Compound with dicycloacetamide structure as well as preparation method and medicinal application thereof

Abstract

The invention discloses a compound with a bi-dicycloacetamide structure as well as a preparation method and medicinal application of the compound. The compound has a structure # imgabs0 # as shown in a general formula (I); wherein C1 represents a benzene ring, indole, pyrrole, furan, thiophene, pyrazole, imidazole, isoxazole, thiazole, pyridine and morpholine; c2 represents pyridine, triazole, imidazolinone or indole; r1 represents one or two of hydrogen, fluorine, chlorine, methyl, hydroxyl, cyano, methoxyl, 2-nitrogen morpholine ethoxyl and 4-morpholinyl; R1 represents one or two of hydrogen, fluorine, chlorine, methyl, hydroxyl, cyano, methoxyl, 2-nitrogen morpholine ethoxyl and 4-morpholinyl; r2 represents hydrogen, fluorine, chlorine, methyl or methoxyl; r3 represents # imgabs 1 #, L is hydrogen, fluorine, chlorine, bromine, methyl or methoxyl, X is C or N, and n is equal to 0, 1, 2 or 3. The compounds have good application in resisting tumors and proliferative diseases except tumors.

Description

Technical Field

[0001] The invention belongs to the field of medicine, and specifically relates to a bicyclic acetamide structural compound, a preparation method thereof and a pharmaceutical use thereof. Background Art

[0002] Microtubules are tubular polymers formed by the polymerization of α and β tubulin heterodimers. They have many functions in cells, especially participating in the mitosis process of cells in the form of spindles. Tumor cells have the ability to proliferate rapidly. If the formation of their spindles is inhibited, mitosis will be blocked, causing tumor cell growth to stagnate in the G2 / M phase and inhibiting cell proliferation.

[0003] Microtubules exist in two forms: cytoplasmic microtubules and spindle microtubules. They participate in cell movement, organelle positioning, intracellular material transport, and the mitotic process of eukaryotic cells. When a cell enters the division phase, its cytoplasmic microtubules depolymerize into tubulin and then assemble into spindle microtubules. In the late stage of division, the spindle microtubules pull the sister chromatids from the equatorial plane to the two poles of the spindle. After the end of mitosis, the spindle microtubules depolymerize and assemble into cytoplasmic microtubules. Therefore, this dynamic conversion (polymerization and depolymerization) between tubulin / microtubules is necessary for normal cell mitosis. If this cycle is disrupted, cells entering mitosis will stop dividing, thereby prolonging the cell cycle or inducing cell apoptosis. Therefore, tubulin has become an important target for anti-tumor drugs.

[0004] Src kinase is a non-receptor tyrosine kinase that is involved in mediating various signal transduction pathways related to cell growth, migration and survival. It includes nine family members, among which c-Src is closely related to tumor growth. The c-Src kinase in normal cells is in a dynamic equilibrium process, but the continuous activation and high expression of c-Src kinase in cancer cells disrupts the signaling pathway, thereby promoting the development of cancer. c-Src promotes tumor survival, proliferation and angiogenesis by regulating downstream effectors, including PI3K / -AKT / mTOR, JAK / -STA and RAS / MAPK, through signaling. Specifically, ① Src overexpression promotes HER2-HER3 dimerization and enhances downstream signaling, and c-Src transfection enhances EGFR-induced tumorigenesis; ② In a hypoxic environment, active c-Src may lead to upregulation of VEGFR, thereby promoting angiogenesis. Therefore, c-Src has long been considered a possible therapeutic target in oncology. In addition, c-Src regulates the nucleation of tubulin during spindle formation. Therefore, combined targeting of tubulin and c-Src will contribute to synergistic effects. Summary of the invention

[0005] The object of the present invention is to provide a bicyclic acetamide structural compound; another object of the present invention is to provide a method for preparing the compound; the third object of the present invention is to provide the use of the compound or a pharmaceutical composition containing the compound in the preparation of a drug for preventing or treating anti-tumor or proliferative diseases other than tumors.

[0006] The specific technical solutions of the present invention are as follows:

[0007] As the first aspect of the present invention, the bicyclic acetamide structure compound and any combination thereof or a pharmaceutically acceptable salt thereof, the general structural formula is as shown in Formula I:

[0008]

[0009] Wherein C1 represents a benzene ring, indole, pyrrole, furan, thiophene, pyrazole, imidazole, isoxazole, thiazole, pyridine; C2 represents pyridine, triazole, imidazolinone, indole; R1 represents one or two of hydrogen, fluorine, chlorine, methyl, hydroxyl, cyano, methoxy, 2-nitromorpholinoethoxy, 4-morpholino; R2 represents hydrogen, fluorine, chlorine, methyl or methoxy; R3 represents L is hydrogen, fluorine, chlorine, bromine, methyl, methoxy, X is C or N, and n=0, 1, 2, 3.

[0010] Further, for

[0011] for R3 is

[0012]

[0013] A biologically acceptable salt formed by the above-mentioned bicyclic acetamide structural compound and at least one of acetic acid, dihydrofolic acid, benzoic acid, citric acid, sorbic acid, propionic acid, oxalic acid, fumaric acid, maleic acid, hydrochloric acid, malic acid, phosphoric acid, diphosphoric acid, hydrobromic acid, nitric acid, sulfurous acid, sulfuric acid, vanillic acid, tartaric acid, succinic acid, ascorbic acid, boric acid, lactic acid, p-toluenesulfonic acid, salicylic acid and ethylenediaminetetraacetic acid.

[0014] As a second aspect of the present invention, the compound of the present invention can be prepared by the following method, and its synthetic route is as follows:

[0015] Synthesis route 1:

[0016]

[0017]

[0018] The specific steps are:

[0019] (1) Preparation of 2-morpholineethanol (2)

[0020] Using morpholine as the starting material, it reacted with 2-bromoethanol through nucleophilic substitution to obtain compound 2;

[0021] (2) Preparation of tert-butyl 4-hydroxyanilinocarbonate (4)

[0022] 4-Aminophenol (compound 3) reacts with di-tert-butyl dicarbonate to obtain compound 4;

[0023] (3) Preparation of tert-butyl 4-(2-morpholinethoxy)anilinocarbonate (5)

[0024] Compound 2 and compound 4 were reacted to obtain compound 5;

[0025] (4) Preparation of 4-(2-morpholinethoxy)aniline hydrochloride (6)

[0026] Compound 5 is subjected to acidic conditions to remove the tert-butyloxycarbonyl protecting group to obtain compound 6;

[0027] (5) Preparation of 4-(2-fluoro-4-nitrophenyl)morpholine (8)

[0028] 3,4-Difluoronitrobenzene (compound 7) reacts with morpholine under alkaline conditions through nucleophilic substitution to give compound 8;

[0029] (6) Preparation of 3-fluoro-4-morpholinoaniline (9)

[0030] Compound 8 is reduced by hydrogen to obtain compound 9;

[0031] (7) Preparation of tert-butyl 5-nitroindole-1-carbonate (11)

[0032] 5-Nitroindole (Compound 10) reacts with di-tert-butyl dicarbonate under alkaline conditions to obtain Compound 11;

[0033] (8) Preparation of tert-butyl 5-aminoindole-1-carbonate (12)

[0034] Compound 11 is reduced by hydrogen to obtain compound 12;

[0035] (9) Preparation of 1-(2-chloroethyl)-3-(4-(2-morpholinethoxy)aryl)urea (14)

[0036] Compound 6 or compound 9 or compound 12 or compound 13 is condensed with 2-chloroethyl isocyanate to obtain compounds 14a, 14b, 14c, and 14d;

[0037] (10) Preparation of 1-(4-(2-morpholinethoxy)aryl)imidazolin-2-one (15)

[0038] Compounds 14a, 14b, 14c, and 14d were subjected to intramolecular cyclization under alkaline conditions to give compounds 15a, 15b, 15c, and 15d;

[0039] (11) Preparation of tert-butyl 2-(3-(4-(2-morpholinethoxy)aryl)-2-oxoimidazolin-1-yl)acetate (16)

[0040] Compound 15a reacts with tert-butyl bromoacetate under alkaline conditions to obtain compound 16 through affinity substitution reaction;

[0041] (12) Preparation of 2-(3-(4-(2-morpholinethoxy)aryl)-2-oxoimidazolin-1-yl)acetic acid (17)

[0042] Compound 16 was hydrolyzed under acidic conditions to obtain compound 17;

[0043] (13) Preparation of N-benzyl 2-(3-(4-(2-morpholinethoxy)aryl)-2-oxoimidazolin-1-yl)acetamide (18)

[0044] Compound 17 is reacted with R3NH2 to obtain compound 18.

[0045] (14) reacting compound 19 with benzylamine or 3-chlorobenzylamine to obtain compound 20a or 20b;

[0046] (15) reacting compound 15b with compound 20a or 20b to obtain 21a or 21b;

[0047] (16) reacting compound 15c or 15d with compound 20a to obtain 21c or 21d;

[0048] Synthesis route 2:

[0049]

[0050] (1) Preparation of 4-(2-morpholinethoxy)phenylboronic acid pinacol ester (22)

[0051] Compound 2 and 4-hydroxyphenylboronic acid pinacol ester were reacted to obtain compound 22;

[0052] (2) Preparation of 2-(4-azidophenoxy)ethoxymorpholine (23)

[0053] Compound 22 reacts with sodium azide to obtain compound 23;

[0054] (3) Preparation of methyl 2-(5-methyl-1-(4-(2-morpholinethoxy)phenyl)-1H-1,2,3-triazol-4-yl)acetate (24)

[0055] Compound 23 reacts with methyl 3-alkynepentanoate to obtain compound 24;

[0056] (4) Preparation of 2-(5-methyl-1-(4-(2-morpholinethoxy)phenyl)-1H-1,2,3-triazol-4-yl)acetic acid (25)

[0057] Compound 24 was hydrolyzed under alkaline conditions to obtain compound 25;

[0058] (5) Preparation of N-benzyl-2-(5-methyl-1-(4-(2-morpholinethoxy)phenyl)-1H-1,2,3-triazol-4-yl)acetamide (26)

[0059] Compound 25 was reacted with benzylamine to give the target product 26.

[0060] Synthesis route three:

[0061]

[0062] (1) Preparation of N-benzyl-4-alkynepentanamide (28)

[0063] 4-Alkynepentanoic acid (compound 27) and benzylamine were condensed to give intermediate 28;

[0064] (2) Preparation of N-benzyl-3-(1-(4-(2-morpholinethoxy)phenyl)-1H-1,2,3-triazol-4-yl)propanamide (29)

[0065] Compound 28 reacts with compound 23 to obtain target compound 29.

[0066] Synthesis route 4:

[0067]

[0068] (1) Preparation of ((4-bromophenoxy)ethyl)-2-morpholine (31)

[0069] Compound 2 and 4-bromophenol (compound 30) were reacted to give compound 31 by Mitsunobu reaction;

[0070] (2) Preparation of (4-((trimethylsilyl)ethynyl)phenoxyethoxy)-2-morpholine (32)

[0071] Compound 31 was coupled with trimethylsilyl acetylene to obtain compound 32;

[0072] (3) Preparation of ((4-ethynylphenoxy)ethyl)-2-morpholine (33)

[0073] Compound 32 was subjected to alkaline conditions to remove the trimethylsilyl group to obtain compound 33;

[0074] (4) Preparation of 2-azido-N-benzylacetamide (34)

[0075] Compound 20a reacts with sodium azide under alkaline conditions to give compound 34;

[0076] (5) Preparation of N-benzyl (33) Preparation of N-benzyl-2-(4-(4-(2-morpholinethoxy)phenyl)-1H-1,2,3-triazol-1-yl)acetamide (35)

[0077] Compound 33 and compound 34 were reacted by click reaction to obtain target compound 35.

