Novel GalNAc targeted delivery fragment and preparation and application thereof

Abstract

The invention discloses a GalNAc delivery molecule shown in the following formula (I), or a stereoisomer, an enantiomer or a pharmaceutically acceptable salt thereof, wherein the definition of each group is described in the specification. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to the field of medicine, and in particular, provides a novel GalNAc targeted delivery fragment and its preparation and application. Background Art

[0002] Hepatitis B virus (HBV) belongs to the Hepaciviridae family of small, enveloped, primarily hepatotropic viruses with a partially double-stranded DNA genome of approximately 3.2 kb in size. HBV infection has been a major public health problem. Despite the availability of safe and effective preventive HBV vaccines, it is estimated that approximately 250 million people worldwide are chronically infected with HBV. Chronic HBV infection puts patients at high risk for cirrhosis and liver cancer, and it is estimated that more than 686,000 people die each year from complications of HBV. (WHO.Global hepatitis report 2017)

[0003] Current treatments are limited to two classes of drugs, which are nucleoside (nucleotide) analogs (lamivudine, adefovir, tenofovir, telbivudine, and entecavir) and interferon alpha (INF-α, both non-PEGylated and PEGylated). Although both therapies can reduce HBV DNA and normalize liver enzymes, neither therapy can provide a high level of clinical cure, which is defined by the loss of HBV surface antigen (HBsAg) with or without seroconversion. Interferon-based therapies are poorly tolerated and are only effective against certain viral genotypes. Although nucleoside (nucleotide) analogs often require prolonged or potentially lifelong treatment, and some have resistance, low efficacy, and tolerability issues. Therefore, there is still a huge medical need to discover and develop effective and safe anti-HBV drugs with novel mechanisms of action to improve disease cure rates. (Marcellin, Patrick, et al. The Lancet 381.9865 (2013): 468-475; Tang, Lydia SY, et al. Jama 319.17 (2018): 1802-1813.)

[0004] GalNAc-mediated liver-targeted delivery of small interfering RNA drugs (GalNAc-siRNA) can inhibit the expression of target genes at the basic level of mRNA translation into protein, and has recently been used to treat a variety of liver-related diseases. GalNAc-siRNA molecules have the characteristics of strong efficacy, high liver targeting specificity, and good safety, so they have unique advantages in the field of anti-hepatitis B, but current experimental drugs cannot effectively reduce HBsAg content and cannot achieve a high level of clinical cure rate. Therefore, it is necessary to optimize this type of molecule, focusing on the structural modification of the targeted delivery fragment containing GalNAc to improve delivery efficiency and drug properties. Summary of the invention

[0005] The object of the present invention is to provide a GalNAc-siRNA molecule and a nucleic acid delivery pharmaceutical composition prepared using the same.

[0006] In the first aspect of the present invention, there is provided a GalNAc delivery molecule as shown in the following formula (I), or a stereoisomer, enantiomer, or a pharmaceutically acceptable salt thereof:

[0007]

[0008] in,

[0009] R1 is selected from the group consisting of H, DMTr, phosphate, phosphorothioate, phosphorothioate-siRNA conjugate or phosphate-siRNA conjugate;

[0010] Linker is a divalent linking group;

[0011] r is 0, 1, 2, 3, 4, 5 or 6;

[0012] R2 is a GalNAc target head portion and has a structure as shown below:

[0013] Wherein, L1, L2 and L3 are each independently a divalent linking group, and R3, R4 and R5 are each independently a substituted or unsubstituted sugar group, preferably a group formed by a substituted or unsubstituted N-acetylgalactosamine molecule;

[0014] The substitution refers to the substitution of one or more hydrogen atoms on the group by a substituent selected from the following group: 1-8 Alkyl, C 2-8 Alkenyl, C 2-8 Alkynyl, C 3-8 Cycloalkyl, 3- to 12-membered heterocyclic group, C 3-8 Aryl, 5- to 7-membered heteroaryl, halogen, hydroxyl, carboxyl (-COOH), C 1-8 Aldehyde, C 2-10 Acyl, C 2-10 Ester group, amino group, C 1-8 Alkoxy, C 1-10 or two substituents located on adjacent ring atoms may together with the connected ring atoms form a group selected from the following group: a 5-7 membered carbon ring or heterocyclic ring, a benzene ring, or a 5-7 membered heteroaromatic ring.

[0015] In another preferred embodiment, the siRNA is selected from the following group: siRNA that inhibits the expression of hepatitis B virus (HBV) gene, siRNA that inhibits the expression of apolipoprotein C3 (APOC3) gene, siRNA that inhibits the expression of PCSK9 gene, siRNA that inhibits the expression of CCR4 gene, and siRNA that inhibits the expression of thyroxine transporter (TTR) gene.

[0016] In another preferred embodiment, the Linker has a structure selected from the following group:

[0017] C4-C20 alkyl,

[0018] wherein n and m are each independently selected from the group consisting of 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12;

[0019] n1 and n2 are each independently selected from the following group: 0 or 1;

[0020] W is selected from the group consisting of a single bond, a substituted or unsubstituted C 1-8 Alkylene, substituted or unsubstituted 7-12 membered fused bicyclic ring, substituted or unsubstituted 7-15 membered spirocyclic ring, substituted or unsubstituted 5-15 membered bridged ring, -NH-substituted or unsubstituted 7-12 membered fused bicyclic ring, -NH-substituted or unsubstituted 7-15 membered spirocyclic ring, -NH-substituted or unsubstituted 5-15 membered bridged ring, -NH-substituted or unsubstituted 7-12 membered fused bicyclic ring-NH-, -NH-substituted or unsubstituted 7-15 membered spirocyclic ring-NH-, -NH-substituted or unsubstituted 5-15 membered bridged ring-NH-; wherein the ring atoms of the fused bicyclic ring, spirocyclic ring and bridged ring may be optionally carbon atoms or heteroatoms;

[0021] The substitution refers to the substitution of one or more hydrogen atoms on the group by a substituent selected from the following group: 1-8 Alkyl, C 2-8 Alkenyl, C 2-8 Alkynyl, C 3-8 Cycloalkyl, 3- to 12-membered heterocyclic group, C 6-10 Aryl, 5- to 10-membered heteroaryl, halogen, hydroxyl, carboxyl (-COOH), C 1-8 Aldehyde, C 2-10 Acyl, C 2-10 Ester group, amino group, C 1-8 Alkoxy, C 1-10 Sulfonyl; or two substituents located on adjacent ring atoms may together with the connected ring atoms form a group selected from the following group: a 5-7 membered carbon ring or heterocyclic ring, a benzene ring, or a 5-7 membered heteroaromatic ring;

[0022] Or W has a structure selected from the group consisting of:

[0023]

[0024]

[0025] wherein s1, s2, s, s4, s5 and s6 are each independently selected from the following group: 0, 1 or 2;

[0026] s7 is selected from the group consisting of 1, 2, 3, 4, 5 or 6;

[0027] Each of M1, M2, M3 and M4 is independently selected from the group consisting of CHR, C(R)R, C(O), O, S, NR;

[0028] M5 and M6 are each independently selected from the following group: (CHR) s6 、C(R)R、C(O)、(CHR) s6 -C(O), O, S, NR;

[0029] R is selected from the following group: H, halogen, methyl; or two R located on adjacent reducing atoms and the carbon atom to which they are connected together form a substituted or unsubstituted 4-8 membered carbocyclic or heterocyclic ring (including saturated, unsaturated or aromatic ring).

[0030] In another preferred embodiment, the W has a structure selected from the following group:

[0031]

[0032]

[0033] In another preferred embodiment, L1, L2 and L3 are each independently selected from the following structures:

[0034]

[0035] wherein each u, v and w are independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or 11; u1 and u2 are independently selected from 0 or 1; Glu is an unmodified or modified 5-6-membered glycosyl;

[0036] X and Y are each independently selected from the following group: -NH-(C1-C6 alkyl)-NH-, -NH-(C3-C8 cycloalkyl)-NH-, -NH-(4-10 membered heterocyclyl)-NH-, Wherein, the ring D is a 4-15 membered heterocyclic group;

[0037] The cycloalkyl and heterocyclic groups may be monocyclic, condensed, bridged or spirocyclic; the heterocyclic group may be saturated or partially unsaturated (but not aromatic).

[0038] In another preferred embodiment, R2 and the linker portion include at least one monocyclic, fused, bridged or spirocyclic structural unit.

[0039] In another preferred embodiment, R2 and the linker portion include at least one fused ring, bridged ring or spiro ring structural unit.

[0040] In another preferred embodiment, X and Y each independently have a structure selected from the following group:

[0041] -NH-(C1-C6 alkyl)-NH-, -NH-(C3-C8 cycloalkyl)-NH-, -NH-(4-10 membered heterocyclyl)-NH-, or a structure selected from the following group:

[0042]

[0043] In another preferred embodiment, the sugar group has a structure shown in the following formula III:

[0044]

[0045] Among them, the R 11 Selected from the group consisting of -NH(C2-C6 acyl), -NH(halogenated C2-C6 acyl), -NH(C2-C6 sulfonyl), -NH(halogenated C2-C6 sulfonyl);

[0046] R 12 , R 13 and R 14 Each is independently selected from the following group: H or C1-C6 acyl.

[0047] In another preferred embodiment, the R 11 Selected from the group consisting of: -NH(C2-C4 acyl); R 12 , R 13 and R 14 Each is independently selected from the following group: H or C1-C4 acyl.

[0048] In another preferred embodiment, the R 11 is -NHAc; R 12 , R 13 and R 14 Each is independently selected from the group consisting of H or Ac.

[0049] In another preferred embodiment, the GalNAc delivery molecule has a structure selected from the group consisting of:

[0050]

[0051] Wherein, X, Y, Z and W are each independently selected from the following groups: -NH-(C1-C6 alkyl)-NH-, -NH-(C3-C8 cycloalkyl)-NH-, -NH-(4-10 membered heterocyclyl)-NH-, Wherein, the ring D is a 4-15 membered heterocyclic group;

[0052] R3, R4, R5 and R5' are each independently a substituted or unsubstituted sugar group, preferably a group formed by a substituted or unsubstituted N-acetylgalactosamine molecule;

[0053] The remaining groups are as defined above.

[0054] In another preferred embodiment, the GalNAc delivery molecule has a structure selected from the following group:

[0055]

[0056]

[0057]

[0058]

[0059]

[0060]

[0061]

[0062]

[0063]

[0064]

[0065]

[0066]

[0067]

[0068] The R1 group is as defined in claim 1.

[0069] In another preferred embodiment, the GalNAc delivery molecule has a structure selected from the following group:

[0070]

[0071]

[0072]

[0073]

[0074]

[0075]

[0076]

[0077]

[0078]

[0079]

[0080]

[0081]

[0082]

[0083]

[0084]

[0085]

[0086]

[0087]

[0088]

[0089]

[0090]

[0091]

[0092]

[0093]

[0094]

[0095]

[0096]

[0097] The second aspect of the present invention provides a pharmaceutical composition, which comprises: one or more of the compound of formula I as described in the first aspect of the present invention, its pharmaceutically acceptable salts, racemates, R-isomers, S-isomers or mixtures thereof, and one or more pharmaceutically acceptable carriers, excipients, adjuvants, auxiliary materials and / or diluents.

[0098] The third aspect of the present invention provides a compound of formula I as described in the first aspect of the present invention, and the use of its pharmaceutically acceptable salt, racemate, R-isomer, S-isomer or a mixture thereof for preparing a pharmaceutical composition for treating or preventing tumors or infections caused by viruses.

[0099] In a fourth aspect, the present invention provides the use of the compound of formula (I) as described in the first aspect of the present invention for preparing a medicament for treating and / or preventing a disease or condition selected from the following group: a disease or condition caused by hepatitis B virus, a disease or condition caused by abnormal expression of ANGPTL3 gene, a disease or condition caused by abnormal expression of TTR gene, a disease or condition caused by abnormal expression of hepatocyte genes, or other liver diseases or conditions.

[0100] In another preferred embodiment, the disease or condition caused by abnormal ANGPTL3 gene expression is cardiovascular disease and / or metabolic disease, including hyperlipidemia, hypertriglyceridemia, hypercholesterolemia, obesity, diabetes and / or ischemic heart disease.

[0101] In another preferred embodiment, the disease or condition caused by abnormal TTR gene expression is selected from the following group: sensory neuropathy, including distal limb paresthesia, hypoesthesia; autonomic neuropathy, including gastrointestinal dysfunction such as gastric ulcer, or postural hypotension; motor neuropathy; epileptic seizures; dementia; myelopathy; polyneuropathy; carpal tunnel syndrome; autonomic defect; cardiomyopathy; vitreous opacities; renal insufficiency; nephropathy; substantial decrease in mBMI; cranial nerve dysfunction and lattice corneal dystrophy.

