Glutamine cyclase inhibitor and application thereof

Abstract

The invention provides a glutamine cyclase inhibitor and application thereof, and relates to the technical field of medicines. The glutamine cyclase inhibitor is a compound as shown in formula I, a stereoisomer thereof, a tautomer thereof, a chiral isomer thereof or a salt thereof. The compound can effectively inhibit the activity of sQC / gQC, can be used as a glutamine cyclase inhibitor, and can be used for preparing medicines for preventing and / or treating diseases (such as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, tumors, cancers, immune diseases and the like) related to glutamine cyclase. More choices are provided for development and application of glutamine cyclase small-molecule inhibitors and medicines for preventing and / or treating diseases (such as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, tumors, cancers, immune diseases and the like) related to glutamine cyclase. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical technology, and particularly relates to a class of glutamine cyclase inhibitors and their uses. Background Art

[0002] Glutamine cyclase (abbreviated as QC) is an important acyltransferase with remarkable cyclization. Its main function is to cyclize glutamine at the amino terminus within a protein or bioactive peptide segment into pyroglutamic acid, releasing NH3 during this cyclization process. This reaction often occurs during the post-translational maturation of various bioactive peptide segments or newly synthesized proteins, mainly to protect their N-terminus and prevent degradation by exogenous peptidases. In addition, QC also plays an important catalytic role in the formation of active peptide segments with specific structures, hormones, and cytokines. Currently, two QC isoenzymes have been confirmed in humans, namely secreted glutamine cyclase (sQC) and Golgi-resident glutamine cyclase (gQC). They are involved in regulating multiple physiological processes and are closely related to diseases such as Alzheimer's disease (AD). Research has shown that sQC and gQC mediate the formation of truncated Aβ pyroglutamate (pE-Aβ) and the maturation of chemokine ligand 2 (CCL2). Compared with the full-length unmodified Aβ peptide, pE-Aβ shows stronger neurotoxicity and aggregation, and can also interact with CCL2 to trigger neuroinflammation and accelerate the progression of AD disease. In addition, sQC and gQC are closely related to various diseases such as Parkinson's syndrome, amyotrophic lateral sclerosis, tumors, cancers, immune diseases, rheumatoid arthritis, non-alcoholic hepatitis, cutaneous melanoma, and lupus erythematosus syndrome. Therefore, sQC / gQC is regarded as a potential target for the treatment of these diseases, and its inhibitors may become an effective strategy for treating these diseases.

[0003] Most of the reported sQC / gQC inhibitors currently involve forming coordination with the zinc ion at the active site; only 1 sQC / gQC inhibitor (PQ912) is in clinical phase II as an anti-AD candidate drug, which is difficult to meet the needs of clinical drug development. Therefore, it is necessary to develop more novel backbone sQC / gQC inhibitors to provide more leads or candidate drugs for the development of innovative drugs targeting sQC / gQC. Summary of the Invention

[0004] The object of the present invention is to provide a class of glutamine cyclase inhibitors and their uses.

[0005] The present invention provides a compound, its stereoisomer, its tautomer, its chiral isomer or its salt, and the structure of the compound is shown in Formula I:

[0006]

[0007] Wherein, X1 is selected from CH or N, X2 is selected from CH or N, X3 is selected from CH or N, and X4 is selected from CH or N;

[0008] m is 0, 1, 2, 3 or 4;

[0009] When m is 0, X1, X2, X3 and X4 are not simultaneously CH;

[0010] Each R1 is independently selected from halogen, C 1-5 alkyl, C 1-5 alkoxy, COR 1a , nitro, hydroxy, NR 1b R 1c , L1L3R 1e , L1L2R 1d ;

[0011] R 1a is selected from C 1-5 alkyl; R 1b is selected from hydrogen, C 1-5 alkyl; R 1c is selected from hydrogen, C 1-5 alkyl;

[0012] L1 is selected from O, S, NH, CO, CS, NHCO, NHCS, NHSO2, NHCONH, NHCSNH;

[0013] L2 is selected from none or C 1-3 alkylene;

[0014] L3 is selected from or C 1-3 alkylene;

[0015] R 1e is selected from C 1-5 alkyl, halo C 1-5 alkyl, COOR 1f ; R 1f is selected from C 1-5 alkyl;

[0016] R 1d is selected from the following groups which are unsubstituted or substituted by 1, 2 or 3 R x : 3-6 membered saturated cycloalkyl, 5-6 membered saturated heterocyclic group, 5-6 membered heteroaryl, phenyl; R x are each independently selected from halogen, C 1-5 alkyl, halo C 1-5 alkyl, C 1-5 alkoxy, halo C 1-5 alkoxy, hydroxy, L4NR 1g R 1h , C 2-4 alkenyl, C 2-4 alkynyl, CORi 、COOR i 、SO2R i ;

[0017] L4 is selected from none or C 1-3 alkylene; R 1g is selected from hydrogen, C 1-5 alkyl; R 1h is selected from hydrogen, C 1-5 alkyl; R i is selected from C 1-5 alkyl, C 1-5 alkyl substituted by C 1-5 alkoxy, unsubstituted or substituted by 1, 2 or 3 R y substituted with the following groups: 5-6 membered saturated heterocyclic group, 5-6 membered heteroaryl, phenyl, 3-6 membered saturated cycloalkyl; R y each independently selected from C 1-5 alkyl, C 1-5 alkoxy;

[0018] R2 is selected from H, hydroxy;

[0019] R3 is selected from H, C 1-5 alkyl, COR 3a ; R 3a is selected from C 1-5 alkyl.

[0020] Furthermore, the structure of the compound is shown in Formula II:

[0021]

[0022] wherein, X1 is selected from CH or N;

[0023] m is 0, 1, 2, 3 or 4;

[0024] When m is 0, X1 is N;

[0025] R1 is as described above;

[0026] R2 is selected from H, hydroxy;

[0027] R3 is selected from H, C 1-3 alkyl, COR 3a ; R 3a is selected from C 1-3 alkyl.

[0028] Furthermore, the structure of the compound is shown in Formula III:

[0029]

[0030] Preferably, the structure of the compound is shown in Formula IV:

[0031]

[0032] Among them, R1 is as described above.

[0033] Further, R1 is selected from halogen, C 1-3 alkyl, C 1-3 alkoxy, COR 1a , nitro, hydroxyl, NR 1b R 1c , L1L3R 1e , L1L2R 1d ;

[0034] R 1a is selected from C 1-3 alkyl; R 1b is selected from hydrogen, C 1-3 alkyl; R 1c is selected from hydrogen, C 1-3 alkyl;

[0035] L1 is selected from O, S, NH, CO, CS, NHCO, NHCS, NHSO2, NHCONH, NHCSNH;

[0036] L2 is selected from none or C 1-2 alkylene;

[0037] L3 is selected from or C 1-2 alkylene;

[0038] R 1e is selected from C 1-3 alkyl, halo C 1-3 alkyl, COOR 1f ; R 1f is selected from C 1-3 alkyl;

[0039] R 1d is selected from the following groups which are unsubstituted or substituted by 1 or 2 R x : 3-6 membered saturated cycloalkyl, 5-6 membered saturated heterocyclic group, 5-6 membered heteroaryl, phenyl; R x are each independently selected from halogen, C 1-3 alkyl, halo C 1-3 alkyl, C 1-3 alkoxy, halo C 1-3 alkoxy, hydroxyl, L4NR 1g R 1h 、C 2-3 alkenyl, C 2-3 alkynyl, COR i 、COOR i 、SO2R i ;

[0040] L4 is selected from none or C 1-2 alkylene; R 1g is selected from hydrogen, C 1-3 alkyl; R 1h is selected from hydrogen, C 1-3 alkyl; R i is selected from C 1-3 alkyl, C 1-3 alkyl substituted by C 1-3 alkoxy, unsubstituted or substituted by 1 or 2 R y substituted with the following groups: 5-6 membered saturated heterocyclic group, 5-6 membered heteroaryl, phenyl, 3-6 membered saturated cycloalkyl; R y each independently selected from C 1-3 alkyl, C 1-3 alkoxy.

[0041] Furthermore, the structure of the compound is shown in Formula V-1, Formula V-2, Formula V-3 or Formula V-4:

[0042]

[0043]

[0044] wherein, R i is selected from C 1-3 alkyl, C 1-3 alkyl substituted by C 1-3 alkoxy, unsubstituted or substituted by 1 or 2 R y substituted with the following groups: 5-6 membered saturated heterocyclic group, 5-6 membered heteroaryl, phenyl, 3-6 membered saturated cycloalkyl; R y each independently selected from C 1-3 alkyl, C 1-3 alkoxy.

[0045] Furthermore, the compound is one of the following compounds:

[0046]

[0047]

[0048]

[0049] The present invention also provides a pharmaceutical composition, which is a preparation obtained by using the above-mentioned compound, its stereoisomer, its tautomer, its chiral isomer or its salt as an active ingredient and adding pharmaceutically acceptable excipients.

[0050] The present invention also provides the use of Compound 5 or the above-mentioned compound, its stereoisomer, its tautomer, its chiral isomer or its salt in the preparation of a glutamine cyclase inhibitor;

[0051]

[0052] Further, the glutamine cyclase is sQC and / or gQC.

[0053] Further, the glutamine cyclase inhibitor is a drug for preventing and / or treating diseases related to glutamine cyclase;

[0054] Preferably, the diseases related to glutamine cyclase are Alzheimer's disease, Parkinson's syndrome, amyotrophic lateral sclerosis, tumors, cancers, immune diseases, etc.;

[0055] More preferably, the tumor is a thyroid tumor, esophageal tumor, lung tumor, kidney tumor or breast tumor, the cancer is lung cancer, gastric cancer, colorectal cancer, liver cancer, breast cancer, esophageal cancer, thyroid cancer, endometrial cancer, brain cancer or pancreatic cancer, and the immune disease is rheumatoid arthritis, non-alcoholic hepatitis, cutaneous melanoma or lupus erythematosus syndrome.

[0056] The present invention provides a class of compounds with a new parent nucleus structure that can effectively inhibit the activities of sQC / gQC. Such compounds can be used as glutamine cyclase inhibitors to prepare drugs for preventing and / or treating diseases related to glutamine cyclase (such as Alzheimer's disease, Parkinson's syndrome, tumors, cancers, amyotrophic lateral sclerosis, immune diseases, etc.).

[0057] The present invention provides more options for the development and application of small molecule inhibitors of glutamine cyclase and drugs for preventing and / or treating diseases related to glutamine cyclase (such as Alzheimer's disease, Parkinson's syndrome, tumors, cancers, amyotrophic lateral sclerosis, immune diseases, etc.).

[0058] Obviously, based on the above content of the present invention, according to the common general knowledge and customary means in the art, without departing from the above basic technical idea of the present invention, various other forms of modifications, substitutions or changes can be made.

[0059] The following is a further detailed description of the above content of the present invention through specific embodiments in the form of examples. However, this should not be construed as limiting the scope of the above subject matter of the present invention to the following examples. All technologies implemented based on the above content of the present invention fall within the scope of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0060] Figure 1 : Results of the rotarod test for each group.

[0061] Figure 2 : Grip strength test results for each group.

[0062] Figure 3 : Open field test results for each group. Detailed implementation method

[0063] The raw materials and equipment used in the specific implementation method of the present invention are all known products, which are obtained by purchasing commercially available products.

[0064] Example 1: Synthesis of Compounds 1-10, 13-18

[0065]

[0066] At room temperature, potassium tert-butoxide (6.72 g, 60 mmol) was added portionwise to a DMF solution of methyl malonate (6.85 mL, 60 mmol). After addition, the mixture was transferred to a preheated oil bath at 90 °C and activated for 30 minutes. After cooling to room temperature, 2-bromo-3-nitroanisole (1a, 2.34 g, 10 mmol) was slowly added, and the mixture was then transferred back to the 90 °C oil bath for reaction for 16 h. After monitoring the reaction to completion by thin-layer chromatography (TLC), the reaction solution was cooled to room temperature, acidified to pH = 7 with 2 M dilute hydrochloric acid, extracted with EA (100 mL × 3), the extraction solutions were combined, dried over anhydrous magnesium sulfate, evaporated to dryness, and directly used in the next step without column chromatography.

[0067] The crude reaction product obtained in the previous step was dissolved in 50 mL of DMSO and transferred to an oil bath at 160 °C for reaction for 6 h. After detecting by TLC that the raw materials had basically reacted completely, the reaction solution was diluted with water and extracted with EA (100 mL × 3). The extraction solutions were combined, dried over anhydrous magnesium sulfate, evaporated to dryness, and purified by column chromatography (PE / EA = 20:1 - 10:1) to obtain 1.62 g of intermediate 1c with a yield of 72%.

[0068] Intermediate 1c was dissolved in 50 mL of glacial acetic acid, iron powder (2.0 g, 36 mmol) was added, and the mixture was transferred to an oil bath at 100 °C for reaction for about 3 h until the raw materials had basically reacted completely. Filter while hot, concentrate under reduced pressure, and purify by column chromatography (PE / EA = 10:1 - 4:1) to obtain 645 mg of yellow solid 1d with a yield of 55%.

[0069] Under an ice bath, lithium aluminum hydride (2.28 g, 60 mmol) was added portionwise to a THF solution of methyl benzimidazole-5-carboxylate (1e, 3.52 g, 20 mmol). After addition, the reaction mixture was transferred to room temperature and stirred for an additional 2 h. After completion of the reaction as monitored by TLC, the reaction was quenched by the sequential addition of 2.28 mL of water, 3 × 2.28 mL of 15% NaOH solution, and 2.28 mL of water. The resulting suspension was stirred for 20 mins and then filtered through diatomaceous earth. The filtrate was collected, concentrated in vacuo, and purified by column chromatography (DCM / MeOH = 50:1 - 5:1) to afford 2.25 g of white solid 1f in 76% yield.

