Five-membered nitrogen heterocyclic derivative as well as preparation method and application thereof

Abstract

The invention belongs to the field of organic compounds, and particularly relates to five-membered nitrogen heterocyclic derivatives as well as a preparation method and application thereof. According to the specific technical scheme, the structural general formula of the five-membered nitrogen heterocyclic compound is # imgabs0, a novel method for preparing the sulfur-containing or oxygen-containing five-membered nitrogen heterocyclic derivative is provided, a series of novel sulfur-containing or oxygen-containing five-membered nitrogen heterocyclic derivatives are synthesized, and the five-membered nitrogen heterocyclic compound has biological activity and anti-tumor activity and has wide medical and treatment application prospects.

Description

Technical Field

[0001] The invention belongs to the field of organic compounds, and specifically relates to a class of five-membered nitrogen heterocyclic derivatives and a preparation method and application thereof. Background Art

[0002] More than 97% of human genome sequences contain non-protein coding DNA, which will be transcribed into RNA and play an important biological function. RNA therapy is a new disease treatment method that has emerged in recent decades. At present, this treatment method is mainly focused on large molecule biological drugs and antisense nucleic acids. There are currently no small molecule drugs on the market in the field of non-coding RNA therapy. Therefore, whether it is basic scientific research or new drug development, the field of non-coding RNA therapy has broad and far-reaching prospects.

[0003] Currently reported chemical small molecule compounds that interfere with TRBP include Enoxacin, Schisandrin (-)-GomisinM1, and CIB-3B compounds that target the interaction between TRBP and Dicer enzyme. Most of them have moderate activity and are in the micromolar range, making it difficult to achieve clinical therapeutic value.

[0004] If new chemical small molecule compounds that interfere with TRBP can be provided, it will have great application potential. Summary of the invention

[0005] The purpose of the present invention is to provide a five-membered nitrogen heterocyclic derivative and a preparation method and application thereof.

[0006] To achieve the above-mentioned purpose, the technical solution adopted by the present invention is: a five-membered nitrogen heterocyclic compound, its conformational isomers, optical isomers and pharmaceutically acceptable salts. The general structural formula of the five-membered nitrogen heterocyclic compound is shown in Formula I:

[0007]

[0008] Wherein, the six-membered ring is selected from an aromatic ring, a cyclohexane ring, and a piperidine ring; X is selected from O and S; W is selected from C, CH, and N; and n is selected from any integer from 0 to 3;

[0009] Y is selected from R5 is selected from hydrogen, alkyl, or is linked to R0 through a linking group in which any one of 1 to 5 methylene groups is replaced by oxygen to form a ring;

[0010] R0 is selected from hydrogen, alkyl, aromatic, acyl, sulfonyl, or is linked to R5 through a linker in which any one of 1-6 methylene groups is replaced by oxygen to form a ring, wherein the alkyl group includes a straight-chain alkyl group, a branched alkyl group, a cyclic alkyl group, an unsaturated alkyl group, an oxygen-containing alkyl group, and a halogen-containing alkyl group; R1 and R2 are each independently selected from hydrogen, substituted or unsubstituted C1-4 alkyl, halogen, cyano, and nitro; R3 and R4 are each independently selected from hydrogen, hydroxyl, halogen, substituted or unsubstituted C1-4 alkyl, trifluoromethyl, -SO3H, -PO(OH)2, -B(OH)2, -COR6, and -CH2R7; R6 and R7 are each independently selected from hydrogen, hydroxyl, -COOH, halogen, substituted or unsubstituted C1-4 alkyl, C1-4 alkoxy, substituted or unsubstituted aromatic benzyl, secondary amine, L0 and L1 are independently selected from 1-4 methylene groups, and R8 and R9 are independently selected from hydrogen, halogen, hydroxyl, substituted or unsubstituted aromatic amine, and secondary amine.

[0011] Preferably, the compound is any one of formula I-1, I-2, I-3 or I-4:

[0012] Based on formula I, W is C, Y is selected from O, SO2, and X is selected from O, S; formula I-1 is:

[0013] Based on formula I, Y is NH, W is C, n=0, X is S, R3 and R4 are selected from methyl and formic acid, R0 is selected from acyl R 01 CO-, sulfonyl R 02 SO2-, where R 01 , R 02 Each is independently selected from an alkyl group, a cycloalkyl group, an aromatic group, and a secondary amine; R1 and R2 are independently selected from a hydrogen group, a substituted or unsubstituted C1-4 alkyl group, a halogen group, a cyano group, and a nitro group; Formula I-2 is:

[0014]

[0015] Based on Formula I, W is C, n is 0, X is S, R3 and R4 are selected from methyl and formic acid respectively, R1 and R2 are independently selected from hydrogen and halogen; m0, m1, m2 are independently selected from integers of 0-6, and m0+m1+m2≤6, Formula I-3 is:

[0016]

[0017] Based on Formula I, W is N, n=0, X is selected from S, R3 and R4 are selected from methyl and formic acid respectively, wherein the six-membered ring is a piperidine ring; Y is selected from n=0; R0 is selected from hydrogen, alkyl, aromatic, acyl, sulfonyl; wherein the alkyl includes straight-chain alkyl, branched-chain alkyl, cyclic alkyl, unsaturated alkyl, oxygen-containing alkyl, halogen-containing alkyl or secondary amine; R1 and R2 are hydrogen; Formula I-4 is:

[0018] Correspondingly, a drug prepared using the compound conformational isomer, or its optical isomer, or its pharmaceutically acceptable salt; or a drug containing the compound conformational isomer, or its optical isomer, or its pharmaceutically acceptable salt.

[0019] The present invention has the following beneficial effects: The present invention provides a new method for preparing sulfur-containing or oxygen-containing five-membered nitrogen heterocyclic derivatives, and synthesizes a series of new sulfur-containing or oxygen-containing five-membered nitrogen heterocyclic derivatives, which have biological activity and anti-tumor activity, and have broad prospects for medicinal and therapeutic applications. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 This is a schematic diagram showing the effect of the compounds prepared in the present invention on the proliferation of SK-HEP-1 cells for 72 hours;

[0021] Figure 2 Schematic diagram showing the effects of the compound prepared in the present invention and sorafenib on the proliferation of HCCLM3 cells for 21 days;

[0022] Figure 3 This is a schematic diagram showing the effect of the compounds prepared in the present invention on the migration ability of SK-HEP-1 cells;

[0023] Figure 4 Schematic diagram of the effect of the compounds prepared in the present invention on the migration ability of SK-HEP-1 cells (wound closure rate);

[0024] Figure 5 Schematic diagram showing the effect of the compounds prepared in the present invention on the migration ability of SK-HEP-1 and HCCLM3 cells;

[0025] Figure 6 Schematic diagram of quantitative analysis of cell migration using ImageJ. DETAILED DESCRIPTION

[0026] The present invention provides a new type of five-membered nitrogen heterocyclic compound, the general structural formula of which is shown in Formula I.

[0027]

[0028] Wherein, the six-membered ring is selected from an aromatic ring, a cyclohexane ring, and a piperidine ring; X is selected from O and S; W is selected from C, CH, and N; and n is selected from any integer of 0-3.

[0029] Y is selected from Wherein R5 is selected from hydrogen, alkyl, or is linked to R0 to form a ring through a linking group in which any one of 1 to 5 methylene groups is replaced by oxygen.

[0030] R0 is selected from hydrogen, alkyl, aromatic, acyl, sulfonyl, or is linked to R5 to form a ring through a linker in which any one of the 1-6 methylene groups is replaced by oxygen, wherein the alkyl group includes a straight-chain alkyl group, a branched-chain alkyl group, a cyclic alkyl group, an unsaturated alkyl group, an oxygen-containing alkyl group, and a halogen-containing alkyl group; a straight-chain alkyl group refers to a fragment containing 1-10 carbon atoms connected by single bonds, and the remaining valence bonds are all combined with hydrogen; a branched-chain alkyl group refers to an alkyl fragment formed on the same or different carbon atoms after one or more hydrogen atoms are replaced at any position on a straight-chain alkane containing 1-10 carbon atoms; a cyclic alkyl group refers to a fragment of a saturated hydrocarbon containing 3-8 carbon atoms and containing one or more rings, which One or more carbon atoms on these rings may be substituted by oxygen atoms, nitrogen atoms, etc.; unsaturated alkyl refers to a carbon chain fragment containing double bonds or triple bonds in the aforementioned straight-chain alkyl, branched-chain alkyl and cyclic alkyl; oxygen-containing alkyl refers to a carbon chain fragment in which one or more carbon atoms at any position in the aforementioned straight-chain alkyl, branched-chain alkyl and cyclic alkyl are substituted by oxygen atoms; halogen-containing alkyl refers to a carbon chain fragment in which one or more hydrogen atoms on carbon at any position in the aforementioned straight-chain alkyl, branched-chain alkyl, cyclic alkyl and oxygen-containing alkyl are substituted by halogen atoms, and halogen atoms refer to fluorine atoms, chlorine atoms, bromine atoms and iodine atoms; aromatic group refers to substituted phenyl, or substituted aromatic heterocyclic group.

[0031] R1 and R2 are each independently selected from hydrogen, substituted or unsubstituted C1-4 alkyl, halogen, cyano, and nitro.

[0032] R3 and R4 are each independently selected from hydrogen, hydroxyl, halogen, substituted or unsubstituted C1-4 alkyl, trifluoromethyl, -SO3H, -PO(OH)2, -B(OH)2, -COR6, -CH2R7, R6 and R7 are each independently selected from hydrogen, hydroxyl, -COOH, halogen, substituted or unsubstituted C1-4 alkyl, C1-4 alkoxy, substituted or unsubstituted aromatic benzyl, secondary amine, L0 and L1 are independently selected from 1-4 methylene groups, and R8 and R9 are independently selected from hydrogen, halogen, hydroxyl, substituted or unsubstituted aromatic amine, and secondary amine.

[0033] Based on Formula I, Y is selected from O, W is selected from C, and the preferred compound structure is shown in Formula I-1:

[0034]

[0035] Y is selected from O and SO2, X is selected from O and S; n is selected from any integer from 0 to 3. The meanings of the remaining substituents are the same as those of compound I.

[0036] Based on Formula I, Y is selected from NH, W is selected from C, n is 0, X is selected from S, R3 and R4 are selected from methyl and formic acid respectively, and the preferred compound structure is shown in Formula I-2:

[0037]

[0038] Wherein R0 is selected from acyl R 01 CO-, sulfonyl R 02 SO2-, where R 01 , R 02 Each is independently selected from an alkyl group, a cycloalkyl group, an aromatic group, and a secondary amine; R1 and R2 are independently selected from a hydrogen group, a substituted or unsubstituted C1-4 alkyl group, a halogen group, a cyano group, and a nitro group.

[0039] Based on Formula I, W is C, n=0, X is S, R3 and R4 are methyl and formic acid respectively, and the preferred compound structure is shown in Formula I-3:

[0040]

[0041] R1, R2 are each independently selected from hydrogen and halogen; m0, m1, m2 are each independently selected from integers of 0-6, m0+m1+m2≤6.

[0042] Based on Formula I, W is N, n=0, X is S, R3 and R4 are selected from methyl and formic acid, respectively. The preferred compound structure is shown in Formula I-4:

[0043]

[0044] Wherein the six-membered ring is a piperidine ring; Y is selected from n=0; R0 is selected from hydrogen, alkyl, aromatic, acyl, sulfonyl; wherein the alkyl includes straight-chain alkyl, branched-chain alkyl, cyclic alkyl, unsaturated alkyl, oxygen-containing alkyl, halogen-containing alkyl or secondary amine; R1 and R2 are hydrogen.

[0045] The present invention also provides a method for preparing the compound I, the reaction equation is:

[0046]

[0047] The specific steps include:

[0048] S1: Compound I-a and a thiolation agent react in an organic solvent at 0°C-100°C to obtain compound I-b. The thiolation agent is selected from sodium hydrosulfide hydrate, phosphorus pentasulfide, hydrogen sulfide, etc., and the organic solvent is selected from methanol, ethanol, DMF, 1,4-dioxane, ethylene glycol dimethyl ether, tetrahydrofuran, etc.

[0049] S2: Compounds Ⅰ-b and Ⅰ-c are refluxed in an organic solvent for more than 6 hours to obtain compound Ⅰ. The organic solvent is selected from methanol, ethanol, isopropanol, acetonitrile, 1,4-dioxane, ethylene glycol dimethyl ether, tetrahydrofuran, etc., and the temperature is selected from 0°C to the solvent reflux temperature. Hal is selected from chlorine, bromine, and iodine.

[0050] Preferably, the reaction equation for preparing compound Ⅰ-1 is:

[0051]

[0052] The specific steps include:

[0053] S1: Compound Ⅰ-1a and a thiolation agent react in an organic solvent at 0°C-100°C to obtain compound Ⅰ-1b. The thiolation agent is selected from sodium hydrosulfide hydrate, phosphorus pentasulfide, hydrogen sulfide, etc., and the organic solvent is selected from methanol, ethanol, DMF, 1,4-dioxane, ethylene glycol dimethyl ether, tetrahydrofuran, etc.

[0054] S2: Compounds Ⅰ-1b and Ⅰ-1c are refluxed in an organic solvent for more than 6 hours to obtain compound Ⅰ-1d. The organic solvent is selected from methanol, ethanol, isopropanol, acetonitrile, 1,4-dioxane, ethylene glycol dimethyl ether, tetrahydrofuran, etc., and the temperature is selected from 0°C to the solvent reflux temperature. Hal is selected from chlorine, bromine, and iodine.

[0055] S3: Compound Ⅰ-1d, R0hal and a base are reacted in an organic solvent at 50°C-140°C to obtain compound Ⅰ-1. The base is selected from potassium carbonate, sodium carbonate, potassium hydroxide, sodium hydroxide, triethylamine, pyridine, diisopropyl ether, etc., and the solvent is selected from DMF, N,N-dimethylacetamide, DMP, ethylene glycol dimethyl ether, ethylene glycol diethyl ether.

[0056] S3': Compound I-1d and an oxidant react in an organic solvent at 0°C-60°C to obtain compound I-1. The oxidant is selected from m-chloroperbenzoic acid, hydrogen peroxide, peracetic acid, tert-butyl peroxide, manganese oxide, etc., and the solvent is selected from dichloromethane, 1,2-dichloroethane, chloroform, carbon tetrachloride, etc.

[0057] Preferably, the reaction equation for preparing compound I-2 is:

[0058]

[0059] Specifically, the process comprises the following steps: S1: Compound I-2a and ammonia water react in alcohol at 0°C-50°C to obtain compound I-2b. The alcohol is selected from methanol, ethanol and isopropanol.