[0102] In another preferred embodiment, the hepatocyte gene is selected from the following group: ApoB, ApoC, ANGPTL3, PCSK9, SCD1, TIMP-1, CollA1, FVII, STAT3, p53, HBV, and HCV.

[0103] In another preferred embodiment, the other liver diseases or disorders are selected from the group consisting of hepatocellular carcinoma (HCC), liver metastasis, hepatoblastoma, diseases involving coagulation factors, complement-mediated inflammation or fibrosis, dyslipidemia and irregularities in glucose regulation.

[0104] The fifth aspect of the present invention provides a method for detecting ASGPR-GalNAc affinity based on primary monkey cells.

[0105] In a sixth aspect of the present invention, there is provided a GalNAc linker as shown in formula (Ia), or a stereoisomer, enantiomer, or a pharmaceutically acceptable salt thereof:

[0106]

[0107] The moiety shown in formula (Ia) has a structure selected from the group consisting of:

[0108]

[0109]

[0110]

[0111]

[0112]

[0113]

[0114]

[0115]

[0116]

[0117]

[0118]

[0119]

[0120]

[0121]

[0122] in, It is the site connected to R1.

[0123] It should be understood that within the scope of the present invention, the above-mentioned technical features of the present invention and the technical features specifically described below (such as embodiments) can be combined with each other to form a new or preferred technical solution. Due to space limitations, they will not be described one by one here. DETAILED DESCRIPTION

[0124] Based on long-term and in-depth research, the inventors have prepared a novel type of GalNAc targeted drug delivery fragment with a novel structure. The drug or drug composition composed of the GalNAc targeted delivery fragment has good targeted delivery effect, good efficacy and higher safety. Based on the above findings, the inventors have completed the present invention.

[0125] the term

[0126] In the present invention, the halogen is F, Cl, Br or I.

[0127] In the present invention, unless otherwise specified, the terms used have the general meanings well known to those skilled in the art.

[0128] In the present invention, the term "C1-C6 alkyl" refers to a straight or branched alkyl group having 1 to 6 carbon atoms, including but not limited to methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl and hexyl, etc.; preferably ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl and tert-butyl.

[0129] In the present invention, the term "C1-C6 alkoxy" refers to a straight or branched alkoxy group having 1 to 6 carbon atoms, including but not limited to methoxy, ethoxy, propoxy, isopropoxy, butoxy and the like.

[0130] In the present invention, the term "C3-C7 cycloalkyl" refers to a cyclic alkyl group having 3 to 7 carbon atoms in the ring, including but not limited to cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, etc. The terms "C5-C6 cycloalkyl" and "C3-C6 cycloalkyl" have similar meanings.

[0131] In the present invention, the term "aromatic ring" or "aryl group" has the same meaning, preferably "aryl group" is "C6-C 12 Aryl" or "C6-C 10 The term "C6-C 12 "Aryl" refers to an aromatic ring group having 6 to 12 carbon atoms without heteroatoms in the ring, such as phenyl, naphthyl, etc. The term "C6-C 10 "Aryl" has a similar meaning.

[0132] In the present invention, the term "aromatic heterocycle" or "heteroaryl" has the same meaning and refers to a heteroaromatic group containing one to multiple heteroatoms. The heteroatoms referred to here include oxygen, sulfur and nitrogen. For example, furanyl, thienyl, pyridyl, pyrazolyl, pyrrolyl, N-alkylpyrrolyl, pyrimidyl, pyrazinyl, imidazolyl, tetrazolyl, etc. The heteroaryl ring can be fused to an aryl, heterocyclic or cycloalkyl ring, wherein the ring connected to the parent structure is a heteroaryl ring. The heteroaryl group can be optionally substituted or unsubstituted.

[0133] In the present invention, the term "3-9 membered carbocyclic group" refers to a saturated or unsaturated (non-aromatic ring, including monocyclic, cyclic, spirocyclic, bridged ring, etc.) 3-9 membered cyclic group, whose ring skeleton structure only includes carbon atoms, such as cyclopentyl, cyclohexyl, etc.

[0134] In the present invention, the term "3-9 membered heterocyclic group" refers to a 3-9 membered cyclic group containing 1 to 3 heteroatoms selected from oxygen, sulfur and nitrogen, which is saturated or unsaturated (non-aromatic ring, including monocyclic, cyclic, spirocyclic, bridged, etc.), such as dioxolanyl, etc. The term "3-7 membered heterocyclic group" has a similar meaning.

[0135] In the present invention, the term "substituted" refers to one or more hydrogen atoms on a specific group being replaced by a specific substituent. The specific substituent is a substituent described above or a substituent appearing in the embodiments. Unless otherwise specified, a substituted group may have a substituent selected from a specific group at any substitutable position of the group, and the substituent may be the same or different at each position. A cyclic substituent, such as a heterocycloalkyl, may be connected to another ring, such as a cycloalkyl, to form a spirobicyclic system, for example, the two rings have a common carbon atom. It will be understood by those skilled in the art that the combinations of substituents contemplated by the present invention are those that are stable or chemically feasible. The substituents are, for example (but not limited to): C 1-8 Alkyl, C 2-8 Alkenyl, C 2-8 Alkynyl, C 3-8 Cycloalkyl, 3- to 12-membered heterocyclic group, aryl, heteroaryl, halogen, hydroxyl, carboxyl (-COOH), C 1-8 Aldehyde, C 2-10 Acyl, C 2-10 Ester group, amino group, alkoxy group, C 1-10 Sulfonyl, etc.

[0136] Pharmaceutical composition containing active ingredients

[0137] The pharmaceutical composition of the present invention comprises a safe and effective amount of the compound of the present invention and a pharmaceutically acceptable excipient or carrier. Wherein "safe and effective amount" means: the amount of the compound is sufficient to significantly improve the condition without causing serious side effects. Usually, the pharmaceutical composition contains 0.01-99.99% by weight of the compound of the present invention / dose, and more preferably, contains 0.1-99.9% by weight of the compound of the present invention / dose. Preferably, the "one dose" is a capsule or tablet.

[0138] "Pharmaceutically acceptable carrier" refers to: one or more compatible solid or liquid fillers or gel substances, which are suitable for human use and must have sufficient purity and sufficiently low toxicity. "Compatibility" here means that the components in the composition can be mixed with the compounds of the present invention and with each other without significantly reducing the efficacy of the compounds. Some examples of pharmaceutically acceptable carriers include cellulose and its derivatives (such as sodium carboxymethyl cellulose, sodium ethyl cellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (such as stearic acid, magnesium stearate), calcium sulfate, vegetable oils (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (such as propylene glycol, glycerol, mannitol, sorbitol, etc.), emulsifiers (such as ), wetting agents (such as sodium lauryl sulfate), colorants, flavoring agents, stabilizers, antioxidants, preservatives, pyrogen-free water, etc.

[0139] There is no particular limitation on the administration of the compound or pharmaceutical composition of the present invention. Representative administration methods include (but are not limited to): oral administration, parenteral administration (intravenous administration, intramuscular administration or subcutaneous administration).

[0140] Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules. In these solid dosage forms, the active compound is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or with the following ingredients: (a) fillers or extenders, for example, starches, lactose, sucrose, glucose, mannitol, and silicic acid; (b) binders, for example, hydroxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone, sucrose, and acacia; (c) humectants, for example, glycerol; (d) disintegrants, for example, agar, calcium carbonate, potato starch or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (e) solubilizers, for example, paraffin; (f) absorption accelerators, for example, quaternary ammonium compounds; (g) wetting agents, for example, cetyl alcohol and glyceryl monostearate; (h) adsorbents, for example, kaolin; and (i) lubricants, for example, talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, or mixtures thereof. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents.

[0141] Solid dosage forms such as tablets, pills, capsules, pills and granules can be prepared using coatings and shell materials, such as enteric coatings and other materials known in the art. They may contain opacifiers, and the release of the active compound or compounds in such compositions can be delayed in a certain part of the digestive tract. Examples of embedding components that can be used are polymeric substances and waxes. If necessary, the active compound can also be formed into microencapsulated form with one or more of the above-mentioned excipients.

[0142] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or tinctures. In addition to the active compound, the liquid dosage form may contain an inert diluent conventionally used in the art, such as water or other solvents, solubilizers and emulsifiers, for example, ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, 1,3-butylene glycol, dimethylformamide and oils, in particular cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil or mixtures of these substances.

[0143] Besides such inert diluents, the composition may also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

[0144] Suspensions, in addition to the active compounds, may contain suspending agents such as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methanol and agar, or mixtures of these substances, and the like.

[0145] Compositions for parenteral injection may include physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous and non-aqueous carriers, diluents, solvents or excipients include water, ethanol, polyols and suitable mixtures thereof.

[0146] The compounds of the present invention may be administered alone or in combination with other pharmaceutically acceptable compounds (eg, anti-HBV agents).

[0147] When administered in combination, the pharmaceutical composition also includes one or more (2, 3, 4, or more) other pharmaceutically acceptable compounds. One or more (2, 3, 4, or more) of the other pharmaceutically acceptable compounds can be used simultaneously, separately or sequentially with the compound of the present invention to prevent and / or treat related diseases or conditions.

[0148] When using a pharmaceutical composition, a safe and effective amount of the compound of the present invention is applied to a mammal (such as a human) in need of treatment, wherein the dosage when applied is a pharmaceutically effective dosage. Of course, the specific dosage should also take into account factors such as the route of administration and the patient's health status, which are all within the skill of a skilled physician.

[0149] The present invention will be further described below in conjunction with specific examples. It should be understood that these examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention. The experimental methods in the following examples that do not specify specific conditions are usually based on conventional conditions or the conditions recommended by the manufacturer. Unless otherwise stated, percentages and parts are calculated by weight.

[0150] In the following examples, the structures of the compounds were determined by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). -6 The unit of (ppm) is given. NMR measurements were performed using a Bruker AVANCE-400 NMR spectrometer, with deuterated dimethyl sulfoxide (DMSO-d6), deuterated chloroform (CDCl3), deuterated methanol (CD3OD) as the measuring solvent, and tetramethylsilane (TMS) as the internal standard.

[0151] SHIMADZU LC system (chromatographic column: CSH TM Prep-C18, 19*150mm, liquid handler LH-40, pump LC-20AP, detector SPD-20A, system controller CBM-20A, solvent system: acetonitrile and 0.05% trifluoroacetic acid aqueous solution).

[0152] LC / MS spectra of the compounds were obtained using LC / MS (Agilent Technologies 1200 Series). LC / MS conditions were as follows (run time was 10 minutes):

[0153] Acidic conditions: A: 0.05% trifluoroacetic acid in water; B: 0.05% trifluoroacetic acid in acetonitrile;

[0154] Alkaline conditions: A: 0.05% NH3·H2O aqueous solution; B: acetonitrile

[0155] Neutral conditions: A: 10 mM NH4OAC in water; B: acetonitrile

[0156] Unless otherwise specified, in the following examples, intermediates and final compounds were purified by silica gel column chromatography or by CSH TM Prep-C18(5μm, OBD TM 19*150mm) column or XBridgeTM Prep Phenyl (5μm, OBD TM 30*100 mm) was purified by preparative HPLC on a reverse phase column.

[0157] Silica gel column chromatography generally uses Yantai Huanghai Silica Gel 200-300 mesh silica gel as the carrier.

[0158] The CombiFlash rapid preparation instrument used Combiflash Rf200 (TELEDYNE ISCO).

[0159] Thin layer chromatography (TLC) silica gel plates use Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plates. The specifications of silica gel plates used in thin layer chromatography detection products are 0.15mm~0.2mm, and the specifications used in thin layer chromatography separation and purification products are 0.4mm~0.5mm.

[0160] The known starting materials of the present invention can be synthesized by methods known in the art, or can be purchased from ABCR GmbH & Co. KG, Acros Organics, Aldrich Chemical Company, AccelaChemBio Inc, Darui Chemicals and other companies.

