PARG inhibitors, methods of making and using the same

Abstract

The present application provides a compound I, and a stereoisomer, a tautomer, a geometric isomer, a deuterated compound or a pharmaceutically acceptable salt of the compound I, a structural formula of the compound I is as follows: -I; the compound I has PARG inhibitory activity, and the compound I, the stereoisomer, the tautomer, the geometric isomer, the deuterated compound or the pharmaceutically acceptable salt of the compound I can be applied to preparation of a PARG inhibitor.

Description

Technical Field

[0001] This invention patent relates to the field of medicinal chemistry technology, and in particular to a PARG inhibitor, its preparation method, and its application. Background Art Tumor cells heavily rely on DNA damage repair mechanisms. Single-strand breaks (SSBs) are the most common type of DNA damage, and their repair mechanisms are involved by PARG (Poly ADP-ribose glycohydrolase) and PARP (Poly(ADP-ribosyl)ation polymerase), as well as other proteins.

[0002] The ability of PARG (Kassab and Yu, 2019) to control PAR reversal, particularly its role as a major enzyme in removing PAR chains at DNA damage sites (Fisher et al., 2007; Brochu et al., 1994), can control damage responses in cancer cells. However, existing natural compounds such as tannins have limitations in their specificity and bioavailability for PARG. Therefore, the development of novel compounds with PARG inhibitory activity remains crucial. Summary of the Invention

[0003] This invention first provides a compound I, or its stereoisomers, tautomers, geometric isomers, deuterated compounds, or pharmaceutically acceptable salts, wherein the structural formula of compound I is shown below:

[0004] I in, Indicates a single bond or a double bond; X1, X2, and X3 are each independently selected from N or C; R 1 R 2 R 3 Each is independently selected from hydrogen, cyano, C1-6 alkyl, or R 1 Selected from hydrogen, cyano, C1-6 alkyl, R 2 R 3 It forms 3-6 membered cycloalkyl or 4-6 membered heterocyclic rings with the carbon atoms attached to it; Ring A is selected from a 5-membered heterocycle or 1-5 R groups. A1 Replaced 5-membered heterocyclic rings; each R A1 Each is independently selected from hydrogen, amino, C1-6 alkyl, and halogen-substituted C1-6 alkyl groups; R 4 Selected from hydrogen or B ring; R 5Selected from hydrogen, C1-6 alkyl, halogen-substituted C1-6 alkyl, -C0-4 alkylene-4-8 membered heterocycle, amino-substituted -C0-4 alkylene-4-8 membered heterocycle, R 5a Substituted -C0-4 alkylene-4-8 membered heterocycles, -C0-4 alkylene-3-8 membered cycloalkyl, amino-substituted -C0-4 alkylene-3-8 membered cycloalkyl, R 5a Substituted -C0-4 alkylene-3-8 membered cycloalkyl, benzene ring, amino-substituted benzene ring, R 5a Substituted benzene ring, R 5a Selected from C1-6 alkyl, hydroxy-substituted C1-6 alkyl, , cyano , , , ;R 6 Selected from oxygen, amino, or amino groups substituted with one or two C1-6 alkyl groups; or R 5 R 6 And the atoms connected to it constitute , , where R D1 Selected from hydrogen or B ring; Ring B is Where X4 is selected from N or C; n1 is selected from any integer from 1 to 4; R B5 Selected from hydrogen, C1-6 alkyl, halogen-substituted C1-6 alkyl, and hydroxyl-substituted C1-6 alkyl; R B6 Selected from hydrogen and C1-6 alkyl groups; R 7 R 8 They are independently selected from hydrogen and C1-6 alkyl groups, respectively.

[0005] Preferably, the structural formula of compound I is selected from the following structures: , , .

[0006] Preferably, R 1 Selected from C1-3 alkyl groups, R 2 R 3 The 4-8 membered heterocycle forms cyclopropane, cyclobutane, or cyclopentane with the carbon atom attached to it; the heterocycle is selected from... , , , , .

[0007] Preferably, R 5 Selected from hydrogen, C1-6 alkyl, halogen-substituted C1-6 alkyl, , , , , Where n2, n3, n4, n5, n6, and n7 are each independently selected from integers from 0 to 8; R 5a1 R 5a2 R 5a3 R 5a4 R 5a5 Each is independently selected from hydrogen, amino, C1-6 alkyl, hydroxyl-substituted C1-6 alkyl, , cyano , , , .

[0008] Preferably, ring A is selected from , , , , R A1 Selected from hydrogen, C1-3 alkyl, and halogen-substituted C1-3 alkyl.

[0009] Preferably, ring A is R A1 It is a C1-3 alkyl group substituted with halogen; the halogen is selected from F, Cl, Br, and I.

[0010] Preferably, R A1 Selected from methyl groups with 1-3 F substitutions.

[0011] Preferably, compound I is selected from one of the following compounds:

[0012]

[0013]

[0014]

[0015]

[0016]

[0017]

[0018]

[0019]

[0020]

[0021]

[0022] .

[0023] Preferably, the pharmaceutically acceptable salt is selected from the hydrochloride, hydrobromide, hydroiodide, perchlorate, sulfate, nitrate, phosphate, formate, acetate, propionate, glycolate, lactate, succinate, maleate, tartrate, malate, citrate, fumarate, gluconate, benzoate, mandelate, methanesulfonate, hydroxyethylsulfonate, benzenesulfonate, oxalate, palmitate, 2-naphthalenesulfonate, p-toluenesulfonate, cyclohexylaminosulfonate, salicylate, hexonate, trifluoroacetate, aluminum salt, calcium salt, chloroprocaine salt, choline salt, diethanolamine salt, ethylenediamine salt, lithium salt, magnesium salt, potassium salt, sodium salt, and zinc salt of compound I.

[0024] In a second aspect, the present invention provides a pharmaceutical composition comprising compound I as described in the first aspect of the present invention, or a stereoisomer, tautomer, geometric isomer, deuterated compound thereof, a pharmaceutically acceptable salt, and a pharmaceutically acceptable excipient.

[0025] In a third aspect of the invention, the invention provides the use of compound I, or a stereoisomer, tautomer, geometric isomer, deuterated compound, pharmaceutically acceptable salt, or pharmaceutical composition of the first aspect of the invention, in the preparation of a PARG inhibitor medicament.

[0026] In a fourth aspect of the invention, the invention provides the use of compound I, or a stereoisomer, tautomer, geometric isomer, deuterated compound, pharmaceutically acceptable salt, or pharmaceutical composition of the second aspect of the invention, in the preparation of an anticancer drug.

[0027] The beneficial effects of this invention are as follows: 1. The present invention provides a compound I, which has PARG inhibitory activity.

[0028] 2. This invention provides the application of this compound in the preparation of PARG inhibitors or anticancer drugs.

[0029] 3. The present invention provides a pharmaceutical composition for preparing PARG inhibitors or anticancer drugs. Detailed Implementation

[0030] The present invention will be further described below with reference to specific embodiments. However, the following embodiments are for illustrative purposes only and should not be considered as limiting the scope of the invention. Unless otherwise specified, specific conditions in the following embodiments are performed under conventional conditions or conditions recommended by the manufacturer. Unless otherwise specified, the methods used are conventional methods known in the art, and the consumables and reagents used are commercially available. Unless otherwise stated, the technical and scientific terms used herein have the same meaning as those familiar with the art. Furthermore, any methods or materials similar to or equivalent to those described herein may also be applied to the present invention.