[0060] Wherein, hal is selected from chlorine, bromine, and iodine; R0 is selected from acyl R 01 CO-, sulfonyl R 02SO2-, where R 01 , R 02 Each is independently selected from an alkyl group, a cycloalkyl group, an aromatic group, and a secondary amine; R1 and R2 are independently selected from a hydrogen group, a substituted or unsubstituted C1-4 alkyl group, a halogen group, a cyano group, and a nitro group.

[0061] S2: Compound Ⅰ-2b and P4S 10 In a solvent, the reaction is carried out at 0°C-100°C to obtain compound I-2c. The thiolation reagent is selected from sodium hydrosulfide hydrate, phosphorus pentasulfide, hydrogen sulfide, etc., and the solvent is selected from methanol, ethanol, DMF, 1,4-dioxane, ethylene glycol dimethyl ether, tetrahydrofuran, etc.

[0062] S3: Compounds I-2c and I-2d are refluxed in a solvent for more than 6 hours to obtain compound I-2e. The solvent is selected from methanol, ethanol, isopropanol, acetonitrile, 1,4-dioxane, ethylene glycol dimethyl ether, tetrahydrofuran, etc., and the temperature is 0°C to the reflux temperature of the solvent.

[0063] S4: Compound Ⅰ-2e reacts with R0hal and K2CO3 in a solvent for more than 4 hours to obtain compound Ⅰ-2f. The base is selected from potassium carbonate, sodium carbonate, potassium hydroxide, sodium hydroxide, triethylamine, pyridine, diisopropylethylamine, etc.; the solvent is selected from DMF, N,N-dimethylacetamide, DMP, ethylene glycol dimethyl ether or ethylene glycol diethyl ether, and the temperature is 50℃-140℃ (preferably 0℃-25℃).

[0064] S5: Compound Ⅰ-2f is hydrolyzed to obtain compound Ⅰ-2. The hydrolysis method is: take 0.9mmol of compound Ⅰ-2f and add it to a 2M base (6mL) solution of H2O / THF / MeOH (volume ratio 1:1:3) at a temperature of 0-60°C. After stirring until the solid is dissolved, slowly heat to 50°C for 1-3h, cool the reaction solution to 0°C, and adjust the pH to 3-4 with 1M hydrochloric acid solution. In this process, a large amount of solid will precipitate. Wash the filter cake 3 times with distilled water (2mL), then wash it twice with methanol and distilled water (v:v=1:1, 2mL), collect it, and dry it in vacuo at 80°C to obtain compound Ⅰ-2. The base is 0.5-10 equivalents of sodium hydroxide, lithium hydroxide, potassium hydroxide, etc. Where not specifically stated in the full text, the hydrolysis method is the same as here.

[0065] Preferably, the reaction equation for preparing compound I-3 is:

[0066]

[0067] The specific steps include:

[0068] S1: Add compound Ⅰ-3a, Ⅰ-3b and a base to a solvent, react at 0℃-25℃ for more than 4h to obtain compound Ⅰ-3c. The base is selected from potassium carbonate, sodium carbonate, potassium hydroxide, sodium hydroxide, triethylamine, pyridine, diisopropylethylamine, etc.; the solvent is selected from methanol, ethanol, isopropanol, acetonitrile, 1,4-dioxane, ethylene glycol dimethyl ether, tetrahydrofuran, etc.

[0069] S2: Compound I-3c and a thiolation agent react in a solvent at 0°C-100°C to obtain compound I-3d. The thiolation agent is selected from sodium hydrosulfide hydrate, phosphorus pentasulfide, hydrogen sulfide, etc., and the solvent is selected from methanol, ethanol, DMF, 1,4-dioxane, ethylene glycol dimethyl ether, tetrahydrofuran, etc.

[0070] S3: Compounds I-3d and I-3e are refluxed in a solvent for more than 6 hours to obtain compound I-3f. The solvent is selected from methanol, ethanol, isopropanol, acetonitrile, 1,4-dioxane, ethylene glycol dimethyl ether, tetrahydrofuran, etc., and the temperature is selected from 0°C to the solvent reflux temperature. Hal is selected from chlorine, bromine, and iodine.

[0071] S4: Compound Ⅰ-3f is hydrolyzed to obtain Ⅰ-3.

[0072] Preferably, the compound is prepared The reaction equation is:

[0073]

[0074] The specific steps include:

[0075] S1: Compounds I-4a and I-4b are refluxed in a solvent for more than 6 hours to obtain compound I-4c. The solvent is selected from methanol, ethanol, isopropanol, acetonitrile, 1,4-dioxane, ethylene glycol dimethyl ether, tetrahydrofuran, etc.; the temperature is selected from 0°C to the solvent reflux temperature. Hal is selected from chlorine, bromine, and iodine.

[0076] S2: Compound I-4c is reacted in a 4M hydrogen chloride solution in 1,4-dioxane at room temperature for 2 hours to obtain compound I-4d. The reagent for removing Boc is selected from 1-4M hydrogen chloride methanol solution, 1-4M hydrogen chloride 1,4-dioxane solution, 1%-38% hydrochloric acid solution, trifluoroacetic acid, etc.

[0077] S3: There are two solutions for this step, as follows:

[0078] Scheme 1: Compound Ⅰ-4d, R0Yhal and a base are reacted in a solvent at 0-50°C for 2-6h to obtain compound Ⅰ-4e. The base is selected from potassium carbonate, sodium carbonate, potassium hydroxide, sodium hydroxide, triethylamine, pyridine, diisopropylethylamine, etc.; the solvent is selected from DMF, N,N-dimethylacetamide, DMP, ethylene glycol dimethyl ether or ethylene glycol diethyl ether.

[0079] Scheme 2: Compound Ⅰ-4d, R0YOH and a condensing agent are reacted in a solvent at 0-50°C for 2-6 hours to obtain compound Ⅰ-4e. The condensing agent is selected from DIC, EDCI, DCC, HATU, HBTU, etc.; the solvent is selected from dichloromethane, tetrahydrofuran, 1,4-dioxane, N,N-dimethylformamide, N,N-dimethylacetamide, dimethyl sulfoxide, etc.

[0080] S4: Compound Ⅰ-4e is hydrolyzed to obtain Ⅰ-4.

[0081] The technical solutions in the embodiments of the present invention will be described clearly and completely below in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, rather than all of the embodiments. Unless otherwise specified, the technical means used in the embodiments are conventional means well known to those skilled in the art. The data obtained are all average values ​​after at least 3 repetitions, and all the data obtained in each repetition are valid data.

[0082] Example 1: Preparation and identification of compounds

[0083] 1. Preparation of compound 1:

[0084] Compound (5-methyl-2-(4-methoxyphenyl)-4-oxazolecarboxylic acid ethyl ester (5.4 g, 20.67 mmol; its synthesis method refers to the synthesis method of compound 3b in CN109810071A) was added to a 25 mL round-bottom flask, and 40 mL of 2M sodium hydroxide in methanol-water-tetrahydrofuran (methanol: water: tetrahydrofuran = 1:1:3 by volume) was added at room temperature, reacted at 50°C for 1 hour, cooled, and adjusted to pH 1-2 with concentrated hydrochloric acid in an ice bath, and a white solid precipitated. 20 mL of water was added, stirred for 30 minutes, filtered, and washed with ether to obtain 2-(4-methoxyphenyl)-5-methyloxazole-4-carboxylic acid (compound 1) as a white solid: 4.05 g, yield 89.8%. The nuclear magnetic resonance data are as follows:

[0085] 1 H NMR (400MHz, DMSO-d6) δ12.97(s,1H),7.90(d,J=8.6Hz,2H),7.01(d,J=8.7Hz,2H),3.84(s,3H),2.63(s,3H); 13 C NMR (100MHz, DMSO-d6) δ163.6,161.6,158.9,155.8,129.0,128.1,119.3,114.9,55.8,12.3; ESI-MS (positive mode)calcd.forC12 H 26 NO4:[M+H] + 234.08; found 234.08.

[0086] 2. Preparation of compound 2:

[0087] 5-Methyl-2-(4-methoxyphenyl)-4-oxazolecarboxylic acid (raw material A, 0.15 g, 0.64 mmol, 1 eq) and bromoethanol (raw material B, 0.322 g, 2.57 mmol, 4 eq) were mixed and dissolved in DMF (2 mL), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDCI) (245.4 mg, 1.28 mmol) was added under ice bath, warmed to room temperature, reacted for 1-2 hours, and after TLC monitoring, 20 mL of water was added, extracted with ethyl acetate, the organic phases were combined and concentrated, and the obtained product was recrystallized from methanol to obtain white solid 2-bromoethyl 2-(4-methoxyphenyl)-5-methyloxazole-4-carboxylate (Compound 2), with a yield of 85%. The NMR data are as follows:

[0088] 1 H NMR (400MHz, CDCl3) δ8.03(d,J=8.9Hz,2H),7.00(d,J=8.9Hz,2H),4.68(t,J=6.1Hz,2H),3.89(s,3H),3.67(t,J=6.1Hz,2H),2.74(s,3H); 13 C NMR (100MHz, CDCl3) δ161.9,161.7,160.0,156.3,128.3,128.0,119.2,114.2,64.0,55.4,28.5,12.4; ESI-MS (positive mode)calcd.for C 14 H 14 BrNO3K:[M+K] + 361.98; found 362.00.

[0089] 3. Referring to the method for preparing compound 2, raw material A was used as (5-methyl-2-(4-methoxyphenyl)-4-oxazolecarboxylic acid ethyl ester, and raw material B was replaced accordingly according to Table 1 to prepare compound 3-7.

[0090] Table 1 Reaction substrate comparison table

[0091]

[0092]

[0093] Compound 3: 1 H NMR (400MHz, CDCl3) δ8.60(d,J=6.1Hz,2H),7.95(d,J=9.2Hz,2H),7.55(t,J=6.1Hz,1H),7. 31(d,J=6.1Hz,2H),6.99(d,J=8.7Hz,2H),4.66(d,J=6.1Hz,2H),3.89(s,3H),2.75(s,3H); 13 C NMR (100MHz, CDCl3) δ162.5,161.6,158.9,152.9,150.0,147.6,129.6,128.0,122.3,119.4,114.3,55.4,41.7,29.7,11.8; ESI-MS (positive mode)calcd.for C 18 H 17 N3O3:[M+H] + 324.13; found 324.13.

[0094] Compound 4: 1 H-NMR(600M Hz, CDCl3) δ10.19(s,1H),8.67(d,J=5.6Hz,1H),8.16(d,J=5.7Hz,1H),8.00(d,J=8.9Hz,2H),7.10(d,J=8.9Hz,2H),3.84(s,3H),2.69(s,3H); 13 CNMR(150MHz, CDCl3)δ162.1,161.8,161.1,159.7,159.4,158.8,129.4,128.6,118.8,115.1,109.1,56.0,12.2; ESI-MS(positive mode)calcd.for C 16 H 13 ClN4O3Na:[M+Na] + 367.06; found 367.06.

[0095] Compound 5: 1 H NMR (400MHz, CDCl3) δ8.02(d,J=8.5Hz,2H),6.98(d,J=9.1Hz,2H),4.48(t,J=5.9Hz, 2H),3.88(s,3H),2.81(t,J=5.4Hz,2H),2.71(s,3H),2.63-2.42(m,8H),2.32(s,3H); 1313C NMR (100 MHz, CDCl3) δ 162.3, 161.6, 159.8, 155.8, 128.2, 119.1, 114.1, 61.5, 56.5, 55.3, 54.8, 53.2, 45.9, 12.1; ESI-MS (positive mode) calcd. for C 19 H 26 N3O4: [M + H] + 360.19; found 360.19.

[0096] Compound 6: 1 1H NMR (400 MHz, CDCl3) δ 8.01 (d, J = 8.7 Hz, 2H), 6.98 (d, J = 8.7 Hz, 2H), 4.49 (t, J = 6.0 Hz, 2H), 3.88 (s, 3H), 2.79 (t, J = 6.0 Hz, 2H), 2.71 (s, 3H), 2.55 (s, 4H), 1.66 (t, J = 5.5 Hz, 4H), 1.48 (t, J = 5.8 Hz, 2H); 13 13C NMR (100 MHz, CDCl3) δ 162.4, 161.9, 159.8, 155.9, 128.3, 119.2, 114.2, 61.5, 57.2, 55.4, 54.7, 29.7, 25.6, 24.1, 12.1; ESI-MS (positive mode) calcd. for C 19 H 25 N2O4: [M + H] + 3345.18; found 345.18.

[0097] Compound 7: 1 1H NMR (400 MHz, CDCl3) δ 8.00 (d, J = 8.7 Hz, 2H), 6.96 (d, J = 9.2 Hz, 2H), 4.45 (t, J = 6.9 Hz, 2H), 3.86 (s, 3H), 2.74 (t, J = 5.6 Hz, 2H), 2.69 (s, 3H), 2.35 (s, 3H); 13 13C NMR (100 MHz, CDCl3) δ 162.5, 161.6, 159.8, 155.8 128.3, 119.3, 114.2, 62.2, 57.6, 55.4, 45.5, 12.2; ESI-MS (positive mode) calcd. for C 16 H 21 N2O4: [M + H] + 305.15; found 305.15.

[0098] 4. Preparation of compound 8:

[0099] 27.7 mg (0.1 mmol) of 5-methyl-2-(4-methoxyphenyl)-4-oxazolecarboxylic acid ethyl ester and 1 mL of dichloromethane were added to a 10 mL round-bottom flask in sequence, and 250 mg (1 mmol) of boron tribromide was added dropwise under a -20 ° C ice-salt bath, and then gradually warmed to room temperature for reaction for 12 hours, and then 2 mL of 5% sulfuric acid solution was added dropwise under an ice bath to quench the reaction, and then the pH was adjusted to 5-6 with a saturated sodium bicarbonate solution, extracted with ethyl acetate, dried and concentrated, and then 3 mL of sodium hydroxide solution (2M NaOH in V (THF / H2O / EtOH): 1 / 1 / 3) was directly added, and the reaction was carried out at 70 ° C for 40 minutes. After the reaction, concentrated hydrochloric acid was added dropwise to adjust the pH to 1-2, the organic phase was extracted with dichloromethane, dried and concentrated, and 2-(4-hydroxyphenyl)-5-methyloxazole-4-carboxylic acid was obtained after dichloromethane beating for 48 hours, as a white solid, with a yield of 59%. The nuclear magnetic resonance data are as follows:

[0100] 1 H NMR (400MHz, DMSO-d6) δ7.79 (d, J = 8.4Hz, 2H), 6.90 (d, J = 8.4Hz, 2H), 2.62 (s, 3H); 13 C NMR (100MHz, DMSO-d6) δ163.7,160.3,159.3,155.5,128.9,128.3,117.8,116.4,12.3; ESI-MS (positive mode)calcd.for C 10 H 10 NO4:[M+H] + 220.06; found 220.06.