[0070] At room temperature, activated manganese dioxide (13.2 g, 152 mmol) was added portionwise to a THF solution of 1H-benzimidazole-5-methanol (1f, 2.25 g, 15.2 mmol). After reacting for 2 h, the reaction was monitored by TLC and the starting material was found to have reacted completely. The reaction mixture was filtered through diatomaceous earth, the filtrate was collected, concentrated in vacuo, and purified by column chromatography (DCM / MeOH = 50:1 - 10:1) to afford 1.33 g of white solid 1g in 60% yield.

[0071] 1d (196 mg, 1.2 mmol) and 1g (146 mg, 1 mmol) were accurately weighed and placed in a 100 mL eggplant-shaped flask. 40 mL of ethanol was added, followed by the dropwise addition of 3 drops of piperidine. The reaction mixture was refluxed in an 80 °C oil bath for 6 h. After completion of the reaction as monitored by TLC, the reaction mixture was concentrated in vacuo and the solid was washed with methanol to afford 195 mg of the final product 1 in 67% yield. 1 H NMR (400 MHz, DMSO-d6): δ 12.69 (br s, 1H), 10.62 (s, 1H), 8.96 (s, 1H), 8.33 (s, 1H), 8.28 (s, 1H), 8.00 - 7.84 (m, 1H), 7.62 (d, J = 8.4 Hz, 1H), 7.17 (t, J = 8.0 Hz, 1H), 6.70 (d, J = 8.5 Hz, 1H), 6.50 (d, J = 7.6 Hz, 1H), 3.98 (s, 1H). 13 C NMR (100 MHz, DMSO-d6): δ 167.18, 155.53, 144.00, 141.54, 141.21, 129.40, 128.56, 127.25, 124.97, 111.31, 104.73, 102.61, 55.53. HRMS m / z: calcd for C 17 H 13 N3O2 [M + H] + 292.1081, found 292.1079.

[0072] Replace 4-methoxyindol-2-one with the corresponding substituted indol-2-one or azaindol-2-one, and prepare Compounds 2-10 and 13-18 respectively according to the synthesis method of Compound 1.

[0073] Compound 2: 1 H NMR(400MHz,DMSO)δ12.66(s,1H),10.53(d,J=14.1Hz,1H),8.32(d,J=5.4Hz,1H),8.01–7.67(m,2H),7.67–7.60(m,2H),7.60–7.43(m,1H),6.61–6.38(m,2H),3.77(d,J=7.0Hz,3H)ppm.

[0074] Compound 3: 1 H NMR(400MHz,DMSO)δ8.33(s,1H),7.96(d,J=11.7Hz,1H),7.79–7.63(m,3H),7.56(dd,J=8.4,1.4Hz,1H),6.74–6.63(m,2H)ppm. Compound 4: 1 H NMR(400MHz,DMSO)δ12.70(s br,1H),10.73(d,J=17.6Hz,1H),8.37(s,1H),8.06–7.83(m,2H),7.66(dd,J=35.8,8.1Hz,2H),7.25–6.93(m,2H)ppm.

[0075] Compound 5: 1 H NMR(400MHz,DMSO)δ12.69(s br,1H),10.59(d,J=17.5Hz,1H),8.35(s,1H),7.97(d,J=14.7Hz,1H),7.79(s,1H),7.77–7.54(m,3H),7.30–7.13(m,1H),7.04–6.80(m,2H)ppm.

[0076] Compound 6: 1 H NMR(400MHz,DMSO)δ12.72(d,J=16.3Hz,1H),11.19(s,1H),8.37(s,1H),8.14–7.90(m,4H),7.87–7.53(m,2H),6.92(dd,J=7.6,5.2Hz,1H)ppm.

[0077] Compound 8: 11H NMR (400 MHz, DMSO) δ 12.72 (s br, 1H), 11.09 (s, 1H), 8.36 (s, 1H), 8.17–7.91 (m, 1H), 7.88 (s, 1H), 7.86–7.47 (m, 3H), 7.24–6.80 (m, 2H) ppm.

[0078] Compound 10: 1 1H NMR (400 MHz, DMSO) δ 12.72 (s br, 1H), 11.09 (s, 1H), 8.36 (s, 1H), 8.17–7.91 (m, 1H), 7.88 (s, 1H), 7.86–7.47 (m, 3H), 7.24–6.80 (m, 2H) ppm.

[0079] Compound 13: 1 1H NMR (400 MHz, DMSO-d6): δ 12.66 (br s, 1H), 10.52 (s, 1H), 10.41 (s, 1H), 8.94 (s, 1H), 8.32 (s, 1H), 8.27 (s, 1H), 7.99 - 7.74 (m, 1H), 7.61 (br s, 1H), 7.00 (t, J=8.0 Hz, 1H), 6.52 (d, J=8.3 Hz, 1H), 6.33 (d, J=7.7 Hz, 1H). 13 13C NMR (100 MHz, DMSO-d6): δ 167.34, 153.61, 141.65, 140.18, 128.96, 127.06, 125.44, 110.47, 109.45, 100.76. HRMS m / z: calcd for C 16 H 11 N3O2 [M + H] + 278.0924, found 278.0920.

[0080] Compound 14: 1 1H NMR (400 MHz, DMSO-d6): δ 12.83 (br s, 1H), 11.14 (s, 1H), 8.87 (s, 1H), 8.40 (s, 1H), 7.97 (s, 1H), 7.86 (d, J=8.4 Hz, 1H), 7.65 (d, J=8.2 Hz, 1H), 7.50 (d, J=7.7 Hz, 1H), 7.42 (t, J=8.0 Hz, 1H), 7.16 (d, J=7.8 Hz, 1H). 13 13C NMR (100 MHz, DMSO-d6): δ. HRMS m / z: calcd for C 16H 10 N4O3[M+H] + 307.0826, found 307.0825.

[0081] Compound 15: 1 H NMR(400 MHz, DMSO-d6): δ 12.61(br s, 1H), 10.54(s, 1H), 8.30(s, 1H), 7.88(s, 1H), 7.55(d, J = 8.3 Hz, 1H), 7.29(dd, J = 8.5, 1.6 Hz, 1H), 7.20(t, J = 7.9 Hz, 1H), 7.06(s, 1H), 6.70(d, J = 8.4 Hz, 1H), 6.49(d, J = 7.4 Hz, 1H), 2.01(s, 6H). 13 C NMR(100 MHz, DMSO-d6): δ. HRMS m / z: calcd for C 18 H 16 N4O[M+H] + 305.1397, found 305.1390.

[0082] Compound 16: 1 H NMR(400 MHz, DMSO-d6): δ 12.75(br s, 1H), 10.90(s, 1H), 9.02(s, 1H), 8.36(s, 1H), 8.02(d, J = 8.1 Hz, 1H), 7.97(d, J = 3.4 Hz, 1H), 7.65(d, J = 8.4 Hz, 1H), 7.23(td, J = 8.1, 5.5 Hz, 1H), 6.87 - 6.82(m, 1H), 6.71(d, J = 7.7 Hz, 1H). 13 C NMR(100 MHz, DMSO-d6): δ 166.56, 158.31, 156.66, 142.33, 142.28, 129.73, 123.75, 118.54, 115.66, 111.71, 111.61, 108.60, 108.46, 105.83. Compound 17: 1 H NMR(400 MHz, DMSO-d6): δ 12.74(brs, 1H), 10.90(s, 1H), 8.95(d, J = 1.6 Hz, 1H), 8.63(s, 1H), 8.36(s, 1H), 7.94(d, J = 8.5 Hz, 1H), 7.64(d, J = 8.4 Hz, 1H), 7.20(t, J = 7.9 Hz, 1H), 7.11 - 6.91(m, 1H), 6.93 - 6.77(m, 1H).13 C NMR (100 MHz, DMSO-d6): δ 166.55, 144.38, 142.73, 142.48, 129.22, 127.74, 127.16, 124.17, 122.95, 120.18, 108.29.

[0083] Compound 18: 1 H NMR (400 MHz, DMSO-d6): δ 12.73 (br s, 1H), 10.87 (s, 1H), 8.91 (s, 1H), 8.81 (s, 1H), 8.36 (s, 1H), 7.99 - 7.84 (m, 2H), 7.21 (dd, J = 8.1, 1.0 Hz, 1H), 7.13 (t, J = 7.9 Hz, 1H), 6.87 (dd, J = 7.6, 1.0 Hz, 1H). 13 C NMR (100 MHz, DMSO-d6): δ 166.56, 158.31, 156.66, 142.33, 142.28, 129.73, 123.75, 118.54, 115.66, 111.71, 111.61, 108.60, 108.46, 105.83.

[0084] Synthesis of Example 2, Compounds 11 and 12

[0085]

[0086] Dissolve 11a (200 mg, 0.9 mmol) in 4 mL of acetic anhydride and react at 130 °C for 4 h. After monitoring the reaction to completion by TLC, adjust the pH to neutral with saturated sodium bicarbonate solution, then extract with ethyl acetate and water (25 mL × 3). Combine the organic layers and wash with saturated sodium chloride solution (25 mL × 3), dry over anhydrous magnesium sulfate, and remove the solvent by rotary evaporation. Then perform column chromatography (5% PE / EA 100 mL, 6% PE / EA 100 mL, 10% PE / EA 200 mL) to obtain intermediate 11b (186 mg, yield 78%, yellowish-brown solid).

[0087] Dissolve 11b (189 mg, 0.7 mmol), benzimidazole-5-carboxylic acid (133 mg, 0.8 mmol), and TBTU (260 mg, 0.8 mmol) in DMF, add triethylamine (163 μL, 1.2 mmol), and react at room temperature for 3 h. After monitoring the reaction to completion by TLC, remove the solvent by rotary evaporation and perform column chromatography (10% PE / EA 100 mL, 12% PE / EA 100 mL, 20% PE / EA 200 mL) to obtain Compound 11 (127 mg, yield 46%, white solid). 11H NMR (400 MHz, DMSO) δ 11.15 (s br, 1H), 9.12 (d, J = 8.7 Hz, 1H), 8.66 (d, J = 8.6 Hz, 1H), 8.38 (d, J = 2.0 Hz, 1H), 7.79–7.64 (m, 2H), 7.50 (dd, J = 8.7, 2.0 Hz, 1H), 7.23 (dd, J = 8.7, 1.9 Hz, 1H), 7.02 (d, J = 1.9 Hz, 1H), 2.67 (s, 3H) ppm.

[0088] Compound 11 (50 mg, 0.1 mmol) was dissolved in EtOH:H2O = 2:1. Lithium hydroxide monohydrate (26 mg, 0.6 mmol) was added, and the reaction was carried out at room temperature for 3 h. After the reaction was monitored by TLC to be complete, the pH was adjusted to neutral with 2N hydrochloric acid solution, and then extracted with ethyl acetate and water (25 mL × 3). The organic layers were combined, dried over anhydrous magnesium sulfate, and the solvent was removed by rotary evaporation. Then column chromatography (10% PE / EA 100 mL, 15% PE / EA 100 mL, 20% PE / EA 100 mL) was carried out to obtain compound 12 (26 mg, yield 63%, white solid). 1 1H NMR (400 MHz, DMSO) δ 12.10 (s br, 1H), 10.52 (s, 1H), 8.52 (s, 1H), 7.33 (dd, J = 8.2, 2.0 Hz, 1H), 7.10 (d, J = 8.2 Hz, 1H), 6.90 (d, J = 1.7 Hz, 1H), 6.83 (dd, J = 8.2, 1.9 Hz, 1H), 5.91 (d, J = 8.2 Hz, 1H) ppm.

[0089] Example 3: Synthesis of Compounds 19 and 20

[0090]

[0091] 4-Bromoindanedione (19a, 1.0 g, 4.42 mmol) was dissolved in DMF. Sodium hydride (180 mg, 4.42 mmol) was added under ice bath, and the mixture was stirred for 30 mins. Then methyl iodide (240 μL, 4.42 mmol) was added, and the reaction was continued under ice bath for 40 mins. After the reaction was completed, the reaction solution was poured into an aqueous solution, and extracted with EA (100 mL x 3). The extraction solutions were combined, dried over anhydrous magnesium sulfate, and evaporated to dryness to obtain 955 mg of intermediate 19b, which was directly used in the next step without purification.

[0092] Accurately weigh 19b (955 mg, 3.98 mmol) and add it to 4 mL of hydrazine hydrate solution. React under sealed tube at 120 °C for 2 h. After the reaction is completed, quench with water, extract with EA (100 mL × 3). Combine the extracts, dry over anhydrous magnesium sulfate, and perform column chromatography (PE / EA = 10:1 - 4:1) to obtain 570 mg of intermediate 19c with a yield of 57%.

[0093] For the last step, refer to the synthesis of compound 1, and the synthesis yield is 73%. 1 H NMR (400 MHz, DMSO-d6): δ 12.75 (d, J = 39.7 Hz, 1H), 8.98 (s, 1H), 8.87 (s, 1H), 8.39 (s, 1H), 8.06 - 7.58 (m, 2H), 7.28 (dd, J = 8.2, 1.1 Hz, 1H), 7.22 (t, J = 7.9 Hz, 1H), 7.07 (dd, J = 7.5, 1.2 Hz, 1H), 3.24 (s, 3H). 13 C NMR (100 MHz, DMSO-d6): δ 164.77, 143.77, 142.03, 129.22, 127.76, 126.88, 123.37, 120.45, 115.39, 107.72, 25.92. HRMS m / z: calcd for C 17 H 12 BrN3O [M + H] + 354.0237, found 354.0232.