[0031] The following preparation example illustrates the preparation of intermediate IM for the synthesis of compound TM. Preparation Example 1: Synthesis of Intermediate IM-1

[0032] Step 1: Synthesis of intermediate IM-1b 3-Fluoro-4-nitrobenzenesulfonyl chloride (10 g, 41.73 mmol) and 1-methylcyclopropylamine hydrochloride (4.49 g, 41.73 mmol) were added to a 250 ml round-bottom flask and dissolved in DCM (150 ml). TEA (12.67 g, 125.19 mmol) was added dropwise under ice bath conditions, and the reaction was slowly brought to room temperature. After 30 minutes, the mixture was extracted with EA using saturated sodium bicarbonate solution (100 ml). The organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated to give crude IM-1b (11.45 g). LCMS: m / z 273.25 [M+H] + . Step 2: Synthesis of intermediate IM-1c IM-1b (4 g, 14.29 mmol) was dissolved in DMF (40 ml), and 5-(difluoromethyl)-2-amino-1,3,4-thiadiazole (2.27 g, 15.00 mmol) and cesium carbonate (6.05 g, 18.58 mmol) were added. The reaction was carried out at 80 °C for 2 hours. After cooling to room temperature, 250 ml of water was added, and the mixture was extracted with EA. The organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated and purified by column chromatography to give IM-1c (2.79 g). LCMS: m / z 406.2 [M+H] + . Step 3: Synthesis of intermediate IM-1 IM-1c (2.7 g, 6.66 mmol) was dissolved in ethanol (40 ml), and water (40 ml) was added. Ammonium chloride (1.77 g, 33.3 mmol) and iron powder (1.86 g, 33.3 mmol) were added with stirring. The reaction mixture was heated to 80 °C. After 2 hours, the mixture was passed through diatomaceous earth while hot, washed with ethanol, and the filtrate was concentrated. Extraction was performed with DCM, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated and purified by column chromatography to obtain IM-1 (2.4 g). LCMS: m / z 376.2 [M+H] + . Preparation Example 2: Synthesis of Intermediate IM-2

[0033] IM-1 (2 g, 5.33 mmol) was dissolved in THF (20 ml), and N-bromosuccinimide (1.5 g, 8.00 mmol) was added. The mixture was reacted at room temperature for 2 hours, then water (200 ml) was added, and the mixture was extracted with EA. The organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated and purified by column chromatography to give IM-2 (1.8 g). LCMS: m / z 455.2 [M+H] + . Preparation Example 3: Synthesis of Intermediate IM-3

[0034] Step 1: Synthesis of intermediate IM-3b 4-Chloro-3-nitrobenzenesulfonyl chloride (11 g, 42.96 mmol) and 1-methylcyclopropylamine hydrochloride (5.08 g, 47.26 mmol) were added to a 500 ml round-bottom flask and dissolved in DCM (200 ml). TEA (13.04 mg, 128.88 mmol) was added dropwise under ice bath conditions, and the reaction was slowly brought to room temperature. After 30 minutes, the mixture was extracted with EA using 100 ml of saturated sodium bicarbonate solution. The organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated to give crude IM-3b (9.03 g, 72.3%). LCMS: m / z 289.13 [M+H] + . Step 2: Synthesis of intermediate IM-3c IM-3b (9 g, 30.96 mmol) was dissolved in DMF (100 ml), and TEA (7.83 g, 77.4 mmol) was added, followed by methylamine hydrochloride (2.19 g, 32.51 mmol). The mixture was heated to 130 °C and reacted for 6 hours. After cooling to room temperature, water (400 ml) was added, and the mixture was extracted with EA. The organic layer was washed with saturated sodium chloride and dried over sodium sulfate. Concentration yielded crude IM-3c (8.23 g, 93.1%). LCMS: m / z 285.2 [M+H] + . Step 3: Synthesis of intermediate IM-3d Intermediate IM-3c (7.5 g, 26.29 mmol) was dissolved in acetonitrile (100 ml), and NBS (5.15 g, 28.92 mmol) was added. The reaction mixture was reacted overnight at 80 °C. The next day, the reaction solution was cooled to room temperature and extracted with EA. The organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated to give crude IM-3d (9.05 g, 94.5%). LCMS: m / z 365.2 [M+H] + . Step 4: Synthesis of intermediate IM-3e IM-3d (8.86 g, 24.33 mmol) was dissolved in ethanol (80 ml), and water (80 ml) was added. Ammonium chloride (6.51 g, 121.65 mmol) and iron powder (6.79 g, 121.65 mmol) were added with stirring. The reaction mixture was heated to 80 °C. After 2 hours, the mixture was passed through diatomaceous earth while hot, washed with ethanol, and the filtrate was concentrated. Extraction was performed with DCM, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated and purified by column chromatography to give IM-3e (7.11 g, 87.4%). LCMS: m / z 335.2 [M+H] + . Step 5: Synthesis of intermediate IM-3f IM-3e (3 g, 8.98 mmol) was dissolved in acetonitrile (40 ml), and CDI (2.62 g, 16.16 mmol) was added. The mixture was reacted at 80 °C for 6 hours. Water (100 ml) was added to the reaction solution, and the mixture was extracted with EA. The organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated to give crude IM-3f (2.7 g, 83.5%). LCMS: m / z 361.2 [M+H] + . Step 6: Synthesis of intermediate IM-3 IM-3f (2.5 g, 6.94 mmol) and 2-bromo-5-(difluoromethyl)-1,3,4-thiadiazole (2.24 g, 10.41 mmol) were dissolved in 1,4-dioxane (30 ml). Potassium carbonate (3.84 g, 27.76 mmol) was added, followed by cuprous iodide (0.26 g, 1.39 mmol) and (1R,2R)-(-)-N,N'-dimethyl-1,2-cyclohexanediamine (0.39 g, 2.77 mmol). The mixture was purged with nitrogen three times and reacted overnight at 80 °C. Water (100 ml) was added to the reaction mixture, and the mixture was extracted with EA. The organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated to give crude IM-3 (2.2 g, 64.1%). LCMS: m / z 495.2 [M+H] + . The following examples illustrate the preparation of compound TM.

[0035] Example 1: Synthesis of TM-1

[0036] Step 1: Synthesis of TM-1a IM-1 (353 mg, 0.94 mmol) was dissolved in THF (10 ml), and N,N'-thiocarbonyldiimidazole (251 mg, 1.41 mmol) was added. The reaction mixture was reacted at 80 °C for 4 hours. The reaction solution was cooled to room temperature, and water (10 ml) was added. The mixture was extracted with EA, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated and purified by column chromatography to obtain TM-1a (355 mg). LCMS: m / z 418.1 [M+H] + . Step 2: Synthesis of TM-1b TM-1a (210 mg, 0.5 mmol) was dissolved in THF (5 ml), potassium carbonate (276 mg, 2 mmol) was added, followed by methyl iodide (191 mg, 1.35 mmol), and the reaction was carried out at 50 °C for 1 hour. The reaction solution was quenched with water (20 ml), extracted with EA, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated to obtain product TM-1b (212 mg). LCMS: m / z 431.2 [M+H] + . Step 3: Synthesis of TM-1c TM-1b (100 mg, 0.116 mmol) and 1-BOC-4-piperidincarbazide (50 mg, 0.2 mmol) were dissolved in pyridine and heated under reflux for 24 hours. The reaction mixture was neutralized with 1N hydrochloric acid, extracted with EA, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated to give product TM-1c (14 mg). LCMS: m / z 609.4 [M+H] + . Step 4: Synthesis of TM-1 TM-1c (5 mg) was dissolved in EA (2 ml), and ethyl acetate hydrochloride (2 ml) was added. The mixture was reacted at room temperature for 1 hour, filtered, and the solid was collected to give product TM-1 (2 mg). LCMS: m / z 509.2 [M+H] + . Example 2: Synthesis of TM-2

[0037] Step 1: Synthesis of TM-2a IM-2 (300.0 mg, 0.660 mmol) was dissolved in THF (10 ml), and N,N'-thiocarbonyldiimidazole (176.5 mg, 0.990 mmol) was added. The reaction mixture was reacted at 80 °C for 4 hours. The reaction solution was cooled to room temperature, and water (10 ml) was added. The mixture was extracted with EA, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated and purified by column chromatography to give TM-2a (322 mg). LCMS: m / z 497.2 [M+H] + .