[0101] 5. Preparation of compound 9:

[0102] Dissolve 4-methoxybenzonitrile (raw material C) (5 grams, 37.55 mmol) in DMF (50 mL) and water (1.35 g, 75.1 mmol, 2.0 eq), add 70% sodium hydrosulfide hydrate (5.26 g, 93.87 mmol, 2.5 eq) under stirring, and stir at room temperature for 1 hour. After most of the solid is dissolved, add anhydrous magnesium chloride (3.94 g, 41.3 mmol, 1.1 eq) in batches under an ice bath, and after the addition, heat to 50 ° C and react overnight. After cooling to room temperature, slowly pour the reaction solution into 1M dilute hydrochloric acid, extract with ethyl acetate, dry and concentrate to obtain thiobenzamide. Add 80 mL of ethanol, add 2-chloroacetoacetic acid ethyl ester (raw material D) under stirring, and heat to reflux overnight. Concentrate under reduced pressure to remove ethanol, add 50 mL of ethyl acetate, and then add 40 mL of saturated sodium bicarbonate solution to separate the organic phase. The aqueous phase is extracted twice with fresh ethyl acetate. The organic phases were combined, washed with saturated brine, dried and concentrated to obtain a crude product, 20 mL of ethyl acetate and 200 mL of petroleum ether were added, and the mixture was slurried for 2 hours. The mixture was filtered to obtain ethyl 2-(4-methoxyphenyl)-4-methylthiazole-5-carboxylate as a light yellow powder (9.5 g) with a yield of 91%.

[0103] 2-(4-methoxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester (4.5 g, 16.22 mmol) was hydrolyzed by the following method: 2-(4-methoxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester (4 g, 14.4 mmol) and 180 mL of sodium hydroxide solution (2M NaOH in VTHF / VH2O / VEtOH: 1 / 1 / 3) were added to a 500 mL round-bottom flask in sequence, and the mixture was reacted at 70°C for 40 min. After the reaction was completed, concentrated hydrochloric acid was added dropwise to adjust the pH to 1-2, the organic phase was extracted with dichloromethane, dried and concentrated, and after beating with dichloromethane for 24 h, compound 9: 2-(4-methoxyphenyl)-4-methylthiazole-5-carboxylic acid, a white solid was obtained. Yield: 88%. The NMR data are as follows:

[0104] 1 H NMR (400MHz, DMSO-d6) δ13.27(s,1H),7.92(d,J=8.3Hz,2H),7.06(d,J=8.6Hz,2H),3.84(s,3H),2.65(s,3H); 13 C NMR (100MHz, DMSO-d6) δ168.9,163.5,162.1,160.0,128.7,125.5,122.1,115.1,55.9,17.6; ESI-MS (positive mode)calcd.forC 12 H 12 NO3S:[M+H] +250.05; found 250.05.

[0105] 6. Referring to the method for preparing compound 2, raw material A was replaced by compound 9, and raw material B was replaced accordingly according to Table 2 to prepare compounds 10, 11, 15-17. Raw material A was replaced by 2-(4-methoxyphenyl)-5-methylthiazole-4-carboxylic acid, and raw material B was replaced accordingly according to Table 2 to prepare compounds 12-14.

[0106] Table 2 Reaction substrate comparison table

[0107]

[0108] Among them, the preparation method of raw material B for synthesizing compound 15 is as follows: 2,4-dichloropyrimidine (3 g, 20.14 mmol) was dissolved in DMF (30 mL), ethanolamine (1.5 g, 24.16 mmol, 1.2 eq) and triethylamine (3 g, 30.21 mmol) were added, stirred overnight, and extracted with ethyl acetate to obtain a white product: 2-((2-chloropyrimidine-4)amino)ethanol.

[0109] Compound 10: 1 H NMR (400MHz, CDCl3) δ7.94(d,J=9.0Hz,2H),6.97(d,J=9.0Hz,2H),4.61(t,J=6.1Hz,2H),3.89(s,3H),3.65(t,J=6.0Hz,2H),2.80(s,3H); 13 C NMR (100MHz, CDCl3) δ170.5,162.1,161.9,161.8,128.5,125.8,119.9,114.4,64.3,55.5,28.6,17.7; ESI-MS (positive mode)calcd.for C 14 H 15 BrNO2S:[M+H] + 355.99; found 356.00.

[0110] Compound 11: 1 H NMR (400MHz, DMSO-d6) δ8.86(t,J=5.9Hz,1H),8.53(d,J=5.9Hz,2H),7.90(d,J=8.7Hz,2H),7 .32(d,J=5.9Hz,2H),7.08(d,J=9.0Hz,2H),4.47(d,J=6.2Hz,2H),3.84(s,3H),2.63(s,3H); 1313C NMR (100 MHz, DMSO-d6) δ 166.5, 161.9, 156.0, 150.0, 148.7, 130.7, 128.4, 125.6, 125.0, 122.6, 115.2, 55.9, 42.4, 17.6; ESI-MS (positive mode) calcd. for C 18 H 18 N3O2S: [M + H] + 340.11; found 340.11.

[0111] Compound 12: 1 1H NMR (400 MHz, CDCl3) δ 7.93 (d, J = 9.6 Hz, 2H), 6.97 (d, J = 9.3 Hz, 2H), 4.43 (t, J = 6.0 Hz, 2H), 3.89 (s, 3H), 2.80 (t, J = 5.6 Hz, 2H), 2.79 (s, 3H), 2.71 (m, 4H), 2.63 (m, 4H), 2.40 (s, 34H); 13 13C NMR (100 MHz, CDCl3) δ 170.1, 162.2, 162.0, 161.3, 128.5, 125.8, 120.5, 114.4, 62.6, 56.4, 55.5, 54.8, 52.6, 45.5, 29.7, 17.6; ESI-MS (positive mode) calcd. for C 19 H 25 N3O3S: [M + H] + 376.17; found 376.17.

[0112] Compound 13: 1 1H NMR (400 MHz, CDCl3) δ 7.96 (d, J = 8.2 Hz, 2H), 6.96 (d, J = 8.6 Hz, 2H), 4.43 (t, J = 6.1 Hz, 2H), 3.87 (s, 3H), 2.77 (s, 3H), 2.74 (t, J = 6.1 Hz, 2H), 2.53 (s, 4H), 1.61 (m, 4H), 1.46 (m, 3H); 13 13C NMR (100 MHz, CDCl3) δ 170.0, 162.2, 162.0, 161.2, 128.4, 125.8, 120.6, 114.4, 62.9, 57.2, 55.5, 54.8, 26.0, 24.1, 17.6; ESI-MS (positive mode) calcd. for C 19 H 25N2O3S:[M+H] + 361.16; found 361.16.

[0113] Compound 14: 1 H NMR (400MHz, CDCl3) δ7.92(d,J=8.5Hz,2H),6.96(d,J=8.9Hz,2H),4.41(t,J=5.2Hz,2H),3.87(s,3H),2.78(s,3H),2.72(t,J=5.6Hz,2H),2.36(s,6H); 13 CNMR(100MHz, CDCl3)δ170.0,162.3,162.0,161.3,128.5,125.8,120.5,114.4,63.1,57.7,55.5,45.8,17.6; ESI-MS(positive mode)calcd.for C 19 H 25 N3O3S:[M+H] + 376.17; found 376.17.

[0114] Compound 15: 1 H NMR (400MHz, CDCl3) δ8.57(d,J=5.5Hz,1H),8.24(d,J=8.6Hz,1H),8.04(s,1H),7.94(d, J=8.8Hz,2H),7.0(d,J=8.8Hz,2H),3.91(s,3H),3.00(s,2H),2.90(s,2H),2.85(s,3H); 13 C NMR (100MHz, CDCl3) δ169.1,162.4,160.8,160.6,160.3,160.0,128.7,128.2,125.2,122.2,114.6,114.4,108.5,55.5,29.7,18.0.

[0115] Compound 16: 1 H NMR (400MHz, CDCl3) δ7.92(d,J=8.3Hz,2H),6.97(d,J=8.7Hz,2H),4.43(t,J=5.6Hz,2H),3. 88(s,3H),3.73(t,J=5.1Hz,4H),2.78(s,3H),2.76(t,J=6.0Hz,2H),2.58(t,J=4.6Hz,4H); 13C NMR (100MHz, CDCl3) δ170.1,162.2,162.0,161.3,128.5,125.8,120.5,114.4,67.0,62.4,57.0,55.5,53.9,17.6; ESI-MS (positive mode)calcd.for C 18 H 23 N2O4S:[M+H] + 363.14; found 363.14.

[0116] Compound 17: 1 H NMR (400MHz, CDCl3) δ7.90(d,J=8.6Hz,2H),6.97(d,J=8.6Hz,2H),5.89(s,2H),3.88(s,3H),,2.76(s,3H); 13 C NMR (100MHz, CDCl3) δ167.9,168.3,161.9,157.2,128.4,125.7,123.9,114.4,55.5,17.6; ESI-MS (positive mode)calcd.forC 12 H 13 N2O2S:[M+H] + 349.07; found 349.07.

[0117] 7. Preparation of compound 18:

[0118] To a 100 mL round-bottom flask, ethyl 2-(4-methoxyphenyl)-4-methylthiazole-5-carboxylate (1.2 g, 4.3 mmol), carbon tetrachloride (12 mL), N-bromosuccinimide (0.847 g, 4.76 mmol, 1.1 eq), and azobisisobutyronitrile (1.55 g, 9.46 mmol, 2.2 eq) were added in sequence and reacted at 70°C for 4 hours. After the reaction was completed by TLC detection, carbon tetrachloride was removed by vacuum distillation, and 30 mL of saturated ammonium chloride solution was added to remove the unreacted N-bromosuccinimide. After filtration, ethyl acetate was added for extraction, and the organic phases were combined, washed with water 3 times, washed with saturated brine once, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography (V 乙酸乙酯 :V 石油醚 =1:10), the target compound: ethyl 2-(4-methoxyphenyl)-4-(bromomethyl)thiazole-5-carboxylate, 1.07 g, reddish brown solid, yield: 84%. Its NMR data are as follows:

[0119] 1H NMR (400MHz, CDCl3) δ7.94(d,J=8.5Hz,2H),6.97(d,J=8.5Hz,2H),4.98(s,2H),4.41(q,J=6.7Hz,2H),3.89(s,3H),1.42(t,J=8.3Hz,3H); 13 C NMR (100MHz, CDCl3) δ170.7,162.3,161.3,158.3,128.6,125.4,119.1,114.6,68.2,61.8,55.5,14.3; ESI-MS (positive mode)calcd.for C 14 H 15 BrNO2S:[M+H] + 355.99; found 356.00.

[0120] Dissolve ethyl 2-(4-methoxyphenyl)-4-(bromomethyl)thiazole-5-carboxylate (0.35 g, 0.98 mmol) in dichloromethane (4 mL), add 30% dimethylamine methanol solution (0.59 g, 3.93 mmol, 4 eq) and tetrabutylammonium iodide (5 mg, 0.014 mmol) dropwise under ice bath, and react at room temperature for 5 h. After TLC detection, remove dichloromethane by vacuum distillation, add 50 mL of water to quench the reaction, extract with dichloromethane, combine the organic phases, wash once with saturated brine, dry over anhydrous sodium sulfate, concentrate, and separate by column chromatography (V 二氯甲烷 :V 甲醇 =15:1) to obtain compound 18: 245 mg of ethyl 4-((dimethylamino)methyl)-2-(4-methoxyphenyl)thiazole-5-carboxylate, yield 78%. The developing system Rf=0.5, EA:PE=1:10. The H NMR spectrum data are as follows:

[0121] 1H NMR (400MHz, CDCl3) δ7.98 (d, J=8.4Hz, 2H), 6.97 (d, J=8.4Hz, 2H), 4.38 (q, J= 7.0Hz,2H),4.05(s,2H),3.89(s,3H),2.45(s,6H),1.41(t,J=6.5Hz,3H); 13C NMR (100MHz, CDCl3) δ170.0,162.0,161.9,160.8,128.7,125.9,123.4,114.3,61.4.56.7.55.5,45.6,14.3; ESI-MS (positive mode)calcd.for C16H21N2O3S:[M+H]+321.13; found 321.13.

[0122] 8. Preparation of compound 19:

[0123] Referring to the method for preparing compound 2, raw material A was replaced by ethyl 2-(4-methoxyphenyl)-4-(bromomethyl)thiazole-5-carboxylate, and raw material B was replaced by morpholine to obtain compound 19: ethyl 2-(4-methoxyphenyl)-4-(morpholinomethyl)thiazole-5-carboxylate, a light yellow solid, with a yield of 79%.

[0124] 1H NMR(400MHz, CDCl3) δ7.95(d,J=8.9Hz,2H),6.98(d,J=8.9Hz,2H),4.37(q,J=7.4Hz,2H),4.1 2(s,2H),3.89(s,3H),3.77(t,J=5.1Hz,4H),2.73(t,J=4.5Hz,4H),1.41(t,J=4.5Hz,3H); 13C NMR (100MHz, CDCl3) δ170.0,162.0,162.0,159.9,128.6,125.8,114.4,67.0,61.4,56.1,55.5,53.6,29.7,14.3; ESI-MS (positive mode)calcd.forC18H23N2O4S:[M+H]+363.14; found 363.14.

[0125] 9. Preparation of compound 20:

[0126] Tetrahydrofuran (10 mL) and lithium aluminum tetrahydride (149 mg, 3.6 mmol, 2 eq) were added to a 25 mL round-bottom flask in sequence, and a solution of ethyl 2-(4-methoxyphenyl)-4-methylthiazole-5-carboxylate (500 mg, 1.8 mmol) in tetrahydrofuran (2 mL) was slowly added dropwise under an ice bath, and then the temperature was gradually raised to room temperature and the reaction was carried out for 1 h. After the reaction was completed by TLC detection, 2 mL of water was added dropwise under an ice bath to quench the reaction, and the mixture was filtered through diatomaceous earth, extracted with dichloromethane, and the organic phases were combined, washed once with saturated brine, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography (V 二氯甲烷 :V 甲醇 =30:1), to obtain compound 20: (2-(4-methoxyphenyl)-4-methylthiazol-5-yl)methanol 313.9 mg, white solid, yield: 74%.

[0127] 1H NMR (400MHz, CDCl3) δ7.84 (d, J = 9.5Hz, 2H), 6.95 (d, J = 9.5Hz, 2H), 4.81 (s, 2H), 3.86 (s, 3H), 2.44 (s, 3H); 13C NMR(100MHz, CDCl3)δ166.2,161.1,150.1,130.0,127.9,126.6,114.2,55.8,55.4,15.1; ESI-MS(positive mode)calcd.for C12H14NO2S:[M+H]+236.07; found 236.07.