[0094] Use 2-bromopropane instead of iodomethane and synthesize compound 20 according to the synthesis method of compound 19. 1 H NMR (400 MHz, DMSO-d6): δ 12.84 (br s, 1H), 8.89 (s, 2H), 8.36 (s, 1H), 7.92 - 7.89 (m, 1H), 7.65 (d, J = 8.5 Hz, 1H), 7.28 - 7.17 (m, 3H), 4.70 - 4.63 (m, 1H), 1.47 (s, 3H), 1.45 (s, 3H). 13 C NMR (100 MHz, DMSO-d6): δ 164.48, 144.31, 142.54, 142.12, 129.21, 127.62, 126.57, 123.62, 120.71, 115.70, 108.79, 43.38, 19.08. HRMS m / z: calcd for C 19 H 16 BrN3O [M + H] +382.0550, found 382.0549.

[0095] Synthesis of Example 4, Compounds 21, 23 - 24, 54, 56

[0096]

[0097] Compound 1d (645 mg, 3.96 mmol) was added to a reaction flask and dissolved in 8 mL of hydrobromic acid. The reaction was carried out at 120 °C for 5 h. After TLC detection showed that the raw material reaction was basically complete, the pH of the reaction solution was adjusted to 6 - 7 with 10 N sodium hydroxide, and extracted with EA (50 mL × 3). The combined extracts were dried over anhydrous magnesium sulfate, concentrated by rotary evaporation, and purified by column chromatography (PE / EA = 4:1 - 1:1) to obtain 383 mg of white solid 21a with a yield of 65%.

[0098] Under an ice bath, 4 - hydroxy - 2 - indolinone (21a, 447 mg, 3 mmol), 3 - hydroxytetrahydrofuran (363 μL, 4.5 mmol), and triphenylphosphine (1.18 g, 4.5 mmol) were added to a 100 - mL two - necked flask. The flask was purged with argon three times, and then 50 mL of ultra - dry THF was added. After stirring for 15 minutes under the ice bath, diisopropyl azodicarboxylate (885 μL, 4.5 mmol) was slowly added dropwise to the above solution. After the addition was complete, stirring was continued under the ice bath for 30 minutes. The ice bath was removed and the reaction was allowed to proceed at room temperature overnight. After TLC monitoring showed that the raw materials were basically completely reacted, the reaction solution was diluted with water and extracted with EA (30 mL × 3). The combined extracts were dried over anhydrous magnesium sulfate, concentrated by rotary evaporation, and purified by column chromatography (PE / EA = 5:1 - 1:1) to obtain 326 mg of intermediate 21b with a yield of 53%.

[0099] For the last step, refer to the synthesis of Compound 1 with a synthesis yield of 48%. 1 H NMR (400 MHz, DMSO - d6): δ 12.68 (br s, 1H), 10.62 (s, 1H), 8.94 (s, 1H), 8.33 (s, 1H), 8.32 (s, 1H), 7.90 (d, J = 8.4 Hz, 1H), 7.63 (d, J = 8.4 Hz, 1H), 7.15 (t, J = 8.0 Hz, 1H), 6.67 (d, J = 8.4 Hz, 1H), 6.49 (d, J = 7.6 Hz, 1H), 5.35 - 5.04 (m, 1H), 4.00 - 3.91 (m, 3H), 3.88 - 3.83 (m, 1H), 2.35 - 2.26 (m, J = 14.2, 8.4, 5.9 Hz, 1H), 2.18 - 2.12 (m, 1H). 1313C NMR(100MHz, DMSO-d6): δ 167.20, 153.17, 144.08, 141.85, 141.15, 129.26, 128.60, 127.28, 124.86, 112.05, 106.61, 102.75, 77.75, 72.57, 66.61, 32.73. HRMS m / z: calcd for C 20 H 17 N3O3 [M+H] + 348.1343, found 348.1342.

[0100] Compounds 23 - 24, 54, and 56 were synthesized according to a similar synthetic method.

[0101] Compound 23: 1 1H NMR(400MHz, DMSO-d6): δ 12.72(br s, 1H), 10.61(s, 1H), 9.05(s, 1H), 8.55(s, 1H), 8.33(s, 1H), 8.10 - 7.61(m, 2H), 7.12(t, J = 8.0Hz, 1H), 6.72(d, J = 8.5Hz, 1H), 6.46(d, J = 7.6Hz, 1H), 4.69 - 4.65(m, 1H), 3.84 - 3.74(m, 3H), 3.64 - 3.58(m, 1H), 2.05 - 1.82(m, 3H), 1.58 - 1.52(m, 1H). 13 13C NMR(100MHz, DMSO-d6): δ 167.35, 153.11, 144.05, 141.86, 141.61, 129.19, 128.93, 127.35, 124.87, 112.60, 106.69, 102.53, 70.53, 69.24, 67.20, 27.02, 22.23. HRMS m / z: calcd for C 21 H 19 N3O3 [M+H] + 362.1499, found 362.1497.

[0102] Compound 24: 11H NMR (400 MHz, DMSO-d6): δ 12.68 (d, J = 34.7 Hz, 1H), 10.62 (s, 1H), 9.04 (s, 1H), 8.41 (s, 1H), 8.35 (s, 1H), 8.04 - 7.59 (m, 2H), 7.14 (t, J = 8.0 Hz, 1H), 6.77 (d, J = 8.5 Hz, 1H), 6.47 (d, J = 7.6 Hz, 1H), 4.88 - 4.82 (m, 1H), 3.94 - 3.88 (m, 2H), 3.64 - 3.58 (m, 2H), 2.13 - 2.06 (m, 2H), 1.84 - 1.76 (m, 2H).

[0103] 13 13C NMR (100 MHz, DMSO-d6): δ 167.28, 153.19, 144.13, 141.94, 141.13, 129.30, 128.70, 127.29, 125.14, 112.18, 106.66, 102.50, 71.35, 64.34, 31.66. HRMS m / z: calcd for C 21 H 19 N3O3 [M + H] + 362.1499, found 362.1495.

[0104] Compound 54: 1 1H NMR (400 MHz, DMSO-d6): δ 12.67 (d, J = 35.7 Hz, 1H), 10.59 (s, 1H), 9.02 (s, 1H), 8.36 (s, 1H), 8.33 (s, 1H), 8.03 - 7.58 (m, 2H), 7.12 (t, J = 8.1 Hz, 1H), 6.73 (d, J = 8.5 Hz, 1H), 6.44 (d, J = 7.5 Hz, 1H), 4.73 - 4.63 (m, 1H), 3.26 (s, 3H), 2.13 - 2.06 (m, 4H), 1.83 - 1.46 (m, 5H). 1313C NMR(100MHz,DMSO-d6):δ167.29,167.26,153.65,153.40,144.03,141.94,141.88,140.87,129.25,128.72,127.20,127.08,125.22,112.17,112.10,106.54,106.42,102.23,76.07,75.90,73.84,72.12,55.19,54.93,27.58,27.33,27.19,26.81.HRMS m / z:calcd for C 23 H 23 N3O3[M+H] + 390.1812,found 390.1814.

[0105] Compound 56: 1 1H NMR(400MHz,DMSO-d6):δ12.70(br s,1H),10.60(s,1H),8.97(s,1H),8.39(s,1H),8.33(s,1H),7.90(s,1H),7.62(d,J=8.4Hz,1H),7.12(t,J=8.1Hz,1H),6.73(d,J=8.6Hz,1H),6.44(d,J=7.6Hz,1H),4.89(br s,1H),3.57(s,3H),2.51-2.50(m,1H),2.03-1.62(m,8H). 13 13C NMR(100MHz,DMSO-d6):δ.HRMS m / z:calcd for C 24 H 23 N3O4[M+H] + 418.1761,found418.1762.

[0106] Synthesis of Example 5, Compounds 22, 29, and 42

[0107]

[0108] Accurately weigh 4-nitroindole (22a, 12.2 g, 75.2 mmol) and N-chlorosuccinimide (6.07 g, 30.1 mmol), dissolve them in 500 mL of chloroform, transfer them to an oil bath, reflux and heat for 30 h. After detecting that the raw materials have completely reacted by TLC, concentrate under reduced pressure. Without purification, directly use it for the next step.

[0109] Dissolve the crude product obtained in the previous step in 100 mL of glacial acetic acid, heat to 100 °C, add dropwise 80 mL of 85% H3PO4. After the addition is complete, continue heating to reflux for 3 h. Detect by TLC that the raw materials have completely reacted. Cool the reaction solution to room temperature, adjust the pH of the reaction solution to 6 - 7 with 10N sodium hydroxide solution, extract with EA (1000 mL x 3), combine the extracts, dry over anhydrous magnesium sulfate, spin-dry, and obtain 9.91 g of intermediate 22b by column chromatography (PE / EA = 15:1 - 4:1), with a yield of 74%.

[0110] Under a hydrogen atmosphere, dissolve 4-nitro-1,3-dihydro-2H-indol-2-one (22b, 1.17 g, 2.84 mmol) and 10% Pd / C (200 mg) in 100 mL of methanol, react at room temperature overnight. After the reaction is completed, filter the mixture through diatomaceous earth, wash it with methanol several times. Concentrate the obtained filtrate under reduced pressure, spin-dry, wash with methanol, and filter to obtain 336 mg of pure product 22c, with a yield of 80%.

[0111] At room temperature, dissolve 4-amino-1,3-dihydroindol-2-one (22c, 336 mg, 2.27 mmol) and tetrahydropyran-4-one (315 μL, 3.41 mmol) in 80 mL of methanol, add glacial acetic acid (585 μL, 10.23 mmol), react for 1 h, then add sodium cyanoborohydride (286 mg, 4.54 mmol). After detecting by TLC that the raw materials have completely reacted, filter and concentrate under reduced pressure. Subject the obtained crude product to column chromatography to obtain 347 mg of intermediate 22-d, with a yield of 56%.

[0112] For the last step, refer to the synthesis of compound 1, with a synthesis yield of 53%. 1 H NMR(400MHz,DMSO-d6):δ12.68(brs,1H),10.49(s,1H),8.36(s,1H),7.76(s,1H),7.66(s,1H),7.34 - 7.22(m,2H),7.08(t,J = 8.0Hz,1H),6.34(d,J = 8.5Hz,1H),6.24(d,J = 7.4Hz,1H),3.42 - 3.39(m,2H),3.18 - 3.09(m,2H),2.86 - 2.84(m,1H),1.39 - 1.36(m,2H),0.41 - 0.32(m,2H). 13 C NMR(100MHz,DMSO-d6):δ170.05,144.21,143.81,141.93,131.49,130.93,126.38,105.27,104.99,98.53,65.51,47.34,32.07.

[0113] Compound 29 and 42 were synthesized according to a similar synthetic method.

[0114] Compound 29: 1 H NMR (400 MHz, DMSO-d6): δ 12.59 (br s, 1H), 10.49 (s, 1H), 8.30 (s, 1H), 7.72 (s, 1H) 7.62 - 7.29 (m, 2H), 7.16 (d, J = 8.2 Hz, 1H), 7.17 - 7.0 (m, 4H), 6.75 - 6.72 (m, 2H), 6.32 - 6.27 (m, 2H), 4.02 (s, 2H). 13 C NMR (100 MHz, DMSO-d6): δ 169.97, 143.82, 143.56, 143.55, 143.16, 138.01, 131.80, 130.68, 128.01, 126.78, 126.69, 125.71, 105.34, 105.20, 98.92, 47.10.

[0115] Compound 42: 1 H NMR (400 MHz, DMSO-d6): δ 12.68 (d, J = 23.6 Hz, 1H), 10.47 (s, 1H), 8.28 (s, 1H), 7.74 - 7.60 (m, 2H), 7.34 - 7.27 (m, 1H), 7.27 - 7.09 (m, 2H), 6.36 (dd, J = 8.6, 3.6 Hz, 1H), 6.24 (d, J = 7.3 Hz, 1H), 3.79 (t, J = 15.0 Hz, 1H), 3.44 - 3.33 (m, 2H), 2.91 - 2.84 (m, 2H), 2.46 - 2.43 (m, 1H), 1.87 (s, 3H), 1.54 - 1.34 (m, 2H), 0.45 - 0.15 (m, 2H). 13 C NMR (100 MHz, DMSO-d6): δ 169.85, 167.60, 144.16, 143.68, 141.81, 131.39, 130.85, 129.06, 126.35, 124.78, 105.25, 104.90, 98.46, 47.84, 44.12, 31.13, 30.50, 21.08. HRMS m / z: calcd for C 23 H 23 N5O2 [M + H] + 402.1925, found 402.1918.

[0116] Synthesis of Example 6, Compounds 25 - 28

[0117]

[0118] 4-Bromoindirubin (5 g, 22.1 mmol) was added to a reaction flask. After adding 100 mL of methanol, hydrazine hydrate (80%) (11 g, 221.2 mmol) was added, and the mixture was placed at 80 °C for reaction for 1 hour. A solid precipitated. The reaction was monitored by TLC and was complete. The pH was adjusted to neutral with 2N hydrochloric acid solution, and then filtered. The solid was collected and dried in an oven at 50 °C to obtain compound 25b (4.1 g, yield 87%, yellow powder).

[0119] Compound 25b (200 mg, 0.9 mmol) was added to a three-necked flask, and 3-furanboronic acid (211 mg, 1.9 mmol), anhydrous sodium carbonate (299 mg, 2.8 mmol), and bis(triphenylphosphine)palladium dichloride (66 mg, 0.1 mmol) were added. Finally, 8 mL of 1,4-dioxane solution and 2 mL of water were added. The reaction was carried out at 100 °C under Ar for 2 hours. After the reaction was monitored by TLC and was complete, it was extracted with ethyl acetate and water (50 mL × 3). The organic layers were combined and washed with saturated sodium chloride solution (50 mL × 3). Finally, the organic layer was dried over anhydrous Na2SO4, filtered, and evaporated under reduced pressure to obtain a crude product. Finally, it was purified by column chromatography (20% PE / EA 120 mL, 30% PE / EA 120 mL, 50% PE / EA 200 mL) to obtain compound 25c (150 mg, yield 80%, greyish-yellow powder).