[0161] Abbreviations:

[0162] TMSOTf: trimethylsilyl trifluoromethanesulfonate; NaIO4: sodium periodate; RuCl3: ruthenium trichloride; BnBr: benzyl bromide; K2CO3: potassium carbonate; DIEPA (DIEA): N,N-diisopropylethylamine; TFA: trifluoroacetic acid; DCM: dichloromethane; TEA: triethylamine; HATU: benzotriazole-N,N,N',N'-tetramethyluronium hexafluorophosphate; CbzCl: benzyl chloroformate; NaOH: sodium hydroxide; DMSO: dimethyl sulfoxide; HOBt: 1-hydroxybenzotriazole; EDCI: 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride; DMF: N,N-dimethylformamide; tert-butyl acrylate: tert-butyl acrylate; Formic acid: formic acid; DMTr: 4,4'-dimethoxytrityl; Ac: acetyl; Boc: tert-butyloxycarbonyl; Bn: benzyl; Cbz: benzyloxycarbonyl

[0163] Preparation Example Preparation of Compound A1

[0164]

[0165] Step 1: Preparation of compound A1-1

[0166] 4A molecular sieve (30 g) and TMSOTf (18.8 g, 84.8 mmol) were slowly added to a dichloromethane (300 ml) solution of compound Int.1 (30 g, 77.1 mmol); after the addition, the reaction mixture was stirred at 55°C for 3 hours. Then the reaction mixture was neutralized with triethylamine, poured into 500 ml of water, and diluted and extracted with dichloromethane. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a yellow oil compound A1-1 (22.8 g). MS: 330.2 (M+H) + .

[0167] Step 2: Preparation of compound A1-2

[0168] Under nitrogen protection, 4A molecular sieve (10 g) and compound Int.2 (3.7 g, 15.2 mmol) were slowly added to a dichloroethane (100 ml) solution of compound A1-1 (11 g, 30.4 mmol). After the addition, the reaction mixture was stirred at 25°C for 30 minutes; then TMSOTf (3.7 g, 15.2 mmol) was slowly added to the above reaction mixture under stirring at 0 degrees, and the reaction mixture was stirred at 25°C for 16 hours. The reaction mixture was filtered, the filter cake was washed with dichloromethane, the filtrate was neutralized with sodium bicarbonate solution and extracted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a yellow oil compound A1-2 (7 g, 48.8%). MS: 430.2 (M+H) + .

[0169] Step 3: Preparation of Compound A1-3

[0170] Under nitrogen protection, a mixed solution of NaIO4 (13.0 g, 60.6 mmol) and compound A1-3 (6.5 g, 15.2 mmol) in dichloromethane (30 ml), acetonitrile (30 ml) and water (45 ml) was stirred at 0°C for 30 minutes; then, RuCl3 3H2O (95 mg, 0.45 mmol) was slowly added to the above reaction mixture under stirring at 0 degrees, and the reaction mixture was stirred at 25°C for 16 hours. The reaction mixture was diluted with 100 ml of water and sodium bicarbonate was added to adjust the pH value to 7.5, and extracted with dichloromethane. The aqueous phase was adjusted to pH 3 with citric acid and extracted with dichloromethane. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a white foam solid compound A1-3 (3.2 g, 45%). MS: 448.2 (M+H) + . 1 H NMR (400MHz, DMSO-d6) δ11.98(s,1H),7.81(d,J=9.2Hz,1H),5.21(d,J=3.2Hz,1H),4.96(dd,J=11.2,3.4Hz,1H),4.49(d,J=8.4Hz,1H),4.02(s, 3H),3.87(d,J=10.0Hz,1H),3.78–3.37(m,2H),2.20(t,J=6.8Hz,2H),2. 10(s,3H),2.00(s,3H),1.89(s,3H),1.77(s,3H),1.49(d,J=2.8Hz,4H).

[0171] Step 4: Preparation of Compound A1

[0172] Under nitrogen protection at 0 degrees, compound Int.3 (10.3 g, 36.8 mmol) was slowly added to a solution of compound A1-3 (11.0 g, 24.6 mmol) and DIEA (6.4 g, 49.2 mmol) in dichloromethane (100 ml), and then the reaction mixture was stirred at 25°C for 3 hours. The reaction mixture was concentrated to obtain a crude product, which was purified to obtain a yellow oil compound A1 (11.2 g, 74.2%). MS: 614.2 (M+H) + . 1 H NMR(400MHz, DMSO-d6)δ7.83(d,J=9.2Hz,1H),5.22(d,J=3.2Hz,1H),4.96(dd, J=11.2,3.4Hz,1H),4.51(d,J=8.4Hz,1H),4.03(s,3H),3.90(dd,J=20.0,9.0H z,1H),3.62(dtd,J=15.8,10.2,6.0Hz,2H),2.79(t,J=7.4Hz,2H),2.11(s,3H) ,2.00(s,3H),1.89(s,3H),1.76(s,3H),1.73–1.64(m,2H),1.63–1.54(m,2H).

[0173] Preparation Example Preparation of Compound A2

[0174]

[0175] Step 1: Preparation of compound A2-1

[0176] Under stirring at room temperature, benzyl bromide (159 g, 0.927 mol) was slowly added to a reaction mixture of compound Int.4 (150 g, 1.03 mol) and potassium carbonate (185 g, 1.34 mol) in DMF (750 ml) and 1,4-dioxane (750 ml). After the addition, the reaction mixture was stirred at 90°C for 16 hours; then the reaction mixture was diluted with 3000 ml of water and extracted with ethyl acetate, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a white solid compound A2-1 (60 g). MS: 235.0 (MH) + .

[0177] Step 2: Preparation of compound A2-3

[0178] The reaction mixture of compound A2-1 (59 g, 0.252 mol), compound Int.5 (25 g, 0.126 mol), DIPEA (49 g, 0.378 mol) and HATU (48 g, 0.126 mol) in dichloromethane (300 ml) was stirred at 25°C for 0.5 hours; then the reaction mixture was diluted with 300 ml of water and extracted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a yellow oil compound A2-3 (25 g). MS: 417.7 (M+H) + . 1 H NMR(400MHz,DMSO-d6)δ7.38–7.32(m,5H),5.08(s,2H),4.18(s,2H),3.95(s,2H),3.93(s,2H),3.67–3.5 7(m,1H),3.19–3.10(m,1H),2.35(t,J=7.2Hz,2H),2.00(t,J=7.2Hz,2H),1.56–1.42(m,4H),1.37(s,9H).

[0179] Step 3: Preparation of Compound A2-4

[0180] Under stirring conditions at 0 degrees, trifluoroacetic acid (10 ml) was slowly added to a solution of compound A2-3 (3 g, 7.2 mmol) in dichloromethane (30 ml), and the reaction mixture was stirred at 25°C for 3 hours; the reaction mixture was then concentrated to obtain compound A2-4 (2.3 g). MS: 317.2 (M+H) + .

[0181] Step 4: Preparation of Compound A2-5

[0182] Under stirring conditions at room temperature, compound Int.3 (34 g, 0.121 mol) was slowly added to a reaction mixture of compound Int.4 (8 g, 55 mmol) and DIPEA (21 g, 0.165 mol) in dichloromethane (100 ml). After the addition, the reaction mixture was heated and stirred for 0.5 hour; then the reaction mixture was diluted with 100 ml of water and extracted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a white solid compound A2-5 (20 g). 1 H NMR (400MHz, CDCl3) δ2.81–2.68(m,4H),2.01–1.85(m,4H).

[0183] Step 5: Preparation of Compound A2

[0184] Under stirring conditions at 0 degrees, compound A2-4 (2.3 g, 7.3 mmol) was slowly added to a reaction mixture of compound A2-5 (8.6 g, 18.2 mmol) and DIPEA (2.8 g, 21.9 mmol) in dichloromethane (100 ml). After the addition, the reaction mixture was stirred at 0°C for 0.5 hours; then the reaction mixture was diluted with 100 ml of water and extracted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a white solid compound A2 (2.2 g). MS: 611.7 (M+H) + .

[0185] Preparation Example Preparation of Compound A3

[0186]

[0187] Step 1: Preparation of compound A3-1

[0188] Tert-butyl acrylate (1060 g, 8260 mmol) and NaOH (5M, 19.8 ml) were slowly added to the DMSO (100 ml) reaction solution of compound Int.6 (100 g, 825.5 mmol) in sequence. After the addition, the reaction mixture was stirred at 20°C for 28 hours; then the reaction mixture was extracted with ethyl acetate, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a light yellow oil compound A3-1 (74 g, 17.7%). MS: 506.4 (M+H) + .

[0189] Step 2: Preparation of compound A3-2

[0190] Under stirring conditions at 0 degrees, CbzCl (22.7 g, 133.6 mmol) was slowly added to a dichloromethane (100 ml) reaction solution of compound A3-1 (15 g, 29.7 mmol) and TEA (9.0 g, 89.1 mmol). After the addition, the reaction mixture was stirred at 20°C for 12 hours; then the reaction mixture was diluted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a yellow oil compound A3-2 (18.8 g). 1 H NMR (400 MHz, DMSO-d6) δ7.36–7.30 (m, 6H), 4.97 (s, 2H), 3.54 (t, J = 6.4 Hz, 6H), 3.51 (s, 6H), 2.38 (t, J = 6.0 Hz, 6H), 1.39 (s, 27H). Step 3: Preparation of compound A3-3

[0191] The reaction mixture of compound A3-2 (18.8 g) and formic acid (200 ml) was stirred at 20°C for 12 hours; then the reaction mixture was concentrated to obtain a yellow oil compound A3-3 (16.5 g). 1H NMR (400 MHz, DMSO) δ12.12 (s, 3H), 7.40–7.32 (m, 5H), 5.17 (s, 1H), 4.99 (s, 2H), 3.57 (t, J=6.4 Hz, 6H), 3.50 (s, 6H), 2.42 (t, J=6.4 Hz, 6H).

[0192] Step 4: Preparation of Compound A3-4

[0193] Under nitrogen protection and stirring at room temperature, HOBt (13.2 g, 97.6 mmol) and EDCI (18.7 g, 97.6 mmol) were slowly added to a DMF (120 ml) solution of compound A3-3 (11.5 g, 24.4 mmol), N-tert-butyloxycarbonyl-1,3-propylenediamine (17.0 g, 97.6 mmol) and DIEA (25 g, 193.6 mmol). After the addition, the reaction mixture was stirred at room temperature for 16 hours. The reaction mixture was diluted with ethyl acetate and washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a yellow oil compound A3-4 (16 g). 1 HNMR (400MHz, CDCl3) δ7.37–7.28(m,5H),6.83(s,3H),5.55(s,1H),5.16(t,J=6.4Hz,3H),5.03(s,2H),3.68(t,J=6. 0Hz, 6H), 3.65 (s, 6H), 3.27–3.24 (m, 6H), 3.14–3.10 (m, 6H), 2.40 (t, J = 5.6Hz, 6H), 1.70–1.47 (m, 6H), 1.43 (s, 27H).

[0194] Step 5: Preparation of Compound A3-5

[0195] A reaction mixture of compound A3-4 (16 g) in dichloromethane (100 ml) and trifluoroacetic acid (20 ml) was stirred at 20°C for 12 hours; the reaction mixture was then concentrated to obtain a yellow oil compound A3-5 (20 g), which was used directly in the next step without purification.

[0196] Step 6: Preparation of Compound A3-6

[0197] Under stirring at 0 degrees and nitrogen protection, compound A1 (5 g) in dichloromethane (20 ml) was slowly added to the reaction mixture of compound A3-5 (5 g) and DIEA (2.8 g) in acetonitrile (20 ml). After the addition, the reaction mixture was stirred at room temperature for 12 hours; then the reaction mixture was diluted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product, which was purified to give a white solid compound A3-6 (3.85 g). 1 H NMR(400MHz,DMSO-d6)δ7.82-7.79(m,6H),7.71(t,J=5.6Hz,3H),7.37-7.27(m,5H),6.51(s,1H),5 .21(d,J=7.6Hz,3H),4.98-4.95(m,5H),4.48(d,J=7.6Hz,3H),4.04-4.00(m,9H),3.90-3.83(m,3H ),3.72-3.67(m,3H),3.56-3.48(m,12H),3.43-3.37(m,3H),3.05-2.99(m,12H),2.27(t,J=6.4Hz, 6H), 2.10 (s, 9H), 2.04 (t, J = 7.6Hz, 6H), 1.99 (s, 9H), 1.89 (s, 9H), 1.77 (s, 9H), 1.51–1.45 (m, 18H).

[0198] Step 7: Preparation of Compound A3

[0199] Compound A3-6 (4.5 g) was dissolved in methanol (25 ml), palladium carbon (1 g) was added to the reaction mixture under stirring, and then the reaction mixture was stirred under hydrogen pressure at room temperature for 1.5 hours; the reaction mixture was filtered and concentrated to obtain white solid compound A3 (4.2 g).