[0078] Synthesis route 5:

[0079]

[0080] (1) Preparation of 2-(4-(1H-indol-4-yl)phenoxy)ethylmorpholine (37)

[0081] 4-Bromoindole (Compound 36) and Compound 22 were reacted by Suzuki reaction to obtain Compound 37;

[0082] (2) Preparation of N-benzyl-2-(4-(4-(2-morpholinethoxy)phenyl)-1H-indol-1-yl)acetamide (38)

[0083] Compound 37 and compound 20a were reacted under alkaline conditions to obtain the target compound 38 through affinity substitution reaction.

[0084] Synthesis route 6:

[0085]

[0086] (1) Preparation of N-benzyl-2-(5-bromopyridin-2-yl)acetamide (40)

[0087] Compound 39 is condensed with benzylamine to give compound 40;

[0088] (2) Preparation of N-benzyl-2-(5-substituted arylpyridin-2-yl)acetamide (41)

[0089] Intermediate 40 is coupled with a substituted heterocyclic aryl boronic acid to give the target compound 41.

[0090] As a third aspect of the present invention, there is also provided the use of any of the above compounds and any combination thereof or pharmaceutically acceptable salts thereof in the preparation of drugs for preventing or treating anti-tumor or other proliferative diseases.

[0091] The tumor disease is breast cancer, ovarian cancer, non-small cell lung cancer, liver cancer, pancreatic cancer, prostate cancer, melanoma or leukemia.

[0092] The present invention also provides a pharmaceutical composition. The compound of general formula (I), pharmaceutically acceptable salt or solvate can be added with a pharmaceutically acceptable carrier to prepare common pharmaceutical preparations, such as tablets, capsules, powders, syrups, liquids, suspensions, injections, and can be added with common pharmaceutical excipients such as flavorings, sweeteners, liquid or solid fillers or diluents.

[0093] Use of the above compound or its pharmaceutically acceptable salt in the preparation of microtubule protein polymerization inhibitors

[0094] Use of the above compound or a pharmaceutically acceptable salt thereof in preparing a Fak protein and a Src protein phosphorylation inhibitor.

[0095] In order to evaluate the anti-tumor activity of the synthetic compound of the present invention and the pharmaceutical composition thereof, the pharmacological activity tests at the cellular and molecular levels were performed on the compound of the present invention.

[0096] The compounds and pharmaceutical compositions of the present invention were subjected to in vitro cell growth inhibition experiments, and the results showed that the compounds generally have strong cell growth inhibition effects. The half inhibitory concentration (IC 50 ) reached the nanomolar level (nM). The cell growth inhibitory activity of some compounds was comparable to that of the positive control drugs KX01 and colchicine.

[0097] The compounds and pharmaceutical compositions of the present invention were subjected to an in vitro microtubule protein polymerization inhibition experiment, and the results showed that compound 41f had a significant inhibitory effect on microtubule protein polymerization.

[0098] The compounds and pharmaceutical compositions of the present invention were subjected to in vitro cell growth cycle experiments, and the results showed that compound 41f at 0.1 μM could significantly arrest cells in the G2 / M phase.

[0099] The compounds and pharmaceutically acceptable carriers of the present invention can be administered orally or intravenously in clinical practice. They can be used as a single drug in clinical practice, or in combination with other clinically used chemotherapeutic drugs and radiotherapy for the treatment of the aforementioned cancers. BRIEF DESCRIPTION OF THE DRAWINGS

[0100] Figure 1This is a graph of the inhibition of tubulin polymerization in vitro. A is the effect of blank control, paclitaxel (3 μM), colchicine (2.5 μM), KX01, and compound 41f (20 μM) on tubulin polymerization;

[0101] Figure 2 This is the effect of compound 41f on the growth cycle of HeLa cells. A is the blank control, BCD are the effects of compound 41f on the cell growth cycle at concentrations of 0.05μM, 0.08μM, and 0.10μM, respectively;

[0102] Figure 3 This is the result of immunoblotting experiment that compound 41f inhibits the phosphorylation of Fak and Src. DETAILED DESCRIPTION

[0103] The technical solution of the present invention is further described in detail below in conjunction with specific examples, but the protection scope of the present invention is not limited thereto. The ratios of the eluents used in the column chromatography in the following examples are all volume ratios.

[0104] Example 1

[0105] Preparation of N-(2-fluorobenzyl)-2-(3-(4-(2-morpholinethoxy)phenyl)-2-oxoimidazolin-1-yl)acetamide (18a)

[0106] Preparation of 1-(4-methoxybenzyl)-3-oxo-1,2,3,4-tetrahydroquinoxaline-6-carboxylic acid methyl ester (18a)

[0107]

[0108] (1) Preparation of 2-morpholinoethanol (2)

[0109] Morpholine (Compound 1, 0.48 mL, 5.5 mmol), acetonitrile (20 mL), 2-bromoethanol (0.47 mL, 6.6 mmol, 1.2 equiv), potassium carbonate (1.0 g, 7.2 mmol, 1.3 equiv) were placed in a 50 mL round-bottom flask in sequence. The mixture was stirred at 30°C for 18 h, and the reaction was monitored by TLC (iodine color development). After the reaction was complete, the mixture was filtered and distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, DCM:MeOH=10:1) was performed to obtain 0.51 g of colorless liquid with a yield of 70.7%. 1 H NMR (300MHz, Chloroform-d) δ3.61(t,J=4.5Hz,4H),3.53(t,J=5.4Hz,2H),3.27(s,1H),2.43(t,J=5.4Hz,2H),2.40(t,J=4.5Hz,4H).

[0110] (2) Preparation of tert-butyl 4-hydroxyanilinocarbonate (4)

[0111] 4-Aminophenol (compound 3, 2.00 g, 18.3 mmol), tetrahydrofuran (20 mL), and triethylamine (12.2 mL, 87.8 mmol, 4.8 equiv) were sequentially placed in a 50 mL round-bottom flask. Subsequently, di-tert-butyl dicarbonate ((Boc)2O, 4.80 g, 22.0 mmol, 1.2 equiv) was dissolved in 5 mL of triethylamine and slowly added to the reaction bottle at a uniform rate through a constant pressure dropping funnel. The mixture was stirred at room temperature for 6 h, and the reaction was monitored by TLC. After the reaction was complete, the solvent was distilled off under reduced pressure, and 3.45 g of colorless liquid was obtained by column chromatography (200-300 mesh silica gel, PE:EA=3:1). The yield was 90.1%. 1 H NMR (300MHz, CD3OD) δ7.17 (d, J = 8.4Hz, 2H), 6.71 (d, J = 8.4Hz, 2H), 4.93 (s, 1H), 1.48 (s, 9H).

[0112] (3) Preparation of tert-butyl 4-(2-morpholinoethoxy)anilinocarbonate (5)

[0113] Intermediate 4 (2.0 g, 9.6 mmol), anhydrous tetrahydrofuran (20 mL), intermediate 2 (2.2 g, 17 mmol, 1.8 equiv), and tributylphosphine (TBUP, 7.2 mL, 29 mmol, 3.0 equiv) were sequentially placed in a 100 mL round-bottom flask. Subsequently, N, N, N', N'-nitrogen dicarbonamide (TMAD, 5.0 g, 29 mmol, 3.0 equiv) was dissolved in a constant pressure dropping funnel filled with 30 mL of anhydrous dichloromethane, the air was replaced by nitrogen, stirred at 0°C for 30 min, and slowly added to the reaction bottle at a uniform speed. After the addition was complete, the mixture was moved to room temperature and stirred for 2 h. The reaction was monitored by TLC. After the reaction was complete, the solvent was distilled off under reduced pressure, and column chromatography (200-300 mesh silica gel, PE:EA=1:1) was performed to obtain 2.8 g of colorless liquid with a yield of 90.5%. 1 H NMR (300MHz, Chloroform-d) δ7.25 (d, J = 8.4Hz, 2H), 6.83 (d, J = 8.4Hz, 2H), 4.07 (t, J = 5. 7Hz,2H),3.7(t,J=4.8Hz,4H),2.78(t,J=5.7Hz,2H),2.56(t,J=4.8Hz,4H),1.50(s,9H).

[0114] (4) Preparation of 4-(2-morpholinethoxy)aniline hydrochloride (6)

[0115] Intermediate 5 (1.0 g, 3.1 mmol) was placed in a 25 mL round-bottom flask. Then 10 mL of hydrochloric acid ethanol solution was added and stirred at 70°C for 1 h. The reaction was monitored by TLC. After the reaction was complete, the oily product was obtained by vacuum distillation without purification and was directly used for the next step.

[0116] (5) Preparation of 1-(2-chloroethyl)-3-(4-(2-morpholinoethoxy)phenyl)urea (14a)

[0117] Intermediate 6 (1.0 g, 3.9 mmol), anhydrous tetrahydrofuran (20 mL), chloroethyl isocyanate (0.43 mL, 5.0 mmol, 1.3 equiv) and triethylamine (1.2 mL, 8.6 mmol, 2.2 equiv) were sequentially placed in a 50 mL round-bottom flask, stirred at room temperature for 4 h, and the reaction was monitored by TLC. After the reaction was complete, vacuum distillation was added to obtain 0.55 g of a white solid with a yield of 43.0%. It was directly used in the next step.

[0118] (6) Preparation of 1-(4-(2-morpholinethoxy)phenyl)imidazolin-2-one (15a)

[0119] Intermediate 14a (0.50 g, 1.5 mmol) and anhydrous tetrahydrofuran (10 mL) were added to a 25 mL round-bottom flask. Subsequently, sodium hydride (0.10 g, 3.0 mmol, 2.0 equiv) was slowly added to the reaction flask, stirred at room temperature for 2 h, and the reaction was monitored by TLC. After the reaction was complete, the solvent was evaporated under reduced pressure and column chromatography (200-300 mesh silica gel, DCM:MeOH=10:1) was performed to obtain 0.31 g of a white solid with a yield of 70.9%. 1 H NMR(300MHz,Chloroform-d)δ7.40(d,J=9.0Hz,2H),6.88(d,J=9.0Hz,2H),5.42(s,1H),4.08(t,J=5.7Hz,2H),3 .88(t,J=8.4Hz,2H),3.73(t,J=4.8Hz,4H),3.54(t,J=8.4Hz,2H),2.78(t,J=5.7Hz,2H),2.56(t,J=4.8Hz,4H).

[0120] (7) Preparation of tert-butyl 2-(3-(4-(2-morpholinethoxy)phenyl)-2-oxoimidazolin-1-yl)acetate (16)

[0121] Intermediate 15a (1.1 g, 3.8 mmol), DMF (20 mL), and tert-butyl bromoacetate (0.81 mL, 5.7 mmol, 1.5 equiv) were sequentially placed in a 50 mL round-bottom flask. Subsequently, sodium hydride (0.13 g, 3.8 mmol, 1.0 equiv) was slowly added to the reaction flask, stirred at room temperature for 4 h, and the reaction was monitored by TLC. After the reaction was complete, dichloromethane and water were added for extraction, and the organic phase was dried over anhydrous sodium sulfate, filtered, and then distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, DCM: MeOH = 20: 1) was performed to obtain 0.78 g of a white solid with a yield of 50.6%. 1 H NMR (300MHz, DMSO-d6) δ7.43(d,J=9.1Hz,2H),6.91(d,J=9.1Hz,2H),4.04(t,J=5.8Hz,2H),3.88(s,2H),3.78(dd,J1=8.4Hz,J 2=6.3,2H),3.57(t,J=4.8Hz,4H),3.50(dd,J1=8.4Hz,J2=6.3,2H),2.66(t,J=5.8Hz,2H),2.45(t,J=4.8Hz,4H),1.42(s,9H).