[0128] 10. Preparation of compound 21:

[0129] 71.2 mg (0.57 mmol) of 2-bromoethanol, 2 mL of dimethylformamide, 94.5 mg (0.63 mmol) of tert-butyldimethylsilyl chloride, and 42.9 mg (0.63 mmol) of imidazole were added to a 100 mL round-bottom flask in sequence, and the mixture was reacted at room temperature for 4 h. After the reaction was completed, 50 mL of water was added to quench the mixture, and the mixture was extracted with ethyl acetate. After column chromatography, intermediate 1 was obtained: (2-(bromoethoxy)-tert-butyldimethylsilane). 5 mL of dimethylformamide was added to intermediate 1, and compound 20 122.6 mg (0.52 mmol), tetrabutylammonium iodide 16 mg (0.052 mmol), 14.9 mg (0.62 mmol) of sodium hydride were added under ice bath, then the temperature was gradually raised to room temperature, and the reaction was carried out for 5 hours. After the reaction was completed, 5 mL of saturated sodium bicarbonate solution was added to quench, extracted with ethyl acetate, and separated by column chromatography to obtain intermediate 2: (5-((3-((tert-butyldimethylsilyl(oxy)propoxy)methyl)-2-(4-methoxyphenyl)-4-methylthiazole). 8 mL of intermediate 2 was added Tetrahydrofuran, 327.6 mg (1.04 mmol) of tetrabutylammonium fluoride was added dropwise under ice bath. After the reaction was completed, 10 mL of saturated ammonium chloride solution was added to quench, extracted with ethyl acetate, the organic phases were combined, washed once with saturated brine, dried over anhydrous sodium sulfate and concentrated, separated by column chromatography (V ethyl acetate: V petroleum ether = 1:5), to obtain compound 21: 2-((2-(4-methoxyphenyl)-4-methylthiazol-5-yl)methoxy)ethane-1-ol, 74.5 mg, light yellow solid, total yield: 51%. The NMR data are as follows:

[0130] 1H NMR(400MHz, CDCl3) δ7.86(d,J=8.8Hz,2H),6.96(d,J=8.8Hz,2H),4.72(s,2H ),3.87(s,3H),3.80(t,J=4.4Hz,2H),3.65(t,J=4.5Hz,2H),2.47(s,3H); 13C NMR(100MHz, CDCl3)δ166.5,161.1,151.1,127.9,126.9,126.6,114.2,74.1,64.6,61.9,55.4,15.3; ESI-MS(positive mode)calcd.for C14H18N1O3S:[M+H]+280.10; found 280.10.

[0131] 11. Preparation of compound 22:

[0132] According to the method for preparing compound 8, 5-methyl-2-(4-methoxyphenyl)-4-oxazolecarboxylic acid ethyl ester was replaced by 2-(4-methoxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester to prepare compound 22: 2-(4-hydroxyphenyl)-5-methylthiazole-4-carboxylic acid with a yield of 83%.

[0133] 1 H NMR (400MHz, DMSO-d6) δ13.22(br s,1H),10.19(br s,1H),7.82(d,J=8.6Hz,2H),6,88(d,J=8.6Hz,2H),2.64(s,3H). 13 C NMR (100MHz, DMSO-d6) δ169.4,163.5,160.9,160.0,128.8,124.1,121.6,116.5,17.6.

[0134] 12. Preparation of compound 23:

[0135] Mix 4-methoxybenzamide (0.5 g, 3.31 mmol) and ethyl 2-chloroacetoacetate (1.36 g, 8.27 mmol, 2.5 eq) in ethanol and seal the tube. React at 110°C for 12 hours, cool and concentrate, extract with ethyl acetate, and separate the crude product with a chromatography column to obtain ethyl 2-(4-methoxyphenyl)-4-methyloxazole-5-carboxylate, 0.467 g, yield 54%.

[0136] Hydrolyze ethyl 2-(4-methoxyphenyl)-4-methyloxazole-5-carboxylate: Add ethyl 2-(4-methoxyphenyl)-4-methyloxazole-5-carboxylate 4.8g, 20.67mmol to a 25mL round-bottom flask, add 2M sodium hydroxide in methanol tetrahydrofuran (volume ratio 1:1:3) (40mL) at room temperature, react at 50°C for 1 hour, cool, adjust pH to 1-2 with concentrated hydrochloric acid in an ice bath, precipitate white solid, add water (20mL) and stir for 30 minutes, filter, and wash with an appropriate amount of ether. Compound 23: 2-(4-methoxyphenyl)-4-methyloxazole-5-carboxylic acid, white solid. Yield 79%.

[0137] 1 H NMR (400MHz, DMSO-d6) δ13.4(s,1H),7.95(d,J=8.9Hz,2H),7.10(d,J=8.9Hz,2H),3.84(s,3H),2.42(s,3H). 13 C NMR (100MHz, DMSO-d6) δ162.4,161.7,159.9,146.5,137.6,128.9,118.9,115.2,55.9,13.6.

[0138] 13. Preparation of compound 24:

[0139] In an ice-water bath, phosphorus pentasulfide (2 eq) was added in batches to stirred anhydrous ethanol (0.44 mmol / mL), and the reaction was slowly heated to room temperature and continued for 1 hour. 4-Hydroxybenzonitrile (raw material E, 1 eq) was added in batches to the reaction system and reacted overnight at room temperature. After reduced pressure concentration and distillation to remove ethanol, the residue was dissolved in ethyl acetate (50 mL), washed with water (20 mL × 2), washed with saturated brine and dried over anhydrous sodium sulfate. After filtration and concentration, the product 4-hydroxythiobenzamide was obtained.

[0140] 4-Hydroxythiobenzamide (raw material F, 12 g, 78.33 mmol) and ethyl 2-chloroacetoacetate (16.76 g, 101.83 mmol, 1.3 eq) were mixed in ethanol and refluxed overnight, then cooled to room temperature, concentrated and evaporated to remove ethanol, 50 mL of ethyl acetate was added, followed by 40 mL of saturated sodium bicarbonate solution, the organic phase was separated, the aqueous phase was extracted twice with fresh ethyl acetate, the combined organic phases were washed with saturated brine, the crude product was dried and concentrated, 20 mL of ethyl acetate was added, 200 ml of petroleum ether was added for slurrying for 2 hours, and a light yellow solid was obtained by filtration: 18.36 g of ethyl 2-(4-hydroxyphenyl)-4-methylthiazole-5-carboxylate, with a yield of 89%.

[0141] To a 5 mL round-bottom flask, ethyl 2-(4-hydroxyphenyl)-4-methylthiazole-5-carboxylate (raw material G, 0.35 g, 1.33 mmol), dimethylformamide (3 mL), ethyl bromide (raw material H, 0.290 g, 2.66 mmol, 2 eq), potassium carbonate (0.55 g, 3.99 mmol) and a catalytic amount of tetrabutylammonium iodide (0.013 g, 0.04 mmol) were added in sequence, and the mixture was reacted at 70 ° C for 3 h. After the reaction was completed by TLC detection, the mixture was cooled, 5 mL of water was added, and the mixture was extracted with ethyl acetate. The organic phases were combined, washed with water 3 times, washed with brine once, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography (V 乙酸乙酯 :V 石油醚 =1:10), to obtain a light yellow solid: 0.362 g of ethyl 2-(4-ethoxyphenyl)-4-methylthiazole-5-carboxylate, with a total yield of 93%.

[0142] 1 H NMR (400MHz, CDCl3) δ7.92 (d, J = 8.7Hz, 2H), 6.96 (d, J = 8.7Hz, 2H), 4.36 (q, J = 6.7Hz, 2H), 4.12 (q, J = 6.7Hz, 2H), 2.78 (s, 3H), 1.47 (t, J = 6.2Hz, 3H), 1.39 (t, J = 7.2Hz, 3H); 13 C NMR (100MHz, CDCl3) δ170.0,162.5,161.3,161.0,128.4(2C),125.7,120.7,114.8(2C),63.8,61.2,17.6,14.8,14.4; ESI-MS(positive mode)calcd.for C 15 H 17 NO3S:[M+H] + 292.10; found 292.10.

[0143] Hydrolysis of ethyl 2-(4-ethoxyphenyl)-4-methylthiazole-5-carboxylate: 4 g (14.4 mmol) of compound ethyl 2-(4-ethoxyphenyl)-4-methylthiazole-5-carboxylate and 180 mL of sodium hydroxide solution (2M NaOH in V (THF / H2O / EtOH): 1 / 1 / 3) were added to a 500 mL round-bottom flask in sequence, and the mixture was reacted at 70°C for 40 min. After the reaction was completed, concentrated hydrochloric acid was added dropwise to adjust the pH to 1-2, and the organic phase was extracted with dichloromethane, dried and concentrated, and slurried with dichloromethane for 24 h to obtain compound 24: 2-(4-ethoxyphenyl)-4-methylthiazole-5-carboxylic acid, a white solid, with a yield of 78%. The developing system Rf = 0.5, DCM / CH3OH = 10:1.

[0144] 1 H NMR (400MHz, DMSO-d6) δ7.90(d,J=8.9Hz,2H),7.03(d,J=8.9Hz,2H),4.10(q,J=7.4Hz,2H),2.65(s,3H),1.35(t,J=6.9Hz,3H); 13 C NMR (100MHz, DMSO-d6) δ168.9,163.5,161.4,160.0,128.7(2C),125.4,122.2,115.5(2C),63.9,17.6,15.0; ESI-MS(positive mode)calcd.for C 13 H 14 NO3S:[M+H] + 264.07; found 264.07.

[0145] 14. Referring to the method for preparing compound 24, the raw material G was replaced with ethyl 2-(4-hydroxyphenyl)-4-methylthiazole-5-carboxylate, and the raw material H was replaced accordingly according to Table 3 to prepare compound 25-35.

[0146] Table 3 Reaction substrate comparison table

[0147]

[0148]

[0149] Compound 25: 1 H NMR (400MHz, DMSO-d6) δ7.90(d,J=8.8Hz,2H),7.04(d,J=8.8Hz,2H),4.00(t,J=6.6Hz,2H),2.64(s,3H),1.75(m,2H),0.99(t,J=7.5Hz,2H); 13 C NMR (100MHz, DMSO-d6) δ168.9,163.5,161.6,160.0,128.6(2C),125.4,122.1,115.5(2C),69.7,22.4,17.6,10.8; ESI-MS(positive mode)calcd.for C 14 H 16 NO3S:[M+H] + 278.08; found 278.08.

[0150] Compound 26: 1H NMR (400MHz, DMSO-d6) δ7.89(d,J=8.3Hz,2H),7.02(d,J=8.4Hz,2H),4.71(m,1H),2.65(s,3H),1.30(s,3H),1.29(s,3H); 13 C NMR (100MHz, DMSO-d6) δ169.0,163.5,160.5,160.0,128.7(2C),125.2,122.1,116.4(2C),70.1,22.2,17.6; ESI-MS(positive mode)calcd.for C 14 H 16 NO3S:[M+H] + 278.08; found 278.08.

[0151] Compound 27: 1 H NMR (400MHz, DMSO-d6) δ7.91(d,J=8.7Hz,2H),7.08(d,J=8.7Hz,2H),6.06(m,1H),5.42(dd,J =17.0Hz, J=0.8Hz, 1H), 5.29 (dd, J=10.7Hz, J=1.2Hz, 1H), 4.66 (d, J=6.2Hz, 2H), 2.65 (s, 3H); 13 C NMR (100MHz, DMSO-d6) δ168.8,163.5,161.0,159.9,133.7,128.6(2C),125.7,118.3,115.8(2C),68.9,17.6; ESI-MS(positive mode)calcd.for C 14 H 14 NO3S:[M+H] + 276.07; found 276.07.

[0152] Compound 28: 1 H NMR (600MHz, DMSO-d6) δ7.89 (d, J = 8.4Hz, 2H), 7.04 (d, J = 8.9Hz, 2H), 4.87 (s, 1H), 4.05 (t, J = 4.7Hz, 2H), 3.72 (t, J = 4.7Hz, 2H), 2.63 (s, 3H); 13C NMR (150MHz, DMSO-d6) δ168.9,163.5,161.6,160.0,128.7(2C),125.5,122.3,115.6(2C),70.3,59.9,17.6; ESI-MS(positive mode)calcd.for C 15 H 16 NO5S:[M+H] + 322.07; found 322.07.

[0153] Compound 29: 1 H NMR(400MHz,DMSO-d6)δ13.24(br s,1H),7.90(d,J=8.8Hz,2H),7.05(d,J=8.8Hz,2H),3.82(d,J=6.4Hz,2H),2.65(s,3H),1.99-2.09(m,1H),0.99(d,J=6.7Hz,6H). 13 C NMR (100MHz, DMSO-d6) δ168.9,163.5,161.6,160.0,128.6(2C),125.4,122.1,115.5(2C),74.4,28.1,19.4(2C),17.5.

[0154] Compound 30: 1 H NMR (400MHz, DMSO-d6) δ7.86 (d, J = 8.4Hz, 2H), 7.04 (d, J = 8.4Hz, 2H), 3.70 (s, 2H), 2.63 (s, 3H), 1.01 (s, 9H). 13 C NMR (100MHz, DMSO-d6) δ168.9, 163.5, 161.9, 159.9, 128. (2C, 125.4, 122.1, 115.6 (2C), 77.8, 32.1, 26.8 (3C), 17.5.

[0155] Compound 31: 1H NMR (400 MHz, DMSO-d6) δ13.24 (br s, 1H), 7.89 (d, J = 8.9 Hz, 2H), 7.06 (d, J = 8.9 Hz, 2H), 4.04 (t, J = 4.0 Hz, 2H), 2.66 (s, 2H), 1.71 (m, 2H), 1.45 (m, 2H), 0.94 (t, 3H), 13C NMR (100 MHz, DMSO-d6) δ168.9, 163.5, 161.6, 160.0, 128.6 (2C), 125.4, 122.1, 115.5 (2C), 68.0, 31.1, 19.1, 17.6, 14.1.

[0156] Compound 32: 1 H NMR (400MHz, DMSO-d6) δ7.90(d,J=8.8Hz,2H),7.05(d,J=8.8Hz,2H),4.07(t,J=6.4 Hz,2H),2.65(s,3H),1.79(m,1H),1.63(q,J=6.8Hz,2H),0.95(s,3H),0.93(s,3H); 13 C NMR (100MHz, DMSO-d6) δ168.9,163.5,161.6,160.0,128.7(2C),125.4,122.2,115.6(2C),66.8,37.8,25.1,22.9,17.6; ESI-MS(positive mode)calcd.forC 16 H 20 NO3S:[M+H] + 306.12; found 306.11.