[0120] Compound 25d (1 g, 5.7 mmol) was added to a reaction flask and dissolved in 20 mL of tetrahydrofuran. Lithium aluminum hydride (646 mg, 17.0 mmol) was slowly added at 0 °C, and then the mixture was transferred to room temperature for reaction. After 1 hour, the reaction was monitored by TLC and was complete. A large amount of water was added to quench the reaction, and then it was extracted with ethyl acetate and water (50 mL × 3). The organic layers were combined and washed with saturated sodium chloride solution (50 mL × 3). Finally, the organic layer was dried over anhydrous Na2SO4, filtered, and evaporated under reduced pressure to obtain a crude product. Finally, it was purified by column chromatography (2% DCM:MeOH 100 mL, 5% DCM:MeOH 200 mL) to obtain compound 25e (712 mg, yield 85%, white solid).

[0121] Compound 25d (712 mg, 4.8 mmol) was added to a reaction flask and dissolved in 20 mL of tetrahydrofuran. Manganese dioxide (4179 mg, 48.1 mmol) was added in batches. After 3 hours, the reaction was monitored by TLC and was complete. The filtrate was collected after filtration and evaporated under reduced pressure to obtain a crude product. Finally, it was purified by column chromatography (2% DCM / MeOH 100 mL, 5% DCM / MeOH 200 mL) to obtain compound 25f (621 mg, yield 88%, white solid).

[0122] Compound 25c (150 mg, 0.8 mmol) was added to a reaction flask, followed by the addition of compound 25f (92 mg, 0.6 mmol) and piperidine (200 μL). After dissolving in 6 mL of absolute ethanol, the mixture was reacted at 80 °C. After 4 hours, obvious yellow solid precipitated out. The solid was collected by filtration and then dried in an oven at 50 °C to obtain the target compound 25 (126 mg, yield 51%, yellow solid). 1 HNMR (400 MHz, DMSO-d6) δ 12.67 (s br, 1H), 10.74 (s, 1H), 8.77 (s br, 1H), 8.33 (s, 1H), 7.93 (d, J = 12.8 Hz, 2H), 7.66 (s, 1H), 7.59 (s, 1H), 7.21 (t, J = 7.7 Hz, 1H), 6.84 (dd, J = 11.4, 7.7 Hz, 2H), 6.76 (d, J = 0.7 Hz, 1H) ppm. HRMS m / z: calcd for C 20 H 13 N3O2 [M+H] + 328.1081, found 328.1077.

[0123] Compounds 26 - 28 were synthesized according to a similar synthetic method.

[0124] Compound 26: 1 H NMR (400 MHz, DMSO-d6) δ 12.63 (s br, 1H), 10.80 (s, 1H), 8.66 (s br, 1H), 8.31 (s, 1H), 7.61 (td, J = 8.1, 6.4 Hz, 1H), 7.54 (s, 1H), 7.42–7.37 (m, 1H), 7.37–7.34 (m, 1H), 7.34–7.31 (m, 1H), 7.26 (t, J = 7.7 Hz, 1H), 7.01 (s, 1H), 6.89 (dd, J = 7.7, 0.8 Hz, 1H), 6.80 (dd, J = 7.7, 0.9 Hz, 1H) ppm. HRMS m / z: calcd for C 22 H 14 FN3O [M+H] + 356.1194, found 356.1192.

[0125] Compound 27: 11H NMR (400 MHz, DMSO-d6) δ 12.65 (s br, 1H), 10.83 (s, 1H), 8.59 (s, 1H), 8.31 (s, 1H), 8.02 (d, J = 7.4 Hz, 1H), 7.84 (dt, J = 14.8, 7.2 Hz, 2H), 7.52 (t, J = 7.4 Hz, 2H), 7.30–7.20 (m, 2H), 6.92 (d, J = 7.7 Hz, 1H), 6.79 (d, J = 7.7 Hz, 1H), 6.36 (s, 1H) ppm. HRMS m / z: calcd for C 23 H 14 F3N3O [M+H] + 406.1162, found 406.1159.

[0126] Compound 28: 1 1H NMR (400 MHz, DMSO-d6) δ 12.58 (s br, 1H), 10.76 (s, 1H), 8.61 (s br, 1H), 8.30 (s, 1H), 7.58–7.53 (m, 3H), 7.53–7.49 (m, 1H), 7.48 (d, J = 1.9 Hz, 1H), 7.46 (d, J = 1.5 Hz, 1H), 7.25 (t, J = 7.7 Hz, 1H), 7.01 (s, 1H), 6.87 (dd, J = 7.7, 0.8 Hz, 1H), 6.78 (dd, J = 7.7, 0.9 Hz, 1H) ppm. HRMS m / z: calcd for C 21 H 14 N4O [M+H] + 339.1240, found 339.1244

[0127] Synthesis of Examples 7, Compounds 30 - 37, 40 - 41

[0128]

[0129] Under argon protection, 4-pyridinecarboxylic acid (30a, 120 μL, 1.5 mmol) and 60 mL of dichloromethane were added to a 100 mL two-necked flask. Then, oxalyl chloride (191 μL, 2.25 mmol) and 2 drops of DMF were added to the reaction. The reaction was carried out at room temperature for 2 h. After the reaction was completed, the excess oxalyl chloride was distilled off. Intermediate 30b was obtained and directly used in the next step without purification, with a yield of 96%.

[0130] Under an ice bath, 22c (178 mg, 1.2 mmol) and 60 mL of ultradry dichloromethane were added to a 100-mL eggplant-shaped flask. Subsequently, triethylamine (499 μL, 3.6 mmol) was added. After addition, the intermediate 30b obtained in the previous step was slowly added dropwise to the above reaction solution. After the addition was complete, the reaction was allowed to proceed at room temperature. After monitoring the reaction by TLC and confirming its completion, the mixture was washed three times with 30 mL of 2N% hydrochloric acid aqueous solution to remove the unreacted intermediate 30b. The mixture was extracted with EA (30 mL × 3), and the combined extracts were dried over anhydrous magnesium sulfate, concentrated by rotary evaporation, and purified by column chromatography (PE / EA = 2:1 - EA) to obtain the intermediate solid 30c (258 mg, 1.02 mmol) with a yield of 85%.

[0131] The last step was carried out according to the synthesis method of compound 1, and the synthesis yield was 83%. 1 H NMR (400 MHz, DMSO-d6): δ 12.68 (br s, 1H), 10.79 (s, 1H), 8.86 (d, J = 6.1 Hz, 2H), 8.80 (s, 1H), 8.32 (s, 1H), 7.99 (d, J = 5.3 Hz, 2H), 7.87 (s, 1H), 7.68 - 7.56 (m, 2H), 7.24 (t, J = 7.9 Hz, 1H), 6.95 (d, J = 7.8 Hz, 1H), 6.81 (dd, J = 7.7, 1.0 Hz, 1H). HRMS m / z: calcd for C 22 H 15 N5O2 [M + H] + 382.1299, found 382.1299.

[0132] Compounds 31 - 37, 40 - 41 were synthesized according to a similar synthetic method.

[0133] Compound 31: 1 H NMR (400 MHz, DMSO-d6): δ 12.71 (br s, 1H), 10.69 (s, 1H), 10.05 (s, 1H), 8.85 (s, 1H), 8.54 (s, 1H), 8.35 (s, 1H), 7.85 - 7.45 (m, 3H), 7.66 - 7.52 (m, 4H), 7.01 (t, J = 7.9 Hz, 1H), 6.71 (d, J = 7.7 Hz, 1H), 6.32 (d, J = 8.1 Hz, 1H). 1313C NMR (100 MHz, DMSO-d6): δ 167.01, 144.23, 142.82, 141.83, 140.79, 132.87, 130.73, 129.28, 128.18, 127.99, 127.63, 126.68, 123.95, 121.13, 120.90, 108.31.

[0134] Compound 32: 1 1H NMR (400 MHz, DMSO-d6): δ 12.69 (d, J = 36.8 Hz, 1H), 10.74 (s, 1H), 10.30 (s, 1H), 8.88 - 8.64 (m, 1H), 8.33 - 8.29 (m, 1H), 8.08 (d, J = 8.8 Hz, 2H), 7.88 (s, 1H), 7.83 - 7.49 (m, 2H), 7.22 (t, J = 7.9 Hz, 1H), 7.13 (d, J = 8.9 Hz, 2H), 6.91 (d, J = 8.0 Hz, 1H), 6.78 (d, J = 7.6 Hz, 1H), 3.87 (s, 3H). 13 13C NMR (100 MHz, DMSO-d6): δ 167.15, 165.17, 162.17, 141.71, 141.55, 132.63, 129.68, 128.22, 127.16, 126.20, 124.94, 121.63, 119.75, 113.93, 107.43, 55.49. HRMS m / z: calcd for C 24 H 18 N4O3 [M + H] + 411.1452, found 411.1450.

[0135] Compound 33: 1 1H NMR (400 MHz, DMSO-d6): δ 12.70 (br s, 1H), 10.75 (s, 1H), 10.31 (s, 1H), 8.79 (s, 1H), 8.32 (s, 1H), 7.92 (s, 1H), 7.76 - 7.56 (m, 4H), 7.22 (t, J = 7.9 Hz, 1H), 7.16 (d, J = 8.5 Hz, 1H), 6.92 (dd, J = 8.0, 1.0 Hz, 1H), 6.78 (dd, J = 7.7, 1.0 Hz, 1H), 3.87 (s, 3H), 3.85 (s, 3H). 1313C NMR (100 MHz, DMSO-d6): δ 167.12, 165.14, 151.86, 148.50, 141.78, 141.54, 132.60, 128.22, 127.17, 126.21, 124.86, 121.63, 121.16, 119.68, 111.17, 110.87, 107.43, 55.72, 55.67. HRMS m / z: calcd for C 25 H 20 N4O4 [M+H] + : 441.1557, found: 441.1556.

[0136] Compound 34: 1 1H NMR (400 MHz, DMSO-d6): δ 12.74 (br s, 1H), 10.70 (s, 1H), 9.87 (s, 1H), 8.82 (s, 1H), 8.50 (s, 1H), 8.36 (s, 1H), 7.83 (d, J = 8.3 Hz, 1H), 7.65 (d, J = 8.5 Hz, 1H), 7.30 (dd, J = 8.4, 2.2 Hz, 1H), 7.17 (d, J = 2.2 Hz, 1H), 7.08 - 7.01 (m, 2H), 6.73 (d, J = 7.7 Hz, 1H), 6.39 (d, J = 8.1 Hz, 1H), 3.74 (s, 3H), 3.66 (s, 3H). 13 13C NMR (100 MHz, DMSO-d6): δ 166.95, 152.18, 148.55, 144.17, 142.74, 141.73, 132.20, 130.94, 128.15, 127.94, 127.47, 123.86, 121.36, 120.85, 120.45, 111.08, 109.31, 108.19, 55.73, 55.55. HRMS m / z: calcd for C 24 H 20 N4O5S [M+H] + 477.1227, found 477.1227.

[0137] Compound 35: 1 1H NMR (400 MHz, DMSO-d6): δ 12.35 (br s, 1H), 10.65 (s, 1H), 8.26 (s, 1H), 8.18 (s, 1H), 7.86 (s, 1H), 7.40 - 7.20 (m, 4H), 7.09 - 7.70 (m, 3H), 6.73 (d, J = 7.5 Hz, 1H), 3.50 (s, 3H).13 C NMR (100 MHz, DMSO-d6): HRMS m / z: calculated for C 21 H 16 N6O2 [M + H] + 385.1408, found 385.1402.

[0138] Compound 36: 1 H NMR (400 MHz, DMSO-d6): δ 13.10 (br s, 1H), 10.69 (s, 1H), 9.81 (s, 1H), 8.83 (s, 1H), 8.47 (s, 1H), 7.90 - 7.83 (m, 2H), 7.68 (d, J = 8.4 Hz, 1H), 7.17 - 7.13 (m, 1H), 6.82 (d, J = 8.0 Hz, 1H), 6.69 (d, J = 7.6 Hz, 1H), 4.61 (s, 1H), 3.42 - 3.17 (m, 1H), 2.51 - 2.40 (m, 1H), 1.95 - 1.92 (m, 3H), 1.74 - 1.47 (m, 3H), 1.27 - 1.18 (m, 2H). 13 C NMR (100 MHz, DMSO-d6): δ 174.24, 166.96, 166.93, 143.88, 141.49, 141.38, 132.47, 128.30, 128.14, 127.26, 125.09, 120.74, 118.90, 118.56, 114.93, 106.83, 68.30, 43.63, 34.75, 31.82, 27.74, 23.58. HRMS m / z: calculated for C 23 H 22 N4O3 [M + H] + 403.1765, found 403.1758.

[0139] Compound 37: 1 H NMR (400 MHz, DMSO-d6): δ 12.71 (d, J = 38.6 Hz, 1H), 10.70 (s, 1H), 9.86 (s, 1H), 8.89 (s, 1H), 8.36 (s, 1H), 7.96 - 7.51 (m, 3H), 7.15 (t, J = 7.9 Hz, 1H), 6.85 (d, J = 8.0 Hz, 1H), 6.70 (d, J = 7.6 Hz, 1H), 3.97 - 3.93 (m, 2H), 3.44 - 3.39 (m, 2H), 2.79 - 2.74 (m, 1H), 1.85 - 1.68 (m, 4H). 1313C NMR (100 MHz, DMSO-d6): δ 173.20, 166.94, 141.71, 141.46, 132.31, 128.07, 127.07, 124.81, 120.74, 118.64, 106.88, 66.42, 41.21, 28.97. HRMS m / z: calcd for C 22 H 20 N4O3 [M+H] + 389.1608, found 389.1609.