[0122] (8) Preparation of 2-(3-(4-(2-morpholinethoxy)phenyl)-2-oxoimidazolin-1-yl)acetic acid (17)

[0123] Intermediate 16 (0.46 g, 1.1 mmol) and dichloromethane (16 mL) were placed in a 50 mL round-bottom flask. 16 mL of trifluoroacetic acid was slowly added to the reaction flask under ice bath conditions, and then stirred at room temperature for 6 h. The reaction was monitored by TLC. When the reaction was complete, dichloromethane was added several times and distilled under low temperature and reduced pressure to evaporate the solvent. Column chromatography (200-300 mesh silica gel, DCM:MeOH=5:1) was performed to obtain 0.26 g of a white solid with a yield of 67.7%. 1 H NMR (300MHz, DMSO-d6) δ7.44(d,J=9.0Hz,2H), 6.92(d,J=9.0Hz,2H), 4.03(t,J=5.7Hz,2H), 4.02(s,2H), 3.80(dd,J1=8 .6Hz, J2=7.5,2H), 3.57(t,J=4.8Hz,4H), 3.51(dd,J1=8.6Hz, J2=7.5,2H), 2.66(t,J=5.7Hz,2H), 2.45(t,J=4.8Hz,4H).

[0124] (9) Preparation of N-(2-fluorobenzyl)-2-(3-(4-(2-morpholinethoxy)phenyl)-2-oxoimidazolin-1-yl)acetamide (18a)

[0125] Intermediate 17 (0.10 g, 0.29 mmol), 2-fluorobenzylamine (0.040 mL, 0.35 mmol, 1.2 equiv), 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU, 0.13 g, 0.35 mmol, 1.2 equiv), N,N-diisopropylethylamine (DIPEA, 0.10 mL, 0.58 mmol, 2.0 equiv) were dissolved in 2 mL DMF. Stir at 40°C for about 4 h. The reaction was monitored by TLC. After the reaction was complete, dichloromethane and water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered, and then distilled under reduced pressure. The solvent was evaporated and column chromatography (200-300 mesh silica gel, DCM:MeOH=10:1) was performed to obtain 0.042 g of a white solid with a yield of 31.7%. HPLC purity: 98.75%, melting point: 153.1-154.7°C. 1 H NMR(300MHz,DMSO-d6)δ8.57(t,J=5.8Hz,1H),7.47(d,J=9.0Hz,2H),7.37-7.28(m,2H), 7.19(d,J=7.5Hz,1H),7.15(dd,J=5.1,2.7Hz,1H),6.93(d,J=9.0Hz,2H),4.35(d,J=5.8H z,2H),4.05(t,J=5.7Hz,2H),3.87(s,2H),3.79(dd,J1=9.0Hz,J2=6.7,2H),3.58(t,J=4 .5Hz,4H),3.51(dd,J1=9.0Hz,J2=6.7,2H),2.67(t,J=5.7Hz,2H),2.46(t,J=4.5Hz,4H). 13 C NMR (75MHz, DMSO) δ168.5, 160.0 (d, J = 243Hz), 157.6, 153.6, 134.1, 129.5 (d, J = 4.4Hz), 128.9 (d, J = 8.1Hz), 125.9 (d, J = 14.7Hz), 124.3 (d, J = 3.5Hz), 118.54(2C),115.1(d,J=21.1Hz),114.6(2C),66.2(2C),65.51,57.1,53.7 (2C),46.7(s),42.3(d,J=5.5Hz),35.8(d,J=4.5Hz).HRMS(ESI)m / z:calcd for C 24 H29 FN4O4Na + [M+Na] + :479.20650,found:497.20896.

[0126] Embodiments 2 to 13

[0127] Except that 3-fluorobenzylamine, 4-fluorobenzylamine, 3-chlorobenzylamine, 4-chlorobenzylamine, 3-bromobenzylamine, 2-methylbenzylamine, 4-methylbenzylamine, 2-methoxybenzylamine, 4-methoxybenzylamine, benzylamine, aniline, and 4-aminopyridine were used in step (9) instead of 2-fluorobenzylamine, the other steps and purification method were the same as those in Example 1, and 18b, 18c, 18d, 18e, 18f, 18g, 18h, 18i, 18j, 18k, 18l, and 18m were prepared respectively.

[0128]

[0129] The 18k NMR data are as follows: 1 H NMR (300MHz, DMSO-d6) δ8.56(t,J=6.0Hz,1H),7.49(d,J=9.0Hz,2H),7.36-7.22(m,5H),6.95(d,J=9.0Hz,2H),4.31(d,J=5.7Hz,2H),4. 12(t,J=5.7Hz,2H),3.87(s,2H),3.80(t,J=7.8Hz,2H),3.65(t,J=4.8Hz,4H),3.51(t,J=7.8Hz,2H),2.89(s,2H),2.68(t,J=4.8Hz,4H). 13 C NMR(75MHz,DMSO-d6)δ168.2,157.5,153.5,139.3,134.1,128.2(2C),127.1(2C),126.7,118.5 (2C),114.6(2C),66.0(2C),65.4,56.9,53.5(2C),46.7,42.3,42.2,41.9.HRMS(ESI)m / z:calcd for C 24 H 30 N4O4Na + [M+Na] + :461.21593,found:461.21832.

[0130] Embodiment 14

[0131] Preparation of N-benzyl-2-(3-(3-fluoro-4-morpholinophenyl)-2-oxoimidazolin-1-yl)acetamide (21a)

[0132]

[0133] (1) Preparation of 4-(2-fluoro-4-nitrophenyl)morpholine (8)

[0134] 3,4-difluoronitrobenzene (compound 7, 2.00 g, 12.6 mmol), DMF (20 mL), morpholine (1.20 mL, 13.8 mmol, 1.1 equiv), potassium carbonate (3.50 g, 25.2 mmol, 2.0 equiv) were sequentially placed in a 50 mL round-bottom flask. Subsequently, the mixture was stirred at 80°C for 6 h, and the reaction was monitored by TLC. After the reaction was complete, dichloromethane and water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered, and then distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, PE:EA=5:1) was performed to obtain 2.52 g of a yellow solid with a yield of 88.4%. 1 H NMR (300MHz, Chloroform-d) δ7.97(dd,J=9.0,2.7Hz,1H),7.88(dd,J=13.2,2.7Hz,1H),6.91(t,J=8.7Hz,1H),3.86(t,J=4.8Hz,4H),3.27(t,J=4.8Hz,4H).

[0135] (2) Preparation of 3-fluoro-4-morpholinoaniline (9)

[0136] The intermediate 8 (1.0 g, 4.4 mmol) was placed in a 25 mL round-bottom flask, and 10 mL of methanol was added to dissolve it. Then 10% palladium carbon (0.094 g, 0.088 mmol, 0.02 equiv) was slowly added to the reaction bottle, and the air was replaced with hydrogen. The mixture was stirred at room temperature for about 4 h. The reaction was monitored by TLC spot plate. After the reaction was complete, the reaction solution was filtered with a suction funnel with diatomaceous earth, and the solvent was evaporated under reduced pressure. Column chromatography (200-300 mesh silica gel, PE:EA=3:1) was performed to obtain 0.70 g of a light pink solid with a yield of 81.1%. 1 H NMR (300MHz, Chloroform-d) δ6.79(t,J=8.7Hz,1H),6.45-6.38(m,2H),3.85(t,J=4.5Hz,4H),3.57(brs,2H),2.96(t,J=4.8Hz,4H).

[0137] (3) Preparation of 1-(2-chloroethyl)-3-(3-fluoro-4-morpholinylphenyl)urea (14b)

[0138] The operation steps are the same as step (5) in Example 1 except that intermediate 6 is replaced by intermediate 9.

[0139] (4) Preparation of 1-(3-fluoro-4-morpholinylphenyl)imidazolin-2-one (15b)

[0140] The operation steps are the same as step (6) in Example 1 except that intermediate 14a is replaced by intermediate 14b.

[0141] 1 H NMR(300MHz,DMSO-d6)δ7.57(dd,J=15.6Hz,2.4Hz,1H),7.10(dd,J=8.4Hz,2.1Hz,1H),7.02-6.94(m,2H ),3.79(dd,J=8.1,7.2Hz,2H),3.72(t,J=4.5Hz,4H),3.37(dd,J=8.1,7.2Hz,2H),2.92(t,J=4.5Hz,4H).

[0142] (5) Preparation of N-benzyl-2-bromoacetamide (20a)

[0143] Benzylamine (2.00 mL, 18.7 mmol), dichloromethane (20 mL), and K2CO3 (5.17 g, 37.4 mmol, 2.0 equiv) were sequentially placed in a 50 mL round-bottom flask, the air was replaced with nitrogen, and the mixture was stirred at 0°C for about 30 min. Bromoacetyl bromide (compound 19, 1.62 mL, 18.7 mmol, 1.0 equiv) was slowly added to the reaction solution through a constant pressure dropping funnel, and the reaction was stirred for 4 h. The reaction was monitored by TLC spot plate. After the reaction was complete, the reaction solution was filtered and distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, PE:EA=5:1) was performed to obtain 2.80 g of a white solid with a yield of 65.6%. 1 H NMR(300MHz,DMSO-d6)δ8.76(brs,1H),7.22–7.37(m,5H),4.29(d,J=6.0Hz,2H),3.91(s,2H)

[0144] (6) Preparation of N-benzyl-2-(3-(3-fluoro-4-morpholinophenyl)-2-oxoimidazolin-1-yl)acetamide (21a)

[0145] Intermediate 15b (0.11 g, 0.40 mmol) and intermediate 20a (0.14 g, 0.60 mmol, 1.5 equiv) were dissolved in 4 mL DMF and stirred at room temperature. Sodium hydride (0.027 g, 0.80 mmol, 2.0 equiv) was slowly added to the reaction solution and stirred at room temperature for about 6 h. The reaction was monitored by TLC spot plate. After the reaction was complete, dichloromethane and water were extracted. The organic phase was dried over anhydrous sodium sulfate, filtered and distilled under reduced pressure. The solvent was evaporated and column chromatography (200-300 mesh silica gel, DCM: MeOH = 20: 1) was performed to obtain 0.042 g of white solid with a yield of 25.5%. HPLC purity: 96.70%, melting point: 155.8-157.0°C. 1 HNMR(300MHz,DMSO-d6)δ8.61(t,J=5.8Hz,1H),7.62(dd,J=15.5,1.8Hz,1H),7.35–7.22(m,5H),7.16(dd,J=8.7,1.8Hz,1H),7.00(t,J=9.4Hz, 1H), 4.32 (d, J = 5.8Hz, 2H), 3.89 (s, 2H), 3.79 (dd, J = 8.5, 6.0Hz, 2H), 3.74 (d, J = 3.9Hz, 4H), 3.51 (dd, J = 8.5, 6.0Hz, 2H), 2.93 (t, J = 3.9Hz, 4H). 13 CNMR(75MHz,DMSO-d6)δ168.28(s),157.3,154.9(d,J=241.1Hz),139.4,136.1(d,J=10.7Hz),134.1(d,J=9.1Hz),128.4(f2C),127.4(2C), 126.9,119.2(d,J=4.3Hz),112.5(d,J=3.2Hz),105.5(d,J=26.0Hz),66.31(2C),50.9(2C),46.7(s),42.2(2C),42.1.HRMS(ESI)m / z:calcd for C 22 H 25 FN4O3Na + [M+Na] + :435.18029,found:435.18229.