[0200] Preparation of Example Compound A4

[0201]

[0202] Step 1: Preparation of compound A4-1

[0203] Under nitrogen protection, a reaction mixture of compound A3 (410 mg, 0.23 mmol), compound A2 (209 mg, 0.34 mmol) and TEA (70 mg, 0.69 mmol) in DMF (5 ml) was stirred at 30 degrees for 12 hours; the reaction mixture was then diluted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain compound A4-1 (370 mg), which was directly used in the next step without purification. MS: 1110.8 (1 / 2M+H) + .

[0204] Step 2: Preparation of compound A4-2

[0205] Compound A4-1 (370 mg) was dissolved in methanol (5 ml), palladium carbon (300 mg) was added to the reaction mixture under stirring, and then the reaction mixture was stirred under hydrogen pressure at room temperature for 1.5 hours; the reaction mixture was filtered and concentrated to obtain compound A4-2 (280 mg), which was directly used in the next step without purification.

[0206] Step 3: Preparation of compound A4-3

[0207] Under nitrogen protection and stirring at 0 degrees, compound Int.3 (72 mg, 0.26 mmol) was slowly added to a solution of compound A4-2 (280 mg, 0.13 mmol) and DIEA (50 mg, 0.39 mmol) in acetonitrile (5 ml), and then the reaction mixture was stirred at 25°C for 12 hours. The reaction mixture was then diluted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain compound A4-3 (280 mg). MS: 1148 (1 / 2M+H) + .

[0208] Step 4: Preparation of Compound A4

[0209] The reaction mixture of compound A4-3 (280 mg, 0.12 mmol), compound Int.7 (100 mg, 0.24 mmol) and DIEA (46 mg, 0.36 mmol) in acetonitrile (5 ml) was stirred at 20 degrees for 12 hours; then the reaction mixture was diluted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was separated and purified to obtain compound A4 (80 mg). MS: 1115.1 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.90–7.78(m,6H),7.74(t,J=5.6Hz,3H),7.38–7.26(m,5H),7.26–7.13(m,5H),7.01(s,1H),6.88(ddd,J=9.0 ,6.6,1.9Hz,4H),5.21(d,J=3.4Hz,3H),4.96(dd,J=11.2,3.4Hz,4H),4.48(d,J=8.5Hz,3H),4.24–4.11(m,6H),4.03(d,J=3.8Hz,9H), 3.93(dd,J=12.6,5.7Hz,5H),3.86(dd,J=11.2,8.8Hz,3H),3.77-3.65(m,10H),3.58-3.47(dd,J=11.1,4.5Hz,14H),3.03(dq,J=12.4, 6.5,5.4Hz,14H),2.27(t,J=6.3Hz,6H),2.10(s,10H),2.08–2.01(m,12H),1.99(s,9H),1.89(s,9H),1.77(s,9H),1.56–1.38(m,26H).

[0210] Preparation of Example Compound A5 and Compound A6

[0211]

[0212] Step 1: Preparation of Compound A5

[0213] Synthesis of Compound A5 Referring to Compound A4, Compound A5 was prepared by using Compound Int.8 instead of Compound Int.5. MS: 1129.9 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.93–7.82(m,6H),7.77(s,3H),7.42(d,J=7.7Hz,1H),7.33(t,J=6.8Hz,5H),7.22(dd,J=10.4,7.7Hz,6H),7 .04(s,1H),6.90(dd,J=8.7,6.0Hz,6H),5.23(d,J=3.4Hz,3H),4.99(dd,J=11.2,3.4Hz,3H),4.50(d,J=8.5Hz,3H),4.42(s,1H),4.1 6(s,1H),4.04(s,8H),3.89(d,J=10.2Hz,3H),3.84–3.68(m,14H),3.55(d,J=7.2Hz,14H),3.48–3.39(m,7H),3.06(q,J=6.3Hz,15H) ,2.30(t,J=6.5Hz,10H),2.12(s,9H),2.06(t,J=7.3Hz,10H),2.02(s,9H),1.91(s,9H),1.79(s,9H),1.51(dt,J=19.5,9.9Hz,32H).

[0214] Step 2: Preparation of Compound A6

[0215] Compound A5 (59 mg, 0.023 mmol) and 7M ammonia in methanol (6 ml) were stirred at 20 degrees for 40 hours; the reaction mixture was then concentrated to obtain a crude product, which was separated and purified to obtain compound A6 (17.5 mg). MS: 941.1 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.88(t,J=5.7Hz,3H),7.78(t,J=5.6Hz,3H),7.66(d,J=9.0Hz,3H),7.37–7.29(m,4H),7.22(td,J=9.5,2.8 Hz,5H),7.06(s,1H),6.90(dt,J=8.9,3.4Hz,4H),5.01-4.90(m,3H),4.67–4.55(m,6H),4.51(d,J=4.3Hz,3H),4.42(d,J=4.4Hz,1H) ,4.24(d,J=8.4Hz,3H),4.20-4.14(m,1H),3.82(d,J=8.5Hz,2H),3.76(s,8H),3.74–3.70(m,5H),3.67(q,J=3.5Hz,4H),3.62–3.39( m,29H),3.32(dd,J=10.9,4.8Hz,6H),3.12-2.95(m,15H),2.39–2.21(m,10H),2.12-1.98(m,12H),1.82(s,9H),1.69–1.38(m,33H).

[0216] Preparation of Example Compound A7

[0217]

[0218] The synthesis of compound A7 was prepared by referring to compound A6 and using compound Int.9 instead of compound Int.8. MS: 954.6 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.89(s,3H),7.78(s,3H),7.66(d,J=9.1Hz,3H),7.38–7.30(m,4H),7.21(dd,J=8.9,2.6Hz,5H),7.06(s,1 H),6.95–6.86(m,4H),5.02(d,J=4.0Hz,1H),4.96–4.89(m,1H),4.66–4.55(m,6H),4.51(d,J=4.3Hz,3H),4.24(d,J=8.4Hz,2H),4. 17(s,1H),3.76(s,8H),3.72(d,J=9.6Hz,4H),3.66(d,J=4.1Hz,4H),3.54(td,J=11.8,11.3,5.4Hz,20H),3.48–3.41(m,8H),3.38 (s,7H),3.34–3.28(m,6H),3.05(q,J=6.2Hz,15H),2.30(t,J=6.3Hz,12H),2.07(t,J=7.3Hz,9H),1.82(s,9H),1.59–1.31(m,36H).

[0219] Preparation of Example Compound A8 and Compound A9

[0220]

[0221] Synthesis of Compound A8 Referring to Compound A4, Compound A8 was prepared by using Compound Int.10 instead of Compound Int.5. MS: 1122.6 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.93–7.82(m,5H),7.78(t,J=5.6Hz,2H),7.38–7.29(m,4H),7.23(td,J=9.6,9.1,2.7Hz,5H),7.06(s,1H),6 .96–6.88(m,4H),5.25(d,J=3.4Hz,2H),5.06–4.91(m,3H),4.52(d,J=8.5Hz,2H),4.43(s,0H),4.32(s,-1H),4.18(s,1H),4.06(s,1 0H),3.97–3.85(m,3H),3.83–3.69(m,12H),3.65–3.52(m,14H),3.44(d,J=9.3Hz,6H),3.35(s,4H),3.20(s,1H),3.08(d,J=7.6Hz,1 4H),2.39–2.21(m,9H),2.14(s,10H),2.08(t,J=7.2Hz,12H),2.03(s,11H),1.93(s,9H),1.81(s,9H),1.52(dt,J=20.3,7.0Hz,26H).

[0222] The synthesis of compound A9 was prepared by referring to compound A6 and using compound A8 instead of compound A5 to obtain compound A9. MS: 933.01 (1 / 2 [M-DMTr] + H) + . 1 H NMR(400MHz, DMSO-d6)δ7.88(s,2H),7.77(s,2H),7.65(d,J=9.0Hz,2H),7.37–7.29(m,4H),7.28–7.17(m, 5H),7.05(s,0H),6.90(d,J=7.5Hz,4H),4.66–4.55(m,6H),4.50(d,J=4.3Hz,3H),4.24(d,J=8.3Hz,3H),4 .08–3.95(m,1H),3.76(s,10H),3.74–3.63(m,11H),3.54(td,J=12.1,11.7,5.5Hz,23H),3.46(d,J=9.5Hz ,8H),3.31(s,6H),3.06(s,16H),2.37–2.21(m,9H),2.13–1.96(m,14H),1.82(s,9H),1.58–1.39(m,29H).

[0223] Preparation of Example Compound A10

[0224]

[0225] Step 1: Preparation of compound A10-1

[0226] Under stirring at room temperature, saturated sodium bicarbonate solution (30 ml) and CbzCl (5.4 g, 32 mmol) were added to a solution of (3-aminopropyl) tert-butyl carbamate (5 g, 29 mmol) in dichloromethane (10 ml). After the addition, the reaction mixture was stirred at 20°C for 1 hour; then the reaction mixture was extracted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain white solid benzyl tert-butyl propane-1,3-diyl dicarbamate (7 g). MS: 309.4 (M+H) + .

[0227] Under stirring conditions at 0 degrees, trifluoroacetic acid (10 ml) was slowly added to a solution of benzyl tert-butyl propane-1,3-diyl dicarbamate (3 g, 9.7 mmol) in dichloromethane (10 ml), and the reaction mixture was stirred at 25°C for 3 hours; the reaction mixture was then concentrated to obtain compound A10-1 (5 g) trifluoroacetate. MS: 209.2 (M+H) + .

[0228] Step 2: Preparation of compound A10-2

[0229] Under stirring conditions at 20 degrees, TEA (5.8 g, 58 mmol) and glutaric anhydride (1.25 g, 12.5 mmol) were slowly added to the dichloromethane (10 ml) reaction solution of the trifluoroacetate salt of compound A10-1 (5 g, 9.6 mmol). After the addition, the reaction mixture was stirred at 20°C for 16 hours; then the reaction mixture was diluted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a white solid compound A10-2 (2.5 g). MS: 323.2 (M+H) + .

[0230] Step 3: Preparation of compound A10-3

[0231] Under stirring conditions at 20 degrees, TEA (2.35 g, 23.3 mmol) and intermediate Int.3 (2.6 g, 9.3 mmol) were slowly added to the dichloromethane (10 ml) reaction solution of compound A10-2 (2.5 g, 7.76 mmol). After the addition, the reaction mixture was stirred at 20°C for 2 hours; then the reaction mixture was diluted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a white solid compound A10-3 (2.5 g). MS: 489.2 (M+H) + .

[0232] Step 4: Preparation of compound A10-4

[0233] Under nitrogen protection, a reaction mixture of compound A3 (1 g, 0.558 mmol), compound A10-3 (410 mg, 0.839 mmol) and TEA (0.17 g, 1.67 mmol) in DMF (5 ml) was stirred at 35 degrees for 16 hours; the reaction mixture was then diluted with ethyl acetate, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a white solid compound A10-4 (0.9 g). MS: 1096.7 (1 / 2M+H) + .

[0234] Step 5: Preparation of Compound A10-5

[0235] Compound A10-4 (900 mg, 0.43 mmol) was dissolved in methanol (20 ml), palladium carbon (600 mg) was added to the reaction mixture under stirring, and then the reaction mixture was stirred under hydrogen pressure at room temperature for 0.5 hours; the reaction mixture was filtered and concentrated to obtain compound A10-5 (750 mg), which was directly used in the next step without purification. MS: 982.8 (1 / 2M+H) + .

[0236] Step 6: Preparation of Compound A10-6

[0237] Under nitrogen protection at 20 degrees, TEA (93 mg, 0.93 mmol) and Cy5-NHS-ester (100 mg, 0.135 mmol) were slowly added to the DMF (2 ml) reaction mixture of compound A10-5 (344 mg, 0.176 mmol). After the addition, the reaction mixture was stirred at room temperature for 16 hours; then the reaction mixture was diluted with dichloromethane, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product compound A10-6 (120 mg). MS: 1295.0 (1 / 2M+H)+ .

[0238] Step 7: Preparation of Compound A10

[0239] The reaction mixture of compound A10-6 (120 mg, 0.046 mmol) and 7M ammonia in methanol (6 ml) was stirred at 20 degrees for 2 hours; the reaction mixture was then concentrated to obtain a crude product, which was separated and purified to obtain compound A10 (22.1 mg). MS: 1105.9 (1 / 2 [M] + H) + . 1 H NMR (400MHz, DMSO-d6) δ8.33(t,J=13.2Hz,2H),7.79(s,2H),7.67(d,J=5.1Hz,2H),7.33(dd,J=8.3,2.7Hz,2H),6.59(t,J=12.3 Hz,1H),6.29(t,J=14.2Hz,2H),4.23(d,J=8.4Hz,3H),4.09(s,2H),3.73–3.66(m,6H),3.65(d,J=3.0Hz,3H),3.60(s,3H),3.54( d,J=2.9Hz,6H),3.49(dd,J=11.1,6.4Hz,11H),3.42(dd,J=10.5,3.1Hz,3H),3.38–3.33(m,3H),3.31(t,J=6.3Hz,3H),3.06–2.9 6(m,17H),2.28(t,J=6.1Hz,6H),2.05(t,J=7.2Hz,12H),1.81(s,9H),1.69(s,12H),1.56–1.37(m,24H),1.28(d,J=26.7Hz,4H).