[0038] Step 2: Synthesis of TM-2b IM-16a (315 mg, 0.634 mmol) was dissolved in THF (5 ml), potassium carbonate (350 mg, 2.54 mmol) was added, followed by methyl iodide (108 mg, 0.761 mmol), and the reaction was carried out at 50°C for 1 hour. The reaction solution was quenched with water (20 ml), extracted with EA, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated to obtain product TM-2b (324 mg). LCMS: m / z 513.2 [M+H] + .

[0039] Step 3: Synthesis of TM-2c TM-2b (100 mg, 0.195 mmol) and formylhydrazine (17.58 mg, 0.293 mmol) were dissolved in pyridine and heated under reflux for 24 hours. The reaction mixture was neutralized with 1N hydrochloric acid, extracted with EA, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated to give product TM-2c (64 mg). LCMS: m / z 505.4 [M+H] + .

[0040] Step 4: Synthesis of TM-2d Under nitrogen protection, cesium carbonate (112 mg, 0.344 mmol), Ruphos (11 mg, 0.024 mmol), and RuphosPdG3 (10 mg, 0.012 mmol) were added sequentially to a solution of TM-2c (57.5 mg, 0.114 mmol) and (2S,6S)-2,6-dimethylpiperazine-1-carboxylic acid tert-butyl ester (49.3 mg, 0.23 mmol) in 1,4-dioxane (2 mL). The reaction mixture was purged with nitrogen three times and stirred at 80 °C for 3 hours under nitrogen protection. After the reaction was complete, the mixture was cooled to room temperature, diluted with ethyl acetate, washed successively with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by preparative HPLC to give TM-2d (17.89 mg, 24.6%). LCMS: m / z 638.5 [M+H] + .

[0041] Step 5: Synthesis of TM-2 TM-2d (12 mg, 0.023 mmol) was dissolved in ethyl acetate (1 mL), and 4M ethyl acetate hydrochloride (1 mL) was added. The mixture was reacted at room temperature for 1 hour. The solid was collected by filtration to give TM-2 (9 mg). LCMS: m / z 538.5 [M+H] + .

[0042] Example 3: Synthesis of TM-3

[0043] Step 1: Synthesis of TM-3a Under nitrogen protection, cesium carbonate (112 mg, 0.344 mmol), Ruphos (11 mg, 0.024 mmol), and RuphosPdG3 (10 mg, 0.012 mmol) were added sequentially to a solution of IM-7 (50 mg, 0.101 mmol) and (2S,6S)-2,6-dimethylpiperazine-1-carboxylic acid tert-butyl ester (49.3 mg, 0.23 mmol) in 1,4-dioxane (2 mL). The reaction solution was purged with nitrogen three times and stirred at 80 °C for 3 hours under nitrogen protection. After the reaction was complete, the reaction solution was cooled to room temperature, diluted with ethyl acetate, washed successively with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by preparative HPLC to give TM-3a (23.06 mg, 0.101 mmol, 36.64%). LCMS: m / z 628.5 [M+H] + .

[0044] Step 2: Synthesis of TM-3 TM-3a (20 mg, 0.031 mmol) was dissolved in ethyl acetate (1 mL), and 4M ethyl acetate hydrochloride (1 mL) was added. The mixture was reacted at room temperature for 1 hour. The solid was collected by filtration to give TM-3 (12 mg). LCMS: m / z 528.5 [M+H] + .

[0045] Example 4: Synthesis of TM-4

[0046] Step 1: Synthesis of TM-4a IM-2 (1.5 g, 3.30 mmol) was dissolved in DCM (20 mL), and TEA (668 mg, 6.60 mmol) was added. Cyanogen bromide (0.6 g, 5.28 mmol) was added with stirring, and the mixture was reacted at room temperature for 12 hours. The solution was concentrated, and methanol (10 mL) and water (10 mL) were added. The reaction was stirred overnight at 50 °C, concentrated, and extracted with EA. The organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated and purified by column chromatography to obtain product TM-4a (1.1 g, 69%). LCMS: m / z 480.2 [M+H] + .

[0047] Step 2: Synthesis of TM-4b TM-4a (200 mg, 0.417 mmol) was dissolved in ethanol (10 mL), sodium bicarbonate (52 mg, 0.625 mmol) was added, followed by chloroacetaldehyde (39 mg, 0.5 mmol). The mixture was purged with nitrogen three times and refluxed overnight. The reaction was monitored for completion by LCMs. The concentrated reaction solution was purified by column chromatography to obtain TM-4b (115 mg, 54%). LCMS: m / z 502.1 [M+H] + .

[0048] Synthesis in steps three and four Replace IM-3 with TM-4b. The synthesis of TM-4 is similar to that of TM-3. TM-4 LCMS: m / z537.6[M+H] + .

[0049] Example 5: Synthesis of TM-5

[0050] Step 1: Synthesis of TM-5a TM-4a (200 mg, 0.417 mmol) was dissolved in DMF (10 mL). NaH (30 mg, 1.25 mmol) was added under ice bath conditions, and the mixture was stirred for 10 minutes. Then, methyl iodide (153 mg, 1.08 mmol) was added, and the mixture was reacted overnight at room temperature. The reaction mixture was diluted with water, extracted with EA, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated and purified by column chromatography to obtain product IM-76a (166 mg, 77%). LCMS: m / z 508.1 [M+H] + .

[0051] Synthesis from step two to step three Replacing TM-2c with TM-5a, the synthesis of TM-5 was similar to that of TM-2. TM-5 (15 mg) was obtained. LCMS: m / z 541.1 [M+H] + .

[0052] Example 6: Synthesis of TM-6

[0053] Step 1: Synthesis of TM-6b 4-Chloro-3-nitrobenzenesulfonyl chloride (11 g, 42.96 mmol) and 1-methylcyclopropylamine hydrochloride (5.08 g, 47.26 mmol) were added to a 250 ml round-bottom flask and dissolved in DCM (150 ml). TEA (17.91 mL, 128.88 mmol) was added dropwise under ice bath conditions, and the reaction was slowly brought to room temperature. After 30 minutes, the mixture was extracted with EA using saturated sodium bicarbonate solution (100 ml). The organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated to give crude TM-6b (9.03 g, 72%). LCMS: m / z 289.17 [M+H] + . Step 2: Synthesis of TM-6c TM-6b (2 g, 6.88 mmol) was dissolved in DMF (20 mL), and TEA (1.43 mL, 10.32 mmol) was added, followed by 1R,2R-N-BOC-cyclohexanediamine (1.47 g, 6.88 mmol). The reaction mixture was heated to 130 °C and reacted for 6 hours. After cooling the reaction mixture to room temperature, it was slowly poured into water, resulting in the precipitation of a yellow solid. The solid was filtered, and the filter cake was collected. The filtrate was extracted with EA, dried over anhydrous sodium sulfate, and concentrated. The filtrate was then combined with the filter cake to obtain a crude yellow solid, which was used directly in the next step. TM-6c (3.3 g). LCMS: m / z 467.38 [M+H] + .