[0157] Compound 33: 1 H NMR (600MHz, DMSO-d6) δ7.88(d,J=8.8Hz,2H),7.03(d,J=8.8Hz,2H),4.05(t,J=6.3Hz,2H),2.63(s,3H),2.38(t,J=7.1Hz,2H),1.95(m,3H); 13 C NMR (150MHz, DMSO-d6) δ174.5,168.8,163.5,161.4,159.9,128.7(2C),125.6,122.4,115.6(2C),67.4,30.5,24.6,17.5; ESI-MS(positive mode)calcd.for C 15 H 16NO5S:[M+H] + 322.07; found 322.07.

[0158] Compound 34: 1 H NMR (400MHz, DMSO-d6) δ7.91(d,J=8.9Hz,2H),7.07(d,J=8.6Hz,2H),4.17(t,J=4.6Hz,2H),3.68(t,J=4.6Hz,2H),3.32(s,3H),2.65(s,3H); 13 C NMR (100MHz, DMSO-d6) δ168.9,163.5,161.4,160.0,128.7(2C),125.6,122.2,115.6(2C),70.7,67.7,58.7,17.6; ESI-MS(positive mode)calcd.for C 14 H 16 NO4S:[M+H] + 294.08; found 294.08.

[0159] Compound 35: 1 H NMR (400MHz, DMSO-d6) δ7.91(d,J=8.7Hz,2H),7.07(d,J=8.7Hz,2H),4.17(t,J=4.6Hz,2H),3 .76(t,J=4.2Hz,2H),3.60(t,J=4.9Hz,2H),3.46(t,J=4.6Hz,2H),3.25(s,3H),2.65(s,3H); 13 C NMR (100MHz, DMSO-d6) δ168.9,163.5,161.4,160.0,128.7(2C),125.6,122.2,115.6(2C),71.7,70.2,69.2,68.0,58.5,17.6; ESI-MS(positive mode)calcd.for C 16 H 20 NO5S:[M+H] + 338.11; found 338.10.

[0160] 15. Preparation of compound 36:

[0161] 2-(4-Hydroxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester (0.5g, 1.9mmol), fluoroethanol (0.365g, 0.57mmol, 3eq), triphenylphosphine (1.0g, 3.8mmol, 2eq) were placed in a dry flask, and after nitrogen exchange in the flask, ultra-dry 1,4-dioxane (5mL) was added, and DEAD (0.661g, 3.8mmol, 2eq) was dropped in an ice bath, and the temperature was raised to room temperature for overnight reaction. The reaction liquid was concentrated, and after adding dichloromethane to dissolve, silica gel was added, concentrated, and separated by column chromatography, and eluted with petroleum ether and ethyl acetate (50:1) to obtain 2-(4-(2-fluoroethoxy)phenyl)-4-methylthiazole-5-carboxylic acid ethyl ester 0.326g, with a yield of 40%.

[0162] Hydrolysis of ethyl 2-(4-(2-fluoroethoxy)phenyl)-4-methylthiazole-5-carboxylate: 0.74 g (2.4 mmol) of ethyl 2-(4-(2-fluoroethoxy)phenyl)-4-methylthiazole-5-carboxylate and 30 mL of sodium hydroxide solution (2M NaOH in V (THF / H2O / EtOH): 1 / 1 / 3) were added to a 250 mL round-bottom flask in sequence, and the reaction was carried out at 70°C for 40 min. After the reaction was completed, concentrated hydrochloric acid was added dropwise to adjust the pH to 1-2, and the organic phase was extracted with dichloromethane, dried and concentrated. After beating with dichloromethane for 24 h, compound 36 was obtained: 2-(4-(2-fluoroethoxyphenyl)-4-methylthiazole-5-carboxylic acid, white solid, yield 82%.

[0163] 1 H NMR(400MHz,DMSO-d6)δ13.24(br s,1H),7.93(d,J=8.9Hz,2H),7.10(d,J=8.9Hz,2H),4.77(dt,J=48.0Hz,J=4.0Hz,2H),4.33(dt,J=30.0Hz,J=4.0Hz,2H),2.66(s,3H). 13 C NMR (100MHz, DMSO-d6) δ168.7,163.5,161.0,160.0,128.7(2C),125.9,122.3,115.6(2C),83.3-81.7(d,1C),67.9-67.7(d,1c),17.5.

[0164] 16. Preparation of compound 37:

[0165] 3-Fluoropropanol (4.0 g, 51.23 mmol) was dissolved in dichloromethane (40 mL). After adding triethylamine (1.5 mL, 102.46 mmol), p-toluenesulfonyl chloride (12.7 g, 66.6 mmol) was added in batches under stirring in an ice bath. After the addition, the temperature was raised to room temperature and the reaction was allowed to proceed overnight. The reaction of the raw material was monitored by TLC. 100 mL of water and 100 mL of dichloromethane were added, and the organic phase was separated by extraction. The aqueous phase was quenched, and the combined organic phases were washed with saturated brine, dried over sodium sulfate, filtered, concentrated, and quickly passed through a column to obtain 11.5 g of 3-fluoropropyl 4-methylbenzenesulfonate with a yield of 96.6%.

[0166] Referring to the method for preparing compound 24, raw material H was replaced with 4-methylbenzenesulfonic acid-3-fluoropropyl ester to prepare compound 37: 2-(4-(3-fluoropropoxy)phenyl)-4-methylthiazole-5-carboxylic acid, a white solid, with a yield of 86%.

[0167] 1 H NMR(400MHz,DMSO-d6)δ13.29(br s,1H),7.92(d,J=8.0Hz,2H),7.04(d,J=8.6Hz,2H),4.56-4.69(dt,J=7.0Hz,2H),4.16(m,2H),2.66(s,3H),2.11-2.17(dt,2H). 13 CNMR(100MHz,DMSO-d6)δ168.9,163.5,161.3,159.9,128.6,125.6,122.2,1 15.6,82.03-80.43(d,1C),64.33-64.39(d,1C),30.02-30.21(d,1C),17.6.

[0168] 17. According to the method for preparing compound 24, replace raw materials G and H as shown in Table 4 to prepare compounds 38-39.

[0169] Table 4 Reaction substrate comparison table

[0170]

[0171]

[0172] Compound 38: 1H NMR (400 MHz, DMSO-d6) δ13.32 (br s, 1H), 7.97 (d, J = 8.9 Hz, 2H), 7.20 (d, J = 8.9 Hz, 2H), 4.87 (m, 2H), 2.66 (s, 3H), 13C NMR (100 MHz, DMSO-d6) δ168.39, 163.43, 161.0, 159.54, 128.7 (2C), 126.97, 125.72-122.96 (d, 1C), 122.68, 115.6.01 (2C), 64.56-65.59 (m, 1C), 17.5.

[0173] Compound 39: 1 H NMR(400MHz,DMSO-d6)δ13.26(br s,1H),7.90(d,J=8.8Hz,2H),7.04(d,J=8.6Hz,2H),3.90(d,J=7.0Hz,2H),2. 65(s,3H),1.22-1.24(m,1H),0.59(d,J=8.1Hz,2H),),0.35(d,J=4.7Hz,2H). 13 C NMR (100MHz, DMSO-d6) δ168.9,163.5,161.5,159.9,128.6(2C),125.4,122.2,115.6(2C),72.8,17.6,10.5,3.6(2C).

[0174] 18. Preparation of compound 40:

[0175] Ethyl 2-(4-(cyclopropylmethoxy)phenyl)-4-methylthiazole-5-carboxylate and hydroxylamine hydrochloride were mixed in ethanol, and sodium ethoxide in an amount equivalent to hydroxylamine hydrochloride was added under ice bath to obtain compound 40: 2-(4-(cyclopropylmethoxy)phenyl)-N-hydroxy-4-methylthiazole-5-carboxamide.

[0176] 19. Preparation of compound 41:

[0177] According to the method for preparing compound 24, ethyl 2-(4-hydroxyphenyl)-4-methylthiazole-5-carboxylate was used instead of raw material G, and bromomethylcyclobutane was used instead of raw material H to prepare compound 41 as a white solid with a yield of 81%.

[0178] 1H NMR(400MHz,DMSO-d6)δ13.28(br s,1H),7.90(d,J=7.1Hz,2H),7.05(d,J=6.6Hz,2H),4.02(s,2H),2.69-2.78(m,1H),2.65(s,3H),2.05-2.12(m,2H),1.82-1.92(m,2H). 13 C NMR (100MHz, DMSO-d6) δ168.9,163.4,161.7,160.0,128.6,125.4,122.1,115.6,72.2,34.3,24.8,18.6,17.6.

[0179] 20. Preparation of compound 42:

[0180] Phosphorus pentasulfide (9.78 g, 44 mmol, 2 eq) was added in batches to pre-cooled anhydrous ethanol (100 mL). After stirring at room temperature for 1.3 h, 4-aminobenzonitrile (2.6 g, 22 mmol) was added and the temperature was raised to 80 ° C for overnight reaction. After cooling to room temperature and concentrating, the reaction solution was slowly poured into saturated sodium bicarbonate and extracted with ethyl acetate. After drying and concentration, 1.5 g of crude product was taken, 30 mL of ethanol was added, 2-chloroacetoacetic acid ethyl ester (2.0 g) was added under stirring, and heated to reflux overnight. Concentrate under reduced pressure, evaporate ethanol, add 50 mL of ethyl acetate and then add 20 mL of saturated sodium bicarbonate solution, separate the organic phase, extract the aqueous phase twice with fresh ethyl acetate, combine the organic phases, wash with saturated brine, dry and concentrate the crude product, add methanol for crystallization, and filter to obtain 1.32 g of ethyl 2-(4-aminophenyl)-4-methylthiazole-5-carboxylate, a light yellow powder.

[0181] 2-(4-aminophenyl)-4-methylthiazole-5-carboxylic acid ethyl ester (0.485g, 1.85mmol) and triethylamine (0.28g, 2.775mmol, 1.5eq) were mixed in dichloromethane (5mL), and cyclopropylcarbonyl chloride (0.232g, 2.22mmol, 1.3eq) was added dropwise under ice bath, and the mixture was heated to room temperature for 1h, and water was added to separate the organic phase. Ethanol crystallization gave 0.342g of 2-(4-(cyclopropanecarboxamido)phenyl)-4-methylthiazole-5-carboxylic acid ethyl ester, with a yield of 56%.

[0182] Hydrolysis of ethyl 2-(4-(cyclopropanecarboxamido)phenyl)-4-methylthiazole-5-carboxylate: 0.79 g (2.4 mmol) of ethyl 2-(4-(cyclopropanecarboxamido)phenyl)-4-methylthiazole-5-carboxylate and 30 mL of sodium hydroxide solution (2M NaOH in V (THF / H2O / EtOH): 1 / 1 / 3) were added to a 250 mL round-bottom flask, and the reaction was carried out at 70° C. for 40 min. After the reaction was completed, concentrated hydrochloric acid was added dropwise to adjust the pH to 1-2, and the organic phase was extracted with dichloromethane, dried and concentrated. After slurrying with dichloromethane for 24 h, compound 42 was obtained: 2-(4-(cyclopropanecarboxamido)phenyl)-4-methylthiazole-5-carboxylic acid, a white solid, with a yield of 56%.

[0183] 1 H NMR(400MHz,DMSO-d6)δ13.28(br s,1H),10.50(s,1H),7.92(d,J=8.7Hz,2H),7.74(d,J=9.0Hz,2H),2.66(s,3H),1.78-1.85(m,1H),0.82-0.84(m,4H). 13 C NMR (100MHz, DMSO-d6) δ172.5,168.8,163.5,160.1,142.5,127.8(2C),127.3,122.4,119.5(2C),17.6,15.2,8.0(2C).

[0184] 21. Preparation of compound 43:

[0185] 4-Fluorobenzonitrile (2.0 g, 16.51 mmol), propanethiol (2.52 g, 33.03 mmol, 2 eq) and potassium carbonate (4.8 g, 34.68 mmol, 2.1 eq) were mixed in DMF (20 mL), heated to 50 ° C, reacted for 4 h, cooled, 60 mL of ethyl acetate and 40 mL of water were added, the organic phase was separated, the aqueous phase was extracted with 40 mL of ethyl acetate, the organic phases were combined, washed twice with 30 mL of water, washed with saturated brine, concentrated and used directly in the next reaction.

[0186] According to the method for preparing compound 9, the raw material C was replaced with 4-propylthiobenzonitrile to prepare ethyl 2-(4-(propylthio)phenyl)-4-methylthiazole-5-carboxylate.

[0187] Dissolve ethyl 2-(4-(propylthio)phenyl)-4-methylthiazole-5-carboxylate (0.456g, 1.42mmol) in dichloromethane (5mL), add m-chloroperbenzoic acid (0.734g, 4.26mmol, 3eq) in batches under ice bath, warm to room temperature and react overnight. Add 40mL of dichloromethane, add 10mL of saturated sodium bicarbonate under ice bath, stir for 1h, separate the organic phase, wash with saturated sodium bicarbonate, and separate through column to obtain ethyl 2-(4-(propylsulfonyl)phenyl)-4-methylthiazole-5-carboxylate 0.431g, yield 86%.

[0188] Hydrolysis of ethyl 4-methyl-2-(4-(propylsulfonyl)phenyl)-4-methylthiazole-5-carboxylate: 0.85 g (2.4 mmol) of compound 4-methyl-2-(4-(propylsulfonyl)phenyl)-4-methylthiazole-5-carboxylate and 30 mL of sodium hydroxide solution (2M NaOH in V (THF / H2O / EtOH): 1 / 1 / 3) were added to a 250 mL round-bottom flask in sequence, and the reaction was carried out at 70°C for 40 min. After the reaction was completed, concentrated hydrochloric acid was added dropwise to adjust the pH to 1-2, and the organic phase was extracted with dichloromethane, dried and concentrated, and compound 43: 2-(4-(propylsulfonyl)phenyl)-4-methylthiazole-5-carboxylic acid was obtained after dichloromethane beating for 24 h. White solid, yield 64%.

[0189] 1 H NMR(400MHz,DMSO-d6)δ13.60(br s,1H),8.24-8.26(m,2H),8.02(d,J=8.1Hz,2H),3.37(t,2H),2.71(s,3H),1.54-1.64(m,2H),0.93(d,J=7.4Hz,3H). 13 C NMR (100MHz, DMSO-d6) δ166.7,163.2,160.3,141.3,137.0,129.2(2C),127.8(2C),125.0,56.6,17.5,16.6,13.0.

[0190] 22. Preparation of compound 44: Compound 45:

[0191] According to the method for preparing compound 43, propyl mercaptan was replaced by cyclopropylmethyl mercaptan to prepare compound 44 as a white solid with a yield of 73%.