[0240] Preparation of Example Compound A11

[0241]

[0242] Synthesis of Compound A11 Referring to Compound A6, Compound A11 was prepared by using Compound Int.11 instead of Compound Int.8. MS: 940.0 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.85(d,J=6.0Hz,2H),7.75(t,J=5.5Hz,2H),7.63(d,J=9.0Hz,2H),7.37-7.26(m,4 H),7.26–7.12(m,5H),7.03(s,1H),6.94–6.78(m,4H),5.02-4.86(m,1H),4.65–4.52(m,5H),4.48(d,J=4.3H z,2H),4.21(d,J=8.3Hz,2H),4.14(s,1H),3.83–3.60(m,19H),3.60-3.46(m,22H),3.45–3.38(m,5H),3.29 -3.13(m,9H),3.10-2.90(m,14H),2.35-2.19(m,10H),2.13-1.92(m,12H),1.79(s,9H),1.61-1.33(m,32H).

[0243] Preparation of Example Compound A12

[0244]

[0245] The synthesis of compound A12 was prepared by referring to compound A6 and using compound Int.12 instead of compound Int.8. MS: 933.2 (1 / 2 [M-DMTr] + H) + . 1 H NMR (400MHz, DMSO-d6) δ7.89(t,J=5.6Hz,3H),7.78(t,J=5.6Hz,3H),7.66(d,J=9.1Hz,3H),7.40–7.29(m,4H),7.2 2(dt,J=9.1,5.1Hz,5H),7.06(s,1H),6.94–6.86(m,4H),5.02(s,1H),4.67–4.57(m,6H),4.51(d,J=4.3Hz,3H),4.4 2(s,1H),4.24(d,J=8.4Hz,3H),4.15(d,J=10.6Hz,1H),4.06–3.92(m,2H),3.79–3.64(m,18H),3.61–3.41(m,26H) ,3.31(t,J=6.2Hz,2H),3.05(q,J=6.3Hz,14H),2.36–2.18(m,11H),2.12(s,15H),1.82(s,9H),1.60–1.37(m,28H).

[0246] Preparation of Example Compound A13

[0247]

[0248] The synthesis of compound A13 was prepared by referring to compound A6 and using compound Int.13 instead of compound Int.8. MS: 926.2 (1 / 2 [M-DMTr] + H) + . 1 H NMR (400MHz, DMSO-d6) δ7.85 (s, 2H), 7.63 (d, J = 8.8Hz, 2H), 7.31 (s, 4H), 6.88 (s, 5H), 5.03-4.87 (m, 1H), 4 .58(dd,J=14.0,6.1Hz,6H),4.48(d,J=4.4Hz,3H),4.21(d,J=8.5Hz,2H),4.16-4.06(m,1H),3.96-3.80(m ,2H),3.73(s,10H),3.69(d,J=9.4Hz,4H),3.64(s,4H),3.57–3.45(m,20H),3.45-3.37(m,4H),3.30(d,J= 9.5Hz,10H),3.10-2.95(s,15H),2.46-2.36(m,1H),2.12-1.94(m,13H),1.79(s,9H),1.59–1.32(m,27H).

[0249] Preparation of Example Compound A14

[0250]

[0251] Synthesis of Compound A14 Referring to Compound A6, Compound A14 was prepared by using Compound Int.14 instead of Compound Int.8. MS: 941.2 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ8.08-8.00(m,1H),7.86(s,2H),7.75(t,J=5.7Hz,3H),7.63(d,J=9.1Hz,2H),7.34-7.26(m,4H),7.24 –7.14(m,5H),7.03(s,1H),6.92–6.83(m,4H),5.03-4.85(m,1H),4.65–4.53(m,6H),4.48(d,J=4.3Hz,3H),4.43-4.30(m,1H), 4.21(d,J=8.4Hz,4H),4.17-3.75(m,4H),3.73(s,8H),3.69(d,J=9.7Hz,5H),3.65(d,J=7.3Hz,4H),3.61–3.38(m,27H),3.32– 3.25(m,7H),3.21-3.14(m,1H),3.11–2.95(m,15H),2.38–2.14(m,9H),2.13-1.93(m,16H),1.79(s,10H),1.61–1.29(m,28H).

[0252] Preparation of Example Compound A15

[0253]

[0254] The synthesis of compound A15 was prepared by referring to compound A6 and using compound Int.15 instead of compound Int.8. MS: 940.1 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.85(t,J=5.6Hz,3H),7.75(t,J=5.6Hz,3H),7.63(d,J=9.1Hz,2H),7.37–7.26(m,4H),7.19(td,J=9.6, 9.1,3.0Hz,5H),7.04(s,1H),6.92–6.83(m,4H),5.05-4.85(m,1H),4.58(s,6H),4.49(s,3H),4.43-4.35(m,1H),4.21(d,J=8.4H z,3H),4.17-4.07(m,1H),3.73(s,9H),3.71-3.61(m,9H),3.61–3.46(m,23H),3.42-3.38(m,6H),3.31-3.23(m,6H),3.20-3.13 (m,1H),3.10-2.96(m,14H),2.28(t,J=6.4Hz,15H),2.04(t,J=7.3Hz,10H),1.95-1.84(m,3H),1.79(s,9H),1.60–1.36(m,28H).

[0255] Preparation of Example Compound A16

[0256]

[0257] Synthesis of Compound A16 Referring to Compound A6, Compound A16 was prepared by using Compound Int.16 instead of Compound Int.8. MS: 971.3 (1 / 2 [M-DMTr] + H) + . 1H NMR(400MHz, DMSO-d6)δ8.13-8.06(m,1H),7.75(t,J=5.4Hz,2H),7.63(d,J=9.0Hz,2H),7.35–7.13(m,14H),7.09-6.97(m, 1H),6.91–6.84(m,6H),5.04-4.87(m,1H),4.58(dd,J=14.3,6.0Hz,6H),4.48(d,J=4.3Hz,2H),4.40(s,1H),4.21(d,J=8.4 Hz,3H),4.15(s,1H),4.06-3.98(m,1H),3.76–3.71(m,11H),3.71–3.60(m,9H),3.59–3.34(m,29H),3.31-3.26(m,6H),3.2 1-3.16(m,1H),3.09-2.95(m,14H),2.38-2.22(m,2H),2.10(s,1H),2.04(t,J=7.3Hz,7H),1.79(s,9H),1.68–1.35(m,29H).

[0258] Preparation of Example Compound A17

[0259]

[0260] The synthesis of compound A17 was prepared by referring to compound A6 and using compound Int.17 instead of compound Int.8. MS: 940.1 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.86(t,J=5.7Hz,3H),7.75(t,J=5.6Hz,2H),7.63(d,J=9.1Hz,2H),7.58-7.51(m,1H),7.35–7.25(m,4H),7. 19(td,J=9.4,3.2Hz,5H),7.03(s,1H),6.92–6.82(m,4H),5.02-4.84(m,1H),4.66–4.53(m,6H),4.48(d,J=4.3Hz,3H),4.39(s,1H),4 .21(d,J=8.4Hz,3H),4.14(d,J=10.8Hz,2H),3.73(s,8H),3.69(d,J=9.6Hz,6H),3.65(d,J=7.2Hz,4H),3.59-3.46(m,21H),3.44-3.3 4(m,7H),3.32-3.25(m,5H),3.20-2.90(m,16H),2.26(q,J=11.5,8.9Hz,10H),2.16–1.96(m,15H),1.79(s,10H),1.60–1.27(m,30H).

[0261] Preparation of Example Compound A18 and Compound A19

[0262]

[0263] Synthesis of Compound A18 Referring to Compound A4, Compound A18 was prepared by using Compound Int.18 instead of Compound Int.5. MS: 1122.7 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.92–7.82(m,6H),7.77(t,J=5.7Hz,3H),7.36–7.31(m,4H),7.22(dd,J=10.3,7.4Hz,5H),7.04(s,1H),6.90 (dd,J=8.5,5.7Hz,4H),5.24(d,J=3.3Hz,3H),4.99(dd,J=11.3,3.5Hz,4H),4.51(d,J=8.4Hz,3H),4.17(s,1H),4.05(s,9H),3.90(q, J=9.3Hz,3H),3.76(s,12H),3.67–3.52(m,18H),3.46–3.41(m,3H),3.29(s,3H),3.19(d,J=10.5Hz,3H),3.06(q,J=6.2Hz,12H),2.30 (t,J=6.4Hz,6H),2.24(s,6H),2.13(s,9H),2.10–2.05(m,8H),2.02(s,9H),1.91(s,9H),1.80(s,9H),1.52(dd,J=15.6,9.2Hz,28H).

[0264] The synthesis of compound A19 was prepared by referring to compound A6 and using compound A18 instead of compound A5 to obtain compound A19. MS: 1106.6 (1 / 2 [M] + Na) + . 1 H NMR (400MHz, DMSO-d6) δ7.89(t,J=5.6Hz,3H),7.78(t,J=5.7Hz,3H),7.66(d,J=9.1Hz,3H),7.34(t,J=6.8Hz,4H),7.31–7.27(m, 1H),7.22(dt,J=9.1,5.0Hz,5H),6.91(td,J=7.4,3.7Hz,6H),5.03(d,J=4.0Hz,1H),4.66–4.57(m,6H),4.51(d,J=4.3Hz,3H),4. 25(d,J=8.4Hz,3H),3.77(s,9H),3.74–3.66(m,9H),3.55(dq,J=11.2,5.5Hz,21H),3.49–3.41(m,6H),3.32(t,J=6.1Hz,6H),3.1 9(d,J=10.4Hz,3H),3.06(t,J=6.2Hz,12H),2.35–2.20(m,12H),2.14–2.00(m,10H),1.83(s,9H),1.50(dt,J=29.8,7.5Hz,28H).

[0265] Preparation of Example Compound A20

[0266]

[0267] The synthesis of compound A20 was prepared by referring to compound A6 and using compound A4 instead of compound A5 to obtain compound A20. MS: 926.0 (1 / 2 [M-DMTr] + H) + . 1 H NMR(400MHz,DMSO-d6)δ7.28(dd,J=6.5,4.3Hz,4H),7.24–7.12(m,5H),6.90–6.82(m,4H) ),4.43-4.25,(m,1H),4.24–4.08(m,8H),3.97–3.86(m,4H),3.72–3.61(m,16H),3.55–3. 45(m,18H),3.40(dd,J=10.7,3.0Hz,4H),3.31(q,J=9.4,6.4Hz,7H),3.01(q,J=6.4Hz,13 H),2.30–2.17(m,8H),2.03(t,J=7.2Hz,14H),1.79(d,J=1.4Hz,9H),1.56–1.34(m,27H).

[0268] Preparation of Example Compound A21

[0269]

[0270] The synthesis of compound A21 was prepared by referring to compound A6 and using compound Int.21 instead of compound Int.8. MS: 926.2 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ8.37–8.29(m,3H),7.84(t,J=5.6Hz,3H),7.74(t,J=5.7Hz,3H),7.62(d,J=9.0Hz,3H),7.33–7.27(m,4H),7.22– 7.15(m,5H),7.02(s,1H),6.90–6.84(m,4H),5.34-5.30(m,1H),5.00-4.86(m,1H),4.61–4.53(m,6H),4.47(d,J=4.3Hz,3H),4.21(d,J=8 .4Hz,3H),3.73(s,6H),3.69(d,J=9.5Hz,5H),3.64(q,J=3.8,3.3Hz,4H),3.58-3.46(m,20H),3.45–3.35(m,6H),3.45-3.35(m,6H),3.29 (t,J=6.0Hz,5H),3.09-2.95(m,15H),2.27(t,J=6.4Hz,6H),2.21(d,J=7.2Hz,2H),2.08-1.96(m,15H),1.80(s,9H),1.55–1.38(m,28H).