[0140] Compound 40: 1 1H NMR (400 MHz, DMSO-d6): δ 13.10 (br s, 1H), 10.68 (s, 1H), 9.97 (s, 1H), 8.92 (d, J = 1.6 Hz, 1H), 8.46 (s, 1H), 8.02 - 7.98 (m, 2H), 7.66 (d, J = 8.4 Hz, 1H), 7.15 (t, J = 7.9 Hz, 1H), 6.88 (d, J = 8.0 Hz, 1H), 6.69 (d, J = 7.6 Hz, 1H), 3.58 (s, 3H), 2.76 - 2.64 (m, 4H). 13 13C NMR (100 MHz, DMSO-d6): δ 173.00, 170.38, 166.96, 143.79, 143.49, 141.68, 141.54, 132.35, 128.75, 128.15, 128.03, 127.94, 125.02, 120.60, 119.02, 118.15, 114.58, 106.76, 51.41, 30.34, 28.56. HRMS m / z: calcd for C 21 H 18 N4O4 [M+H] + 391.1401, found 391.1397 Compound 41: 1 1H NMR (400 MHz, DMSO-d6): δ 12.72 (d, J = 46.2 Hz, 1H), 10.73 (s, 1H), 10.29 (s, 1H), 8.94 (s, 1H), 8.34 (s, 1H), 8.07 - 7.59 (m, 3H), 7.19 (t, J = 7.9 Hz, 1H), 6.92 (d, J = 8.1 Hz, 1H), 6.74 (d, J = 7.6 Hz, 1H), 3.68 (q, J = 11.3 Hz, 2H). 1313C NMR(100MHz,DMSO-d6):δ167.00,162.29,144.19,142.19,141.67,131.21,128.35,128.00,127.45,126.44,124.50,123.69,120.50,118.39,107.49.HRMS m / z:calcd for C 19 H 13 F3N4O2[M+H] + 387.1063,found 387.1063.

[0141] Example 8. Synthesis of Compounds 38 and 39

[0142]

[0143] Under argon protection, trans-4-(((tert-butoxycarbonyl)amino)methyl)cyclohexanecarboxylic acid (38a, 514 mg, 2 mmol) and 50 mL of DMF were added to a 100 mL two-necked flask. Subsequently, HATU (912 mg, 2.4 mmol) and N,N-diisopropylethylamine (1.04 mL, 6 mmol) were added to the reaction solution. After activation at room temperature for 1 h, intermediate 22c was added, and the reaction was carried out overnight. After the reaction solution was diluted with water, it was extracted with EA (30 mL x 3). The combined extracts were dried over anhydrous magnesium sulfate, evaporated to dryness, and purified by column chromatography (EA / DCM = 5:1 - 1:1) to obtain intermediate 38b (615 mg, 1.64 mmol) with a yield of 82%. Compound 38 was synthesized using intermediate 38b according to the synthesis of Compound 1, and the synthesis yield was 76%. 1 1H NMR(400MHz,DMSO-d6):δ12.72(br s,1H),10.68(s,1H),9.78(s,1H),8.84(s,1H),8.34(s,1H),7.90 - 7.64(s,3H),7.14(t,J = 7.9Hz,1H),6.83(d,J = 8.0Hz,2H),6.69(d,J = 7.6Hz,1H),2.82(t,J = 6.3Hz,2H),2.47 - 2.41(m,1H),1.99 - 1.96(m,2H),1.81 - 1.78(m,2H),1.50 - 1.38(s,12H),1.03 - 0.93(m,2H). 1313C NMR(100MHz, DMSO-d6): δ 174.40, 166.98, 155.81, 144.12, 141.66, 141.45, 132.46, 128.04, 127.01, 124.86, 120.80, 118.67, 106.81, 77.35, 46.11, 44.49, 37.42, 29.58, 28.83, 28.31. HRMS m / z: calcd for C 29 H 33 N5O4 [M + H] + 516.2605, found 516.2599.

[0144] The intermediate 38 was dissolved in 4 mL of TFA / DCM = 1 / 1 solution, stirred at room temperature for 30 minutes, and then the reaction solution was evaporated to dryness. The solid was washed with methanol to obtain the final product 39 (129 mg, 0.5 mmol), with a yield of 30%. 1 1H NMR(400MHz, DMSO-d6): δ 10.75(s, 1H), 9.87(s, 1H), 9.10(s, 1H), 8.91(s, 1H), 7.92 - 7.79(m, 5H), 7.18(t, J = 7.9Hz, 1H), 6.86(d, J = 8.1Hz, 1H), 6.71(d, J = 7.7Hz, 1H), 2.74 - 2.67(m, 2H), 2.01 - 1.85(m, 4H), 1.58 - 1.38(m, 4H), 1.10 - 1.00m, 2H). 13 13C NMR(100MHz, DMSO-d6): δ 174.22, 166.81, 159.31, 158.98, 158.66, 158.33, 142.60, 141.97, 139.52, 133.67, 132.94, 132.59, 130.78, 129.15, 128.90, 126.98, 120.73, 118.04, 117.53, 115.38, 114.26, 107.03, 44.38, 43.92, 35.11, 28.92, 28.43.

[0145] Example 9, Synthesis of Compounds 43 - 53

[0146]

[0147] Triphosgene (144 mg, 0.5 mmol) was added to a reaction flask, dissolved in 6 mL of dry dichloromethane, and pre-cooled to 0 °C. p-Chloroaniline (204 mg, 1.6 mmol) was added to a 10 mL centrifuge tube, dissolved in 8 mL of dry dichloromethane, and slowly dropped into the dichloromethane solution of triphosgene. After the addition was complete, the mixture was stirred at room temperature for 30 minutes and then concentrated by rotary evaporation to obtain p-chlorophenyl isocyanate. 22c (200 mg, 1.3 mmol) was added to another reaction flask, dissolved in 8 mL of dry dichloromethane, triethylamine (1871 μL, 13.5 mmol) was added, and the mixture was stirred at room temperature for 30 minutes and then dropped into p-chlorophenyl isocyanate. The reaction mixture was then heated at 40 °C for 4 hours. After the reaction was monitored by TLC to be complete, it was evaporated under reduced pressure to obtain the crude product, which was finally purified by column chromatography (100 mL of DCM, 100 mL of 2% DCM / MeOH, 200 mL of 3% DCM / MeOH) to obtain intermediate 43a (260 mg, yield 66%, gray-violet solid).

[0148] 43a (260 mg, 0.9 mmol) was dissolved in 6 mL of absolute ethanol, 1 g (105 mg, 0.7 mmol), 200 μL of piperidine were added, and the reaction was carried out at 80 °C. After 4 hours, obvious yellow solid precipitated. The solid was collected by filtration and dried in an oven at 50 °C to obtain the target compound 43 (147 mg, yield 38%, yellow solid). 1 H NMR (400 MHz, DMSO-d6) δ 12.71 (sbr, 1H), 10.65 (s, 1H), 9.26 (s, 1H), 8.86 (d, J = 37.3 Hz, 1H), 8.50 (s, 1H), 8.32 (s, 1H), 8.05 (s, 1H), 7.91 (d, J = 85.6 Hz, 1H), 7.66 (s, 1H), 7.55–7.50 (m, 2H), 7.36–7.30 (m, 2H), 7.19–7.10 (m, 2H), 6.65 (dd, J = 7.0, 1.5 Hz, 1H) ppm. HRMS m / z: calcd for C 23 H 16 ClN5O2 [M+H] + 430.1065, found 430.1071.

[0149] Compounds 44 - 53 were synthesized according to a similar synthetic method.

[0150] Compound 44: 11H NMR (400 MHz, DMSO-d6) δ 12.78 (s br, 1H), 10.62 (s, 1H), 8.86 (s, 1H), 8.63 (s, 1H), 8.35 (s, 1H), 8.25 (d, J = 7.5 Hz, 2H), 7.97 (d, J = 8.2 Hz, 1H), 7.64 (d, J = 8.5 Hz, 1H), 7.17–7.11 (m, 1H), 7.09–7.03 (m, 4H), 6.62 (dd, J = 7.5, 1.0 Hz, 1H), 2.25 (s, 6H) ppm. HRMS m / z: calcd for C 25 H 21 N5O2 [M+H] + 424.1768, found 424.1770.

[0151] Compound 45: 1 1H NMR (400 MHz, DMSO-d6) δ 12.69 (d, J = 43.1 Hz, 1H), 10.68 (s, 1H), 9.29 (s, 1H), 8.88 (d, J = 42.8 Hz, 1H), 8.56 (s, 1H), 8.33 (s, 1H), 8.04 (s, 1H), 7.83 (dd, J = 6.9, 2.1 Hz, 1H), 7.61 (dd, J = 24.2, 16.0 Hz, 2H), 7.38–7.31 (m, 2H), 7.19–7.13 (m, 1H), 7.10 (d, J = 7.5 Hz, 1H), 6.66 (dd, J = 7.4, 1.0 Hz, 1H) ppm. HRMS m / z: calcd for C 23 H 15 ClFN5O2 [M+H] + 448.0971, found 448.0980.

[0152] Compound 46: 11H NMR (400 MHz, DMSO-d6) δ 12.74 (s br, 1H), 10.69 (s, 1H), 9.09 (s, 1H), 8.92 (s, 1H), 8.42 (s, 1H), 8.34 (s, 1H), 8.11 (s, 1H), 7.95 (d, J = 8.4 Hz, 1H), 7.84 (d, J = 8.3 Hz, 1H), 7.63 (d, J = 8.5 Hz, 1H), 7.47 (d, J = 1.3 Hz, 1H), 7.15 (dt, J = 7.4, 4.7 Hz, 2H), 7.08 (d, J = 7.5 Hz, 1H), 6.66 (dd, J = 7.5, 0.8 Hz, 1H), 2.26 (s, 3H) ppm. HRMS m / z: calcd for C 24 H 18 BrN5O2 [M+H] + 488.0717, found 488.0716.

[0153] Compound 47: 1 1H NMR (400 MHz, DMSO-d6) δ 12.76 (s br, 1H), 10.61 (s, 1H), 8.82 (s, 1H), 8.33 (s, 1H), 8.18 (s, 1H), 7.96 (s, 1H), 7.89 (d, J = 8.4 Hz, 1H), 7.63 (d, J = 8.5 Hz, 1H), 7.15–7.03 (m, 2H), 6.90 (d, J = 2.8 Hz, 1H), 6.59 (dd, J = 7.3, 1.1 Hz, 1H), 2.67–2.55 (m, 1H), 0.70–0.60 (m, 2H), 0.52–0.42 (m, 2H) ppm. HRMS m / z: calcd for C 20 H 17 N5O2 [M+H] + 360.1455, found 360.1456.

[0154] Compound 48: 11H NMR (400 MHz, DMSO-d6) δ 12.47 (s br, 1H), 10.62 (s, 1H), 8.11 (s, 1H), 7.85 (s, 1H), 7.48 (d, J = 34.5 Hz, 2H), 7.37–7.27 (m, 2H), 7.25–7.17 (m, 2H), 6.76 (s, 1H), 6.69 (d, J = 8.7 Hz, 1H), 6.62 (dt, J = 7.5, 3.7 Hz, 1H), 6.49 (d, J = 2.4 Hz, 1H), 6.25 (dd, J = 8.6, 2.4 Hz, 1H), 3.68 (s, 6H) ppm. HRMS m / z: calcd for C 25 H 21 N5O4 [M+H] + 456.1666, found 456.1667.

[0155] Compound 49: 1 1H NMR (400 MHz, DMSO-d6) δ 12.69 (s br, 1H), 10.68 (s, 1H), 9.32 (s, 1H), 8.87 (s, 1H), 8.53 (s, 1H), 8.32 (s, 1H), 8.05 (s, 1H), 7.91 (s br, 1H), 7.62 (s, 1H), 7.52 (d, J = 8.7 Hz, 2H), 7.40 (d, J = 8.7 Hz, 2H), 7.20–7.11 (m, 2H), 6.65 (dd, J = 6.9, 1.6 Hz, 1H), 4.04 (s, 1H) ppm. HRMS m / z: calcd for C 25 H 17 N5O2 [M+H] + 420.14556, found 420.1450.

[0156] Compound 50: 11H NMR (400 MHz, DMSO-d6) δ 12.69 (s br, 1H), 10.68 (s, 1H), 9.04 (s, 1H), 8.87 (s, 1H), 8.64 (s, 1H), 8.33 (s, 1H), 8.07 (s, 1H), 7.91 (s br, 1H), 7.81 (d, J = 3.0 Hz, 1H), 7.62 (d, J = 8.0 Hz, 1H), 7.20–7.13 (m, 1H), 7.09 (d, J = 7.4 Hz, 1H), 6.95 (d, J = 8.9 Hz, 1H), 6.65 (dd, J = 7.4, 0.9 Hz, 1H), 6.51 (dd, J = 8.9, 3.1 Hz, 1H), 3.86 (s, 3H), 3.66 (s, 3H) ppm. HRMS m / z: calcd for C 25 H 21 N5O4 [M+H] + 456.1666, found 456.1669.