[0192] 1H NMR (400MHz, DMSO-d6) δ8.22(d,J=8.5Hz,2H),8.02(d,J=8.5Hz,2H),3.33(d,J=6 .5Hz,2H),2.69(s,3H),0.83-0.91(m,1H),0.43-0.48(m,2H),0.10-0.13(m,2H). 13 CNMR(100MHz,DMSO-d6)δ166.7,163.2,160.3,141.6,137.0,129.5(2C),127.6(2C),125.0,59.7,17.5,5.0,4.3(2C).

[0193] According to the method for preparing compound 43, propyl mercaptan was replaced by isobutyl mercaptan to prepare compound 45. White solid, yield 75%.

[0194] 1 H NMR(400MHz,DMSO-d6)δ13.60(br s,1H),8.25(d,J=8.0Hz,2H),8.04(d,J=8.6Hz,2H),3.29(d,J=6.1Hz,2H),2.71(s,3H),1.99-2.09(m,1H),0.99(d,J=6.7Hz,3H). 13 C NMR (100MHz, DMSO-d6) δ166.7,163.2,160.3,142.2,136.9,129.0(2C),127.8(2C),125.0,62.3,24.2,22.6(2C),17.5.

[0195] 23. Preparation of compound 46:

[0196] According to the method for preparing compound 43, propylthiol was replaced by cyclopropylmethanol to obtain 4-(cyclopropylmethoxy)benzonitrile.

[0197] According to the method for preparing compound 9, raw material C was replaced by 4-(cyclopropylmethoxy)benzonitrile, and raw material D was replaced by ethyl chloroformyl acetate to prepare compound 46: 2-(4-(cyclopropylmethoxy)phenyl)thiazole-5-carboxylic acid, a white solid, with a yield of 36%.

[0198] 1H NMR (400MHz, DMSO-d6) δ13.06(s,1H),8.41(s,1H),7.89(d,J=8.4Hz,2H),7.06(d,J=8.9 Hz,2H),3.98(d,J=7.2Hz,2H),1.22-1.28(m,1H),0.56-0.61(m,2H),0.33-0.36(m,2H). 13 C NMR (100MHz, DMSO-d6) δ167.8,162.5,161.1,148.3,128.5,128.3(2C),125.6,115.6(2C),72.8,10.5,3.6(2C).

[0199] 24. Preparation of compound 47:

[0200] Dissolve ethyl 3-oxopentanoate (2.0 g, 13.87 mmol) in dichloromethane (25 mL), add sulfonyl chloride (2.0 g, 14.57 mmol, 1.05 eq) dropwise under ice bath, react at room temperature overnight. Add 60 mL of dichloromethane, wash with water, separate the organic phase, and concentrate to obtain ethyl 2-chloro-3-oxopentanoate.

[0201] According to the method for preparing compound 9, raw material C was replaced with 4-(cyclopropylmethoxy)benzonitrile, and raw material D was replaced with ethyl 2-chloro-3-oxopentanoate to prepare 2-(4-(cyclopropylmethoxy)phenyl)-4-ethylthiazole-5-carboxylic acid, a white solid with a yield of 52%.

[0202] 1 H NMR (400MHz, DMSO-d6) δ13.28(s,1H),7.91(d,J=8.6Hz,2H),7.04(d,J=9.0Hz,2H),3.90(d,J=6.8Hz,2H ),3.08(q,J=7.3Hz,2H),1.25(t,J=7.3Hz,3H),1.23-1.27(m,1H),0.57-0.61(m,2H),0.33-0.37(m,2H). 13 C NMR (100MHz, DMSO-d6) δ169.2,165.5,163.3,161.5,128.6(2C),125.4,121.5,115.5(2C),72.8,24.0,14.1,10.5,3.6(2C).

[0203] 25. Preparation of compound 48: Compound 49:

[0204] According to the method for preparing compound 9, raw material C was replaced by 4-(cyclopropylmethoxy)benzonitrile, and raw material D was replaced by ethyl 2-chloro-4,4,4-trifluoroacetoacetate to prepare compound 48: 2-(4-(cyclopropylmethoxy)phenyl)-4-(trifluoromethyl)thiazole-5-carboxylic acid, a white solid, with a yield of 82%.

[0205] H NMR (400MHz, DMSO-d6) δ7.94(d,J=8.0Hz,2H),7.07(d,J=8.0Hz,2H),3.91(d ,J=4.0Hz,2H),1.20-1.29(m,1H),0.55-0.61(m,2H),0.33-0.39(m,2H).13C NMR(101MHz,DMSO-d6)δ170.16,162.19,160.30,144.75,130.72,129.05,124.23,120.37,115.77,72.95,10.45,3.59.HRMS(ESI)m / z[M+H]+calcd.for C15H13F3NO3S:344.0568; found344.0574.

[0206] According to the method for preparing compound 9, raw material C was replaced by 4-(cyclopropylmethoxy)benzonitrile, and raw material D was replaced by diethyl chloromalonate to prepare compound 49: 2-(4-(cyclopropylmethoxy)phenyl)-4-methoxythiazole-5-carboxylic acid, a white solid, with a yield of 88%.

[0207] 1 H NMR(400MHz,DMSO-d6)δ7.91(d,J=9.0Hz,2H),7.06(d,J=8.7Hz,2H),4.01(s,3H) ,3.9(d,J=5.2Hz,4H),1.23-1.26(m,1H),0.59-0.60(dd,2H),0.34-0.36(dd,2H). 13 CNMR(100MHz,DMSO-d6)δ167.04,165.33,162.44.1,161.84,128.2(2C),125.2,115.6(2C),100.5,72.9,58.1,10.46,3.6(2C).

[0208] 26. Preparation of compound 50:

[0209] According to the method for preparing compound 9, raw material C was replaced by 4-(cyclopropylmethoxy)benzonitrile, and raw material D was replaced by diethyl chlorosuccinate to prepare compound 50: 2-(4-(cyclopropylmethoxy)phenyl)-4-ethoxythiazole-5-carboxylic acid was obtained as a white solid with a yield of 84%.

[0210] 1 H NMR(400MHz,DMSO-d6)δ12.67(br s,1H),7.89(d,J=8.3Hz,2H),7.05(d,J=8.7Hz,2H),4.52(q,J=6.4Hz,2H),3.90(d,J=7.2Hz ,2H),1.37(t,J=6.8Hz,3H),1.20-1.28(m,1H),0.59(d,J=7.2Hz,2H),0.35(d,J=4.2Hz,2H). 13 C NMR (100MHz, DMSO-d6) δ167.0,164.9,162.5,161.8,128.2,125.3,115.6,100.7,72.9,66.7,15.4,10.5,3.6.

[0211] 27. Preparation of compound 51:

[0212] Tert-butyl acetoacetate (3.0 g, 18.97 mmol), potassium carbonate (4 g, 28.46 mmol, 1.5 eq) and TBAI (0.1 g) were mixed in DMF (20 mL), ethyl bromoacetate (3.5 g, 20.86 mmol, 1.1 eq) was added, heated to 80 ° C, reacted for 4 h, cooled, extracted, and passed through a column to obtain 2.85 g of 1-(tert-butyl)-4-ethyl-2-acetyl succinate, with a yield of 61.5%. 1-(tert-butyl)-4-ethyl-2-acetyl succinate and sulfonyl chloride were used to prepare 1-(tert-butyl)-4-ethyl-2-acetyl-2-chlorosuccinate. Referring to the preparation method of compound 24, raw material F and ethyl 2-chloroacetoacetate were replaced by 4-(cyclopropylmethoxy)thiobenzamide and 1-(tert-butyl)-4-ethyl-2-acetyl-2-chlorosuccinate, respectively, to prepare compound 51: 2-(2-(4-(cyclopropylmethoxy)phenyl)-4-methylthiazol-5-yl)acetic acid (51), a white solid, with a yield of 57%.

[0213] 1H NMR(400MHz,DMSO-d6)δ12.7(s,br,1H),7.8(d,2H),7.0(d,2H),3.86(t,2H),3.8 1(s,2H),2.3(s,3H),1.22-1.26(m,1H),0.58-0.61(dd,2H),0.32-0.36(dd,2H), 13 C NMR (100MHz, DMSO-d6) δ171.9,164.3,160.4,150.5,127.6(2C),126.4,123.6,115.4(2C),72.7,32.0,15.3,15.3,10.5,3.6(2C).

[0214] 28. Preparation of compound 52:

[0215] According to the method for preparing compound 9, raw material C was replaced with 4-(cyclopropylmethoxy)benzylacetonitrile to prepare compound 52: 2-(4-(cyclopropylmethoxy)benzyl)-4-methylthiazole-5-carboxylic acid, a white solid, with a yield of 63%.

[0216] 1 H NMR(400MHz,DMSO-d6)δ13.14(br s,1H),7.24(d,J=8.6Hz,2H),6.90(d,J=8.6Hz,2H),4.19(s,2H),3.79(d,J=6.4 Hz,2H),2.57(s,3H),1.16-1.26(m,1H),0.54-0.58(dd,2H),0.29-0.33(dd,2H). 13 C NMR (100MHz, DMSO-d6) δ174.2,163.4,159.1,158.2,130.7(2C),129.7,123.0,115.1(2C),72.4,38.4,17.3,10.6,3.5(2C).

[0217] 29. Preparation of compound 53:

[0218] According to the preparation method of compound 43, morpholine was used to replace propanethiol to prepare 4-morpholinobenzonitrile. According to the preparation method of compound 9, raw material C was replaced with 4-morpholinobenzonitrile to prepare compound 53: 2-(4-morpholinophenyl)-4-methylthiazole-5-carboxylic acid, white solid, yield 89%.

[0219] 1H NMR (400MHz, DMSO-d6) δ7.82(d,J=9.0Hz,2H),7.02(d,J=8.7Hz,2H),3.74(t,J=5.2Hz,4H),3.25(t,J=4.6Hz,4H),2.64(s,3H). 13 C NMR (100MHz, DMSO-d6) δ169.4,163.5,160.1,153.3,128.2(2C),122.9,121.2,114.7(2C),66.3(2C),47.6(2C),17.6.

[0220] 30. Using the method for preparing compound 24, replace raw material E accordingly according to Table 5, replace raw material F with the prepared compound, then replace raw material G with the corresponding compound further prepared, and replace raw material H accordingly according to Table 5 to prepare compounds 54-65.

[0221] Table 5 Reaction substrate comparison table

[0222]

[0223]

[0224] The raw material of compound 60, 3,5-dichloro-4-hydroxybenzonitrile, was prepared according to Industrial & Engineering Chemistry Research, 51(10), 3881-3886; 2012.

[0225] The raw material of compound 63, 3-bromo-5-fluoro-4-hydroxybenzonitrile, was prepared according to the reference Journal of Medicinal Chemistry, 61(17), 7796-7813; 2018.

[0226] The raw material of compound 64, 3-bromo-5-chloro-4-hydroxybenzonitrile, was prepared according to ACS Infectious Diseases, 4(4), 592-604; 2018.

[0227] The raw material of compound 65, 3,5-dibromo-4-hydroxybenzonitrile, was prepared according to patent CN105859583.

[0228] Compound 54: 1H NMR (400MHz, DMSO-d6) δ13.34(s,1H),7.68-7.73(m,2H),7.10(t,J=9.4Hz,1H),3.75(t,J=4.9Hz,4H),3 .12(t,J=4.9Hz,4H),2.65(s,3H),2.23(s,3H),1.22-1.29(m,1H),0.57-0.61(m,2H),0.34-0.38(m,2H). 13 C NMR (100MHz, DMSO-d6) δ167.6,163.3,160.0,142.4,126.5,123.9,122.6,119.6,114.4,114.1,66.5,50.4,17.5.

[0229] Compound 55: 1 H NMR(400MHz,DMSO-d6)δ13.35(br s,1H),7.73-7.79(m,2H),7.27(t,J=8.7Hz,1H),4.08(t,J=6.6Hz,2H),2.65(s,3H),1.75-1.80(m,2H),0.99(t,J=7.4Hz,3H). 13 C NMR(100MHz,DMSO-d6)δ167.56-167.58(d,1C),163.3,160.0,153.2,150.8-149.4(t,1C),125.67 -125.6(d,1C,123.96-124.0(d,1H),122.9,115.5,114.26-114.06(d,1C),70.8,22.3,17.5,10.6.

[0230] Compound 56: 1 H NMR (400MHz, DMSO-d6) δ13.35(br s,1H),7.74-7.80(m,2H),7.26-7.31(m,1H),3.80(s,2H),2.66(s,3H),1.03(s,9H). 13 C NMR(100MHz,DMSO-d6)δ167.54-,167.57(d,1C),163.3,160.0,153.3,149.87-150.86(t,1C),125.63-125.7( d,1C),123.95-123.98(d,1C),122.9,115.64-115-66(d,1C),114.03-114.23(1C),78.8,32.2,26.6(3C),17.5

[0231] Compound 57: 1 H NMR(400MHz,DMSO-d6)δ13.29(br s,1H),7.78(d,J=12.2Hz,1H),7.74(d,J=9.3Hz,1H),7.25(t,J=8.7Hz,1H),3.98(d, J=7.1Hz,2H),2.65(s,3H),1.25-1.28(m,1H),0.58-0.63(m,2H),0.35-0.38(m,2H),. 13 C NMR (101MHz, DMSO-d6) δ167.6,163.4,160.0,153.2,150.8,149.5,149.4,125.6,124.0-,122.9,115.7,114.2,73.9,17.5,10.4,3.6.HRMS (ESI) m / z calcd.for C 15 H 15 FNO3S[M+H] + :308.0751; found:308.0761.

[0232] Compound 58: 1 H NMR (400MHz, DMSO-d6) δ7.96 (d, J = 2.2Hz, 1H), 7.86 (dd, J = 8.6Hz, J = 2.2Hz, 1H), 7.21 (J = 8.7Hz, 1H), 3.78 (s, 2H), 2.65 (s, 3H), 1.04 (s, 9H). 13 C NMR (100MHz, DMSO-d6) δ167.3,163.3,160.0,156.9,127.9,127.4,126.1,122.9,122.8,114.5,78.8,32.3,26.7(2C),17.5.

[0233] Compound 59: 1 H NMR(400MHz,DMSO-d6)δ13.29(br s,1H),7.96-8.00(m,1H),7.82-7.90(m,1H),7.17-7.26(m,1H),4.00(d,2H),2.6 5(s,3H),1.28(m,1H),0.59-0.64(dd,J=7.5Hz,2H),0.37-0.40(dd,J=4.3Hz,2H). 13C NMR (100MHz, DMSO-d6) δ167.3,163.3,160.0,156.6,127.9,127.3,126.1,122.9,122.6,114.6,73.9,17.5,10.3,3.6(2C).