[0271] Preparation of Example Compound A22 (L96-deAc)

[0272]

[0273] The synthesis of compound A22 was prepared by referring to compound A6 and using compound A29-1 instead of compound A5-1 to obtain compound A22. MS: 864.7 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.88(t,J=5.7Hz,3H),7.78(t,J=5.7Hz,3H),7.66( d,J=9.0Hz,3H),7.36–7.29(m,4H),7.21(td,J=8.8,3.0Hz,5H),7.03(s,1H) ,6.91–6.87(m,4H),5.01(d,J=4.0Hz,1H),4.65–4.56(m,6H),4.51(d,J=4. 3Hz,3H),4.23(d,J=8.4Hz,3H),4.15(d,J=11.9Hz,1H),3.75(s,6H),3.73–3 .63(m,9H),3.54(td,J=11.7,11.3,5.6Hz,21H),3.44(ddd,J=9.9,6.2,2.9 Hz,3H),3.31(q,J=6.2,5.1Hz,4H),3.17(s,1H),3.05(p,J=6.4Hz,15H),2.3 0(t,J=6.4Hz,6H),2.22(t,J=7.4Hz,2H),2.05(q,J=11.3,9.2Hz,10H),1.82 (s, 9H), 1.52 (dd, J = 12.7, 6.6 Hz, 16H), 1.43 (t, J = 7.2 Hz, 6H), 1.23 (s, 12H).

[0274] Preparation of Example Compound A23

[0275]

[0276] The synthesis of compound A23 was prepared by referring to compound A6 and using compound Int.23 instead of compound Int.8. MS: 926.2 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.88(t,J=5.6Hz,3H),7.78(t,J=5.6Hz,3H),7.66(d,J=9.0Hz,3H),7.37– 7.30(m,4H),7.22(td,J=9.4,3.1Hz,5H),7.06(s,1H),6.96–6.86(m,4H),6.06(s,3H),4.52(s,1H ),4.42(s,1H),4.33–4.13(m,6H),3.81–3.63(m,20H),3.63–3.45(m,33H),3.43–3.30(m,18H),3. 12–2.98(m,15H),2.30(t,J=6.5Hz,11H),2.07(t,J=7.2Hz,11H),1.82(s,9H),1.58–1.38(m,28H).

[0277] Preparation of Example Compound A24

[0278]

[0279] The synthesis of compound A24 was prepared by referring to compound A6 and using compound Int.24 instead of compound Int.8. MS: 960.1 (1 / 2 [M-DMTr] + H) + . 1 H NMR (400MHz, DMSO-d6) δ7.87(t,J=5.6Hz,3H),7.76(t,J=5.6Hz,2H),7.63(d,J=9.1Hz,2H),7.37–7.26(m,6H),7.25–7.14(m,5H),7.02(s,1H), 6.88(ddd,J=8.8,6.3,1.7Hz,4H),5.06-4.87(m,1H),4.64–4.53(m,6H) ,4.49(d,J=4.3Hz,3H),4.43-4.36(m,1H),4.21(d,J=8.4Hz,2H),4.16-

[0280] 4.08(m,1H),3.73(d,J=1.0Hz,8H),3.69(d,J=9.8Hz,4H),3.67-3.63(m, 4H),3.61–3.38(m,22H),3.32-3.23(m,5H),3.18-3.13(m,1H),3.06-2.94 (m,14H),2.27(t,J=6.4Hz,6H),2.22(d,J=8.0Hz,1H),2.12(s,2H),2.09 –1.93(m,16H),1.84(d,J=16.3Hz,8H),1.79(s,10H),1.56–1.31(m,30H).

[0281] Preparation of Example Compound A25

[0282]

[0283] The synthesis of compound A25 was prepared by referring to compound A6 and using compound Int.25 instead of compound Int.8. MS: 933.4 (1 / 2 [M-DMTr] + H) + . 1 H NMR (400MHz, DMSO-d6) δ8.10–8.00(m,1H),7.89(t,J=5.6Hz,3H),7.78(t,J=5.7Hz,3H),7.66(d,J=9.1Hz,3H),7.40–7.31(m,4H),7 .30–7.14(m,5H),7.06(s,1H),6.96–6.86(m,4H),5.06–4.90(m,1H),4.66–4.57(m,6H),4.52(d,J=4.3Hz,3H),4.43(d,J=4.8Hz,1H) ,4.25(d,J=8.4Hz,3H),4.23–3.96(m,3H),3.87(s,1H),3.80–3.64(m,17H),3.65–3.42(m,23H),3.33(dt,J=12.4,6.2Hz,5H),3.20 (s,1H),3.17–2.95(m,13H),2.43(d,J=9.3Hz,2H),2.36–2.21(m,8H),2.06(q,J=8.8,7.9Hz,16H),1.83(s,9H),1.59–1.35(m,27H).

[0284] Preparation of Example Compound A26

[0285]

[0286] Synthesis of Compound A26 Referring to Compound A6, Compound A26 was prepared by using Compound Int.26 instead of Compound Int.8. MS: 933.4 (1 / 2[M-DMTr]+H) + . 1 H NMR (400MHz, DMSO-d6) δ8.07(dd,J=20.3,7.7Hz,1H),7.88(t,J=5.7Hz,3H),7.78(t,J=5.6Hz,3H),7.66(d,J=9.0Hz,3H), 7.39–7.29(m,4H),7.27–7.16(m,5H),7.05(s,1H),6.96–6.85(m,4H),5.05–4.90(m,1H),4.67–4.57(m,6H),4.51(d,J=4. 3Hz,3H),4.24(d,J=8.4Hz,4H),3.99(d,J=18.3Hz,1H),3.80–3.65(m,16H),3.62–3.42(m,24H),3.36–3.16(m,8H),3.13– 2.98(m,14H),2.37–2.20(m,9H),2.07(t,J=7.2Hz,12H),1.89(s,1H),1.82(s,9H),1.61–1.34(m,27H),0.78–0.50(m,3H).

[0287] Preparation of Example Compound A27

[0288]

[0289] Synthesis of Compound A27 Referring to Compound A29, Compound A27 was prepared by using Compound Int.18 instead of Compound Int.5. MS: 1094.9 (1 / 2 [M] + Na) + . 1H NMR (400MHz, DMSO-d6) δ7.63(d,J=9.0Hz,3H),7.33(t,J=7.3Hz,4H),7.21(dd,J=8.8,2.5Hz,5H),7.11(s,1H),6.92–6.87(m,4 H),5.00(d,J=4.0Hz,1H),4.63–4.55(m,6H),4.49(d,J=4.3Hz,3H),4.42(s,1H),4.25(d,J=8.4Hz,3H),4.16(s,1H),3.78–3.65 (m,22H),3.56(td,J=12.0,6.3Hz,25H),3.51–3.37(m,12H),3.31(t,J=6.1Hz,6H),3.20(d,J=11.8Hz,6H),3.00(d,J=10.1Hz,3 H), 2.90 (s, 3H), 2.75 (d, J = 39.9Hz, 2H), 2.47 (s, 6H), 2.27–2.17 (m, 8H), 2.08 (s, 2H), 1.81 (s, 9H), 1.50 (s, 16H), 1.24 (s, 12H).

[0290] Preparation of Example Compound A28

[0291]

[0292] Synthesis of Compound A28 Referring to Compound A29, Compound A28 was prepared by using Compound Int.21 instead of Compound Int.5. MS: 1051.7 (1 / 2 [M] + H) + . 1H NMR (400MHz, DMSO-d6) δ8.44(s,3H),8.35(s,3H),7.64(d,J=9.0Hz,3H),7.34(dt,J=8.6,4.4Hz,4H),7.22(td,J=9.2,8.8,3.7Hz,5H ),7.01(s,1H),6.92–6.87(m,4H),5.01(d,J=3.8Hz,1H),4.63–4.56(m,6H),4.50(d,J=4.3Hz,3H),4.42(s,1H),4.23(d,J=8.4Hz,3H) ,4.15(d,J=15.1Hz,1H),3.73(d,J=24.0Hz,12H),3.67(t,J=3.9Hz,3H),3.54(q,J=5.8Hz,21H),3.46–3.42(m,3H),3.33–3.16(m,5H ), 3.01 (d, J = 7.5Hz, 1H), 2.27 (t, J = 6.6Hz, 6H), 2.17 (s, 21H), 2.03 (t, J = 6.9Hz, 10H), 1.82 (s, 9H), 1.52–1.40 (m, 16H), 1.24 (s, 12H).

[0293] Preparation of Example Compound A29

[0294]

[0295]

[0296] Synthesis of Compound A29-1 Referring to Compound A1, Compound A29-1 was prepared by using Compound Int.29 instead of Compound A1-3. MS: 509.1 (M+Na) + .

[0297] The synthesis of compound A29-2 was prepared by referring to compound A3 and using compound Int.5 instead of N-tert-butyloxycarbonyl-1,3-propylenediamine to obtain compound A29-2. MS: 934.1 (M / 2+H) + .

[0298] Synthesis of Compound A29 Referring to Compound A20, Compound A29 was prepared by using Compound A29-2 instead of Compound A3 and Compound A29-1 instead of Compound A2. MS: 1051.7 (1 / 2 [M] + H) +. 1H NMR (400MHz, DMSO-d6) δ7.63(d,J=9.1Hz,3H),7.33(t,J=7.3Hz,4H),7.25–7.19(m,5H),7.08(s,1H),6.92–6.87(m,4H),5.00(d,J =4.1Hz,1H),4.62–4.55(m,6H),4.49(d,J=4.3Hz,3H),4.42(s,1H),4.30–4.22(m,15H),4.15(d,J=13.6Hz,1H),4.02–3.96(m,12H ),3.76(s,6H),3.72(d,J=9.0Hz,5H),3.67(s,3H),3.58–3.53(m,15H),3.53–3.48(m,3H),3.48–3.40(m,6H),3.31(t,J=6.3Hz,6H ), 3.19 (dd, J = 8.7, 5.1Hz, 1H), 3.01 (d, J = 7.7Hz, 2H), 2.25 (s, 8H), 2.02 (d, J = 6.4Hz, 9H), 1.82 (s, 9H), 1.47 (s, 16H), 1.25 (s, 12H).

[0299] Preparation of Example Compound A30

[0300]

[0301] Synthesis of Compound A30 Referring to Compound A6, Compound A30 was prepared by using Compound Int.30 instead of Compound Int.8. MS: 940.7 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.89(t,J=5.6Hz,3H),7.78(t,J=5.6Hz,3H),7.66(d,J=9.0Hz,3H),7.37–7.32(m,4H),7.22(dt,J=9.0 ,5.6Hz,5H),7.06(s,1H),6.94–6.88(m,4H),5.03(d,J=4.1Hz,1H),4.65–4.57(m,6H),4.51(d,J=4.3Hz,3H),4.43(s,1H),4.2 5(d,J=8.4Hz,3H),4.18(s,1H),3.79–3.65(m,18H),3.61–3.42(m,27H),3.32(t,J=6.2Hz,4H),3.07(p,J=6.1Hz,15H),2.35–2 .23(m,10H),2.14–2.02(m,12H),1.91(d,J=12.6Hz,1H),1.83(s,9H),1.56–1.44(m,26H),1.27(s,2H),0.59(d,J=96.6Hz,2H).

[0302] Preparation of Example Compound A31

[0303]

[0304] Synthesis of Compound A31 Referring to Compound A20, Compound A31 was prepared by using Compound Int.31 instead of Compound Int.4. MS: 912.5 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.84(t,J=5.6Hz,3H),7.74(t,J=5.7Hz,3H),7.62(d,J=9.0Hz,3H),7.31(s,4H),7.19(dt,J=9.2,4.9Hz,5H),7. 11(s,1H),6.91–6.84(m,4H),5.34-5.30(m,1H),5.00-4.85(m,1H),4.63–4.51(m,6H),4.47(d,J=4.3Hz,3H),4.21(t,J=7.5Hz,6H),3.94 (d,J=11.2Hz,4H),3.73(s,6H),3.69(d,J=9.5Hz,4H),3.64(d,J=4.1Hz,4H),3.56-3.46(m,18H),3.44-3.35(m,5H),3.29(t,J=6.1Hz,4H ),3.20-3.13(m,1H),3.08-2.98(m,13H),2.27(t,J=6.5Hz,8H),2.24-2.13(m,4H),2.08-1.92(m,11H),1.80(s,9H),1.55–1.37(m,24H).