[0054] Step 3: Synthesis of TM-6d TM-6c crude product (3.3 g) was dissolved in ethanol (40 mL), water (40 mL) was added, and ammonium chloride (1.88 g, 35.2 mmol) was added with stirring. Iron powder (1.97 g, 35.2 mmol) was then added, and the reaction was carried out at 70 °C for 2 hours. The mixture was filtered through diatomaceous earth while hot, washed with methanol, concentrated, extracted with EA, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated and purified by column chromatography to obtain TM-6d (2.3 g). LCMS: m / z 439.39 [M+H] + .

[0055] Step 4: Synthesis of TM-6e TM-6d (2.3 g, 5.24 mmol) was dissolved in acetonitrile (25 mL), and CDI (1.70 g, 10.48 mmol) was added. The reaction was carried out at 80 °C for 2 hours. The reaction solution was diluted with water, extracted with EA, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated and purified by column chromatography to obtain TM-6e (2 g). LCMS: m / z 463.44 [M+H] + .

[0056] Step 5: Synthesis of TM-6f TM-6e (2.4 g, 5.17 mmol) was dissolved in 1,4-dioxane (30 mL), followed by 2-bromo-5-(difluoromethyl)-1,3,4-thiadiazole (1.11 g, 5.17 mmol) and potassium carbonate (2.86 g, 20.68 mmol). Then, cuprous iodide (0.2 g, 1.03 mmol) and (1R,2R)-(-)-N,N'-dimethyl-1,2-cyclohexanediamine (0.29 g, 2.07 mmol) were added. The mixture was purged three times with nitrogen and reacted at 80 °C for 4 hours. The reaction solution was diluted with water, extracted with EA, and the organic layer was washed with saturated sodium chloride and dried over sodium sulfate. The solution was concentrated and purified by column chromatography to obtain TM-6f (1.8 g, 60%). LCMS: m / z 599.29 [M+H] + .

[0057] Step Six: Synthesis of TM-6 TM-6f (500 mg) was dissolved in 1,4-dioxane (5 mL), and 4M ethyl acetate hydrochloride solution (5 mL) was added. The mixture was reacted at room temperature for 2 hours, and then concentrated to obtain TM-6 (450 mg). LCMS: m / z 499.36 [M+H] + . 1 H NMR (600 MHz, DMSO) δ 8.82 (s, 1H), 8.35 (s, 3H), 8.24 (s, 1H), 7.81 (s, 1H), 7.66 (t, J =53.2 Hz, 1H), 4.47 (m, J = 11.6, 4.3 Hz, 1H), 3.94 (s, 1H), 2.21 (d, J = 12.2 Hz,2H), 1.99 – 1.91 (m, 1H), 1.80 (m, J = 12.4 Hz, 2H), 1.62 (s, 1H), 1.42 (s, 2H), 1.13 (s, 3H), 0.68 – 0.59 (m, 2H), 0.44 – 0.35 (m, 2H).

[0058] Example 7-15: Synthesis of TM-7-15 The preparation methods for TM-7 to TM-15 are similar to the synthesis of the final product TM-6. The raw materials used in the preparation of TM-6c from TM-6b are... Replace with , , , , , , , , Compounds TM-7 to TM-15 were obtained sequentially, and the specific structures of TM-7 to TM-15 are shown in Table 1.

[0059] Table 1. Compound Numbers and Structures

[0060] Example 16: Synthesis of TM-16

[0061] TM-13 (18.88 mg, 0.039 mmol) was dissolved in DCM (1 mL), TEA (5.97 mg, 0.59 mmol) was added, followed by cyanogen bromide (5.42 mg, 0.51 mmol). The mixture was reacted at room temperature for 4 hours. The reaction solution was concentrated, and TM-16 (8.90 mg) was purified by preparative HPLC. LCMS: m / z 510.1 [M+H] + . 1 H NMR (600 MHz, DMSO) δ8.82 (d, J = 1.8 Hz, 1H), 8.21 (s, 1H), 7.88 (d, J = 8.5 Hz, 1H), 7.79 (dd, J= 8.4, 1.9 Hz, 1H), 7.64 (t, J = 53.2 Hz, 1H), 4.58 (m, J = 16.2, 11.9, 4.3Hz, 1H), 3.77 (t, J = 11.9 Hz, 1H), 3.59 (dd, J = 12.3, 4.4 Hz, 1H), 3.41 (d,J = 12.6 Hz, 1H), 3.19 (m, J = 12.6, 3.3 Hz, 1H), 2.32 (m, J = 12.6, 4.3 Hz, 1H), 2.01 (m, J = 9.8 Hz, 1H), 1.90 – 1.79 (m, 2H), 1.11 (s, 3H), 0.62 (q, J= 4.5 Hz, 2H), 0.43 – 0.36 (m, 2H).

[0062] Examples 17-21: Synthesis of TM-17-21 The preparation methods of TM-17 to TM-21 are similar to the synthesis of the final product TM-16. TM-13 is used as a starting material and reacted with acryloyl chloride; TM-12 is used as a starting material and reacted with cyanogen bromide; TM-12 is used as a starting material and reacted with acryloyl chloride; TM-8 is used as a starting material and reacted with cyanogen bromide, and compounds TM-17 to TM-21 are obtained sequentially. The specific structures of TM-17 to TM-21 are shown in Table 2.

[0063] Table 2. Compound Numbers and Structures

[0064] Examples 22-27: Synthesis of TM-22-27 The preparation method of TM-22 to TM-27 is similar to the synthesis of the final product TM-6, except that in step two, 1R,2R-N-BOC-cyclohexanediamine is replaced with... , , , , , This leads to TM-22 to TM-27, and the specific structures of TM-22 to TM-27 are shown in Table 3.

[0065] Table 3. Compound Numbers and Structures

[0066] Example 28: Synthesis of TM-28

[0067] The preparation method of TM-28 is similar to that of the final product TM-16, except that the cyano group is replaced with 2-chloroethanesulfonyl chloride, which is then reacted with TM-6 to obtain TM-28. LCMS: m / z (M+H)+ = 589.24. 1 H NMR (600 MHz, DMSO) δ 8.81(s, 1H), 8.15 (s, 1H), 7.85 – 7.47 (m, 4H), 7.42 – 7.34 (m, 1H), 7.26 – 7.23(m, 1H), 6.38 (s, 1H), 5.66 (m, 1H), 4.13 (s, 1H), 3.71 (s, 1H), 1.96 (m, J =15.0 Hz, 2H), 1.77 (dd, J= 36.2, 10.6 Hz, 2H), 1.50 (s, 1H), 1.40 – 1.33 (m, 2H), 1.08 (s, 3H), 0.66 – 0.58 (m, 2H), 0.38 – 0.32 (m, 2H).

[0068] Examples 29-41: Synthesis of TM-29-41 The preparation method of TM-29 to TM-41 is similar to the synthesis of the final product TM-28. The following reactions were conducted using TM-6 as a starting material with acryloyl chloride, TM-6 with 2-chloroethanesulfonyl chloride, TM-14 with 2-chloroethanesulfonyl chloride, TM-14 with cyanogen bromide, TM-15 with cyanogen bromide, TM-15 with 2-chloroethanesulfonyl chloride, TM-12 with 2-chloroethanesulfonyl chloride, TM-13 with 2-chloroethanesulfonyl chloride, TM-24 with 2-chloroethanesulfonyl chloride, TM-25 with 2-chloroethanesulfonyl chloride, TM-26 with 2-chloroethanesulfonyl chloride, TM-27 with 2-chloroethanesulfonyl chloride, and TM-14 with ethylsulfonyl chloride, successively yielding TM-29 to TM-41. The specific structures of TM-29 to TM-41 are shown in Table 4.