[0234] Compound 60: 1 H NMR (400MHz, DMSO) δ7.96 (s, 2H), 3.91 (d, J = 7.2Hz, 2H), 2.65 (s, 3H), 1.26–1.30 (m, 1H), 0.56–0.60 (m, 2H), 0.30–0.34 (m, 2H). 13 C NMR(101MHz,DMSO-d6)δ165.3,163.2,160.0,153.0,130.1,127.2,124.5,78.8,17.5,11.1,3.7.HRMS(ESI)m / zcalcd.for C 15 H 14 35 Cl2NO3S[M+H] + :358.0066; found:358.0057.calcd.forC 15 H 14 35 Cl 37 ClNO3S[M+H] + :360.0036; found:360.3236.calcd.for C 15 H 14 37 Cl2NO3S[M+H] + :362.0007; found:362.3267.

[0235] Compound 61: 1 H NMR(400MHz,DMSO-d6)δ13.31(br s,1H),8.11(m,1H),7.88(m,1H),7.16(m,1H),4.00(d,2H),2.65(s,3H),1. 26-1.30(s,1H),0.59-0.63(dd,J=7.5Hz,2H),0.37-0.41(dd,J=4.3Hz,2H). 13C NMR (100MHz, DMSO-d6) δ167.1,163.3,160.0,157.5,130.9,128.0,126.5,122.9,114.4,112.2,73.8,17.5,10.3,3.5(2C).

[0236] Compound 62: 1 H NMR(400MHz,DMSO-d6)δ13.46(br s,1H),7.98(s,1H),7.84(d,J=11.8Hz,1H),4.00(d,J=7.1Hz,2H),2.65(s,3H),1.74-1.83(m,2H),1.02(t,J=7.5Hz,3H). 13 C NMR (100MHz, DMSO-d6) δ167.2,163.3,160.0,157.5,130.9,128.1,126.5,122.9,114.3,112.2,70.1,22.3,17.5,10.8.

[0237] Compound 63: 1 H NMR(400MHz,DMSO-d6)δ13.44(br s,1H),7.98(s,1H),7.84(d,J=11.6Hz,1H),4.00(d,J=7.1Hz,2H),2.65(s,3H),1.19-1.27(m,1H),0.54-0.85(m,2H),0.29-0.33(m,1H). 13 C NMR(100MHz,DMSO-d6)δ165.5,163.2,160.0,154.6-157.1(d,1C),146.18-146.31(d,1c),129.53-129.61(d, 1C),126.57-126.60(d,1C),124.3,118.64-118.68(d,1C),114.8-115.07(d,1c),79.2,17.4,11.2,3.6(2C).

[0238] Compound 64: 1 H NMR(400MHz,DMSO-d6)δ13.50(br s,1H),8.02-8.17(d,2H),3.91(d,J=6.9Hz,2H),2.67(s,3H),1.27-1.36(m,1H),0.54(dd,J=7.7Hz,2H),0.35(dd,J=4.5Hz,2H). 13C NMR (100MHz, DMSO-d6) δ165.1,163.2,160.0,154.0,130.7,130.0,129.8,127.9,124.5,119.7,78.7,17.5,11.1,3.7(2C)

[0239] Compound 65: 1 H NMR (400MHz, DMSO-d6) δ13.30(br s,1H),7.65(s,2H),3.66(d,J=7.0Hz,2H),2.67(s,3H),1.3-1.32(m,1H),0.58-0.63(dd,2H),0.37-0.39(dd,2H). 13 C NMR (100MHz, DMSO-d6) δ165.0,163.2,160.0,154.9,131.2,130.8(2C),124.5,119.3(2C),78.5,17.5,11.1,3.8(2C).

[0240] 31. Preparation of compound 66:

[0241] According to the method for preparing compound 9, the raw material C was replaced with 3-bromo-5-fluoro-4-hydroxybenzonitrile to prepare the compound 2-(3-bromo-5-fluoro-4-hydroxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester. 2-(3-bromo-5-fluoro-4-hydroxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester and 1-bromo-2,2-dimethylpropane were dissolved in DMF (0.35mmol / mL), potassium carbonate (2.1eq.) was added, and heated to 80℃ for 5h. After the reaction was completed by TLC detection, the reaction solution was cooled to room temperature, and water (20mL) and ethyl acetate (EA, 40mL) were added for extraction. The organic layer was washed with water (20mL×3) and brine, and dried with Na2SO4. Then it was hydrolyzed to obtain compound 66 as a white solid with a yield of 73%.

[0242] 1 H NMR (400MHz, DMSO-d6) δ13.52(br s,1H),8.01(s,2H),3.70(s,2H),2.67(s,3H),1.08(s,9H). 13 C NMR (100MHz, DMSO-d6) δ165.2,163.1,160.0,153.0,130.2,129.8,127.3,124.5,82.7,32.8,26.6,17.5.

[0243] 32. Referring to the method for preparing compound 24, replace raw material E according to Table 6, replace raw material F with the corresponding compound obtained, and then replace raw material G with the corresponding compound. At the same time, replace raw material H according to Table 5 to prepare compounds 67-70.

[0244] Table 6 Reaction substrate comparison table

[0245]

[0246] Compound 67: 1 H NMR (400MHz, DMSO-d6) δ13.25(s,1H),7.77(s,2H),7.76(d,J=7.1Hz,1H),7.00(d,J=9.4Hz,1H),3.92( d,J=6.8Hz,2H),2.64(s,3H),2.23(s,3H),1.22-1.29(m,1H),0.57-0.61(dd,2H),0.34-0.38(dd,2H). 13 C NMR (100MHz, DMSO-d6) δ169.2,163.5,160.0,159.7,128.9,127.2,126.3,124.9,121.9,112.1,72.7,17.5,16.4,10.5,3.4(2C).

[0247] Compound 68: 1 H NMR(400MHz,DMSO-d6)δ13.30(br s,1H),7.65(s,2H),3.66(d,J=7.0Hz,2H),2.65(s,3H),2.30(s,6H),1.20-1.30(m,1H),0.55-0.59(m,2H),0.28-0.32(m,2H). 13 C NMR (100MHz, DMSO-d6) δ168.9,163.4,159.9,158.6,132.2(2C),128.1,127.3(2C),122.7,77.0,17.5,16.5(2C),11.5,3.5(2C).

[0248] Compound 69: 1 H NMR(400MHz,DMSO-d6)δ13.39(br s,1H),8.10-8.20(m,2H),7.32-7.39(m,1H),4.06-4.11(m,2H),2.67(s,3H),1.24-1.28(m,1H),0.59(dd,J=7.9Hz,2H),0.38(dd,J=4.6Hz,2H).13 C NMR(100MHz,DMSO-d6)δ167.2,163.3,160.0,158.9,132.9,125.06 124.9,123.1,122.3,118.1-118.4(m,1C),115.2,73.6,17.5,10.2,3.3(2C).

[0249] Compound 70: 1 H NMR (400MHz, DMSO-d6) δ13.37(br s,1H),8.23(s,1H),8.17(d,1H),7.33(d,2H),2.64(s,3H),2.05-2.12(m,1H),1.02(d,6H).

[0250] 33. Preparation of compound 71

[0251] N-Boc-piperidine-4-thiocarboxamide (5.0g, 25.5mmol, 1eq.) and ethyl 2-chloroacetoacetate (4.4g, 26.6mmol, 1.3eq.) were mixed in ethanol (50mL) and heated to reflux for 6h. After the reaction solution was cooled to room temperature, ethanol was distilled off under reduced pressure, and ethyl acetate (2×100mL) and saturated sodium bicarbonate (NaHCO3) solution were added to extract the product. Wash with water (2×40mL), wash with saturated brine, and dry over anhydrous sodium sulfate. Filter and concentrate. The obtained compound was directly dissolved in dichloromethane (60mL), and 4M hydrogen chloride 1,4-dioxane solution (30mL) was added dropwise under ice-water bath. After addition, the reaction was continued at room temperature for 2h. Concentrate under reduced pressure until no liquid drops. After adding diethyl ether (40 mL), the mixture was stirred at room temperature for half an hour, and petroleum ether (50 mL) was added. The mixture was filtered to collect the white solid 4-methyl-2-(piperidin-4-yl)thiazole-5-carboxylic acid ethyl ester hydrochloride, which was directly used in the next step.

[0252] 4-Methyl-2-(piperidin-4-yl)thiazole-5-carboxylic acid ethyl ester hydrochloride (raw material I, 0.4 g, 1.36 mmol) was dissolved in DCM (4 mL), triethylamine (1 mL) was added under ice bath, and then isovaleryl chloride (raw material J, 0.333 g, 2.76 mmol, 2.0 eq) was added dropwise, and the mixture was heated to room temperature for 30 minutes. 30 mL of dichloromethane and 20 mL of water were added, and the organic phase was separated, washed with saturated ammonium chloride, and concentrated and passed through a column to obtain 0.371 g of 2-(1-(3-methylbutyryl)piperidin-4-yl)-4-methylthiazole-5-carboxylic acid ethyl ester. Hydrolysis of ethyl 2-(1-(3-methylbutyryl)piperidin-4-yl)-4-methylthiazole-5-carboxylate gave compound 71: 2-(1-(3-methylbutyryl)piperidin-4-yl)-4-methylthiazole-5-carboxylic acid as a white solid in a yield of 82%.

[0253] 1 H NMR (400MHz, DMSO-d6) δ13.23(s,1H),4.46(d,J=13.0Hz,1H),3.20-3.27(m,1H),3.10-33.17(t,J=12.5Hz,1H),2.65-2.72(t,J=11 .7Hz,1H),2.58(s,3H),2.20-2.22(d,J=7.0Hz,2H),1.93-2.07(m,3H),1.53-1.63(m,1H),1.45-1.51(m,1H),0.90(d,J=6.6Hz,1H). 13 C NMR (100MHz, DMSO-d6) δ177.0,170.3,163.4,158.8,122.2,45.2,41.6,41.0,32.8,32.1,25.6,23.0,23.0,17.4.

[0254] 34. According to the method for preparing compound 71, replace raw materials I and J accordingly according to Table 7 to prepare compounds 72-78.

[0255] Table 7 Reaction substrate comparison table

[0256]

[0257] Compound 72: 11H NMR (400 MHz, DMSO-d6) δ 13.22 (s, 1H), 4.44 (d, J = 13.6 Hz, 1H), 3.93 (d, J = 13.6 Hz, 1H), 3.20 - 3.26 (m, 1H), 3.14 (t, J = 11.3 Hz, 1H), 2.68 (dt, J = 11.3 Hz, 1H), 2.58 (s, 3H), 2.30 (t, J = 6.8 Hz, 2H), 2.01 (t, J = 14.7 Hz, 2H), 1.41 - 1.65 (m, 4H), 0.89 (t, J = 7.9 Hz, 3H). 13 13C NMR (100 MHz, DMSO-d6) δ 177.0, 170.8, 163.4, 158.8, 122.2, 45.0, 41.0, 34.7, 32.7, 32.1, 18.7, 17.4, 14.3.

[0258] Compound 73: 1 1H NMR (400 MHz, DMSO-d6) δ 13.20 (s, 1H), 3.57 (d, J = 13.2 Hz, 2H), 3.09 - 3.17 (m, 1H), 3.12 (dt, J = 7.4 Hz, 2H), 2.78 - 2.84 (dt, J = 11.7 Hz, 2H), 2.73 (s, 3H), 2.58 (s, 3H), 1.99 (dt, J = 11.0 Hz, 2H), 1.61 - 1.65 (dt, 2H), 1.05 (t, J = 7.1 Hz, 3H). 13 13C NMR (100 MHz, DMSO-d6) δ 177.3, 164.1, 163.4, 158.8, 122.1, 46.7 (2C), 44.6, 40.4, 35.6, 32.0 (2C), 17.4, 12.9.

[0259] Compound 74: 1 1H NMR (400 MHz, DMSO-d6) δ 13.25 (s, 1H), 3.65 (d, J = 12.5 Hz, 2H), 3.11 - 3.16 (m, 1H), 3.03 (t, J = 7.8 Hz, 2H), 2.95 (dt, J = 10.9 Hz, 2H), 2.59 (s, 3H), 2.11 (dt, J = 10.9 Hz, 2H), 1.63 - 1.74 (m, 4H), 0.99 (t, J = 7.5 Hz, 3H). 13 13C NMR (100 MHz, DMSO-d6) δ 176.7, 163.4, 158.9, 122.3, 50.2, 45.2 (2C)), 39.4, 31.9 (2C), 17.4, 16.9, 13.3.

[0260] Compound 75: 1 H NMR (400MHz, DMSO-d6) δ13.23(s,1H),3.67(dd,J=12.2Hz,2H),3.12-3.18(m,1H),2.99(dt,J=11.9Hz,J=2.0 Hz,2H),2.59-2.65(m,1H),2.59(s,3H),2.11-2.15(dt,J=11.0Hz,2H),1.65-1.77(m,2H),0.92-1.02(m,4H). 13 C NMR (100MHz, DMSO-d6) δ176.7,163.4,158.9,122.3,45.9(2C),39.4,31.7(2C),25.8,17.4,4.4(2C).

[0261] Compound 76: 1 H NMR(400MHz,DMSO-d6)δ13.24(s,1H),3.61-3.67(m,6H),3.13-3.19(m,1H),3.11-3.13(m,4H),2.98-3 .05(dt,J=11.7Hz,2H),2.59(s,3H),20.1(dd,J=11.0Hz,2H),1.63-1.73(m,2H),1.05(t,J=7.1Hz,3H). 13 C NMR (100MHz, DMSO-d6) δ176.7,163.4,158.9,122.2,66.1(2C),46.6(2C),46.1(2C),39.4,31.8(2C),17.4.

[0262] Compound 77: 1 H NMR (400MHz, DMSO-d6) δ13.22(s,1H),8.94(d,J=2.4Hz,1H),8.91(d,J=5.0Hz,1H),8.20(dd,J=7.8Hz,1H),7.72(dd,J=7.8Hz,J=5 .5Hz,1H),3.76(dt,J=11.6Hz,2H),3.00-3.07(m,1H),2.56(s,3H),2.48-2.55(m,2H),2.10(d,J=11.7Hz,2H),1.64-1.74(m,2H). 13C NMR (100MHz, DMSO-d6) δ176.4,163.3,158.8,154.2,148.2,136.0,132.8,125.0,122.3,45.8(2C),38.8,31.3(2C),17.4.

[0263] Compound 78: 1 H NMR (400MHz, DMSO-d6) δ13.23(s,1H),7.70(d,J=8.8Hz,2H),7.17(d,J=8.8Hz,2H),3.86(s,3H),3.67(d,J=12. 1Hz,2H),2.94-3.02(m,1H),2.56(s,3H),2.37(dt,J=10.0Hz,2H),2.09(dd,J=10.0Hz,2H),1.63-1.73(m,2H). 13 C NMR (100MHz, DMSO-d6) δ176.5,163.3,163.2,158.8,130.2(2C),127.4,122.3,115.0(2C),56.2,46.0(2C),39.0,31.3(2C),17.4.

