH ２ CH ２ CH ２ CH ２ ), ｃ -(OCH ２ CH ２ CH ２ CH ２ CH ２ ), ｃ - or -NHC(O)-(CH ２ ), ｄ and is In the formula, a=0, 1, 2 or 3; b=1, 2, 3, 4 or 5; c=1, 2, 3, 4 or 5; d=1, 2, 3, 4, 5, 6, 7 or 8; L is a chemical bond, -CH ２ -O- or -NHC(O)-, L' is -O(CH ２ CH ２ O) ｅ - and In the formula, e is 1, 2, 3, 4 or 5; T is a chemical bond, -CH ２ -, -C(O)-, -M-, -CH ２ -M-, or -C(O)-M-, In the formula, M is 【Chemistry 41】 and R １ and R ２ Both are -CH ２ CH ２ O- or -CH ２ forming CH(R)-O- and R3 is H; Or, R １ and R ３ Both are -C １～２ alkylene-, and R ２ is H, In the formula, R is -OR', -CH ２ OR' or -CH ２ CH ２ OR', where R' is H, a hydroxyl protecting group or a solid support, said hydroxyl protecting group is preferably -C(O)CH ２ CH ２ C(O)OH or 4,4′-dimethoxytrityl; m=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10; The ligand of claim 1, wherein n=0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

19. 2. The ligand of claim 1, wherein the conjugate group is selected from the following: 【Table 1-1】 【Table 1-2】 【Table 1-3】 【Table 1-4】 【Table 1-5】

20. 2. The ligand of claim 1, wherein the conjugate group is selected from the following: 【Table 2-1】 【Table 2-2】 【Table 2-3】 【Table 2-4】 【Table 2-5】 【Table 2-6】

21. A ligand according to any one of claims 1 to 20 which targets ASGPR.

22. A nucleic acid molecule comprising a nucleic acid and a ligand according to any one of claims 1 to 21 connected thereto.

23. 23. The nucleic acid molecule of claim 22, wherein the nucleic acid is selected from DNA, RNA and a DNA / RNA hybrid.

24. 24. The nucleic acid molecule of claim 23, wherein the nucleic acid molecule is single-stranded or double-stranded.

25. 24. The nucleic acid molecule of claim 23, wherein the nucleic acid molecule is selected from a small interfering RNA (siRNA) and a short hairpin RNA (shRNA).

26. The nucleic acid molecule of claim 23, wherein the nucleic acid molecule is a double-stranded RNA molecule and comprises a sense strand and an antisense strand, and the conjugate group is attached to the 3'-end or 5'-end, preferably the 3'-end, of the sense strand of the double-stranded RNA molecule.

27. 27. The nucleic acid molecule according to claim 26, wherein the sense and antisense strands of the double-stranded RNA molecule each have 10 to 30, preferably 15 to 25, more preferably 20 to 25 nucleotides.

28. 28. The nucleic acid molecule of claim 26 or 27, wherein one or more nucleotides of the sense strand and / or the antisense strand of the double-stranded RNA molecule comprise a chemical modification.

29. 28. The nucleic acid molecule of claim 26 or 27, wherein one or more nucleotides in the sense strand and / or the antisense strand of the double-stranded RNA molecule are replaced with a nucleotide analogue.

30. 30. The nucleic acid molecule according to any one of claims 22 to 29, which specifically binds to the asialoglycoprotein receptor (ASGPR) on the surface of liver cells.

31. A pharmaceutical composition comprising a nucleic acid molecule according to any one of claims 22 to 30 and a pharma- ceutically acceptable carrier or excipient.

32. A kit comprising a nucleic acid molecule according to any one of claims 22 to 30.

33. An intermediate compound, which is a compound of formula (V): 【Chemistry 42】 During the ceremony, Pg is, for example, a hydroxyl protecting group selected from 【Chemistry 43】 Q' is independently H; 【Chemistry 44】 and The intermediate compound, wherein the other groups are as defined in any one of claims 1 to 18.

34. An intermediate compound selected from: 【Table 3-1】 【Table 3-2】 【Table 3-3】 【Table 3-4】 【Table 3-5】

35. An intermediate compound selected from: 【Table 4-1】 【Table 4-2】 【Table 4-3】 【Table 4-4】 【Table 4-5】 【Table 4-6】 【Table 4-7】

36. A process for preparing a compound of formula (I), comprising treating a compound of formula (V) under conditions to remove a hydroxyl protecting group, 【Chemistry 45】 A process for the preparation of a compound of formula (I), wherein each group is as defined in any one of claims 1 to 18.

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