[0305] Preparation of Example Compound A32

[0306]

[0307] The synthesis of compound A32 was prepared by referring to compound A6 and using compound Int.32 instead of compound Int.8. MS: 1106.7 (1 / 2 [M] + Na) + . 1H NMR (400MHz, DMSO-d6) δ7.90(t,J=5.6Hz,3H),7.80(t,J=5.5Hz,3H),7.68(d,J=9.1Hz,3H),7.36–7.30(m,4H),7.21(dt,J=9.1,5.1Hz ,5H),7.07(s,1H),6.93–6.87(m,4H),5.07(d,J=3.9Hz,1H),4.67(t,J=5.6Hz,3H),4.61(d,J=6.3Hz,3H),4.52(d,J=4.3Hz,3H),4.23( d,J=8.4Hz,3H),4.16(s,1H),3.99(s,1H),3.71(d,J=35.9Hz,21H),3.55(d,J=8.7Hz,27H),3.42–3.29(m,15H),3.10–3.02(m,12H),2. 30(t,J=6.4Hz,6H),2.23(d,J=8.0Hz,3H),2.07(t,J=7.3Hz,10H),1.82(s,9H),1.49(dq,J=25.1,7.2Hz,25H),0.59(t,J=41.8Hz,1H).

[0308] Preparation of Example Compound A33

[0309]

[0310] The synthesis of compound A33 was prepared by referring to compound A6 and using compound Int.33 instead of compound Int.8. MS: 926.2 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.99(s,1H),7.88(t,J=5.6Hz,3H),7.77(t,J=5.6Hz,3H),7.65(d,J=9.2Hz,3H),7.39–7.30(m,4H),7.2 2(dt,J=9.1,5.1Hz,5H),7.04(s,1H),6.95–6.85(m,4H),5.01(d,J=4.8Hz,1H),4.65–4.56(m,6H),4.50(d,J=4.3Hz,4H),4.24( d,J=8.4Hz,4H),3.79–3.65(m,15H),3.55(tt,J=11.8,5.6Hz,21H),3.49–3.37(m,9H),3.32(t,J=6.2Hz,4H),3.16(d,J=5.4Hz, 1H),3.05(q,J=6.1Hz,14H),2.34–2.22(m,8H),2.07(t,J=7.3Hz,11H),1.87(d,J=9.9Hz,1H),1.83(s,9H),1.60–1.35(m,28H).

[0311] Preparation of Example Compound A34

[0312]

[0313] The synthesis of compound A34 was prepared by referring to compound A6 and using compound Int.34 instead of compound Int.8. MS: 940.4 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.88(t,J=5.6Hz,3H),7.77(t,J=5.5Hz,3H),7.65(d,J=9.1Hz,3H),7.33(q,J=8.4,7.7Hz,4H),7.2 8–7.17(m,5H),7.06(s,1H),6.90(dd,J=8.7,2.5Hz,4H),5.06–4.87(m,2H),4.66–4.57(m,7H),4.51(d,J=4.3Hz,3H),4.38( d,J=40.5Hz,2H),4.27–4.13(m,4H),3.80–3.64(m,18H),3.55(td,J=11.5,5.4Hz,20H),3.47–3.38(m,8H),3.34–3.17(m,6 H),3.12–2.96(m,14H),2.30(q,J=11.1,8.7Hz,10H),2.07(t,J=7.2Hz,11H),1.89(s,1H),1.83(s,9H),1.60–1.37(m,27H).

[0314] Preparation of Example Compound A35

[0315]

[0316]

[0317] Step 1: Preparation of compound A35-1

[0318] Potassium carbonate (7.72 g, 55.9 mmol), 6-bromohexanoic acid (5.45 g, 27.9 mmol) and benzyl bromide (9.56 g, 55.9 mmol) in DMF (60 ml) were stirred at 20°C for 16 hours. The reaction mixture was then poured into ice water and diluted with petroleum ether for extraction. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain compound A35-1 (7.22 g, 91%). MS: 285.2 & 287.2 (M + H) + .

[0319] Step 2: Preparation of compound A35-2

[0320] Potassium carbonate (0.977 g, 7.07 mmol), intermediate Int.35 (0.8 g, 3.53 mmol) and compound A35-1 (1.21 g, 4.24 mmol) in DMF (30 ml) were stirred at 50°C for 1 hour. The reaction mixture was then poured into ice water and diluted with dichloromethane for extraction. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain compound A35-2 (2.5 g, 99%). MS: 431.5 (M+H) + .

[0321] Step 3: Preparation of compound A35-3

[0322] A solution of trifluoroacetic acid (0.429 g, 3.76 mmol) and compound A35-2 (2.7 g, 3.76 mmol) in dichloromethane (30 ml) was stirred at 20°C for 0.5 hours. The reaction mixture was then concentrated to give a crude product, compound A35-3 (5.8 g, 97%). MS: 331.5 (M+H) + .

[0323] Step 4: Preparation of compound A35-4

[0324] Potassium carbonate (5.01 g, 36.2 mmol), compound A35-3 (2.02 g, 3.62 mmol) and tert-butyl 6-bromohexanoate (2.6 g, 3.63 mmol) in DMF (30 ml) were stirred at 80°C for 2 hours. The reaction mixture was then poured into ice water and diluted with dichloromethane for extraction. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain compound A35-4 (1.2 g, 33%). MS: 501.6 (M+H) + .

[0325] Step 5: Preparation of compound A35-5

[0326] A solution of trifluoroacetic acid (0.137 g, 1.198 mmol) and compound A35-4 (1.2 g, 1.198 mmol) in dichloromethane (30 ml) was stirred at 20°C for 2 hours. The reaction mixture was then concentrated to obtain a crude product, which was purified to obtain compound A35-5 (470 mg, 88%). MS: 445.4 (M+H) + .

[0327] Step 6: Preparation of Compound A35-6

[0328] Under nitrogen protection at 0°C, a solution of intermediate Int.3 (850 mg, 3.04 mmol), DIEA (654 mg, 5.06 mmol) and compound A35-5 (45 mg, 1.012 mmol) in dichloromethane (20 ml) was stirred at 25°C for 16 hours. The reaction mixture was concentrated to obtain a crude product, which was purified to obtain compound A35-6 (540 mg, 87%). MS: 611.5 (M+H) + .

[0329] Step 7: Preparation of Compound A35

[0330] The synthesis of compound A35 was prepared by referring to compound A6 and using compound A35-6 instead of compound A5-1. MS: 926.6 (1 / 2 [M-DMTr] + H) + . 1 H NMR (400MHz, DMSO-d6) δ7.88(t,J=5.6Hz,3H),7.78(t,J=5.7Hz,3H),7.66(d,J=9.0Hz,3H),7.33(t,J=6.8Hz,4H),7.22(td,J =8.7,2.7Hz,5H),7.03(s,1H),6.96–6.84(m,4H),5.40-5.27(m,4H),5.06–4.83(m,8H),4.67–4.54(m,6H),4.51(d,J=4.3Hz,3 H),4.41(s,3H),4.32(s,2H),4.24(d,J=8.4Hz,3H),4.16(s,3H),3.82–3.63(m,13H),3.63–3.42(m,24H),3.32–3.14(m,4H), 3.12-2.92(m,13H),2.74-2.66(m,1H),2.43-2.15(m,11H),2.13-1.97(m,10H),1.82(s,9H),1.63(s,1H),1.58-1.29(m,38H).

[0331] Preparation of Example Compound A36

[0332]

[0333] Synthesis of Compound A36 Referring to Compound A6, Compound A36 was prepared by using Compound Int.36 instead of Compound Int.8. MS: 954.3 (1 / 2 [M-DMTr] + H) + . 1H NMR(400MHz, Methanol-d4)δ7.40–7.33(m,2H),7.32–7.16(m,8H),6.86(t,J=9.0Hz,4H),4.35 (d,J=8.4Hz,3H),3.96–3.86(m,8H),3.83(d,J=3.3Hz,3H),3.79–3.70(m,16H),3.69-3.62(m, 13H),3.58(dd,J=10.7,3.3Hz,4H),3.55–3.46(m,9H),3.25–3.17(m,13H),2.46-2.36(m,10H) ,2.20(t,J=7.3Hz,10H),1.98(s,9H),1.73–1.53(m,30H),1.44-1.27(dt,J=28.2,14.6Hz,7H).

[0334] Preparation of Example Compound A37

[0335]

[0336] Synthesis of Compound A37 Referring to Compound A29, Compound A37 was prepared by using Compound Int.16 instead of Compound Int.5. 1 H NMR (400MHz, Methanol-d4) δ8.14(d,J=7.9Hz,3H),7.35(d,J=7.5Hz,2H),7.28-7.13(m,13H),7.04(t,J=7.4Hz,3H),6.86–6.79( m,4H),4.55(d,J=12.6Hz,4H),4.37(d,J=8.4Hz,3H),4.12(s,6H),4.01(d,J=14.5Hz,3H),3.92(t,J=9.3Hz,6H),3.82(d,J=2.9Hz ,3H),3.78–3.67(m,26H),3.60–3.45(m,12H),3.17-3.05(m,1H),3.07(d,J=7.0Hz,1H),2.88-2.74(m,9H),2.46(s,6H),2.30(d, J=7.0Hz,1H),2.24–2.16(m,2H),2.05(d,J=10.7Hz,4H),1.97(s,9H),1.90-1.82(m,3H),1.80–1.57(m,24H),1.46–1.24(m,12H).

[0337] Preparation of Example Compound A38

[0338]

[0339] The synthesis of compound A38 was prepared by referring to compound A6 and using compound Int.38 instead of compound Int.8. MS: 947.7 (1 / 2 [M-DMTr] + H) + . 1H NMR (400MHz, DMSO-d6) δ7.85(t,J=5.6Hz,3H),7.74(t,J=5.6Hz,3H),7.62(d,J=9.0Hz,3H),7.30(q,J=8.2,7.7Hz,4H),7.20(t d,J=9.7,9.1,3.0Hz,5H),7.02(s,1H),6.92–6.84(m,4H),4.94(dd,J=35.3,4.2Hz,1H),4.64–4.52(m,6H),4.51–4.28(m,4H), 4.25–4.09(m,4H),3.89(d,J=19.2Hz,2H),3.78–3.60(m,19H),3.58-3.46(m,19H),3.45–3.36(m,8H),3.29(t,J=6.1Hz,6H),3 .03(p,J=6.1Hz,14H),2.34–2.18(m,10H),2.12–1.95(m,12H),1.80(s,9H),1.58–1.35(m,27H),0.83(dt,J=21.2,7.8Hz,3H).

[0340] Example: Amingalactose compound III-LA20-CPG attached to a solid support

[0341]

[0342] Step 1: Preparation of compound III-LA20-1

[0343] Under stirring at 20 degrees, succinic anhydride (32 mg, 0.32 mmol) was slowly added to the acetonitrile (10 ml) reaction solution of compound A4 (400 mg, 0.16 mmol) and DMAP (117 mg, 0.96 mmol). After the addition, the reaction mixture was stirred at 20°C for 12 hours; then the reaction mixture was diluted with ethyl acetate, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified to obtain a yellow solid compound III-LA20-1 (150 mg). MS: 1165.1 (1 / 2 [M-DMTr] + H) + .

[0344] Step 2: Preparation of aminogalactose compound III-LA20-CPG attached to a solid support

[0345] Compound III-LA20-1 (50 mg, 0.021 mmol) was added to acetonitrile (1.5 ml), and HATU (10 mg, 0.026 mmol) and DIEA (10 μl) were added in sequence; then lcaa CPG (500°A, 125 mg) was added to the above reaction solution, and the reaction was stirred at room temperature for 2 hours. After the reaction was completed, 0.5 ml of CapA (acetic anhydride: tetrahydrofuran = 1:9, v / v) and 0.5 ml of CapB (n-methylimidazole: pyridine: acetonitrile = 15:10:75, v / v / v) were added to lcaa CPG after washing with acetonitrile, and then stirring was continued at room temperature for 1 hour. The mixture was then filtered and washed with acetonitrile 3 times. The target product, compound III-LA20-CPG (210 mg, white powder) connected to the solid phase carrier was obtained by freeze drying.

[0346] Example Synthesis of siRNA conjugated with aminogalactose clusters

[0347] The siRNA used for the test, siRNA targeting mouse TTR gene mRNA (Nucleic Acids Res. 2020 Dec 2; 48(21): 11827-11844) is as follows: an aminogalactose molecule cluster M is covalently linked to the 3' end of the SS chain,

[0348] SS strand (5'-3'): a·a·caguGuUCUugcucuauaaL

[0349] AS chain (5'-3'):u·U·auaGaGCaagaAcAcuguu·u·u

[0350] Capital letters and lowercase letters represent monomers modified with 2′-deoxy-2′-fluoro (2′-F) and 2′-O-methyl (2′-OMe), respectively. · represents thio, and L represents N-acetylgalactosamine ligand.