[0146] Embodiment 15

[0147] Preparation of N-(3-chlorobenzyl)-2-(3-(3-fluoro-4-morpholinophenyl)-2-oxoimidazolin-1-yl)acetamide (21b)

[0148]

[0149] Except that intermediate 20b was used instead of intermediate 20a in step (6), the other steps and purification method were the same as in Example 14, and 0.046 g of white solid was obtained with a yield of 25.7%. HPLC purity: 96.75%, melting point: 115.8-117.0°C. 1 H NMR (300MHz, DMSO-d6) δ8.62(t,J=5.8Hz,1H),7.60(d,J=15.5Hz,1H),7.39–7.27(m,3H),7.22(d,J=7.1Hz,1H),7.16(d,J=8.8Hz,1H),7.01(t,J=9. 3Hz,1H),4.30(d,J=5.8Hz,2H),3.87(s,2H),3.80(dd,J=8.5,7.3Hz,2H), 3.72(t,J=4.3Hz,4H), 3.51(dd,J=8.5,7.3Hz,2H), 2.93(t,J=4.3Hz,4H). 13 C NMR (75MHz, DMSO-d6) δ168.5,157.3,154.8(d,J=241.0Hz),142.0,136.1(d,J=10.7Hz),134.1(d,J=9.0Hz),133.0,130.2,127.0,126.8 ,125.9,119.2(d,J=4.3Hz),112.5(d,J=3.0Hz),105.4(d,J=26.0Hz),66.3(2C),50.9(2C),46.7,42.2,42.1,41.5.HRMS(ESI)m / z:calcd for C 22 H 24 ClFN4O3Na + [M+Na] + :469.14132,found:469.14260.

[0150] Example 16

[0151] Preparation of N-benzyl-2-(3-(1H-indol-5-yl)-2-oxoimidazolin-1-yl)acetamide (21c)

[0152]

[0153] (1) Preparation of tert-butyl 5-nitro-1H-indole-1-carbonate (11)

[0154] 5-Nitroindole (Compound 10, 1.50 g, 9.30 mmol), DMF (15 mL), di-tert-butyl dicarbonate (4.06 g, 18.6 mmol, 2.0 equiv), triethylamine (2.59 mL, 18.6 mmol, 2.0 equiv), and 4-dimethylaminopyridine (DMAP, 0.227 g, 1.86 mmol, 0.2 equiv) were sequentially placed in a 50 mL round-bottom flask. The mixture was stirred at 40°C for 4 h and the reaction was monitored by TLC. After the reaction was complete, dichloromethane and water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered, and then distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, PE:EA=6:1) was performed to obtain 1.95 g of a brown solid with a yield of 79.9%. 1 H NMR (300MHz, DMSO-d6) δ8.56(s,1H),8.17(s,2H),7.87(s,1H),6.92(s,1H),1.63(s,9H).

[0155] (2) Preparation of tert-butyl 5-amino-1H-indole-1-carbonate (12)

[0156] The intermediate 11 (0.50 g, 1.9 mmol) was placed in a 25 mL round-bottom flask, and 10 mL of methanol was added to dissolve it. 10% palladium carbon (0.04 g, 0.38 mmol, 0.02 equiv) was slowly added to the reaction solution, and the air was replaced by hydrogen three times. The solution was stirred at room temperature for 2 h and monitored by TLC. After the reaction was complete, the reaction solution was filtered with a funnel with diatomaceous earth, and the solution was distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, PE:EA=4:1) was performed to obtain 0.35 g of a brown solid with a yield of 79.9%. 1 HNMR(300MHz,DMSO-d6)δ7.73(d,J=8.6Hz,1H),7.47(d,J=3.3Hz,1H),6.72(s, 1H),6.63(d,J=8.6Hz,1H),6.44(d,J=3.3Hz,1H),4.87(brs,2H),1.58(s,9H).

[0157] (3) Preparation of 1-(2-chloroethyl)-3-(1H-indole-1-carbonic acid tert-butyl ester-5-yl)urea (14c)

[0158] The operation steps are the same as step (5) in Example 1 except that intermediate 12 is used to replace intermediate 6.

[0159] (4) Preparation of 1-(1H-indole-1-tert-butyl carbonate-5-yl)imidazolin-2-one (15c)

[0160] The operation steps are the same as step (6) in Example 1 except that intermediate 14c is used to replace intermediate 14a.

[0161] 1 H NMR(300MHz,DMSO-d6)δ7.95(d,J=9.0Hz,1H),7.71(s,1H),7.66–7.57(m,2H),6.90(s ,1H),6.67(d,J=3.5Hz,1H),3.89(t,J=7.9Hz,2H),3.41(t,J=7.9Hz,2H),1.62(s,9H).

[0162] (5) Preparation of N-benzyl-2-(3-(1H-indol-5-yl)-2-oxoimidazolin-1-yl)acetamide (21c)

[0163] Except that intermediate 15c was used instead of intermediate 15b in step (6), the other steps and purification method were the same as in Example 14, and 0.032 g of brown solid was obtained with a yield of 45.9%. HPLC purity: 97.75%, melting point: 147.8-149.0°C. 1 H NMR (300MHz, DMSO-d6) δ11.0(s,1H),8.55(t,J=5.6Hz,1H),7.53(s,1H),7.46(d,J=8.9Hz,1H) ,7.37–7.20(m,7H),6.37(s,1H),4.31(d,J=5.6Hz,2H),3.89–3.81(m,4H),3.57–3.48(m,2H). 13 C NMR(75MHz,DMSO-d6)δ168.6,158.0,139.4,133.1,132.3,128.3(2C),127.6,127.3(2C),12 7.2,126.8,125.9,113.7,111.1,109.0,101.0,47.0,43.2,42.5,42.0.HRMS(ESI)m / z:calcd for C 20 H 20 N4O2Na + [M+Na] + :371.14785,found:371.14876.

[0164] Embodiment 17

[0165] Preparation of N-benzyl-2-(2-oxo-3-phenylimidazolin-1-yl)acetamide (21d)

[0166]

[0167] (1) Preparation of 1-(2-chloroethyl)-3-phenylurea (14d)

[0168] The operation procedure is the same as step (5) in Example 1 except that aniline (compound 13) is used to replace intermediate 6.

[0169] (2) Preparation of 1-(2-chloroethyl)-3-(3-fluoro-4-morpholinylphenyl)urea (15d)

[0170] The operation steps are the same as step (6) in Example 1 except that intermediate 14d is used to replace intermediate 14a.

[0171] 1 H NMR (300MHz, DMSO-d6) δ7.53(d,J=8.1Hz,2H),7.22(t,J=7.8Hz,2H),6.84(t,J=7.1Hz,1H),3.70(dd,J=8.1,7.6Hz,2H),3.36(dd,J=8.1,7.6Hz,2H).

[0172] (3) Preparation of N-benzyl-2-(2-oxo-3-phenylimidazolin-1-yl)acetamide (21d)

[0173] Except that intermediate 15d was used instead of intermediate 15b in step (6), the other steps and purification methods were the same as those in Example 14, and 0.43 g of white solid was obtained with a yield of 44.8%. HPLC purity: 91.53%, melting point: 141.9-143.8°C. 1 H NMR (300MHz, DMSO-d6) δ8.59(t,J=4.7Hz,1H),7.59(d,J=7.6Hz,2H),7.37–7.23(m,7H),7.00(t,J=6. 8Hz, 1H), 4.31 (d, J = 4.7Hz, 2H), 3.88 (s, 2H), 3.82 (dd, J = 7.7, 7.6Hz, 2H), 3.53 (dd, J = 7.7, 7.6Hz, 2H). 13 C NMR(75MHz,DMSO-d6)δ168.3,157.4,140.7,139.4,128.7(2C),128.5(2C),128.3(2C) ,127.5,127.2,126.8,121.7,116.8(2C),48.7,42.2,42.0(2C).HRMS(ESI)m / z:calcd for C 18 H 19 N3O2Na + [M+Na] +:332.13694,found:332.13718.

[0174] Embodiment 18

[0175] Preparation of N-benzyl-2-(5-methyl-1-(4-(2-morpholinethoxy)phenyl)1H-1,2,3-triazol-4-yl)acetamide (26)

[0176]

[0177] (1) Preparation of 4-(2-morpholinoethoxy)phenylboronic acid pinacol ester (22)

[0178] Intermediate 2 (3.00 g, 22.9 mmol, 1.8 equiv), anhydrous tetrahydrofuran (30 mL), 4-hydroxyphenylboronic acid pinacol ester (2.80 g, 12.7 mmol), and tributylphosphine (9.50 mL, 38.1 mmol, 3.0 equiv) were placed in a 100 mL round-bottom flask in sequence. Subsequently, N,N,N',N'-tetramethylazodicarbonamide (6.60 g, 38.1 mmol, 3.0 equiv) was dissolved in 30 mL of anhydrous dichloromethane and added to a constant pressure dropping funnel. The air was replaced by nitrogen. The mixture was stirred at 0°C for 30 min and slowly added dropwise to the reaction bottle at a uniform rate. After the addition was complete, the mixture was moved to room temperature and stirred for 3 h. The reaction was monitored by TLC. After the reaction was complete, the solvent was distilled off under reduced pressure and subjected to column chromatography (200-300 mesh silica gel, DCM:MeOH=25:1) to obtain 3.8 g of a colorless oil with a yield of 89.8%. 1 HNMR(300MHz,Chloroform-d)δ7.72(d,J=8.2Hz,2H),6.87(d,J=8.2Hz,2H),4.10(t,J=5.6 Hz,2H),3.70(t,J=4.0Hz,4H),2.77(t,J=5.6Hz,2H),2.55(t,J=4.0Hz,4H),1.30(s,12H).

[0179] (2) Preparation of 4-(2-(4-azidophenoxy)ethyl)morpholine (23)

[0180] The intermediate 22 (1.8 g, 5.4 mmol) and copper acetate (0.54 g, 2.7 mmol, 0.5 equiv) were sequentially placed in a 100 mL round-bottom flask, and 15 mL of acetonitrile and 15 mL of water were added to dissolve. Sodium azide (1.8 g, 27.0 mmol, 5.0 equiv) was slowly added to the reaction flask and stirred at room temperature for 6 h. The reaction was monitored by TLC. After the reaction was complete, dichloromethane and water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered and then distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, DCM:MeOH=30:1) was performed to obtain 0.73 g of a reddish brown liquid with a yield of 54.4%. 1 H NMR(300MHz,Chloroform-d)δδ6.88(d,J=9.0Hz,2H),6.82(d,J=9.0Hz,2H),4.01(t ,J=5.7Hz,2H),3.67(t,J=4.7Hz,4H),2.72(t,J=5.7Hz,2H),2.50(t,J=4.7Hz,4H).

[0181] (3) Preparation of methyl 2-(5-methyl-1-(4-(2-morpholinethoxy)phenyl)-1H-1,2,3-triazol-4-yl)acetate (24)

[0182] Intermediate 23 (0.57 g, 2.3 mmol), dimethyl sulfoxide (5 mL), methyl levulinate (0.99 mL, 8.0 mmol, 3.5 equiv), and tetrahydropyrrole (0.19 mL, 2.3 mmol, 1.0 equiv) were sequentially placed in a 25 mL round-bottom flask. Stirred at 80°C for 48 h, and the reaction was monitored by TLC. When the reaction was complete, dichloromethane and a large amount of water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered, and then distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, DCM:MeOH=20:1) was performed to obtain 0.29 g of a white solid with a yield of 35.0%. 1 H NMR(300MHz,Chloroform-d)δ7.35(d,J=8.6Hz,2H),7.02(d,J=8.6Hz,2H),4.16(t,J=5.6Hz,2H),3.79 (s,2H),3.75(t,J=4.1Hz,4H),3.73(s,3H),2.84(t,J=5.6Hz,2H),2.60(t,J=4.1Hz,4H),2.25(s,3H).