[0157] Compound 51: 1 1H NMR (400 MHz, DMSO-d6) δ 12.68 (d, J = 42.1 Hz, 1H), 10.60 (s, 1H), 8.83 (d, J = 47.4 Hz, 1H), 8.33 (s, 1H), 8.15 (s, 1H), 7.98 (s, 1H), 7.90–7.45 (m, 2H), 7.09 (dd, J = 12.2, 5.2 Hz, 2H), 6.67 (t, J = 5.6 Hz, 1H), 6.57 (dd, J = 6.8, 1.8 Hz, 1H), 3.02 (d, J = 6.2 Hz, 2H), 1.03–0.90 (m, 1H), 0.46–0.39 (m, 2H), 0.23–0.17 (m, 2H) ppm. HRMS m / z: calcd for C 21 H 19 N5O2 [M+H] + 374.1612, found 374.1614.

[0158] Compound 52: 11H NMR (400 MHz, DMSO-d6) δ 12.70 (s br, 1H), 10.71 (s, 1H), 9.31 (s, 1H), 8.98 (s, 1H), 8.54 (s, 1H), 8.34 (s, 1H), 8.11 (s, 1H), 8.08 (d, J = 8.2 Hz, 1H), 7.91 (s br, 1H), 7.64 (s br, 1H), 7.47 (dd, J = 7.7, 1.4 Hz, 1H), 7.39–7.31 (m, 1H), 7.19 (d, J = 8.0 Hz, 1H), 7.07 (d, J = 8.0 Hz, 1H), 7.01 (td, J = 7.6, 0.9 Hz, 1H), 6.71–6.66 (m, 1H), 4.68 (s, 1H) ppm.

[0159] Compound 53: 1 1H NMR (400 MHz, DMSO-d6) δ 12.72 (s br, 1H), 10.67 (s, 1H), 9.18 (s, 1H), 8.87 (s, 1H), 8.50 (s, 1H), 8.33 (s, 1H), 8.08 (s, 1H), 7.92 (d, J = 8.4 Hz, 1H), 7.61 (d, J = 8.5 Hz, 1H), 7.50 (d, J = 7.6 Hz, 2H), 7.29 (t, J = 7.9 Hz, 2H), 7.16 (d, J = 2.1 Hz, 1H), 7.15 (s, 1H), 6.98 (t, J = 7.4 Hz, 1H), 6.64 (dd, J = 5.3, 3.3 Hz, 1H) ppm.

[0160] Example 10. Synthesis of Compound 55

[0161]

[0162] Under an ice bath, tetrabutylammonium fluoride (2 mL, 1 M in THF) was added dropwise to a THF solution of intermediate 55a (489 mg, 1 mmol). After the addition was complete, the ice bath was removed and the reaction was continued at room temperature for 4 h. After TLC detection showed that the raw materials had completely reacted, the mixture was concentrated under reduced pressure and purified by column chromatography to obtain 251 mg of yellow solid 55 in a yield of 67%. 11H NMR (400 MHz, DMSO-d6): δ 12.69 (br s, 1H), 10.60 (s, 1H), 8.96 (s, 1H), 8.43 (s, 1H), 8.33 (s, 1H), 7.90 - 7.62 (m, 2H), 7.12 (t, J = 8.0 Hz, 1H), 6.72 (d, J = 9.0 Hz, 1H), 6.44 (d, J = 7.6 Hz, 1H), 4.72 4.57 (m, 2H), 3.70 - 3.60 (m, 1H), 2.17 - 1.86 (m, 3H), 1.76 - 1.39 (m, 5H). 13 13C NMR (100 MHz, DMSO-d6): δ 167.26, 153.74, 153.52, 144.00, 141.89, 141.83, 140.90, 140.86, 129.28, 128.71, 127.22, 127.15, 125.21, 125.19, 112.10, 112.03, 106.64, 106.43, 102.20, 102.15, 74.39, 72.00, 66.84, 66.54, 31.56, 30.49, 28.27, 27.38. HRMS m / z: calcd for C 22 H 21 N3O3 [M + H] + 376.1656, found 376.1653.

[0163] Example 11. Synthesis of Compound 57

[0164]

[0165] Under an ice bath, 2 mL of TAF was added dropwise to a dichloromethane solution (2 mL) of intermediate 57a (124 mg, 0.27 mmol). After the addition was complete, the ice bath was removed and the reaction was continued at room temperature for 1 h. After TLC detection showed that the raw materials had completely reacted, the mixture was concentrated under reduced pressure and purified by column chromatography (DCM / MeOH = 50:1 - 10:1) to obtain 120 mg of yellow solid 57, with a yield of 81%. 11H NMR (400 MHz, DMSO-d6): δ 10.72 (s, 1H), 9.16 (s, 1H), 9.08 (s, 1H), 8.79 (br s, 1H), 8.68 (br s, 1H), 8.26 (s, 1H), 7.98 (dd, J = 8.6, 1.6 Hz, 1H), 7.80 (d, J = 8.5 Hz, 1H), 7.20 (t, J = 8.1 Hz, 1H), 6.81 (d, J = 8.8 Hz, 1H), 6.51 (d, J = 7.6 Hz, 1H), 4.96 - 4.90 (m, 1H), 3.30 - 3.28 (m, 2H), 3.21 - 3.16 (m, 2H), 2.27 - 2.20 (m, 2H), 2.06 - 1.98 (m, 2H). 13 13C NMR (100 MHz, DMSO-d6): δ 166.98, 159.46, 159.13, 158.80, 158.48, 153.20, 142.52, 138.72, 133.36, 132.49, 131.28, 130.27, 129.64, 127.17, 118.29, 117.36, 115.34, 114.20, 111.64, 106.76, 103.03, 69.54, 40.80, 27.38.

[0166] Example 12, Synthesis of Compounds 58 - 71

[0167]

[0168] Under an ice bath, 4 - hydroxypiperidine (60a, 505 mg, 5 mmol) and 60 mL of ultradry dichloromethane were added to a 100 - mL two - necked flask. Subsequently, triethylamine (2.08 mL, 15 mmol) was added. After addition, methanesulfonyl chloride (389 μL, 5 mmol) was slowly added dropwise to the above reaction solution. After the addition was complete, the reaction was allowed to proceed at room temperature. After monitoring the reaction to completion by TLC, the reaction solution was first washed three times with 30 mL of 2N% hydrochloric acid aqueous solution to remove the unreacted methanesulfonyl chloride, extracted with EA (30 mL × 3), the combined extracts were dried over anhydrous magnesium sulfate, concentrated by rotary evaporation, and purified by column chromatography (DCM / MeOH = 50:1 - 10:1) to obtain 258 mg of intermediate 60b with a yield of 87%. Using compound 60b, compound 60 was synthesized according to the synthesis method of compound 1. 11H NMR (400 MHz, DMSO-d6): δ 12.68 (br s, 1H), 10.63 (s, 1H), 8.99 (s, 1H), 8.36 (s, 1H), 8.33 (s, 1H), 7.92 (d, J = 8.5 Hz, 1H), 7.63 (d, J = 8.4 Hz, 1H), 7.15 (t, J = 8.1 Hz, 1H), 6.77 (d, J = 8.5 Hz, 1H), 6.48 (d, J = 7.6 Hz, 1H), 4.86 - 4.81 (m, 1H), 3.43 - 3.34 (m, 2H), 3.28 - 3.22 (m, 2H), 2.85 (s, 3H), 2.17 - 2.10 (m, 2H), 1.99 - 1.91 (m, 2H). 13 13C NMR (100 MHz, DMSO-d6): δ 167.25, 153.05, 144.08, 141.96, 141.26, 129.35, 128.61, 127.38, 125.05, 112.26, 106.63, 102.66, 70.96, 42.69, 34.27, 29.96. HRMS m / z: calcd for C 22 H 22 N4O4S [M + H] + 439.1435, found 439.1434.

[0169] Compounds 58, 59, and 61 - 71 were synthesized according to a similar synthetic method.

[0170] Compound 58: 1 1H NMR (400 MHz, DMSO-d6): δ 12.69 (br s, 1H), 10.63 (s, 1H), 8.96 (d, J = 1.5 Hz, 1H), 8.36 (s, 1H), 8.33 (s, 1H), 7.49 (dd, J = 8.4, 1.2 Hz, 1H), 7.63 (d, J = 8.4 Hz, 1H), 7.14 (t, J = 8.1 Hz, 1H), 6.77 (d, J = 8.5 Hz, 1H), 6.47 (d, J = 7.6 Hz, 1H), 4.87 (tt, J = 7.5, 3.6 Hz, 1H), 3.86 - 3.67 (m, 2H), 3.49 - 3.40 (m, 2H), 2.10 - 1.98 (m, 5H), 1.88 - 1.69 (m, 2H). 13CNMR(100MHz, DMSO-d6): δ 168.67, 167.66, 153.63, 144.57, 142.37, 141.58, 129.72, 128.99, 127.64, 125.47, 112.54, 106.96, 102.93, 72.46, 43.38, 38.52, 31.37, 30.81, 21.74. HRMS m / z: calcd for C 23 H 22 N4O3 [M + H] + 403.1765, found 403.1764.

[0171] Compound 59: 1 H NMR(400MHz, DMSO-d6): δ 12.68(br s, 1H), 10.62(s, 1H), 8.91(s, 1H), 8.34(s, 1H), 8.25(d, J = 7.4Hz, 1H), 7.80 - 7.51(m, 2H), 7.117 - 7.11m, 1H), 6.80 - 6.71(m, 1H), 6.48(t, J = 7.5Hz, 1H), 4.84 - 4.66(m, 1H), 4.09 - 3.81(m, 3H), 3.37 - 3.30(m, 1H), 1.97(s, 3H), 1.81 - 1.55(m, 4H). 13 C NMR(100MHz, DMSO-d6): δ 168.75, 168.48, 167.19, 167.12, 153.18, 153.04, 143.88, 142.03, 141.92, 141.28, 129.46, 129.30, 128.56, 127.16, 124.96, 112.17, 112.15, 106.37, 106.09, 102.68, 102.52, 70.54, 70.14, 48.98, 45.85, 44.51, 41.15, 28.55, 28.23, 22.06, 21.42, 21.10, 20.98. HRMS m / z: calcd for C 23 H 22 N4O3 [M + H] + 403.1765, found 403.1760.

[0172] Compound 61: 11H NMR (400 MHz, DMSO-d6): δ 12.73 (br s, 1H), 10.66 (s, 1H), 8.95 (s, 1H), 8.35 (s, 1H), 8.23 (s, 1H), 7.84 (d, J = 8.2 Hz, 1H), 7.62 (dd, J = 8.4, 4.0 Hz, 1H), 7.15 (t, J = 8.0 Hz, 1H), 6.72 (dd, J = 8.5, 3.9 Hz, 1H), 6.50 (d, J = 7.6 Hz, 1H), 5.32 - 5.22 (m, 1H), 3.86 - 3.40 (m, 4H), 2.35 - 2.10 (m, 2H), 1.94 (s, 3H). 13 13C NMR (100 MHz, DMSO-d6): δ 168.56, 168.37, 167.16, 152.84, 152.82, 144.05, 141.91, 141.88, 141.16, 141.08, 129.31, 129.28, 128.54, 128.49, 127.29, 124.84, 124.80, 112.25, 112.15, 106.69, 106.49, 102.86, 76.79, 75.57, 52.49, 51.07, 45.00, 43.54, 31.31, 29.94, 22.34, 22.07. HRMS m / z: calcd for C 22 H 20 N4O3 [M + H] + 389.1608, found 389.1608.

[0173] Compound 62: 1 1H NMR (400 MHz, DMSO-d6): δ 12.67 (d, J = 41.1 Hz, 1H), 10.62 (s, 1H), 9.02 (s, 1H), 8.37 - 8.31 (m, 2H), 8.06 7.56 (m, 2H), 7.46 - 7.43 (m, 5H), 7.14 (t, J = 8.1 Hz, 1H), 6.78 (d, J = 8.5 Hz, 1H), 6.47 (d, J = 7.6 Hz, 1H), 4.94 - 4.88 (m, 1H), 3.99 (br s, 1H), 3.60 - 3,47 (m, 3H), 2.10 (br s, 2H), 1.86 (br s, 2H). 13¹³C NMR (100 MHz, DMSO-d₆): δ 169.14, 167.19, 153.19, 144.02, 141.92, 141.17, 136.14, 129.47, 129.30, 128.45, 127.27, 126.79, 125.02, 112.07, 106.52, 102.52, 72.10, 44.35, 30.40. HRMS m / z: calcd for C 28 H 24 N₄O₃ [M+H]⁺ + 465.1921, found 465.1922.

[0174] Compound 63: 1 ¹H NMR (400 MHz, DMSO-d₆): δ 12.69 (br s, 1H), 10.63 (s, 1H), 9.01 (s, 1H), 8.67 (d, J = 5.9 Hz, 2H), 8.36 (s, 1H), 8.33 (s, 1H), 8.05 - 7.66 (m, 2H), 7.45 (d, J = 5.9 Hz, 2H), 7.14 (t, J = 8.1 Hz, 1H), 6.78 (d, J = 8.6 Hz, 1H), 6.47 (d, J = 7.6 Hz, 1H), 4.94 - 4.89 (m, 1H), 3.66 - 3.49 (m, 2H), 3.58 (dd, J = 56.1, 12.0 Hz, 2H), 2.16 - 1.85 (m, 4H). 13 ¹³C NMR (100 MHz, DMSO-d₆): δ 167.22, 166.85, 153.18, 150.10, 144.06, 143.68, 141.95, 141.23, 129.33, 128.61, 127.34, 125.05, 121.14, 112.08, 106.50, 102.56, 71.94, 48.66, 44.14, 30.71, 30.21. HRMS m / z: calcd for C 27 H 23 N₅O₃ [M+H]⁺ + 466.1874, found 466.1872.