[0069] Table 4. Compound Numbers and Structures

[0070] Example 42: Synthesis of TM-42

[0071] Step 1: Synthesis of TM-42b 2-Fluoro-4-bromo-6-nitroaniline (10 g, 42.55 mmol) was dissolved in anhydrous acetonitrile (100 mL) at room temperature, and copper dichloride (17.16 g, 127.65 mmol) was added. Then, tert-butyl nitrite (10.65 mL, 85.10 mmol) was slowly added dropwise. After reacting at room temperature for 1 hour, the reaction was monitored by TLC to indicate completion. The reaction solution was quenched with water (100 mL), extracted three times with ethyl acetate, and the organic phase was washed with saturated sodium chloride, dried over anhydrous sodium sulfate, concentrated under reduced pressure to remove the organic phase, and purified by column chromatography using petroleum ether / ethyl acetate (50 / 1) as eluent to obtain a yellow solid TM-42b (8.8 g, 81.28%).

[0072] Step 2: Synthesis of TM-42c At room temperature, TM-42b (2.1 g, 8.25 mmol) was dissolved in DMF (15 mL), and (S)-1-tert-butoxycarbonyl-3-aminopyrrolidine (3.07 g, 16.5 mmol) and potassium carbonate (2.28 g, 16.5 mmol) were added. The mixture was stirred overnight at 70 °C. The reaction solution was diluted with water, extracted three times with ethyl acetate, and the organic phase was washed with saturated sodium chloride, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to remove the organic phase. TM-42c (2.8 g, orange oil, 70% purity, 58.75%) was obtained using petroleum ether / ethyl acetate (2 / 1 to 1 / 1) as eluent. LCMS: m / z 406.81 [M+H] + . Step 3: Synthesis of TM-42d TM-42c (2.8 g, 6.93 mmol) was dissolved in ethanol (30 mL) and water (30 mL). Ammonium chloride (1.48 g, 27.72 mmol) and iron powder (1.55 g, 27.72 mmol) were added under stirring, and the mixture was reacted at 80 °C for 2 hours. The reaction solution was filtered through diatomaceous earth, washed with methanol, concentrated, and extracted with DCM. The organic phase was concentrated and purified by column chromatography using petroleum ether / ethyl acetate (1 / 1) as eluent to obtain a black solid TM-42d (1.7 g, 65.58%). LCMS: m / z 335.96 [M+H] + . Step 4: Synthesis of TM-42e TM-42d (2.5 g, 6.68 mmol) was dissolved in anhydrous acetonitrile (20 mL), and CDI (2.17 g, 13.36 mmol) was added. The reaction was carried out overnight at 70 °C. The organic phase was removed by concentration under reduced pressure, and TM-42e (1.0 g, 37.40%) was obtained as a yellow oil using petroleum ether / ethyl acetate (1 / 1) as the eluent. LCMS: m / z 399.98 [M+H] + . Step 5: Synthesis of TM-42f TM-42e (1.0 g, 2.20 mmol) was dissolved in 1,4-dioxane solution (10 mL), and 2-bromo-5-(difluoromethyl)-1,3,4-thiadiazole (0.81 g, 3.75 mmol), potassium carbonate (1.38 g, 10 mmol), cuprous iodide (0.095 g, 0.50 mmol), and (1R,2R)-(-)-N,N'-dimethyl-1,2-cyclohexanediamine (0.14 g, 1.0 mmol) were added. After purging with nitrogen three times, the reaction was carried out overnight at 80 °C. The reaction solution was diluted with water, extracted three times with ethyl acetate, and the organic phase was washed with saturated sodium chloride, dried over anhydrous sodium sulfate, concentrated under reduced pressure to remove the organic phase, and eluented with petroleum ether / ethyl acetate (1 / 1) to obtain a yellow oily TM-42f. LCMS: m / z 479.84 [M+H] + . Step Six: Synthesis of TM-42g TM-42f (337 mg, 0.63 mmol) was dissolved in 1,4-dioxane (5 mL), and DIEA (310 μL, 0.69 mmol), 4,5-bis(diphenylphosphine-9,9-dimethyloxanthracene) (36 mg, 0.063 mmol), and tris(dibenzylacetone)dipalladium (58 mg, 0.063 mmol) were added. The mixture was purged with nitrogen three times, and benzyl mercaptan (81 μL, 0.69 mmol) was added dropwise over 30 minutes with stirring at 100 °C. After the addition was complete, the reaction was monitored for completion 6 hours later. The reaction solution was cooled to room temperature and extracted three times with ethyl acetate. The organic phase was washed with saturated sodium chloride, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to remove the organic phase. The solution was eluent with petroleum ether / ethyl acetate (1 / 1) to obtain a yellow oily TM-42g (300 mg, 82.35%). LCMS: m / z 521.99 [M+H] + . Step 7: Synthesis of TM-42h TM-42 g (100 mg, 0.17 mmol) was dissolved in acetonitrile (2 mL). Acetic acid (48 μL, 0.85 mmol) and water (45 μL, 2.55 mmol) were added under ice bath conditions, followed by NCS (45.40 mg, 0.34 mmol). The mixture was slowly heated to room temperature and reacted for 6 hours. Two drops of ice water were added dropwise under ice bath conditions, followed by extraction with ethyl acetate and anhydrous sodium sulfate to obtain crude TM-42 h.

[0073] Step 8: Synthesis of TM-42i 1-Methylcyclopropylamine hydrochloride (22.59 mg, 0.21 mmol) was dissolved in anhydrous DCM, and DIEA (54 mg, 0.42 mmol) was added dropwise. TM-42h (60 mg, 0.11 mmol) DCM solution was then added dropwise under ice bath conditions. After 30 minutes, the reaction was monitored by LCMS to indicate completion. The reaction solution was purified by column chromatography to obtain a pale yellow solid TM-42i (30 mg, 47.06%). LCMS: m / z 586.89 [M+H] + . Step Nine: Synthesis of TM-42j TM-42i (25 mg, 0.042 mmol) was dissolved in ethyl acetate (2 mL), and 4M HCl (in 1 mL of EA) was added. After reacting at room temperature for 2 hours, the solution was concentrated to give a yellow solid product TM-42j (14 mg, 67.48%). LCMS: m / z 489.09 [M+H] + . Step 10: Synthesis of TM-42 TM-42j (14 mg, 0.029 mmol) was dissolved in dichloromethane (3 mL), TEA (8.80 mg, 0.087 mmol) was added, followed by 2-chloroethanesulfonyl chloride (7.09 mg, 0.044 mmol), and the reaction was carried out at room temperature for 1 hour. The organic phase was removed by concentration under reduced pressure, and TM-42 (3 mg, 18.1%) was obtained by eluing with petroleum ether / ethyl acetate (2 / 1 to 1 / 1). LCMS: m / z 578.9 [M+H] + . 1 H NMR (600 MHz, DMSO) δ 8.73 (d, J = 1.6 Hz, 1H), 8.37 (s, 1H), 7.75 – 7.55 (m, 2H), 6.97 (dd, J = 16.5, 10.0 Hz, 1H), 6.29 – 6.14 (m, 2H), 5.30 (m, J = 8.4 Hz, 1H), 3.68 (m, J = 10.1 Hz, 1H), 3.63 – 3.52 (m, 2H), 3.41 –3.35 (m, 1H), 2.54 – 2.51 (m, 1H), 2.39 (dd, J = 7.8, 4.6 Hz, 1H), 1.14 (s,3H), 0.64 (m, J= 4.5 Hz, 2H), 0.45 – 0.40 (m, 2H).