[0183] (4) Preparation of 2-(5-methyl-1-(4-(2-morpholinethoxy)phenyl)-1H-1,2,3-triazol-4-yl)acetic acid (25)

[0184] Intermediate 24 (0.22 g, 0.60 mmol), methanol (9 mL), water (3 mL), and lithium hydroxide (0.072 g, 3.0 mmol, 5.0 equiv) were sequentially placed in a 25 mL round-bottom flask. The temperature was raised to 60°C and stirred for 12 h. The reaction was monitored by TLC. After the reaction was complete, the mixture was evaporated under reduced pressure, and then ethyl acetate and water were added. The pH was adjusted to acidic conditions, and the organic phase was dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure. The solvent was evaporated and column chromatography (200-300 mesh silica gel, DCM:MeOH=2:1) ​​was performed to obtain 0.06 g of a white solid with a yield of 28.9%. 1 H NMR(300MHz,D2O-d2)δ7.50(d,J=8.8Hz,2H),7.23(d,J=8.8Hz,2H),4.37(t,J=4.8Hz,2H),3 .88(t,J=4.4Hz,4H),3.67(s,2H),3.14(t,J=4.8Hz,2H),2.92(t,J=4.4Hz,4H),2.24(s,3H).

[0185] (5) Preparation of N-benzyl-2-(5-methyl-1-(4-(2-morpholinethoxy)phenyl)1H-1,2,3-triazol-4-yl)acetamide (26)

[0186] Intermediate 25 (0.050 g, 0.14 mmol), DMF (4 mL), benzylamine (0.032 mL, 0.29 mmol, 2.0 equiv), 1-hydroxybenzotriazole (HOBt, 0.039 g, 0.29 mmol, 2.0 equiv), 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCI, 0.056 g, 0.29 mmol, 2.0 equiv), and triethylamine (0.081 mL, 0.58 mmol, 4.0 equiv) were placed in a 25 mL round-bottom flask in sequence. The mixture was heated to 40°C and stirred for 6 hours. The reaction was monitored by TLC. After the reaction was complete, the mixture was distilled under reduced pressure, and then dichloromethane and water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered, and distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, DCM:MeOH=20:1) was performed to obtain 0.020 g of a white solid with a yield of 32.8%. HPLC purity: 100.00%, melting point: 115.6-116.7°C. 1H NMR(300MHz,Chloroform-d)δ7.39–7.27(m,7H),7.07(d,J=8.7Hz,2H),4.49(d,J=5.8Hz,2H),4.20(t,J= 5.6Hz,2H),3.78(t,J=4.5Hz,2H),3.73(s,2H),2.87(t,J=5.6Hz,2H),2.63(t,J=4.5Hz,2H),2.30(s,3H). 13 C NMR(75MHz,Chloroform-d)δ168.9,158.9,139.4,139.3,131.7,129.3,128.3(2C),127.3(2C),126 .8,126.3(2C),115.2(2C),66.2(2C),65.7,57.0,53.6(2C),42.3,32.4,8.5.HRMS(ESI)m / z:calcd for C 24 H 29 N5O3H + [M+H] + :436.23794, found:436.23701.

[0187] Embodiment 19

[0188] Preparation of N-benzyl-3-(1-(4-(2-morpholinethoxy)phenyl)-1H-1,2,3-triazol-4-yl)propanamide (29)

[0189]

[0190] (1) Preparation of N-benzyl-4-ynylpentanamide (28)

[0191] 1-Hydroxybenzotriazole (1.0 g, 7.6 mmol, 1.5 equiv), anhydrous dichloromethane (25 mL), 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (1.5 g, 7.6 mmol, 1.5 equiv) were placed in a 50 mL eggplant-shaped bottle and stirred at room temperature for 20 min. 4-Pentynoic acid (Compound 27, 0.50 g, 5.1 mmol) was dissolved in 2 mL anhydrous dichloromethane and added to the reaction solution and stirred for 10 min. Benzylamine (2 equiv) was dissolved in 5 mL dichloromethane and added to the reaction solution and stirred for 10 min. Triethylamine (1.42 mL, 10.2 mmol, 2.0 equiv) was added to the reaction bottle at 0°C and stirred for 1 h. Subsequently, the mixture was stirred at 40°C for 24 h. The reaction was monitored by TLC. After the reaction was complete, the solvent was distilled off under reduced pressure to obtain 0.90 g of a white solid with a yield of 94.2%. The mixture was directly used for the next step.

[0192] (2) Preparation of N-benzyl-3-(1-(4-(2-morpholinethoxy)phenyl)-1H-1,2,3-triazol-4-yl)propanamide (29)

[0193] Intermediate 28 (0.27 g, 1.4 mmol, 1.2 equiv), tert-butyl alcohol (4 mL), water (1 mL), intermediate 23 (0.30 g, 1.2 mmol), copper sulfate pentahydrate (0.15 g, 0.60 mmol, 0.5 equiv), sodium ascorbate (0.36 g, 1.8 mmol, 1.5 equiv) were sequentially placed in a 25 mL round-bottom flask and stirred at room temperature for 12 h. The reaction was monitored by TLC. After the reaction was complete, dichloromethane and water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered, and then distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, DCM:MeOH=20:1) was performed to obtain 0.035 g of a white solid with a yield of 6.7%. HPLC purity: 99.62%, melting point: 144.3-145.4°C. 1 H NMR(300MHz,Chloroform-d)δ7.69(s,1H),7.53(d,J=8.8Hz,2H),7.25–7.11(m,5H),6.99(d,J=8.8Hz,2H),6.40(t,J=4.5Hz,1H),4.38(d,J=5.7 Hz,2H),4.14(t,J=5.6Hz,2H),3.73(t,J=4.4Hz,4H),3.11(t,J=6.8Hz,2H),2.82(t,J=5.6Hz,2H),2.68(t,J=6.8Hz,2H),2.58(t,J=4.4Hz,4H). 13 C NMR(75MHz,Chloroform-d)δ171.9,158.8,147.0,138.3,130.7,128.6(2C),127.6(2C),127.4,122 .0(2C),120.0,115.4(2C),66.9(2C),66.2,57.6,54.2(2C),43.5,35.7,21.5.HRMS(ESI)m / z:calcd for C 24 H 29 N5O3H + [M+H] + :436.23794,found:436.23674.

[0194] Embodiment 20

[0195] Preparation of N-benzyl-2-(4-(4-(2-morpholinethoxy)phenyl)-1H-1,2,3-triazol-1-yl)acetamide (35)

[0196]

[0197] (1) Preparation of ((4-bromophenoxy)ethyl)-2-morpholine (31)

[0198] 4-Bromophenol (compound 30, 1.5 g, 8.7 mmol), anhydrous tetrahydrofuran (20 mL), 2-morpholinoethanol (compound 2, 1.7 g, 13 mmol, 1.5 equiv), and tributylphosphine (6.0 mL, 24.0 mmol, 3.0 equiv) were sequentially placed in a 100 mL round-bottom flask. Subsequently, N,N,N',N'-tetramethylazodicarbonamide (4.1 g, 24.0 mmol, 3.0 equiv) was dissolved in a constant pressure dropping funnel filled with 30 mL of anhydrous dichloromethane. The air was replaced by nitrogen. The mixture was stirred at 0°C for 30 min and then slowly added dropwise to the reaction flask at a uniform speed. After the addition was complete, the mixture was moved to room temperature and stirred for 3 h. The reaction was monitored by TLC. After the reaction was complete, the solvent was distilled off under reduced pressure and column chromatography (200-300 mesh silica gel, DCM:MeOH=25:1) was performed to obtain 2.0 g of a colorless oil with a yield of 80.3%. 1 H NMR(300MHz,Chloroform-d)δ7.35(d,J=8.9Hz,2H),6.77(d,J=8.9Hz,2H),4.05(t ,J=5.7Hz,2H),3.71(t,J=4.6Hz,4H),2.77(t,J=5.7Hz,2H),2.55(t,J=4.6Hz,4H).

[0199] (2) Preparation of (4-((trimethylsilyl)ethynyl)phenoxyethoxy)-2-morpholine (32)

[0200] Intermediate 31 (0.20 g, 0.70 mmol), trimethylsilyl acetylene (0.99 mL, 7.0 mmol, 10.0 equiv), 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl (X-Phos, 0.33 g, 0.7 mmol, 1.0 equiv), cuprous iodide (0.067 g, 0.35 mmol, 0.5 equiv), and tris(dibenzylideneacetone)dipalladium (Pd2(dba)3, 0.064 g, 0.070 mmol, 0.1 equiv) were placed in a 25 mL round-bottom flask in sequence. Subsequently, 10 mL of anhydrous triethylamine was added to dissolve, and the air was replaced by nitrogen. The mixture was stirred at 80° C. for 12 h and the reaction was monitored by TLC. When the reaction was complete, dichloromethane and water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered and distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, PE:EA=1:1) was performed to obtain 0.10 g of a white solid with a yield of 49.0%. 1 H NMR(300MHz,Chloroform-d)δ7.39(d,J=8.4Hz,2H),6.81(d,J=8.4Hz,2H),4.10(t,J=5.7 Hz,2H),3.73(t,J=4.8Hz,4H),2.79(t,J=5.7Hz,2H),2.57(t,J=4.8Hz,4H),0.23(s,9H).

[0201] (3) Preparation of ((4-ethynylphenoxy)ethyl)-2-morpholine (33)

[0202] Intermediate 32 (0.104 g, 0.34 mmol) and potassium carbonate (0.19 g, 1.4 mmol, 4.0 equiv) were sequentially placed into a 25 mL round-bottom flask, and 5 mL of methanol was added to dissolve. The mixture was stirred at room temperature for 4 h and the reaction was monitored by TLC. After the reaction was complete, the solvent was evaporated under reduced pressure and the mixture was purified by column chromatography (200-300 mesh silica gel, DCM:MeOH=25:1) to obtain 0.060 g of a white solid with a yield of 76.3%. 1 H NMR(300MHz,Chloroform-d)δ7.42(d,J=8.4Hz,2H), 6.84(d,J=8.4Hz,2H), 4.11(t,J=5.7 Hz,2H), 3.74(t,J=4.5Hz,4H), 3.00(s,1H), 2.80(t,J=5.7Hz,2H), 2.58(t,J=4.5Hz,4H).

[0203] (4) Preparation of 2-azido-N-benzylacetamide (34)

[0204] Intermediate 20a (0.50 g, 2.2 mmol) was placed in a 25 mL round-bottom flask, and then 5 mL of DMF was added to dissolve it. Sodium azide (0.45 g, 6.6 mmol, 3.0 equiv) was slowly added to the reaction flask and stirred at room temperature for 6 h. The reaction was monitored by TLC. After the reaction was complete, dichloromethane and water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered and then distilled under reduced pressure. The solvent was evaporated and distilled under reduced pressure to obtain 0.36 g of a colorless oily liquid with a yield of 86.0%. 1 H NMR (300MHz, DMSO-d6) δ8.64(t,J=6.0Hz,1H),7.38–7.21(m,5H),4.32(d,J=6.0Hz,2H),3.90(s,2H).