[0175] Compound 64: 11H NMR (400 MHz, DMSO-d6): δ 12.68 (br s, 1H), 10.62 (s, 1H), 8.96 (s, 1H), 8.37 (s, 1H), 8.33 (s, 1H), 7.91 - 7.62 (m, 2H), 7.41 (d, J = 8.7 Hz, 2H), 7.14 (t, J = 8.0 Hz, 1H), 6.98 (d, J = 8.7 Hz, 2H), 6.78 (d, J = 8.5 Hz, 1H), 6.47 (d, J = 7.6 Hz, 1H), 4.93 - 4.88 (m, 1H), 3.79 (s, 5H), 3.52 (br s, 2H), 2.09 (br s, 2H), 1.85 (br s, 2H). 13 13C NMR (100 MHz, DMSO-d6): δ 169.19, 167.26, 160.21, 153.23, 144.04, 141.97, 141.20, 129.32, 128.94, 128.66, 128.63, 128.00, 127.34, 125.10, 113.65, 112.14, 106.51, 102.55, 72.15, 55.21, 48.67, 30.64.

[0176] Compound 65: 1 1H NMR (400 MHz, DMSO-d6): δ 12.67 (d, J = 39.0 Hz, 1H), 10.62 (s, 1H), 9.04 (s, 1H), 8.39 - 8.32 (m, 2H), 8.07 - 7.57 (m, 2H), 7.14 (t, J = 8.1 Hz, 1H), 7.02 - 6.70 (m, 3H), 6.78 (d, J = 8.5 Hz, 1H), 6.47 (d, J = 7.6 Hz, 1H), 4.93 - 4.88 (m, 1H), 3.79 (s, 4H), 3.77 (s, 4H), 3.54 (br s, 2H), 2.10 (br s, 2H), 1.87 (br s, 2H). 13 13C NMR (100 MHz, DMSO-d6): δ 169.13, 167.22, 153.23, 149.81, 148.44, 141.94, 141.17, 129.31, 128.17, 127.26, 125.06, 119.81, 112.11, 111.14, 110.95, 106.54, 102.52, 72.17, 55.55, 48.64, 30.63. HRMS m / z: calcd for C 30 H 28 N4O5 [M + H] +525.2132, found 525.2131.

[0177] Compound 66: 1 H NMR(400MHz, DMSO-d6): δ12.64(br s, 1H), 10.63(s, 1H), 8.98(s, 1H), 8.37(s, 1H), 8.32(s, 1H), 7.91(br s, 1H), 7.76(d, J = 5.0Hz, 1H), 7.63(d, J = 8.4Hz, 1H), 7.46 - 7.45(m, 1H), 7.17 - 7.12(m, 2H), 6.79(d, J = 8.5Hz, 1H), 6.47(d, J = 7.6Hz, 1H), 4.96 - 4.91(m, 1H), 3.99 - 3.93(m, 2H), 3.70 - 3.64(m, 2H), 2.17 - 2.10(m, 2H), 1.94 - 1.85(m, 2H). 13 C NMR(100MHz, DMSO-d6): δ167.28, 162.48, 153.22, 144.11, 141.98, 141.26, 137.30, 129.40, 129.36, 128.98, 127.39, 127.13, 125.10, 112.15, 106.53, 102.60, 72.00, 30.73. HRMS m / z: calcd for C 26 H 22 N4O3S [M + H] + 471.1485, found 471.1481.

[0178] Compound 67: 1 H NMR(400MHz, DMSO-d6): δ12.68(br s, 1H), 10.62(s, 1H), 8.96(s, 1H), 8.36(s, 1H), 8.32(s, 1H), 8.07(s, 1H), 7.89(br s, 1H), 7.68(s, 1H), 7.62(d, J = 8.4Hz, 1H), 7.15(t, J = 8.1Hz, 1H), 6.80(d, J = 8.6Hz, 1H), 6.47(d, J = 7.6Hz, 1H), 4.96 - 4.90(m, 1H), 3.96 - 3.90(m, 2H), 3.85(s, 3H), 3.63 - 3.58(m, 2H), 2.14 - 2.07(m, 2H), 1.88 - 1.80(m 2H). 13¹³C NMR (100 MHz, DMSO-d₆): δ 167.19, 162.68, 153.20, 144.00, 141.92, 141.14, 139.20, 132.36, 129.30, 128.56, 127.28, 125.03, 116.38, 112.08, 106.55, 102.49, 72.12, 48.63, 30.80. HRMS m / z: calcd for C 26 H 24 N₆O₃ [M + H] + 469.1983, found 469.1982.

[0179] Compound 68: 1 ¹H NMR (400 MHz, DMSO-d₆): δ 12.68 (br s, 1H), 10.62 (s, 1H), 8.95 (s, 1H), 8.35 (s, 1H), 8.32 (s, 1H), 7.88 (s, 1H), 7.62 (d, J = 8.4 Hz, 1H), 7.14 (t, J = 8.1 Hz, 1H), 6.78 (d, J = 8.5 Hz, 1H), 6.47 (d, J = 7.6 Hz, 1H), 4.90 - 4.85 (m, 1H), 3.85 - 3.78 (m, 4H), 3.57 - 3.38 (m, 4H), 2.96 - 2.89 (m, 1H), 2.09 - 1.99 (m, 2H), 1.83 - 1.71 (m, 2H), 1.62 - 1.51 (m, 4H). 13 ¹³C NMR (100 MHz, DMSO-d₆): δ 172.27, 167.18, 153.20, 144.00, 141.92, 141.06, 129.31, 128.57, 127.22, 125.05, 112.11, 106.59, 102.51, 72.12, 66.31, 41.85, 38.39, 36.30, 31.36, 30.43, 29.05, 28.92. HRMS m / z: calcd for C 27 H 28 N₄O₄ [M + H] + 473.2183, found 473.2181. Compound 70: 11H NMR (400 MHz, DMSO-d6): δ 12.67 (br s, 1H), 10.62 (s, 1H), 8.96 (s, 1H), 8.37 (s, 1H), 8.33 (s, 1H), 7.89 (s, 1H), 7.63 (d, J = 8.3 Hz, 1H), 7.15 (t, J = 8.0 Hz, 1H), 6.79 (d, J = 8.6 Hz, 1H), 6.47 (d, J = 7.6 Hz, 1H), 4.92 - 4.87 (m, 1H), 3.97 - 3.84 (m, 2H), 3.71 - 3.42 (m, 2H), 2.09 - 1.98 (m, 3H), 1.84 - 1.75 (m, 2H), 0.75 - 0.68 (m, 4H). 13 13C NMR (100 MHz, DMSO-d6): δ 171.02, 167.24, 153.22, 144.39, 141.95, 141.12, 129.32, 128.78, 127.26, 125.09, 112.15, 106.56, 102.53, 72.15, 56.12, 42.05, 31.20, 30.40, 18.60, 10.38, 6.97. HRMS m / z: calcd for C 25 H 24 N4O3 [M + H] + 429.1921, found 429.1920.

[0180] Compound 71: 1 1H NMR (400 MHz, DMSO-d6): δ 12.69 (br s, 1H), 10.62 (s, 1H), 8.96 (s, 1H), 8.35 (s, 1H), 8.33 (s, 1H), 7.88 (d, J = 8.5 Hz, 1H), 7.62 (d, J = 8.4 Hz, 1H), 7.14 (t, J = 8.1 Hz, 1H), 6.78 (d, J = 8.5 Hz, 1H), 6.47 (d, J = 7.6 Hz, 1H), 4.91 - 4.85 (m, 1H), 3.87 - 3.70 (m, 2H), 3.56 (t, J = 6.6 Hz, 2H), 3.51 - 3.43 (m, 2H), 3.22 (s, 3H), 2.62 (t, J = 6.6 Hz, 2H), 2.09 - 1.99 (m, 2H), 1.84 - 1.70 (m, 2H). 1313C NMR (100 MHz, DMSO-d6): δ 168.78, 167.20, 153.20, 144.02, 141.92, 141.06, 129.30, 128.59, 127.25, 125.07, 112.09, 106.56, 102.49, 72.03, 68.40, 57.99, 42.22, 38.26, 32.55, 31.03, 30.40. HRMS m / z: calcd for C 25 H 26 N4O4 [M + H] + 447.2027, found 447.2019.

[0181] Example 13, Synthesis of Compound 72

[0182]

[0183] Weigh 72a (300 mg, 1.1 mmol), tris(dibenzylideneacetone)dipalladium(0) (201 mg, 0.22 mmol), 1,1'-bis(diphenylphosphino)ferrocene (88 mg, 0.16 mmol) into a reaction flask. After dissolving in 6 mL of DMF, add triethylamine (305 μL, 2.2 mmol). Stir at room temperature for 10 min under argon protection, then add methyl 3-mercaptopropionate (264 mg, 2.2 mmol) and react at 80 °C for 2 h. After monitoring the reaction to completion by TLC, extract with ethyl acetate and water (25 mL × 3). Combine the organic layers and wash with saturated sodium chloride solution (50 mL × 3). Dry over anhydrous magnesium sulfate, rotary evaporate to remove the solvent, and then perform column chromatography (2% DCM / MeOH 200 mL) to obtain intermediate 72b (302 mg, yield 91%, yellow solid).

[0184] Dissolve intermediate 72b (302 mg, 1.0 mmol) in 6 mL of methanol, add 80% hydrazine hydrate solution (488 μL, 10.1 mmol), and react at 80 °C for 1 h. After monitoring the reaction to completion by TLC, adjust the pH to neutral with 2N hydrochloric acid solution, then extract with ethyl acetate and water (25 mL × 3). Combine the organic layers, dry over anhydrous magnesium sulfate, rotary evaporate to remove the solvent, and then perform column chromatography (2% DCM / MeOH 300 mL) to obtain intermediate 72c (224 mg, yield 89%, yellow solid).

[0185] 72c (224 mg, 0.9 mmol) was dissolved in 6 mL of anhydrous ethanol. After adding 25f (109 mg, 0.7 mmol) and 200 μL of piperidine, the reaction was carried out at 80 °C for 5 h. After the reaction was monitored by TLC to be complete, the solvent was removed by rotary evaporation and then column chromatography (2% DCM / MeOH 100 mL, 3% DCM / MeOH 100 mL, 5% DCM / MeOH 200 mL) was performed to obtain compound 72 (158 mg, yield 46%, yellow solid). 1 1H NMR (400 MHz, DMSO) δ 12.15 (s br, 1H), 10.73 (s, 1H), 8.84 (s, 1H), 8.56 (d, J = 3.6 Hz, 1H), 8.34 (s, 1H), 7.64 (s, 1H), 7.20 (t, J = 7.9 Hz, 1H), 7.04 (d, J = 8.1 Hz, 1H), 6.74 (d, J = 7.2 Hz, 1H), 4.03 (q, J = 7.1 Hz, 2H), 3.33 (s, 3H), 2.70 (dd, J = 11.8, 4.9 Hz, 2H) ppm.

[0186] The beneficial effects of the present invention are demonstrated by the following experimental examples.

[0187] Experimental Example 1, Inhibitory Activity of the Compounds of the Present Invention against sQC / gQC Proteins

[0188] (1) Experimental Materials

[0189] Recombinant human sQC (33 - 361aa) and human gQC (53 - 382aa) proteins were expressed in the prokaryotic system using the pET32a vector (method reference: Protein Expr Purif. 2005 43(1):65 - 72); recombinant Bacillus amyloliquefaciens PGP - I (1 - 215aa) protein was expressed using the pET28 vector (method reference: J Biomol Struct Dyn. 2011 29(2):267 - 82); the fluorescent substrate H - Gln - AMC (Nanjing Lion Biotechnology Co., Ltd.); the synthetic fluorescent substrate pGA, the positive control compound PQ912; the target compounds synthesized in the examples of the present invention.

[0190]

[0191] (2) Experimental Method

[0192] The activity test of sQC / gQC protein was carried out in a black 96-well plate. In each well of the test plate, 10 μL of compounds with different concentrations, 30 μL of buffer (25 mM Tris-HCl, 150 mM NaCl, 10% glycerol, pH 8.0), and 10 μL of sQC / gQC enzyme (final concentration 30 nM) were added. After incubation at room temperature for 20 minutes, 10 μL of PGP-1 enzyme (final concentration 100 nM) and 10 μL of fluorescent substrate H-Gln-AMC (final concentration 3 μM) were added, and the fluorescence change value of the system within 10 min was immediately detected (λex = 380 nm, λem = 460 nm).

[0193] In addition, to exclude the false positive of the final fluorescence result caused by the possible inhibition of PGP-1 by the compound, an activity test experiment of PGP-1 was carried out. The whole reaction system was 60 μL. In the test wells, 0.6 μL of compound (final concentration 100 μM), 39.4 μL of buffer (25 mM Tris-HCl, 150 mM NaCl, 10% glycerol, pH 8.0), 10 μL of PGP-1 (final concentration 0.05 μM), and 10 μL of substrate pGA (final concentration 1 μM) were added in sequence. After adding the substrate, the fluorescence change value of the system within 10 min was immediately detected (λex = 380 nm, λem = 460 nm).

[0194] Each experiment was carried out with three independent replicates, and the reaction wells without compound were used as the blank control group. The obtained dose-effect relationship was fitted with GraphPad to obtain the corresponding half-maximal inhibitory concentration (IC 50 ).

[0195] (3) Experimental results

[0196] Through the above experimental method, the inhibitory activity of the compounds of the present invention against sQC / gQC was tested. The half-maximal inhibitory concentration (IC 50 ) of the compounds against sQC / gQC is shown in Table 1.

[0197] Table 1. Inhibitory activity of the compounds of the present invention against sQC / gQC a

[0198]

[0199]

[0200] a****: represents good inhibitory activity (IC 50 <1 μM); ***: represents medium inhibitory activity (1 μM < IC 50 <100 μM); **: represents weak inhibitory activity (IC 50 > 100 μM).