[0074] Example 43: Synthesis of TM-43 The preparation method of TM-43 is similar to that of the final product TM-42, except that S-3-aminopyrrolidine is replaced with cis-1,4-cyclohexanediamine to obtain TM-43. The specific structure is shown in Table 5.

[0075] Table 5. Compound Numbers and Structures

[0076] Example 44: Synthesis of TM-44

[0077] Step 1: Synthesis of TM-44b 3-Fluoro-4-nitrobenzenesulfonyl chloride (10 g, 41.73 mmol) and 1-methylcyclopropylamine hydrochloride (4.49 g, 41.73 mmol) were added to a 250 mL round-bottom flask and dissolved in DCM (150 mL). TEA (17.40 mL, 125.19 mmol) was added dropwise under ice bath conditions, and the reaction was slowly brought to room temperature. After 30 minutes, the mixture was extracted three times with ethyl acetate using 100 mL of saturated sodium bicarbonate solution. The organic phase was washed with saturated sodium chloride and dried over anhydrous sodium sulfate. The solution was concentrated to give a crude, pale yellow solid, TM-44b (11.98 g). LCMS: m / z 273.35 [M+H] + . Step 2: Synthesis of TM-44c TM-44b (4 g, 14.29 mmol) and 5-(difluoromethyl)-2-amino-1,3,4-thiadiazole (2.27 g, 15.00 mmol) were dissolved in DMF (40 mL), and cesium carbonate (6.05 g, 18.58 mmol) was added. The mixture was reacted at 80 °C for 3 h. The mixture was extracted three times with ethyl acetate, and the organic phase was washed with saturated sodium chloride and dried over anhydrous sodium sulfate. The solution was concentrated and purified by column chromatography using petroleum ether / ethyl acetate (6 / 1 to 1 / 1) as eluent to give an orange solid TM-44c (2.79 g, 48.15%). LCMS: m / z 406.16 [M+H] + . Step 3: Synthesis of TM-44d TM-44c (2.7 g, 6.66 mmol) was dissolved in ethanol (30 mL) and water (30 mL). Ammonium chloride (1.78 g, 33.3 mmol) and iron powder (1.86 g, 33.3 mmol) were added under stirring, and the mixture was reacted at 80 °C for 2 hours. The reaction solution was filtered through diatomaceous earth, washed with methanol, concentrated, and extracted with DCM. The organic phase was concentrated and purified by column chromatography using petroleum ether / ethyl acetate (1 / 1) as eluent to obtain a yellow solid TM-44d (2.0 g, 76.2%). LCMS: m / z 374.23 [M+H] + . Step 4: Synthesis of TM-44e TM-44d (1.80 g, 4.79 mmol) was dissolved in anhydrous acetonitrile (20 mL), and CDI (1.06 g, 6.68 mmol) was added. The reaction was carried out overnight at 70 °C. The organic phase was removed by concentration under reduced pressure, and solid TM-44e (1.6 g, 83.13%) was obtained by eluing with petroleum ether / ethyl acetate (1 / 1). LCMS: m / z 399.98 [M+H] + . Step 5: Synthesis of TM-44f TM-44e (0.5 g, 1.25 mmol) was dissolved in DCM-free (10 mL), and 3-nitrophenylboronic acid (250.40 mg, 1.5 mmol) and copper acetate (227 mg, 1.25 mmol) and pyridine (148.93 mg, 1.88 mmol) were added. The mixture was reacted overnight at room temperature under air. The reaction solution was diluted with DCM, washed once with saturated sodium bicarbonate solution, dried, and concentrated. The solid TM-44f (105 mg, 16.13 mmol) was obtained by eluing with petroleum ether / ethyl acetate (3 / 1). LCMS: m / z 522.97 [M+H] + . Step Six: Synthesis of TM-44g TM-44f (105 mg, 0.20 mmol) was dissolved in ethanol (10 mL) and water (10 mL). Ammonium chloride (42.8 mg, 0.80 mmol) and iron powder (44.6 mg, 0.80 mmol) were added under stirring, and the mixture was reacted at 80 °C for 2 hours. The reaction solution was filtered through diatomaceous earth, washed with methanol, concentrated, and extracted with DCM. The organic phase was concentrated and purified by column chromatography using petroleum ether / ethyl acetate (1 / 1) as eluent to obtain a yellow solid TM-44g (80 mg, 80.8%). LCMS: m / z 493.00 [M+H] + . 1H NMR (600 MHz, DMSO) δ 8.87 (d, J = 1.8 Hz, 1H), 8.23 ​​(s, 1H), 7.79 (dd, J = 8.4, 1.8Hz, 1H), 7.65 (t, J = 53.2 Hz, 2H), 7.30 (d, J = 8.4 Hz, 1H), 7.26 (t, J = 7.9 Hz, 1H), 6.77 (t, J = 2.1 Hz, 1H), 6.74 – 6.69 (m, 2H), 5.53 (s, 2H), 1.12 (s, 3H), 0.63 (q, J = 4.5 Hz, 2H), 0.42 – 0.36 (m, 2H).

[0078] Step 7: Synthesis of TM-44 TM-44 g (30 mg, 0.061 mmol) was dissolved in DCM (5 mL), TEA (12.35 mg, 0.12 mmol) was added, followed by 2-chloroethanesulfonyl chloride (12.93 mg, 0.079 mmol), and the mixture was reacted at room temperature for 1 hour. The reaction solution was concentrated, purified by preparative liquid chromatography, and lyophilized to give a white solid TM-44 (1.1 mg, 3.06%). LCMS: m / z 580.98 [M+H] + . 1 HNMR (600 MHz, DMSO) δ 8.88 (d, J = 1.8 Hz, 1H), 8.37 – 8.27 (m, 2H), 8.24 (s,1H), 7.80 (dd, J = 8.4, 1.9 Hz, 1H), 7.65 (t, J = 53.2 Hz, 2H), 7.46 (t, J = 8.0Hz, 1H), 7.33 – 7.26 (m, 2H), 7.21 – 7.12 (m, 2H), 6.79 – 6.71 (m, 1H), 6.06(d, J = 17.0 Hz, 1H), 5.92 (d, J = 10.3 Hz, 1H), 1.13 (s, 3H), 0.64 (d, J= 2.1 Hz, 2H), 0.42 – 0.37 (m, 2H).

[0079] Example 45: Synthesis of TM-45

[0080] Replace 1-methylcyclopropylamine hydrochloride with 1-amino-1-cyclopropylcyanocyanate; the remaining raw materials, reagents, and preparation methods are the same as for TM-31. LCMS: m / z 572.12 [M+H] + . Example 46: Synthesis of TM-46

[0081] Replace 1-methylcyclopropylamine hydrochloride with 3-methyloxetane-3-amine hydrochloride; the remaining raw materials, reagents, and preparation methods are the same as those for TM-31. LCMS: m / z 577.11 [M+H] + . Example 47: Synthesis of TM-47

[0082] Step 1: Synthesis of TM-47b Ethyl 5-bromo-1H-indazole-3-carboxylate (3 g, 11.15 mmol) was dissolved in ethanol (20 mL) in a 100 mL round-bottom flask, and hydrazine hydrate (1.74 g, 27.88 mmol) was added. The reaction was carried out overnight at 90 °C. After cooling to room temperature, the mixture was filtered, washed with ethanol, and the solid was collected to give a white solid powder TM-47b (2.40 g).