[0205] (5) Preparation of N-benzyl-2-(4-(4-(2-morpholinethoxy)phenyl)-1H-1,2,3-triazol-1-yl)acetamide (35)

[0206] Intermediate 33 (0.10 g, 0.43 mmol, 1.2 equiv), intermediate 34 (0.069 g, 0.36 mmol), copper sulfate pentahydrate (0.045 g, 0.18 mmol, 0.5 equiv), sodium ascorbate (0.107 g, 0.54 mmol, 1.5 equiv) were sequentially placed in a 25 mL round-bottom flask, 4 mL of tert-butyl alcohol and 1 mL of water were added for dissolution, and the mixture was stirred at room temperature for 12 h. The reaction was monitored by TLC. After the reaction was complete, dichloromethane and water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered, and then distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, DCM:MeOH=20:1) was performed to obtain 0.035 g of a white solid with a yield of 23.1%. HPLC purity: 98.39%, melting point: 138.9-140.6°C. 1 H NMR(300MHz,Chloroform-d)δ8.83(t,J=5.8Hz,1H),8.42(s,1H),7.77(d,J=8.4Hz,2H),7.38–7.23(m,5H),7.02(d,J=8.0Hz,2H ), 5.19 (s, 2H), 4.34 (d, J = 5.8Hz, 2H), 4.13 (t, J = 5.3Hz, 2H), 3.58 (t, J = 5.8Hz, 4H), 3.24 (t, J = 5.8Hz, 4H), 2.71 (t, J = 5.3Hz, 2H). 13C NMR(75MHz,Chloroform-d)δ165.3,159.1,148.3,137.3,128.9(2C),127.8(2C),127.2(2C),123 .0,120.6,115.1(2C),77.4,67.0(2C),66.0,57.72,54.2(2C),53.2,43.9.HRMS(ESI)m / z:calcd for C 23 H 27 N5O3H + [M+H] + :422.22202,found:422.22026.

[0207] Embodiment 21

[0208] Preparation of N-benzyl-2-(4-(4-(2-morpholinethoxy)phenyl)-1H-indol-1-yl)acetamide (38)

[0209]

[0210] (1) Preparation of 2-(4-(1H-indol-4-yl)phenoxy)ethylmorpholine (37)

[0211] Compound 36 (0.30 g, 1.5 mmol), intermediate 22 (1.0 g, 3.0 mmol, 2.0 equiv), 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl (0.14 g, 0.30 mmol, 0.2 equiv), tris(dibenzylideneacetone)dipalladium (0.14 g, 0.15 mmol, 0.1 equiv) and cesium fluoride (1.14 g, 7.5 mmol, 5.0 equiv) were added sequentially into a 50 mL round-bottom flask. Subsequently, 1,4-dioxane (12 mL) and water (3 mL) were added, and the air was replaced by nitrogen. The mixture was heated and stirred at 90° C. for 12 h. The reaction was monitored by TLC. After the reaction was complete, dichloromethane and water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered, and then distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, PE:EA=1:1) was performed to obtain 0.40 g of a white solid with a yield of 82.7%. 1H NMR(300MHz,Chloroform-d)δ8.28(brs,1H),7.63(d,J=8.4Hz,2H),7.37(d,J=8.1Hz,1H),7.26(t,J=7.8Hz,3H),7.03( d,J=8.4Hz,2H),6.72(s,1H),4.21(t,J=5.7Hz,2H),3.77(t,J=4.8Hz,4H),2.87(t,J=5.7Hz,2H),2.65(t,J=4.8Hz,4H).

[0212] (2) Preparation of N-benzyl-2-(4-(4-(2-morpholinethoxy)phenyl)-1H-indol-1-yl)acetamide (38)

[0213] Intermediate 37 (0.40 g, 1.2 mmol) and intermediate 20a (0.68 g, 3.0 mmol, 2.5 equiv) were sequentially placed in a 25 mL round-bottom flask. Subsequently, 4 mL of DMF was added for dissolution, and sodium hydride (0.25 g, 7.20 mmol, 6.0 equiv) was slowly added to the reaction flask, stirred at room temperature for 12 h, and the reaction was monitored by TLC. When the reaction was complete, dichloromethane and water were added, extracted, and the organic phase was dried over anhydrous sodium sulfate, filtered, and then distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, PE:EA=1:6) was performed to obtain 0.20 g of a white solid with a yield of 35.5%. HPLC purity: 95.77%, melting point: 128.2-129.5°C. 1 H NMR(300MHz,DMSO-d6)δ7.65(d,J=8.0Hz,2H),7.41–7.21(m,6H),7.15–7.05(m,5H),6.79(s,1H),6.00(brs,1 H), 4.85 (s, 2H), 4.40 (d, J = 5.4Hz, 2H), 4.22 (t, J = 4.8Hz, 2H), 3.79 (m, 4H), 2.90 (t, J = 4.8Hz, 2H), 2.65 (m, 4H). 13 C NMR(75MHz,DMSO-d6)δ168.2,158.1,137.6,136.7,134.7,133.4,129.8(2C),128.6(2C),128.4,127.5,127.3(2C) ,127.0,123.1,120.2,114.7(2C),108.0,103.1,66.9(2C),65.9,57.7,54.1(2C),50.2,43.1.HRMS(ESI)m / z:calcd forC 29 H 31N3O3H + [M+H] + :470.24384,found:470.24364.

[0214] Embodiment 22

[0215] Preparation of N-benzyl-2-(5-(thiophen-2-yl)pyridin-2-yl)acetamide (41a)

[0216]

[0217] (1) Preparation of N-benzyl-2-(5-bromopyridin-2-yl)acetamide (40)

[0218] 2-(5-Bromopyridin-2-yl)acetic acid (Compound 39) (1.0 g, 4.63 mmol), benzylamine (1.5 mL, 13.9 mmol, 3.0 equiv), 1-hydroxybenzotriazole (0.81 g, 6.02 mmol, 1.3 equiv), 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (1.3 g, 6.9 mmol, 1.5 equiv), and 4-dimethylaminopyridine (0.17 g, 1.4 mmol, 0.3 equiv) were placed in a 50 mL round-bottom flask in sequence. Subsequently, 20 mL of DMF was added to dissolve, and the mixture was stirred at 80° C. for 12 h. The reaction was monitored by TLC. When the reaction was complete, dichloromethane and water were added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered, and distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, PE:EA=1:1) was performed to obtain 1.00 g of a white solid with a yield of 70.8%. 1 H NMR (300MHz, DMSO-d6) δ8.57(s,1H),7.95(d,J=8.4Hz,1H),7.33–7.16(m,6H),4.24(d,J=5.7Hz,2H),3.64(s,2H).

[0219] (2) Preparation of N-benzyl-2-(5-(thiophen-2-yl)pyridin-2-yl)acetamide (41a)

[0220] Intermediate 40 (0.10 g, 0.33 mmol), 2-thiopheneboronic acid (0.051 g, 0.40 mmol, 1.2 equiv), 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl (0.031 g, 0.066 mmol, 0.2 equiv), tris(dibenzylideneacetone)dipalladium (0.030 g, 0.033 mmol, 0.1 equiv) and cesium fluoride (0.25 g, 1.7 mmol, 5.0 equiv) were placed in a 25 mL round-bottom flask in sequence. Then, 4 mL of 1,4-dioxane and 1 mL of water were added to dissolve, and the air was replaced by nitrogen. Then, the mixture was heated and stirred at 90°C for 12 h. The reaction was monitored by TLC. When the reaction was complete, dichloromethane and water were added to extract the mixture. The organic phase was dried over anhydrous sodium sulfate, filtered, and then distilled under reduced pressure to remove the solvent. Column chromatography (200-300 mesh silica gel, PE:EA=1:1) was performed to obtain 0.035 g of a white solid with a yield of 34.4%. HPLC purity: 100.00%, melting point: 132.7-134.0°C. 1 H NMR(300MHz,Chloroform-d)δ8.79(s,1H),7.87(d,J=6.0Hz,1H),7.72(brs,1H),7.40–7.28(m,8H),7.16(s,1H),4.51(s,2H),3.83(s,2H). 13 C NMR(75MHz,Chloroform-d)δ169.2,154.4,146.3,134.0,138.4,134.2,129.0,128.8,128.7(2C),12 8.4,127.87,127.6(2C),127.5,127.4,126.1,124.3,124.1,77.4,45.0,43.6.HRMS(ESI)m / z:calcd for C 18 H 16 N2OSH + [M+H] + :309.10616,found:309.09891.

[0221] Embodiment 23

[0222] Preparation of N-benzyl-2-(5-(furan-3-yl)pyridin-2-yl)acetamide (41b)

[0223]

[0224] The same procedure as step (2) in Example 22 was followed except that 3-furanboronic acid was used instead of 2-thiopheneboronic acid to obtain 0.032 g of a white solid with a yield of 33.2%. HPLC purity: 99.39%, melting point: 127.7-129.1°C. 1 H NMR(300MHz,Chloroform-d)δ8.70(s,1H),7.85–7.69(m,3H),7.58(s,1H),7.40–7.28(d,J=8.4Hz,6H),6.75(s,1H),4.53(d,J=4.9Hz,2H),3.86(s,2H). 13 C NMR(75MHz,Chloroform-d)δ169.3,154.1,146.5,144.3,139.0,138.5,134.3,128.7(2C ),127.6(2C),127.4,127.0,124.2,122.9,108.5,77.4,45.1,43.6.HRMS(ESI)m / z:calcd for C 18 H 16 N2O2H + [M+H] + :293.12900,found:293.12210.

[0225] Embodiment 24

[0226] Preparation of N-benzyl-2-(5-(furan-2-yl)pyridin-2-yl)acetamide (41c)

[0227]

[0228] The same procedure as step (2) in Example 22 was followed except that 2-furanboronic acid was used instead of 2-thiopheneboronic acid to obtain 0.028 g of a white solid with a yield of 29.0%. HPLC purity: 99.40%, melting point: 119.8-121.1°C. 1 H NMR(300MHz,Chloroform-d)δ8.88(s,1H),7.97(dd,J=7.9,2.4Hz,1H),7.73(brs,1H),7.5 8(s,1H),7.41–7.27(m,6H),6.79(s,1H),6.57(s,1H),4.54(d,J=5.7Hz,2H),3.86(s,2H). 13C NMR(75MHz,Chloroform-d)δ169.2,154.1,150.8,144.8,143.1,138.4,132.0,128.7( 2C),127.6(2C),127.4,125.5,124.0,112.0,106.5,45.1,43.6.HRMS(ESI)m / z:calcd forC 18 H 16 N2O2Na + [M+Na] + :315.11047,found:315.11133.

[0229] Embodiment 25

[0230] Preparation of N-benzyl-2-(5-(thiophen-3-yl)pyridin-2-yl)acetamide (41d)

[0231]

[0232] The same procedure as step (2) in Example 22 was followed except that 3-thiopheneboronic acid was used instead of 2-thiopheneboronic acid to obtain 0.030 g of a white solid with a yield of 29.5%. HPLC purity: 100.00%, melting point: 142.4-143.8°C. 1 H NMR(300MHz,Chloroform-d)δ8.80(s,1H),7.88(dd,J=8.1,2.4Hz,1H),7.71(brs,1H),7.54(d,J= 2.4Hz,1H),7.50-7.47(m,1H),7.41(m,1H),7.37-7.30(m,6H),4.52(d,J=5.7Hz,2H),3.85(s,2H). 13 CNMR(75MHz,Chloroform-d)δ169.3,154.1,147.0,138.4,138.4,134.8,130.2,128.7 (2C),127.6(2C),127.4,127.2,125.9,124.2,121.5,45.0,43.6.HRMS(ESI)m / z:calcd for C 18 H 16 N2OS + [M+Na] + :331.08767,found:331.08882.