[0264] 35. Preparation of compound 79:

[0265] 4-Methyl-2-(piperidin-4-yl)thiazole-5-carboxylic acid ethyl ester hydrochloride (0.53g, 1.83mmol) and 2-(tetrahydrofuran-2-yl)acetic acid (0.25g, 1.92mmol, 1.05eq) were mixed in DMF (5mL), EDCI.HCl (0.454g, 2.38mmol, 1.3eq) was added under ice bath, and then 0.3ml of triethylamine was added, and the mixture was heated to room temperature and stirred for 1h. 60ml of ethyl acetate was added, and the mixture was washed with water three times, washed with saturated brine, and passed through a column to obtain 0.57g of 2-(1-(2-(tetrahydrofuran-2-yl)acetyl)piperidin-4-yl)-4-methylthiazole-5-carboxylic acid ethyl ester, with a yield of 85%. Ethyl 2-(1-(2-(tetrahydrofuran-2-yl)acetyl)piperidin-4-yl)-4-methylthiazole-5-carboxylate was hydrolyzed to obtain compound 79: 2-(1-(2-(tetrahydrofuran-2-yl)acetyl)piperidin-4-yl)-4-methylthiazole-5-carboxylic acid, a white solid, with a yield of 88%.

[0266] 1H NMR (400MHz, DMSO-d6) δ13.22(s,1H),4.43(d,J=13.3Hz,1H),3.93(d,J=13 .3Hz,1H),3.82(t,J=7.6Hz,1H),3.68-3.73(m,1H),3.61(q,J=7.8Hz,1H), 3.21-3.26(m,1H),3.10-3.17(m,2H),2.69(t,J=12.0Hz,1H),2.58(s,3H), 2.39-2.53(m,3H),1.97-2.06(m,3H),1.56-1.66(m,1H),1.42-1.52(m,2H). 13 C NMR (100MHz, DMSO-d6) δ177.0,170.1,163.4,158.8,122.2,72.9,67.2,44.9,41.1,40.18,36.6,35.7,32.6,32.0,17.4.

[0267] 36. According to the method for preparing compound 71, replace raw materials I and J accordingly according to Table 8 to prepare compounds 80-81.

[0268] Table 8 Reaction substrate comparison table

[0269]

[0270] Compound 80: 1 H NMR (400MHz, DMSO-d6) δ13.22(s,1H),4.43(d,J=13.1Hz,1H),3.96(d,J=13.6Hz,1H),3.55(t,J=6.2Hz,2H),3.20-3.28(m,1H),3.23(s,3H ),3.14(t,J=11.5Hz,1H),2.70(t,J=11.8Hz,1H),2.56-2.61(m,2H),2.58(s,3H),1.97-2.06(m,2H),1.56-1.66(m,1H),1.42-1.52(m,2H). 13 C NMR (100MHz, DMSO-d6) δ177.0,169.1,163.4,158.8,122.2,68.8,58.4,45.1,41.1,40.21,33.0,32.6,32.0,17.4.

[0271] Compound 81: 1H NMR (400MHz, DMSO-d6) δ13.24(s,1H),4.44(d,J=13.0Hz,1H),3.91(d,J=13 .6Hz,1H),3.20-3.26(m,1H),3.13(t,J=12.3Hz,1H),2.69(t,J=11.1Hz,1H) ,2.58(s,3H),2.28(d,J=6.5Hz,2H),2.03(m,2H),1.55-1.65(m,1H),1.42-1 .53(m,1H),0.92-0.99(m,1H),0.45(d,J=7.2Hz,2H),0.11(d,J=3.9Hz,2H). 13 C NMR (100MHz, DMSO-d6) δ177.0,170.6,163.4,158.8,122.2,45.2,41.0,38.0(2C),32.7,32.1,17.4,7.76,4.77,4.73.

[0272] Example 2: Demonstration of the inhibitory activity of compounds on miR-21 biosynthesis in activity screening experiments

[0273] Hela-luciferase-miR-21 stable cervical cancer cell lines were cultured in DMEM high glucose medium containing 300 μg / mL G418, 10% fetal bovine serum (FBS) and 1% penicillin / streptomycin. Hepatoma cells SK-HEP-1 (from ATCC) and HCCLM3 (from Procell) were cultured in DMEM (Hyclone, Utah, USA) supplemented with 10% FBS (Gibco, New York, USA) and 1% penicillin / streptomycin. TRBP knockout cells SK-KOTRBP and HCCLM3-KOTRBP were cultured in DMEM supplemented with 10% FBS, 1% penicillin / streptomycin and 1 μg / mL puromycin. All cells were cultured in a 37°C, 5% CO2 incubator. To ensure the authenticity and integrity of the cell line, it was identified using the short tandem repeat sequence mapping technology of Suzhou Gene Detection Biotechnology Co., Ltd., China. In addition, the cells were checked for mycoplasma contamination to ensure the validity of the experimental results.

[0274] HeLa cells were transfected with the pCIneo-luciferase-miR-21 plasmid, and G418-resistant cells were isolated, cultured and cloned to establish a stable cell line. The cells were treated with different concentrations of compounds to evaluate the activity of different compounds (the HPLC purity of all compounds was above 95%). The relative luciferase expression level was then measured using the luciferase reporter assay system from Promega according to the manufacturer's instructions. The experiment used 0.1% DMSO as a control. The data obtained represent the relative expression of luciferase and can be used to calculate the half effective concentration (EC 50 Finally, the concentration-response curves were fitted using GraphPad Prism version 8.0 (GraphPad Software, San Diego, USA).

[0275] The half effective concentration EC of the compound prepared by the present invention for inhibiting miR-21 biosynthesis in the activity screening experiment 50 As shown in Table 9. The numbers under the compounds represent the specific compounds in Example 1, for example, "1" represents compound 1. In Table 9, the RLU value represents the concentration of the compound required to double the luciferase signal in HeLa-luciferase-miR-21 cells.

[0276] Table 9 List of inhibitory activities of each compound on miR-21 biosynthesis

[0277]

[0278]

[0279] The results showed that the compounds prepared by the present invention can significantly improve the inhibitory activity on miR-21 biosynthesis.

[0280] Example 3: Demonstration of the effects of compounds on tumor cells

[0281] To further verify the anti-tumor effect of the compound, EC 50 Compounds with values ​​less than 15 nM were subjected to this example.

[0282] 1. Effects on cell viability and proliferation ability.

[0283] 2D cell viability assay: SK-HEP-1 cells were seeded into 96-well plates at a density of 1000 cells per well and allowed to adhere overnight. They were then treated with 0.1% dimethyl sulfoxide (DMSO) and 10 μM of the test compound. After 72 h of incubation, 10% (v / v) CCK-8 solution from Beyotime, Shanghai, China was added to each well. The cells were then incubated at 37°C for 30 minutes. The fluorescence intensity was then measured at a wavelength of 450 nm using a Thermo Fisher Scientific Varioskan Flash multimode reader.

[0284] 3D cell proliferation assay: SK-HEP-1 cells were seeded at a density of 5000 cells per well in ultra-low attachment 96-well plates. The day after seeding, cells were treated with various compounds, with DMSO as a control. After 72 h of incubation, cell viability was assessed using the CellTiter-Lumi method (Beyotime, Shanghai, China) according to the manufacturer's instructions. The results were graphically analyzed using GraphPadPrism 8.0.

[0285] 3D soft agar colony formation test: 1.5 mL of medium (DMEM, 10% FBS) containing 0.8% soft agar was added to a 6-well plate to prepare the bottom layer, and then 1 mL of HCCLM3 cell suspension (5000-10,000 cells / well) containing 0.36% soft agar was inoculated in the well plate to form a cell layer. The cells were incubated overnight in 3D, and then 1 mL of medium containing 25 μM test compound or 50 μM sorafenib was added to the top layer of the well plate. After 14-21 days of culture, cells were stained with 0.05% crystal violet, and the number of clones was analyzed by colony counting using Image J software.

[0286] The results are as follows Figure 1-2 The results showed that in a three-dimensional (3D) cell culture environment, most compounds, especially compounds 24, 29, 30, 37, 39, 41, 34, 53, 67, 68 and 47, showed significant anti-proliferative ability ( Figure 1 In the 3D soft agar cell colony assay, compounds 24, 37, 34, 53, 68, and 47 had an inhibitory effect on HCCLM3 cell proliferation comparable to that of sorafenib ( Figure 2 ). This suggests that these compounds have the potential to be used as alternative therapeutic drugs to inhibit tumor growth.

[0287] 2. Impact on wound healing.

[0288] Wound healing assay: SK-HEP-1 cells were cultured at 2×10 5Cells were seeded in 12-well plates at a density of 100 μL. A straight scratch was made on the cell monolayer using a 200 μL sterile pipette tip. The resulting cell debris was rinsed with phosphate buffered saline (PBS, Hyclone). After the scratch was made, the cells were incubated with 10 μM of the different compounds or DMSO in the culture medium. Images of the scratched area were captured at 0 and 24 hours using a Canon digital camera or similar imaging system. The wound healing rate was analyzed using Image J software.

[0289] Cell migration analysis: SK-HEP-1 cells and HCCLM3 cells were seeded into the upper chamber of a Transwell chamber (Corning, NY, USA). 5 Cells were seeded at a density of 100 μL in 200 μL serum-free medium. 600 μL DMEM containing 10% FBS was added to the lower chamber to form a chemotactic gradient. After treatment with 10 μM compound, cells migrated overnight at 37°C. The upper chamber was wiped with cotton, fixed with 4% methanol, and stained with 0.1% crystal violet. Images were taken under a microscope and analyzed with Image J software.

[0290] The results are as follows Figure 3-6 The results showed that most of the compounds had an inhibitory effect on the migration of tumor cells, among which compounds 24, 37, 34, 53 and 47 showed good anti-migratory activity against SK-HEP-1 cells. The results showed that the sulfur-containing or oxygen-containing five-membered nitrogen heterocyclic derivatives obtained by structural optimization can effectively inhibit the proliferation and migration of liver cancer cells.

[0291] The embodiments described above are only descriptions of the preferred modes of the present invention and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various deformations, modifications, and substitutions made to the technical solutions of the present invention by ordinary technicians in this field should all fall within the protection scope determined by the claims of the present invention.

Claims

1. A five-membered nitrogen heterocyclic compound, characterized in that: The general structural formula is shown in Formula I: Wherein, the six-membered ring is selected from an aromatic ring, a cyclohexane ring, and a piperidine ring; X is selected from O and S; W is selected from C, CH, and N; and n is selected from any integer from 0 to 3; Y is selected from R5 is selected from hydrogen, alkyl, or is linked to R0 through a linking group in which any one of 1 to 5 methylene groups is replaced by oxygen to form a ring; R0 is selected from hydrogen, alkyl, aromatic, acyl, sulfonyl, or is linked to R5 through a linker in which any one of 1-6 methylene groups is replaced by oxygen to form a ring, wherein the alkyl group includes a straight-chain alkyl group, a branched alkyl group, a cyclic alkyl group, an unsaturated alkyl group, an oxygen-containing alkyl group, and a halogen-containing alkyl group; R1 and R2 are each independently selected from hydrogen, substituted or unsubstituted C1-4 alkyl, halogen, cyano, and nitro; R3 and R4 are each independently selected from hydrogen, hydroxyl, halogen, substituted or unsubstituted C1-4 alkyl, trifluoromethyl, -SO3H, -PO(OH)2, -B(OH)2, -COR6, and -CH2R7; R6 and R7 are each independently selected from hydrogen, hydroxyl, -COOH, halogen, substituted or unsubstituted C1-4 alkyl, C1-4 alkoxy, substituted or unsubstituted aromatic benzyl, secondary amine, L0 and L1 are independently selected from 1-4 methylene groups, and R8 and R9 are independently selected from hydrogen, halogen, hydroxyl, substituted or unsubstituted aromatic amine, and secondary amine.

2. The five-membered nitrogen heterocyclic compound according to claim 1, characterized in that: The compound is any one of formula I-1, I-2, I-3 or I-4: Based on formula I, W is C, Y is selected from O, SO2, and X is selected from O, S; formula I-1 is: Based on formula I, Y is NH, W is C, n=0, X is S, R3 and R4 are selected from methyl and formic acid, R0 is selected from acyl R 01 CO-, sulfonyl R 02 SO2-, where R 01 , R 02 Each is independently selected from an alkyl group, a cycloalkyl group, an aromatic group, and a secondary amine; R1 and R2 are independently selected from a hydrogen group, a substituted or unsubstituted C1-4 alkyl group, a halogen group, a cyano group, and a nitro group; Formula I-2 is: Based on Formula I, W is C, n is 0, X is S, R3 and R4 are selected from methyl and formic acid respectively, R1 and R2 are independently selected from hydrogen and halogen; m0, m1, m2 are independently selected from integers of 0-6, and m0+m1+m2≤6, Formula I-3 is: Based on Formula I, W is N, n=0, X is selected from S, R3 and R4 are selected from methyl and formic acid respectively, wherein the six-membered ring is a piperidine ring; Y is selected from n=0; R0 is selected from hydrogen, alkyl, aromatic, acyl, sulfonyl; wherein the alkyl includes straight-chain alkyl, branched-chain alkyl, cyclic alkyl, unsaturated alkyl, oxygen-containing alkyl, halogen-containing alkyl or secondary amine; R1 and R2 are hydrogen; Formula I-4 is:

3. The five-membered nitrogen heterocyclic compound according to claim 1, characterized in that: The compound is as follows Any of the following compounds:

4. A conformational isomer of the five-membered nitrogen heterocyclic compound according to any one of claims 1 to 3, or an optical isomer thereof, or a pharmaceutically acceptable salt thereof.

5. The method for preparing the five-membered nitrogen heterocyclic compound according to claim 1, characterized in that: The reaction equation of the preparation method is as follows:

6. The preparation method according to claim 5, characterized in that: The thiolation reagent is any one of sodium hydrosulfide hydrate, phosphorus pentasulfide, and hydrogen sulfide; the solvent is any one of methanol, ethanol, DMF, 1,4-dioxane, ethylene glycol dimethyl ether, and tetrahydrofuran; and hal is any one of chlorine, bromine, and iodine.

7. The method for preparing the five-membered nitrogen heterocyclic compound according to claim 2, characterized in that: The reaction equation for preparing compound Ⅰ-1 is: The reaction equation for preparing compound I-2 is: The reaction equation for preparing compound I-3 is: The reaction equation for preparing compound I-4 is:

8. A medicament prepared using a conformational isomer of the compound according to any one of claims 1 to 4, or an optical isomer thereof, or a pharmaceutically acceptable salt thereof.

9. A medicament comprising a conformational isomer of the compound according to any one of claims 1 to 4, or an optical isomer thereof, or a pharmaceutically acceptable salt thereof.

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