[0083] Step 2: Synthesis of TM-47c TM-47b (2.9 g, 11.37 mmol) was dissolved in DCM (20 mL), and TEA (3.45 g, 34.11 mmol) was added. Difluoroacetic anhydride (2.37 g, 13.64 mmol) was added dropwise under ice bath conditions. The reaction was allowed to proceed for 2 hours, quenched with water, and extracted with DCM. The organic phase was washed with saturated sodium chloride and dried over anhydrous sodium sulfate. The concentrate yielded a yellow oil, TM-47c (2.5 g, 66.01%). LCMS: m / z 332.93 [M+H] + . Step 3: Synthesis of TM-47d TM-47c (2.0 g, 6.00 mmol) was dissolved in toluene (15 mL), and phosphorus pentasulfide (2.67 g, 12.00 mmol) was added. The reaction mixture was reacted at 130 °C for 2 hours. The reaction solution was concentrated, diluted with water, extracted with ethyl acetate, and the organic phase was washed with saturated sodium chloride and dried over anhydrous sodium sulfate. Purification by column chromatography yielded TM-47d (1.6 g, 80.47%). LCMS: m / z 332.31 [M+H] + . Steps four through nine: Synthesis of TM-47 Replace TM-42f with TM-47d; the remaining raw materials, reagents, and preparation methods are the same as for TM-42. LCMS: m / z 545.12 [M+H] + . Example 48: Synthesis of TM-48

[0084] Step 1: Synthesis of TM-48b (S)-1-BOC-pyrrolidine-3-carboxylic acid (5 g, 23.23 mmol) was dissolved in DCM (20 mL). 1-Ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (5.34 g, 27.88 mmol) was added under ice bath conditions. After stirring for 5 minutes, 1-hydroxybenzotriazole (3.77 g, 27.88 mmol) was added, followed by stirring for 5 minutes. Then, dimethylhydroxylamine hydrochloride (3.40 g, 34.84 mmol) and TEA (8.23 g, 11.30 mmol) were added, and the reaction was carried out at room temperature for 2 hours. The reaction solution was diluted with water, extracted with DCM, and the organic phase was washed with saturated sodium chloride and dried over anhydrous sodium sulfate. Purification by column chromatography yielded a colorless oil, TM-48b (5.3 g, 88.33%). LCMS: m / z 259.57 [M+H] + . Step 2: Synthesis of TM-48c 1,4-Dibromo-2-fluorobenzene (10 g, 39.39 mmol) was dissolved in anhydrous THF (100 mL) under nitrogen protection. Butyllithium (24.60 mL, 261.16 mmol) was added dropwise at -78 °C. After reacting at this low temperature for 1 hour, a THF solution of TM-48b (9.16 g, 35.45 mmol) was added dropwise, and the reaction was continued at room temperature for 3 hours. The reaction solution was quenched dropwise with an aqueous solution of saturated ammonium chloride, extracted with ethyl acetate, and the organic phase was washed with saturated sodium chloride and dried over anhydrous sodium sulfate to obtain crude TM-48c (10 g). LCMS: m / z 372.98 [M+H] + . Step 3: Synthesis of TM-48d Crude TM-48c (10 g) was dissolved in THF (10 mL), and hydrazine hydrate (2.45 mL, 40.30 mmol) was added. The mixture was reacted overnight at 100 °C. The reaction solution was diluted with water, extracted with ethyl acetate, and the organic phase was washed with saturated sodium chloride, dried over anhydrous sodium sulfate, and purified by column chromatography to give a yellow solid TM-48d (2.6 g, 26.41%). LCMS: m / z 366.12 [M+H] + . Steps four through nine: Synthesis of TM-48 Replace TM-42e with TM-48d; the remaining raw materials, reagents, and preparation methods are the same as for TM-42. LCMS: m / z 545.13 [M+H] + . 1 H NMR (600 MHz, DMSO) δ 8.95 – 8.89 (m, 1H), 8.44 (s, 1H), 8.30 (dd, J = 8.4, 0.8 Hz, 1H), 7.86 (dd, J = 8.4, 1.5 Hz, 1H), 7.60 (t, J = 53.1Hz, 2H), 6.95 (dd, J = 16.5, 10.0 Hz, 1H), 6.20 – 6.13 (m, 2H), 4.07 (p, J =7.7 Hz, 1H), 3.86 (dd, J = 10.2, 7.7 Hz, 1H), 3.58 – 3.47 (m, 2H), 3.43 –3.36 (m, 1H), 2.53 (dd, J = 7.3, 4.8 Hz, 1H), 2.26 (dq, J = 12.4, 8.0 Hz, 1H), 1.09 (s, 3H), 0.62 (q, J = 4.4 Hz, 2H), 0.44 – 0.38 (m, 2H).

[0085] The following test examples illustrate the beneficial effects of the compounds of the present invention.

[0086] The positive compound used in this invention is PDD00017273, and its structure is as follows:

[0087] PDD00017273 Test Example 1: Determination of the inhibitory activity of the PARG inhibitor compound of the present invention on the proliferation of PARG-sensitive cell lines. PARG-sensitive cell lines NCI-H460 and MDA-MB-436 were collected and seeded at 300 cells / well in 96-well white plates (180 μL). After 24 hours of adhesion, 20 μL of the compound was added (starting at a final concentration of 20 μM, with 8 concentration gradients of 5-fold dilution). After culturing at 37°C in a 5% CO2 incubator for 7 days, 100 μL of the original culture medium was aspirated from each well, and 30 μL of CTG detection reagent was added in the dark. After shaking for 3-5 min in the dark and incubating for 10 min, the cells were detected using a microplate reader. Cell viability was calculated using the following formula, and cell survival curves were fitted.

[0088] Cell viability (%) = (Number of cells in pores containing the PARG inhibitor compound of this invention - Number of cells in pores containing pure culture medium) / (Number of cells in pores containing DMSO - Number of cells in pores containing pure culture medium) * 100%.

[0089] Table 6. Inhibitory activity of the PARG inhibitor compounds of the present invention against NCI-H460 cells.

[0090] In the table, IC 50 >20000nM is represented by "-".

[0091] Table 7. Inhibitory activity of the PARG inhibitor compounds of the present invention against MDA-MB-436 cells.

[0092] Cell viability assay results showed that the compound TM-1-48 obtained in the examples of the present invention had varying degrees of inhibitory activity against PARG-sensitive cell lines.

[0093] Test Example 2: Determination of the activity of the invented compound on PARG luciferase PARG was co-incubated with an ADP-ribose substrate (BPS 82138) containing a fluorescent group, where the fluorophore was quenched by the presence of ribose. Under the action of PARG, the fluorescent group was released, and fluorescence was detectable at λ=502 nm (excitation wavelength λmax=385 nm). The fluorescence intensity was directly proportional to the PARG enzyme activity. The specific operation is as follows: 1. Experimental Reagents: Assay Buffer: 50 mM Tris, pH 7.4, 0.1 mg / mL BSA, 3 mM EDTA, 0.4 mM EGTA, 1 mM DTT, 0.01% Tween 20, 50 mM KCl. PARG enzyme (BPS 101726), luciferin PARG substrate (BPS 82138), PARG inhibitor compound prepared in the embodiments of this invention, DMSO (solvent). II. Experimental Procedures 1) Dilute the PARG enzyme to 1 ng / μL with Assay Buffer, mix well and place on ice for later use.

[0094] 2) Grouping and Sampling: To eliminate fluorescence interference from the compounds themselves, a fluorescence interference group was set up. This group contained only compounds of different concentrations, diluted with Assay Buffer. A DMSO control well was also included: a buffer containing 1% DMSO.