[0233] Embodiment 26

[0234] Preparation of N-benzyl-2-(5-(isoxazol-4-yl)pyridin-2-yl)acetamide (41e)

[0235]

[0236] The same procedure as step (2) of Example 22 was used except that isoxazole-4-boronic acid was used instead of 2-thiopheneboronic acid, 1,1'-bis(di-tert-butylphosphino)ferrocenedichloropalladium was used instead of tris(dibenzylideneacetone)dipalladium and 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl, and the temperature was changed to 40°C to obtain 0.022 g of a white solid with a yield of 22.7%. HPLC purity: 99.64%, melting point: 210.7-211.9°C. 1 H NMR(300MHz,DMSO-d6)δ9.53(s,1H),9.23(s,1H),8.85(s,1H),8.60(brs,1H),8.05(dd,J =8.1,2.3Hz,1H),7.43(d,J=8.1Hz,1H),7.29(m,5H),4.30(d,J=5.9Hz,2H),3.72(s,2H). 13 C NMR(75MHz,DMSO-d6)δ169.1,155.7,155.5,148.1,146.6,139.4,133.9,128.3 (2C),127.2(2C),126.8,124.0,122.7,117.7,44.6,42.3.HRMS(ESI)m / z:calcd for C 17 H 15 N3O2Na + [M+Na] + :316.10567,found:316.10783.

[0237] Embodiment 27

[0238] Preparation of N-benzyl-2-(5-(N-methylpyrrol-2-yl)pyridin-2-yl)acetamide (41f)

[0239]

[0240] The same procedure as step (2) in Example 22 was followed except that N-methylpyrrole-2-boronic acid pinacol ester was used instead of 2-thiopheneboronic acid to obtain 0.031 g of a white solid with a yield of 30.8%. HPLC purity: 100.00%, melting point: 65.2-66.4°C. 1HNMR(300MHz,Chloroform-d)δδ8.65(s,1H),7.79–7.73(m,2H),7.39(d,J=8.2Hz,2H),7.44– 7.29(m,4H),6.84(s,1H),6.39–6.26(m,2H),4.56(d,J=5.8Hz,2H),3.88(s,2H),3.73(s,3H). 13 C NMR(75MHz,Chloroform-d)δ169.3,153.6,148.6,138.5,136.6,130.5,128.7(2C),127. 9,127.6(2C),127.4,125.0,123.7,110.0,108.4,45.1,43.6,35.2.HRMS(ESI)m / z:calcd for C 19 H 19 N3OH + [M+H] + :306.16012,found:306.16188.

[0241] Embodiment 28

[0242] Preparation of N-benzyl-2-(5-(pyrazol-4-yl)pyridin-2-yl)acetamide (41 g)

[0243]

[0244] The same procedure as step (2) in Example 22 was followed except that 4-pyrazole boronic acid pinacol ester was used instead of 2-thiophene boronic acid to obtain 0.035 g of a white solid with a yield of 36.3%. HPLC purity: 100.00%, melting point: 215.3-216.3°C. 1 H NMR (300MHz, DMSO-d6) δ13.05(s,1H),8.78(s,1H),8.60(t,J=6.0Hz,1H),8.15(s,1H ),7.94(dd,J=8.1,2.3Hz,1H),7.34-7.20(m,7H),4.30(d,J=6.0Hz,2H),3.68(s,2H). 13 C NMR(75MHz,DMSO-d6)δ169.8,154.0,146.1,139.9,133.1,128.7(2C),127.7(2C),127.3,127.2,124.2,118.2,45.0,42.7.HRMS(ESI)m / z:calcd for C 17 H 16 N4A+ [M+Na] + :315.12167,found:315.12366.

[0245] Embodiment 29

[0246] Preparation of N-benzyl-2-(5-(3,5-dimethylisoxazol-4-yl)pyridin-2-yl)acetamide (41h)

[0247]

[0248] The same operation steps as step (2) in Example 22 were carried out except that 3,5-dimethylisoxazole-4-boric acid was used instead of 2-thiopheneboric acid, tetrakis(triphenylphosphine)palladium and sodium carbonate were used instead of tris(dibenzylideneacetone)dipalladium and 2-dicyclohexylphosphine-2',4',6'-triisopropylbiphenyl, and sodium carbonate was used instead of cesium fluoride to obtain 0.038 g of a white solid with a yield of 35.8%. HPLC purity: 96.86%, melting point: 79.4-80.5°C. 1 H NMR (300MHz, DMSO-d6) δ8.67(t,J=6.0Hz,1H),8.53(s,1H),7.80(d,J=8.0Hz,1H),7.46(d,J= 8.0Hz,1H),7.36-7.20(m,5H),4.31(d,J=6.0Hz,2H),3.74(s,2H),2.42(s,3H),2.24(s,3H). 13 C NMR(75MHz,DMSO-d6)δ169.1,166.0,158.4,155.5,148.7,139.5,136.7,128.4(2C), 127.3(2C),126.9,124.0,124.0,113.0,44.7,42.3,11.4,10.5.HRMS(ESI)m / z:calcd for C 19 H 19 N3O2Na + [M+Na] + :344.13750,found:344.13693.

[0249] Embodiment 30

[0250] Preparation of N-benzyl-2-(5-(thiazol-5-yl)pyridin-2-yl)acetamide (41i)

[0251]

[0252] The same operation steps as step (2) in Example 22 were carried out except that thiazole-5-boronic acid pinacol ester was used instead of 2-thiopheneboric acid, tetrakis(triphenylphosphine)palladium was used instead of tris(dibenzylideneacetone)dipalladium and 2-dicyclohexylphosphine-2',4',6'-triisopropylbiphenyl, and sodium carbonate was used instead of cesium fluoride to obtain 0.032 g of a white solid with a yield of 31.3%. HPLC purity: 100.00%, melting point: 151.7-153.3°C. 1 H NMR(300MHz,Chloroform-d)δ8.90(s,1H),8.80(s,1H),8.18(s,1H),7.91(d,J=7 .8Hz,1H),7.64(brs,1H),7.50–7.28(m,9H),4.55(d,J=5.3Hz,2H),3.89(s,2H). 13 CNMR(75MHz,Chloroform-d)δ168.9,155.7,153.2,147.1,140.1,138.4,135.3,128.7(2C),127.7(2C),127.5,126.0,124.3,45.2,43.7.HRMS(ESI)m / z:calcd for C 27 H 15 N3OS + [M+Na] + :332.08335,found:332.08270.

[0253] Embodiment 31

[0254] Preparation of N-benzyl-2-(5-(1-methyl-1H-imidazol-5-yl)pyridin-2-yl)acetamide (41j)

[0255]

[0256] The same operation steps as step (2) in Example 22 were carried out except that 1-methyl-1-H-imidazole-5-boronic acid pinacol ester was used instead of 2-thiopheneboric acid, tetrakis(triphenylphosphine)palladium was used instead of tris(dibenzylideneacetone)dipalladium and 2-dicyclohexylphosphine-2',4',6'-triisopropylbiphenyl, and sodium carbonate was used instead of cesium fluoride to obtain 0.035 g of a white solid with a yield of 34.6%. HPLC purity: 100.00%, melting point: 119.0-120.7°C. 1H NMR(300MHz,Chloroform-d)δ8.65(d,J=2.3Hz,1H),7.77(dd,J=8.0,2.3Hz,1H),7.65(s,1H),7.47 (d,J=8.0Hz,1H),7.41–7.30(m,7H),7.24(s,1H),4.56(d,J=5.8Hz,2H),3.91(s,2H),3.75(s,3H). 13 C NMR(75MHz,Chloroform-d)δ167.9,154.0,147.3,139.0,137.3,135.5,128.2,12 7.6(2C),126.6(2C),126.3,123.4,123.0,44.1,42.6,31.6.HRMS(ESI)m / z:calcd for C 18 H 18 N4A + [M+Na] + :329.13783,found:329.13765.

[0257] Embodiment 32

[0258] Preparation of N-benzyl-2-(5-(2-methylpyridin-4-yl)pyridin-2-yl)acetamide (41k)

[0259]

[0260] The same operation steps as step (2) in Example 22 were carried out except that 2-methyl-4-pyridineboronic acid was used instead of 2-thiopheneboronic acid, tetrakis(triphenylphosphine)palladium was used instead of tris(dibenzylideneacetone)dipalladium and 2-dicyclohexylphosphine-2',4',6'-triisopropylbiphenyl, and sodium carbonate was used instead of cesium fluoride to obtain 0.034 g of a white solid with a yield of 32.5%. HPLC purity: 100.00%, melting point: 116.3-117.7°C. 1 H NMR(300MHz,Chloroform-d)δ8.82(d,J=2.4Hz,1H),8.65(d,J=5.3Hz,1H),7.97(dd,J=8.1,2.4Hz,1H),7.77(t,J= 5.8Hz,1H),7.50(d,J=8.1Hz,1H),7.41(s,1H),7.39–7.28(m,6H),4.55(d,J=5.8Hz,2H),3.92(s,2H),2.71(s,3H). 13C NMR(75MHz,Chloroform-d)δ168.9,159.4,156.2,149.9,147.5,145.1,138.3,135.5,132.6 ,128.7(2C),127.6(2C),127.4,124.3,121.0,118.6,45.1,43.6,24.6.HRMS(ESI)m / z:calcd forC 20 H 19 N3O + [M+Na] + :340.14258,found:340.14202.

[0261] Embodiment 33

[0262] Preparation of N-benzyl-2-(5-(N-methylpyrazol-4-yl)pyridin-2-yl)acetamide (411)

[0263]

[0264] The same operation steps as step (2) in Example 22 were carried out except that 1-methyl-1H-pyrazole-4-boric acid was used instead of 2-thiopheneboric acid, tetrakis(triphenylphosphine)palladium and tris(dibenzylideneacetone)dipalladium and 2-dicyclohexylphosphine-2',4',6'-triisopropylbiphenyl were used, and sodium carbonate was used instead of cesium fluoride to obtain 0.037 g of a white solid with a yield of 36.6%. HPLC purity: 100.00%, melting point: 145.7-147.0°C. 1 H NMR (300MHz, Chloroform-d) δ8.63 (s, 1H), 7.80–7.62 (m, 4H), 7.37–7.22 (m, 6H), 4.49 (d, J = 5.6Hz, 2H), 3.97 (s, 3H), 3.80 (s, 2H). 13 C NMR(75MHz,Chloroform-d)δ169.4,153.3,146.2,138.5,136.8,133.8,128.7(2C),1 27.6(2C),127.3,127.2,127.2,124.2,119.3,45.0,43.6,39.3.HRMS(ESI)m / z:calcd for C 18 H 18 N4A + [M+Na] + :329.13783,found:329.13691.

[0265] Below are some pharmacological experiments and results described in the present invention:

[0266] Experimental Example 1: Experiment on inhibition of tumor cell proliferation (MTT experiment)

[0267] (1) Experimental methods

[0268] The cell proliferation inhibitory activity of the compounds of the present invention on human liver cancer cell line (HepG2) and human cervical cancer cell line (HeLa) was tested. The above cell lines were frozen and passaged by the laboratory. Methods: The cells were inoculated in a 96-well flat-bottom plate (about 4000-6000 cells per well), incubated at 37°C, 5% CO2 for 24 hours, and then different concentrations of the test compound or positive control drug were added to each well. After culturing for another 72 hours, 50 μL of MTT PBS buffer (concentration of 1 mg / mL) was added to each well, incubated at 37°C for 4 hours, MTT and culture medium were removed, and 100 μL of DMSO was added to each well, and detected at 570 nm (Power Wave XS, Bio Tek, USA). Tirbanibulin (KX01) and colchicine were selected as positive control drugs. The data were statistically analyzed.