[0201] The data in Table 1 show that the compounds of the present invention can effectively inhibit the activities of sQC / gQC. Among them, the inhibitory activities of compounds 6, 18, 21, 23, 24, 33, 54, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, and 72 against sQC and gQC are both less than 1 μM, which is comparable to the activity of the positive control compound PQ912. This indicates that these compounds can be used as glutaminyl cyclase inhibitors to prepare drugs for preventing and / or treating diseases related to glutaminyl cyclase.

[0202] Experimental Example 2. Therapeutic effect of compound 63 of the present invention on a Parkinson's animal model

[0203] (1) Experimental materials

[0204] The animals used were C57BL / 6J mice; the experimental conditions were a SPF-class mouse room, SPF maintenance feed, SPF-class animal drinking water, SPF corncob bedding, and 0.9% sodium chloride injection; the materials and instruments used included disposable sterile syringes with needles (1 mL / 20 ml), 1.5 ml microcentrifuge tubes, 10 mL round-bottom centrifuge tubes with lids, 50 ml centrifuge tubes, ZB-200 fatigue rotarod, 40 cm × 40 cm × 35 cm mouse spontaneous activity detection box, video tracking analysis system, electronic balance, sealing film, tin foil, timer, iron stand, string, ophthalmic forceps (10 cm, straight with hook, matte), ophthalmic forceps (10 cm, curved with hook, matte), hemostatic forceps (14 cm, straight full-tooth), experimental surgical scissors (12.5 cm, straight pointed), experimental surgical scissors (12.5 cm, curved pointed); the compound solvents included PEG 300 and DMSO; the compounds included MPTP (used to induce a Parkinson's disease mouse model), compound 63, and the positive drug Madopar (abbreviated as Ma).

[0205] (2) Experimental method

[0206] The in vivo efficacy of compound 63 in a MPTP-induced Parkinson's disease mouse model. The procedures and protocols of all animal experiments complied with the guidelines for the use and care of laboratory animals and relevant guidelines and policies. Rodent experiments were approved by the Animal Experiment Committee of West China Medical Center, Sichuan University. Animal experiments followed the guidelines established by the Animal Ethics Committee of Sichuan University. A total of 42 male C57BL / 6J mice (6 - 8 weeks old, 19 - 20 g) were purchased from Hunan SJA Laboratory Animal Co., Ltd. and were housed under standard conditions of seven per cage, including a 12-hour light / dark cycle, maintained at a temperature of 22 - 25 °C, and with free access to food and water. An adaptation period of one week was carried out before the start of the experiment.

[0207] Mice were divided into different groups: control group, MPTP-treated group, MPTP + 63 10 mg / kg group, MPTP + 63 25 mg / kg group, MPTP + 63 40 mg / kg group, and MPTP + positive drug Madopar 50 mg / kg group (as positive control), with 7 mice in each group. During the first 7 days, Parkinson's disease models were induced in mice by intraperitoneal injection of 30 mg / kg of MPTP at a fixed time every day (9:00 am). In the latter five groups; meanwhile, mice in the control group received saline injection. In the subsequent four groups, 63 or Madopar was intraperitoneally injected once a day for an additional 21 days, while the vehicle and MPTP groups continued to receive saline. All experimental procedures were conducted in a blinded manner. After the behavioral tests, the mice were sacrificed and their tissues were used for further analysis and processing.

[0208] Before the behavioral tests, mice first underwent a familiarization phase with the test room, equipment, and experimenters. Then, the rotarod and grip strength tests were performed according to the established protocol. For the rotarod test to evaluate global motor coordination and balance ability, mice were placed on a rotarod with a rotational speed of 25 rpm, and the time of the first fall was recorded, with a maximum execution time of 180 seconds. The rotarod test was conducted on days 0, 7, 14, and 21. The grip strength test was conducted on day 21. Mice were placed in the center of a rope and evaluated according to the following criteria: 0, dropped; 1, hung on the rope for more than 60 seconds; 2, escaped in 40 - 60 seconds; 3, escaped in 20 - 40 seconds; 4, escaped in 10 - 20 seconds; 5, escaped within 10 seconds. Each test was repeated three times.

[0209] The open field experiment used a rectangular box (40 cm × 40 cm × 35 cm) equipped with a tracking detector inside. Mice were acclimated to the experimental environment for 30 minutes and an additional 5 minutes before the test. Each mouse was placed on the central stage, and its movement and behavior were recorded by a video analysis system for 5 minutes. Parameters such as the total moving distance, the proportion of stationary and active time, and the distance covered in the central area were calculated from the recorded data.

[0210] (3) Experimental results

[0211] Before treatment, significant motor function impairment was observed in the MPTP-induced Parkinson's disease mouse model, especially in the rotarod test, where the latency time was significantly reduced compared to mice not receiving MPTP injection. In the 63 treatment group at a low dose (10 mg / kg), a trend towards improvement was observed compared to the MPTP group as the treatment time extended. In contrast, 63 treatment at doses of 25 and 40 mg / kg significantly improved MPTP-related motor deficits, similar to (even better than, see Figure 1)。The results of the grip strength test were consistent with those of the rotarod test, showing that 63 significantly improved the decline in motor ability observed in the MPTP group, indicating that 63 can enhance muscle strength and balance ability in Parkinson's disease mice. Figure 2 )。In addition, the open field test was used to evaluate the effects of 63 on the motor performance and exploratory ability of Parkinson's disease mice. Compared with the control group, the mice in the MPTP group showed higher immobile behavior in terms of exploratory ability and reduced movement in the central area. After treatment with 63, the exploratory behavior of the mice was significantly improved. Specifically, even at a dose of 10 mg / kg, 63 significantly increased the total distance traveled by the mice in the rectangular box, and this response was comparable to that achieved with the treatment of the positive drug levodopa at a dose of 50 mg / kg. Figure 3 )。The above experimental results indicate that the compound 63 of the present invention can effectively alleviate the motor deficits in the MPTP-induced Parkinson's disease mouse model.

[0212] In summary, the present invention provides a class of glutaminyl cyclase inhibitors represented by Formula I and their uses. The compounds of the present invention can effectively inhibit the activities of sQC / gQC and can be used as glutaminyl cyclase inhibitors to prepare drugs for preventing and / or treating diseases related to glutaminyl cyclase (such as Alzheimer's disease, Parkinson's syndrome, amyotrophic lateral sclerosis, tumors, cancers, immune diseases, etc.). The present invention provides more options for the development and application of small molecule inhibitors of glutaminyl cyclase and drugs for preventing and / or treating diseases related to glutaminyl cyclase (such as Alzheimer's disease, Parkinson's syndrome, amyotrophic lateral sclerosis, tumors, cancers, immune diseases, etc.).

Claims

1. A compound, its stereoisomer, its tautomer, its chiral isomer or its salt, characterized in that, The structure of the said compound is shown in Formula I: Wherein, X1 is selected from CH or N, X2 is selected from CH or N, X3 is selected from CH or N, and X4 is selected from CH or N; m is 0, 1, 2, 3 or 4; When m is 0, X1, X2, X3 and X4 are not simultaneously CH; Each R1 is independently selected from halogen, C 1-5 alkyl, C 1-5 alkoxy, COR 1a , nitro, hydroxy, NR 1b R 1c , L1L3R 1e , L1L2R 1d ; R 1a selected from C 1-5 alkyl; R 1b selected from hydrogen, C 1-5 alkyl; R 1c selected from hydrogen, C 1-5 alkyl; L1 is selected from O, S, NH, CO, CS, NHCO, NHCS, NHSO2, NHCONH, NHCSNH; L2 is selected from none or C 1-3 alkylene; L3 is selected from or C 1-3 alkylene; R 1e selected from C 1-5 alkyl, halo C 1-5 alkyl, COOR 1f ; R 1f selected from C 1-5 alkyl; R 1d selected from unsubstituted or 1-, 2- or 3-substituted groups of: 3- to 6-membered saturated cycloalkyl, 5- to 6-membered saturated heterocyclic group, 5- to 6-membered heteroaryl, phenyl; R x each independently selected from halogen, C x alkyl, halo C 1-5 alkyl, C 1-5 alkoxy, halo C 1-5 alkoxy, hydroxy, L4NR 1-5 R 1g R 1h 、C 2-4 alkenyl, C 2-4 alkynyl, COR i 、COOR i 、SO2R i ; L4 is selected from none or C 1-3 Alkylene; R 1g is selected from hydrogen, C 1-5 Alkyl; R 1h is selected from hydrogen, C 1-5 Alkyl; R i is selected from C 1-5 Alkyl, C 1-5 Alkyl substituted with alkoxy, C 1-5 Alkyl, the following groups which are unsubstituted or substituted with 1, 2 or 3 R y Substituted: 5-6 membered saturated heterocyclic group, 5-6 membered heteroaryl, phenyl, 3-6 membered saturated cycloalkyl; R y Each is independently selected from C 1-5 Alkyl, C 1-5 Alkoxy; R2 is selected from H, hydroxyl; R3 is selected from H, C 1-5 alkyl, COR 3a ; R 3a is selected from C 1-5 alkyl.

2. The compound according to claim 1, its stereoisomers, its tautomers, its chiral isomers or its salts, characterized in that, The structure of the said compound is shown in Formula II: Wherein, X1 is selected from CH or N; m is 0, 1, 2, 3 or 4; When m is 0, X1 is N; R1 is as described in claim 1; R2 is selected from H, hydroxyl; R3 is selected from H, C 1-3 alkyl, COR 3a ; R 3a is selected from C 1-3 alkyl.

3. The compound according to claim 2, its stereoisomer, its tautomer, its chiral isomer or its salt, characterized in that, The structure of the said compound is shown in Formula III: Preferably, the structure of the said compound is shown in Formula IV: Wherein, R1 is as described in claim 2.

4. The compound, stereoisomer, tautomer, chiral isomer or salt thereof according to claim 3, characterized in that, R1 is selected from halogen, C 1-3 alkyl, C 1-3 alkoxy, COR 1a , nitro, hydroxy, NR 1b R 1c , L1L3R 1e , L1L2R 1d ; R 1a selected from C 1-3 alkyl; R 1b selected from hydrogen, C 1-3 alkyl; R 1c selected from hydrogen, C 1-3 alkyl; L1 is selected from O, S, NH, CO, CS, NHCO, NHCS, NHSO2, NHCONH, NHCSNH; L2 is selected from none or C 1-2 an alkylene group; L3 is selected from or C 1-2 Alkylene; R 1e selected from C 1-3 alkyl, halo C 1-3 alkyl, COOR 1f ; R 1f selected from C 1-3 alkyl; R 1d selected from unsubstituted or 1- or 2-R x substituted groups: 3- to 6-membered saturated cycloalkyl, 5- to 6-membered saturated heterocyclic group, 5- to 6-membered heteroaryl, phenyl; R x each independently selected from halogen, C 1-3 alkyl, halo-C 1-3 alkyl, C 1-3 alkoxy, halo-C 1-3 alkoxy, hydroxy, L4NR 1g R 1h 、C 2-3 alkenyl, C 2-3 alkynyl, COR i 、COOR i 、SO2R i ; L4 is selected from none or C 1-2 alkylene; R 1g is selected from hydrogen, C 1-3 alkyl; R 1h is selected from hydrogen, C 1-3 alkyl; R i is selected from C 1-3 alkyl, C 1-3 alkyl substituted by C 1-3 alkoxy, unsubstituted or substituted by 1 or 2 R y substituted with the following groups: 5-6 membered saturated heterocyclic group, 5-6 membered heteroaryl, phenyl, 3-6 membered saturated cycloalkyl; R y each independently selected from C 1-3 alkyl, C 1-3 alkoxy.

5. The compound, stereoisomer, tautomer, chiral isomer or salt thereof according to claim 4, characterized in that, The structure of the said compound is shown in Formula V-1, Formula V-2, Formula V-3 or Formula V-4: Among them, R i is selected from C 1-3 alkyl, C 1-3 alkoxy-substituted C 1-3 alkyl, unsubstituted or substituted by 1 or 2 R y groups: 5-6 membered saturated heterocyclic group, 5-6 membered heteroaryl group, phenyl group, 3-6 membered saturated cycloalkyl group; R y is each independently selected from C 1-3 alkyl, C 1-3 alkoxy.

6. The compound according to claim 1, its stereoisomers, its tautomers, its chiral isomers or its salts, characterized in that, The said compound is one of the following compounds:

7. A pharmaceutical composition, characterized in that, It is a preparation prepared from the compound described in any one of claims 1-6, its stereoisomer, its tautomer, its chiral isomer or its salt as the active ingredient, plus pharmaceutically acceptable excipients.

8. Use of compound 5 or the compound described in any one of claims 1-6, its stereoisomer, its tautomer, its chiral isomer or its salt in the preparation of a glutamine cyclase inhibitor; 9. The use according to claim 8, characterized in that, The said glutamine cyclase is sQC and / or gQC.

10. The use according to claim 8 or 9, characterized in that, The said glutamine cyclase inhibitor is a drug for preventing and / or treating diseases related to glutamine cyclase; Preferably, the diseases related to glutamine cyclase are Alzheimer's disease, Parkinson's syndrome, amyotrophic lateral sclerosis, tumors, cancers, immune diseases, etc.; More preferably, the said tumor is a thyroid tumor, esophageal tumor, lung tumor, kidney tumor or breast tumor, the said cancer is lung cancer, gastric cancer, colorectal cancer, liver cancer, breast cancer, esophageal cancer, thyroid cancer, endometrial cancer, brain cancer or pancreatic cancer, and the said immune disease is rheumatoid arthritis, non-alcoholic hepatitis, cutaneous melanoma or lupus erythematosus syndrome.

Images