[0095] The official test group settings are as follows: Positive Control Well: 5 μL PARG enzyme + 2.5 μL Buffer (containing 5% DMSO) + 5 μL substrate.

[0096] Inhibitor Control well: 5 μL PARG enzyme + 2.5 μL serially diluted PARG inhibitor prepared in this invention (concentration 5 times the final concentration, DMSO final concentration 1%) + 5 μL substrate Blank wells: 5 μL Buffer (PARG-free) + 2.5 μL Buffer (containing 5% DMSO) + 5 μL Substrate 3) Incubate at room temperature for 15 minutes to allow the PARG enzyme and PARG inhibitor compounds to fully interact.

[0097] 4) Substrate-initiated reaction: Dilute the fluorescein PARG substrate (1 mM) 800 times with Assay Buffer, add 5 μL to each well to make the final reaction volume 12.5 μL, and incubate at room temperature in the dark for 1 hour.

[0098] 5) Fluorescence detection: The fluorescence intensity of each well was read using a fluorescence microplate reader at an excitation wavelength of 385 nm and an emission wavelength of 502 nm.

[0099] Part Three: Data Processing and Result Analysis 1) If there is no fluorescence interference, the enzyme activity inhibition rate = 1 - (fluorescence intensity of PARG inhibitor well (Inhibitor Control well) at 502 nm - fluorescence intensity of Blank well at 502 nm) / (fluorescence intensity of Positive Control well at 502 nm - fluorescence intensity of Blank well at 502 nm) * 100% 2) If the compound itself causes interference, the test value minus the interference value is the correction value; Enzyme activity inhibition rate = 1 - (Corrected value of fluorescence intensity at 502 nm for PARG inhibitor wells - Fluorescence intensity at 502 nm for Blank wells - Fluorescence intensity at 502 nm for DMSO wells) / (Fluorescence intensity at 502 nm for Positive Control wells - Fluorescence intensity at 502 nm for Blank wells - Fluorescence intensity at 502 nm for DMSO wells) * 100% 3) Inhibition rate and IC 50 Analysis: By fitting dose-response curves using relative enzyme activity data of a series of inhibitor concentrations, the half-maximal inhibitory concentration (IC50) of the compound on PARG was calculated. 50 ).

[0100] Table 8. Inhibitory activity of the compounds of the present invention against PARG luciferase

[0101] The results of PARG luciferase activity assay showed that the compound TM-1-48 obtained in the examples of the present invention had varying degrees of inhibitory activity against PARG enzyme.

[0102] Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Those skilled in the art can make changes, modifications, substitutions and variations to the above embodiments within the scope of the present invention, all of which should be included within the protection scope of the present invention.

Claims

1. A compound I, or its stereoisomers, tautomers, geometric isomers, deuterated compounds, or pharmaceutically acceptable salts, characterized in that, The structural formula of compound I is shown below: I in, Indicates a single bond or a double bond; X1, X2, and X3 are each independently selected from N or C; R 1 R 2 R 3 Each is independently selected from hydrogen, cyano, C1-6 alkyl, or R 1 Selected from hydrogen, cyano, C1-6 alkyl, R 2 R 3 It forms 3-6 membered cycloalkyl or 4-6 membered heterocyclic rings with the carbon atoms attached to it; Ring A is selected from a 5-membered heterocycle or 1-5 R groups. A1 Replaced 5-membered heterocyclic rings; each R A1 Each is independently selected from hydrogen, amino, C1-6 alkyl, and halogen-substituted C1-6 alkyl groups; R 4 Selected from hydrogen or B ring; R 5 Selected from hydrogen, C1-6 alkyl, halogen-substituted C1-6 alkyl, -C0-4 alkylene-4-8 membered heterocycle, amino-substituted -C0-4 alkylene-4-8 membered heterocycle, R 5a Substituted -C0-4 alkylene-4-8 membered heterocycles, -C0-4 alkylene-3-8 membered cycloalkyl, amino-substituted -C0-4 alkylene-3-8 membered cycloalkyl, R 5a Substituted -C0-4 alkylene-3-8 membered cycloalkyl, benzene ring, amino-substituted benzene ring, R 5a Substituted benzene ring, R 5a Selected from C1-6 alkyl, hydroxy-substituted C1-6 alkyl, , cyano , , , , ;R 6 Selected from oxygen, amino, or amino groups substituted with one or two C1-6 alkyl groups; or R 5 R 6 And the atoms connected to it constitute , , where R D1 Selected from hydrogen or B ring; Ring B is Where X4 is selected from N or C; n1 is selected from any integer from 1 to 4; R B5 Selected from hydrogen, C1-6 alkyl, halogen-substituted C1-6 alkyl, and hydroxyl-substituted C1-6 alkyl; R B6 Selected from hydrogen and C1-6 alkyl groups; R 7 R 8 They are independently selected from hydrogen and C1-6 alkyl groups, respectively.

2. The compound I according to claim 1, or its stereoisomers, tautomers, geometric isomers, deuterated compounds, or pharmaceutically acceptable salts, characterized in that, The structural formula of compound I is selected from the following structures: 、 、 。 3. Compound I according to claim 1, or its stereoisomers, tautomers, geometric isomers, deuterated compounds, or pharmaceutically acceptable salts, characterized in that, R 1 Selected from C1-3 alkyl groups, R 2 R 3 The 4-8 membered heterocycle forms cyclopropane, cyclobutane, or cyclopentane with the carbon atom attached to it; the heterocycle is selected from... , , , , .

4. Compound I according to claim 1, or its stereoisomers, tautomers, geometric isomers, deuterated compounds, or pharmaceutically acceptable salts, characterized in that, R 5 Selected from hydrogen, C1-6 alkyl, halogen-substituted C1-6 alkyl, , , , , Where n2, n3, n4, n5, n6, and n7 are each independently selected from integers from 0 to 8; R 5a1 R 5a2 R 5a3 R 5a4 R 5a5 Each is independently selected from hydrogen, amino, C1-6 alkyl, hydroxyl-substituted C1-6 alkyl, , cyano , , , .

5. Compound I according to claim 1, or its stereoisomers, tautomers, geometric isomers, deuterated compounds, or pharmaceutically acceptable salts, characterized in that, R A1 It is a C1-3 alkyl group substituted with halogen; the halogen is selected from F, Cl, Br, and I.

6. Compound I according to claim 5, or its stereoisomers, tautomers, geometric isomers, deuterated compounds, or pharmaceutically acceptable salts, characterized in that, Ring A is selected from , , , , R A1 Selected from hydrogen, C1-3 alkyl, and halogen-substituted C1-3 alkyl.

7. Compound I according to claim 1, or its stereoisomers, tautomers, geometric isomers, deuterated compounds, or pharmaceutically acceptable salts, characterized in that, Compound I is selected from one of the following compounds: 。 8. A pharmaceutical composition comprising compound I as described in any one of claims 1-7, or a stereoisomer, tautomer, geometric isomer, deuterated compound, pharmaceutically acceptable salt, and pharmaceutically acceptable excipient.

9. The use of compound I according to any one of claims 1-7, or its stereoisomers, tautomers, geometric isomers, deuterated compounds, pharmaceutically acceptable salts, or the pharmaceutical composition according to claim 8, in the preparation of a PARG inhibitor medicament.

10. The use of compound I according to any one of claims 1-7, or its stereoisomers, tautomers, geometric isomers, deuterated compounds, pharmaceutically acceptable salts, or the pharmaceutical composition according to claim 8, in the preparation of an anticancer drug.

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