[0351] The siRNA conjugates disclosed herein are synthesized by methods well known to those skilled in the art or by methods described in detail in the prior art. For example, WO2015006740A2 describes in detail the preparation methods of various siRNA conjugates.

[0352] Synthesize aminogalactose cluster-conjugated siRNA, and the siRNA used in the experiment targets mouse TTR mRNA.

[0353]

[0354] Table X. siRNA numbers and sequences for evaluation of targeting ligand activity

[0355]

[0356] Among them, S-L96 was purchased from Jima Gene; C, G, A, U or c, g, a, u represent the base composition of nucleotides. The uppercase and lowercase letters represent monomers modified with 2′-deoxy-2′-fluoro (2′-F) and 2′-O-methyl (2′-OMe), respectively. · represents thio, and L represents N-acetylgalactosamine ligand.

[0357] After the preparation of the siRNA conjugate disclosed above is completed, it is freeze-dried into solid powder using standard means and stored for later use. When in use, it can be redissolved into a solution of desired concentration using, for example, water for injection, physiological saline, phosphate buffer or phosphate buffer.

[0358] (I) ASGPR-GalNAc TM affinity detection test based on monkey primary cells

[0359] Principle: The test compound and GalNAc-Cy5 compete with the ASGPR receptor on the surface of monkey primary cells (PCMH, purchased from Miaoshun Biotechnology Co., Ltd.). The fluorescence intensity of Cy5 detected by flow cytometry can reflect the affinity of the test compound to ASGPR. The weaker the Cy5 fluorescence intensity, the stronger the affinity of the test compound to ASGPR, and vice versa.

[0360] Experimental process: Monkey primary cells were plated in 96-well plates at 40,000 / well and incubated with GalNAc-Cy5 (final concentration 100 nM) and the test compound (final concentration starting from 1 μM, 10-fold dilution for initial screening) in a 37°C, 5% CO2 incubator for 4 hours. The cells were then collected and the fluorescence intensity of Cy5 in the cells was analyzed by flow cytometry.

[0361] Conclusion: The results of affinity detection experiments show that the affinity of most of the compounds currently tested, such as A11, A14, A15, A16, A17, A19, A21, A23, A24, A25, A26, A27, A28, A31, A32, A33, A34, A35, A36, A37, and A38, to ASGPR shows activity comparable to or slightly superior to that of the L96-DeAc molecule.

[0362] (II) Inhibition of mRNA expression in vivo by siRNA conjugated with aminogalactose clusters

[0363] C57 / BL6 mice aged 6 to 8 weeks were randomly divided into 3 groups, 3 mice in each group, and 1 group received no treatment. The GalNac-siRNA was injected subcutaneously at a dose of 1 mg / kg in the control group (S-L96), the positive control group (S-L96), and the drug-treated group (S-LA20). Blood was collected from the eye sockets to separate serum before administration (day 0) and 3 days after administration (day 3), and the mice were killed on day 7 to obtain liver samples. The TTR levels in mouse serum and liver samples were detected using the Mouse Prealbumin ELISA Kit (cat#ALPCO(41-PALMS-E01)), and the results are as follows:

[0364]

[0365] Conclusion: The in vivo efficacy test data showed that S-LA20 exhibited excellent mTTR gene expression inhibition efficiency, and the activity level of 1 mpk was comparable to that of the control group S-L96.

[0366] All documents mentioned in the present invention are cited as references in this application, just as each document is cited as reference individually. In addition, it should be understood that after reading the above teachings of the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms also fall within the scope defined by the claims attached to this application.

Claims

1. A GalNAc delivery molecule as shown in the following formula (I), or a stereoisomer, enantiomer, or a pharmaceutically acceptable salt thereof: in, R1 is selected from the group consisting of H, DMTr, phosphate, phosphorothioate, phosphorothioate-siRNA conjugate or phosphate-siRNA conjugate; Linker is a divalent linking group; r is 0, 1, 2, 3, 4, 5 or 6; R2 is a GalNAc target head portion and has a structure as shown below: Wherein, L1, L2 and L3 are each independently a divalent linking group, and R3, R4 and R5 are each independently a substituted or unsubstituted sugar group, preferably a group formed by a substituted or unsubstituted N-acetylgalactosamine molecule; The substitution refers to the substitution of one or more hydrogen atoms on the group by a substituent selected from the following group: 1-8 Alkyl, C 2-8 Alkenyl, C 2-8 Alkynyl, C 3-8 Cycloalkyl, 3- to 12-membered heterocyclic group, C 3-8 Aryl, 5- to 7-membered heteroaryl, halogen, hydroxyl, carboxyl (-COOH), C 1-8 Aldehyde, C 2-10 Acyl, C 2-10 Ester group, amino group, C 1-8 Alkoxy, C 1-10 or two substituents located on adjacent ring atoms may together with the connected ring atoms form a group selected from the following group: a 5-7 membered carbon ring or heterocyclic ring, a benzene ring, or a 5-7 membered heteroaromatic ring.

2. The GalNAc delivery molecule according to claim 1, or a stereoisomer, an enantiomer, or a pharmaceutically acceptable salt thereof, wherein: The linker has a structure selected from the following group: C4-C20 alkyl, wherein n and m are each independently selected from the group consisting of 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12; n1 and n2 are each independently selected from the following group: 0 or 1; W is selected from the group consisting of a single bond, a substituted or unsubstituted C 1-8 Alkylene, substituted or unsubstituted 7-12 membered fused bicyclic ring, substituted or unsubstituted 7-15 membered spirocyclic ring, substituted or unsubstituted 5-15 membered bridged ring, -NH-substituted or unsubstituted 7-12 membered fused bicyclic ring, -NH-substituted or unsubstituted 7-15 membered spirocyclic ring, -NH-substituted or unsubstituted 5-15 membered bridged ring, -NH-substituted or unsubstituted 7-12 membered fused bicyclic ring-NH-, -NH-substituted or unsubstituted 7-15 membered spirocyclic ring-NH-, -NH-substituted or unsubstituted 5-15 membered bridged ring-NH-; wherein the ring atoms of the fused bicyclic ring, spirocyclic ring and bridged ring may be optionally carbon atoms or heteroatoms; The substitution refers to the substitution of one or more hydrogen atoms on the group by a substituent selected from the following group: 1-8 Alkyl, C 2-8 Alkenyl, C 2-8 Alkynyl, C 3-8 Cycloalkyl, 3- to 12-membered heterocyclic group, C 6-10 Aryl, 5- to 10-membered heteroaryl, halogen, hydroxyl, carboxyl (-COOH), C 1-8 Aldehyde, C 2-10 Acyl, C 2-10 Ester group, amino group, C 1-8 Alkoxy, C 1-10 Sulfonyl; or two substituents located on adjacent ring atoms may together with the connected ring atoms form a group selected from the following group: a 5-7 membered carbon ring or heterocyclic ring, a benzene ring, or a 5-7 membered heteroaromatic ring; Or W has a structure selected from the group consisting of: wherein s1, s2, s, s4, s5 and s6 are each independently selected from the following group: 0, 1 or 2; s7 is selected from the group consisting of 1, 2, 3, 4, 5 or 6; Each of M1, M2, M3 and M4 is independently selected from the group consisting of CHR, C(R)R, C(O), O, S, NR; M5 and M6 are each independently selected from the following group: (CHR) s6 、C(R)R、C(O)、(CHR) s6 -C(O), O, S, NR; R is selected from the following group: H, halogen, methyl; or two R located on adjacent reducing atoms and the carbon atom to which they are connected together form a substituted or unsubstituted 4-8 membered carbocyclic or heterocyclic ring (including saturated, unsaturated or aromatic ring).

3. The GalNAc delivery molecule according to claim 1, or a stereoisomer, an enantiomer, or a pharmaceutically acceptable salt thereof, wherein: Said W has a structure selected from the following group:

4. The GalNAc delivery molecule according to claim 1, or a stereoisomer, an enantiomer, or a pharmaceutically acceptable salt thereof, wherein: The L1, L2 and L3 are each independently selected from the following structures: wherein each u, v and w are independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or 11; u1 and u2 are independently selected from 0 or 1; Glu is an unmodified or modified 5-6-membered glycosyl; X and Y are each independently selected from the following group: -NH-(C1-C6 alkyl)-NH-, -NH-(C3-C8 cycloalkyl)-NH-, -NH-(4-10 membered heterocyclyl)-NH-, Wherein, the ring D is a 4-15 membered heterocyclic group; The cycloalkyl and heterocyclic groups may be monocyclic, condensed, bridged or spirocyclic; the heterocyclic group may be saturated or partially unsaturated (but not aromatic).

5. The GalNAc delivery molecule according to claim 4, or a stereoisomer, an enantiomer, or a pharmaceutically acceptable salt thereof, wherein: The X and Y each independently have a structure selected from the following group: -NH-(C1-C6 alkyl)-NH-, -NH-(C3-C8 cycloalkyl)-NH-, -NH-(4-10 membered heterocyclyl)-NH-, or a structure selected from the following group:

6. The GalNAc delivery molecule according to claim 1, or a stereoisomer, an enantiomer, or a pharmaceutically acceptable salt thereof, wherein: The sugar group has a structure shown in the following formula III: Among them, the R 11 Selected from the group consisting of -NH(C2-C6 acyl), -NH(halogenated C2-C6 acyl), -NH(C2-C6 sulfonyl), -NH(halogenated C2-C6 sulfonyl); R 12 , R 13 and R 14 Each is independently selected from the following group: H or C1-C6 acyl.

7. The GalNAc delivery molecule according to claim 6, or a stereoisomer, an enantiomer, or a pharmaceutically acceptable salt thereof, wherein: The R 11 Selected from the group consisting of: -NH(C2-C4 acyl); R 12 , R 13 and R 14 Each is independently selected from the following group: H or C1-C4 acyl.

8. The GalNAc delivery molecule according to claim 6, or a stereoisomer, an enantiomer, or a pharmaceutically acceptable salt thereof, wherein: The R 11 is -NHAc; R 12 , R 13 and R 14 Each is independently selected from the group consisting of H or Ac.

9. The GalNAc delivery molecule according to any one of claims 1 to 8, or a stereoisomer, enantiomer, or a pharmaceutically acceptable salt thereof, characterized in that: The GalNAc delivery molecule has a structure selected from the group consisting of: Wherein, X, Y, Z and W are each independently selected from the following groups: -NH-(C1-C6 alkyl)-NH-, -NH-(C3-C8 cycloalkyl)-NH-, -NH-(4-10 membered heterocyclyl)-NH-, Wherein, the ring D is a 4-15 membered heterocyclic group; R3, R4, R5 and R5' are each independently a substituted or unsubstituted sugar group, preferably a group formed by a substituted or unsubstituted N-acetylgalactosamine molecule; The remaining groups are as defined above.

10. The GalNAc delivery molecule according to any one of claims 1 to 8, or a stereoisomer, enantiomer, or a pharmaceutically acceptable salt thereof, characterized in that: The GalNAc delivery molecule has a structure selected from the group consisting of: The R1 group is as defined in claim 1.

11. The GalNAc delivery molecule according to any one of claims 1 to 8, or a stereoisomer, an enantiomer, or a pharmaceutically acceptable salt thereof, characterized in that: The GalNAc delivery molecule has a structure selected from the group consisting of:

12. A pharmaceutical composition, characterized in that The pharmaceutical composition comprises: one or more of the compound of formula I according to any one of claims 1 to 11, its pharmaceutically acceptable salt, racemate, R-isomer, S-isomer or mixture thereof, and one or more pharmaceutically acceptable carriers, excipients, adjuvants, auxiliary materials and / or diluents.

13. The use of a compound of formula I as claimed in any one of claims 1 to 11, its pharmaceutically acceptable salt, racemate, R-isomer, S-isomer or mixture thereof, characterized in that: Used for preparing a pharmaceutical composition for treating or preventing tumors or infections caused by viruses.

14. Use of a compound of formula (I) according to any one of claims 1 to 11 for preparing a medicament for treating and / or preventing a disease or condition selected from the group consisting of a disease or condition caused by hepatitis B virus, a disease or condition caused by abnormal expression of ANGPTL3 gene, a disease or condition caused by abnormal expression of TTR gene, a disease or condition caused by abnormal expression of hepatocyte genes, or other liver diseases or conditions.

15. A method for detecting ASGPR-GalNAc affinity based on monkey primary cells.

16. A GalNAc delivery molecule linker as shown in formula (Ia), or a stereoisomer, enantiomer, or a pharmaceutically acceptable salt thereof: The moiety shown in formula (Ia) has a structure selected from the group consisting of: in, It is the site connected to R1.

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