[0269] (2) Experimental results

[0270] The experimental results are shown in Table 1. The results show that this type of compounds generally have a strong cell growth inhibitory effect. The half inhibitory concentration (IC 50 ) reached the nanomolar level (nM). The cell growth inhibitory activity of these compounds was significantly better than that of doxorubicin.

[0271] Table 1 MTT test results of compounds

[0272]

[0273]

[0274] Note: Compounds KX01 and Cochicine in the table are positive control drugs and are not included in the scope of patent protection.

[0275] 2. Tubulin polymerization inhibition experiment

[0276] (1) Experimental methods

[0277] The inhibitory effect of the compounds of the present invention on tubulin polymerization was tested using the BK011P kit of cytoskeleton. A 96-well plate was placed on a multifunctional microplate reader (CLARIOstar, BMG LABTECH, Germany) and preheated at 37°C for 10 minutes. According to the instructions, paclitaxel solution (10×), CA-4 solution (10×), test compound solution (10×), buffer 1, GTP mother solution, tubulin glycerol buffer, and tubulin mother solution were prepared for use. According to the experiment, a suitable ratio was selected, and buffer 1, GTP mother solution, tubulin glycerol buffer, and tubulin mother solution were mixed to prepare a microtubule reaction solution, which was placed on an ice bath. Add 5 μL of control buffer, paclitaxel solution (3 μM), cochicine solution (5 μM), KX01 solution (5 μM), and test compound solution (41f, 5 μM, set up 1 replicate) to a 96-well plate, preheat at 37°C for 1 min on a microplate reader, quickly add the prepared microtube reaction solution, and record the change in fluorescence intensity within 60 minutes.

[0278] Preparation of buffer 1: ① Dissolve the substances in each bottle of buffer 1 with 10 mL of Milli-Q ultrapure water (two bottles in total); ② Mix the substances in the two bottles; ③ Divide the above buffer into 13 cryovials, with 1.5 mL in each tube, and store at -70°C.

[0279] Tubulin glycerol buffer, stored at 4°C.

[0280] Preparation of GTP mother solution: ① Dissolve the substance in each tube of GTP mother solution with 100 μL of ice-cold sterile distilled water (3 tubes in total); ② Place it on ice; ③ Mix the substances in the three tubes; ④ Divide the above solution into 13 cryotubes, 20 μL in each tube, and store at -70°C.

[0281] Preparation of tubulin solution: ① Place the vial containing freeze-dried tubulin on ice and place liquid nitrogen in a Dewar flask; ② Label 12 cryovials and place them on ice for use; ③ Defrost a 20μL GTP stock solution; ④ Mix 1.5mL of ice-cold buffer 1 with 15μL GTP stock solution; ⑤ Use 1.1mL of the above mixture to dissolve the freeze-dried tubulin powder; ⑥ Place on ice for 2 minutes until the powder is completely dissolved and suspended; ⑦ Divide the above tubulin solution into 12 cell cryovials, 88μL per tube, and quickly freeze in liquid nitrogen; ⑧ Store at -70℃.

[0282] Preparation of paclitaxel stock solution: dissolve paclitaxel in 100 μL DMSO and store at -70°C.

[0283] (2) Experimental results

[0284] The results showed that compound 41f had a significant inhibitory effect on tubulin polymerization. Figure 1 shown.

[0285] 3. Cell cycle experiment

[0286] (1) Experimental methods

[0287] The effect of the compound of the present invention on the cancer cell cycle (human cervical cancer cell line HeLa was selected) was tested by PI (propidium iodide) staining. Method: The cells were inoculated in a 6-well flat-bottom plate (about 100,000 cells per well), incubated at 37°C, 5% CO2 for 24 hours, and then different concentrations of 41f were added to each well (the final concentrations were 50nM, 80nM, and 100nM, respectively), and a blank control group was set up (the blank control group did not add compound 41f), and incubated at 37°C, 5% CO2 for 24 hours. The cells were collected, washed with precooled PBS (4°C, 0.01mol / L), and the cells were added to precooled (4°C) 70% (v / v) ethanol, and the fixed cells were placed at -20°C overnight. The cells were centrifuged, ethanol was removed, and the cells were washed with pre-cooled PBS (4°C, 0.01 mol / L). PI dye with RNase was added and stained at 4°C in the dark for 1 hour. The cells were centrifuged, the dye was removed, and the cells were washed with PBS. The cells were filtered through a 300-mesh filter and tested on a flow cytometer (BD FACSVerse, USA).

[0288] (2) Experimental results

[0289] The results are as follows Figure 2 As shown, Figure 2 It can be seen that compound 41f can significantly arrest cells in the G2 / M growth phase at 100 nM.

[0290] IV. Immunoblotting

[0291] Immunoblotting was used to test the effect of drugs on the content of phosphorylated Fak and phosphorylated Src proteins. The cells were seeded in a 6-well flat-bottom plate (about 100,000 cells per well), incubated at 37°C, 5% CO2 for 24 hours, and KX01, 41f (final drug concentration was 100 nM) were added to each well, and a blank control group (no compound was added to the blank control group) was set up and incubated at 37°C, 5% CO2 for different time periods (12, 24, 26, 48 hours). ① Extraction of total protein: HepG2 cells were lysed. ② Determination of protein content: The method used was BCA (Bicinchoninic Acid Assay). ③ SDS-PAGE (polyacrylamide gel electrophoresis) was used to separate proteins: the first step was to prepare separation gel and concentration gel; the second step was to load the sample and perform electrophoresis. ④ Transfer to membrane: Since the gel was too fragile to operate, the protein was transferred to a PVDF (polyvinylidene fluoride) membrane by electrotransfer. ⑤ Antigen-antibody immune reaction: add primary antibody after blocking, and then add secondary antibody for incubation. ⑥ Chemiluminescence, development, and fixation: add developer to develop the protein bands washed with TBST.

[0292] The results are as follows Figure 3 As shown, the levels of phosphorylated Fak protein and phosphorylated Src protein decreased with the extension of drug addition time within 48 hours.

Claims

1. A compound having a bicyclic acetamide structure, characterized in that: The general structural formula is shown in Formula I: Wherein C1 represents a benzene ring, indole, pyrrole, furan, thiophene, pyrazole, imidazole, isoxazole, thiazole, pyridine; C2 represents pyridine, triazole, imidazolinone, indole; R1 represents one or two of hydrogen, fluorine, chlorine, methyl, hydroxyl, cyano, methoxy, 2-nitromorpholinoethoxy, 4-morpholino; R2 represents hydrogen, fluorine, chlorine, methyl or methoxy; R3 represents L is hydrogen, fluorine, chlorine, bromine, methyl, or methoxy, X is C or N, and n=0, 1, 2, or 3.

2. The bicyclic acetamide structure compound according to claim 1, characterized in that for for R3 is 3. A biologically acceptable salt formed by the bicyclic acetamide structural compound described in claim 1 or 2 and at least one of acetic acid, dihydrofolic acid, benzoic acid, citric acid, sorbic acid, propionic acid, oxalic acid, fumaric acid, maleic acid, hydrochloric acid, malic acid, phosphoric acid, diphosphoric acid, hydrobromic acid, nitric acid, sulfurous acid, sulfuric acid, vanillic acid, tartaric acid, succinic acid, ascorbic acid, boric acid, lactic acid, p-toluenesulfonic acid, salicylic acid and ethylenediaminetetraacetic acid.

4. The method for preparing the bicyclic acetamide structure compound according to claim 1 or 2, characterized in that: The synthetic route of this type of compound is as follows: Synthesis route 1: The specific steps are: (1) Using morpholine as the starting material, it reacts with 2-bromoethanol through a nucleophilic substitution reaction to obtain compound 2; (2) Compound 3 reacts with di-tert-butyl dicarbonate to obtain compound 4; (3) Compound 2 and compound 4 are reacted to obtain compound 5; (4) Compound 5 is subjected to acidic conditions to remove the tert-butyloxycarbonyl protecting group to obtain compound 6; (5) Compound 7 reacts with morpholine under alkaline conditions to give compound 8 through nucleophilic substitution reaction; (6) Compound 8 is reduced by hydrogen to obtain compound 9; (7) Compound 10 reacts with di-tert-butyl dicarbonate under alkaline conditions to obtain compound 11; (8) Compound 11 is reduced by hydrogen to obtain compound 12; (9) Compound 6 or Compound 9 or Compound 12 or Compound 13 is condensed with 2-chloroethyl isocyanate to obtain Compounds 14a, 14b, 14c, and 14d; (10) Compounds 14a, 14b, 14c, and 14d were subjected to intramolecular cyclization under alkaline conditions to give compounds 15a, 15b, 15c, and 15d; (11) Compound 15a reacts with tert-butyl bromoacetate under alkaline conditions to give compound 16 through affinity substitution reaction; (12) Compound 16 is hydrolyzed under acidic conditions to obtain compound 17; (13) Compound 17 is reacted with R3NH2 to obtain intermediate 18; (14) reacting compound 19 with benzylamine or 3-chlorobenzylamine to obtain compound 20a or 20b; (15) reacting compound 15b with compound 20a or 20b to obtain 21a or 21b; (16) reacting compound 15c or 15d with compound 20a to obtain 21c or 21d; Synthesis route 2: (1) Compound 2 and 4-hydroxyphenylboronic acid pinacol ester are reacted to obtain compound 22; (2) Compound 22 reacts with sodium azide to obtain intermediate 23; (3) Compound 23 reacts with methyl 3-pentyne valerate to obtain compound 24; (4) Compound 24 is hydrolyzed under alkaline conditions to obtain compound 25; (5) Compound 25 is reacted with benzylamine to obtain the target product 26; Synthesis route three: (1) Compound 27 is reacted with benzylamine to obtain compound 28; (2) Compound 28 reacts with compound 23 to obtain target compound 29; Synthesis route 4: (1) Compound 2 and compound 30 were reacted to obtain compound 31; (2) Compound 31 is coupled with trimethylsilyl acetylene to obtain compound 32; (3) Compound 32 was subjected to alkaline conditions to remove the trimethylsilyl group to obtain compound 33; (4) Compound 20a reacts with sodium azide under alkaline conditions to obtain compound 34; (5) Compound 33 and compound 34 were subjected to click reaction to obtain target compound 35; Synthesis route 5: (1) Compound 36 and compound 22 were reacted by Suzuki reaction to obtain compound 37; (2) Compound 37 and compound 20a were reacted under alkaline conditions to obtain the target compound 38; Synthesis route 6: (1) Compound 39 is condensed with benzylamine to obtain compound 40; (2) Compound 40 is subjected to coupling reaction with a substituted heterocyclic aromatic boronic acid to obtain the target compound 41.

5. A pharmaceutical composition, characterized in that Containing the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

6. Use of the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof in the preparation of a microtubule polymerization inhibitor.

7. Use of the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for preventing and / or treating a tumor or a proliferative disease other than a tumor.

8. Use of the compound according to claim 7 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for preventing and / or treating tumors or proliferative diseases other than tumors, characterized in that: The tumor disease is breast cancer, ovarian cancer, non-small cell lung cancer, liver cancer, pancreatic cancer, prostate cancer, melanoma or leukemia.

9. Use of the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof in the preparation of a Fak protein and a Src protein phosphorylation inhibitor.

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