Pyrazole derivative compounds and uses thereof

Abstract

The application discloses a pyrazole derivative compound and application thereof, and belongs to the field of pharmacy. The pyrazole derivative compound can improve the activity of microglia cells, and has a wide application prospect in the preparation of drugs for preventing and treating Alzheimer's disease.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to a class of pyrazole-derived compounds and their uses. Background Technology

[0002] Dementia has become a global health challenge. According to the World Health Organization's 2022 Dementia Research Blueprint, an estimated 55.2 million people worldwide are affected. Prevalence varies by region among people aged 60 and over. While the incidence of dementia is generally rising, it is declining in some high-income countries. By 2030, the estimated number of people with dementia is projected to surge to 78 million. Furthermore, the global economic burden associated with healthcare, social services, and informal care for people with dementia is expected to exceed US$2.8 trillion. This situation will have profound impacts on individuals, families, and societies. Alzheimer's disease (AD), the leading form of dementia, exhibits similar epidemiological trends and is an urgent and escalating challenge facing the world.

[0003] Microglia are immune cells unique to the central nervous system, accounting for 5% of the glial cells in the cerebral cortex. They play a crucial role in maintaining brain homeostasis and are considered the first line of defense for the central nervous system. Resting microglia monitor the functional state of neuronal synapses through direct contact and respond efficiently to changes in the surrounding environment. When acute inflammation, infection, or trauma occurs in the brain, microglia are rapidly activated, altering their morphology, accelerating proliferation, and promoting inflammatory responses. Full-length TREM2 is a 230-amino acid, 40 kDa membrane glycoprotein constitutively expressed in microglia. Upon activation by ligands, it regulates phagocytosis, synaptic pruning, and energy metabolism.

[0004] This invention provides a class of pyrazole-derived compounds, pharmaceutical compositions thereof, and uses. The pyrazole-derived compounds of this invention can induce TREM2 to bind to DAP12, thereby promoting SYK phosphorylation and enhancing the phagocytic activity of microglia. These compounds have broad application prospects in the preparation of drugs for the prevention and treatment of Alzheimer's disease. Summary of the Invention

[0005] Technical problem solved: This invention provides a class of pyrazole-derived compounds that can promote the binding of TREM2 and DAP12, enhance microglial cell viability, and can be used to prepare drugs for the treatment of Alzheimer's disease.

[0006] Technical solution: A class of compounds as shown in Formula I and Formula II, or their pharmaceutically acceptable salts, deuterated compounds, solvates, racemic mixtures, enantiomers, diastereomers, and tautomers, as shown in Formula I and Formula II below.

[0007]

[0008]

[0009] in,

[0010] X is selected from -O-, -S-, or -NH-;

[0011] R8 is selected from hydrogen or C1-C6 alkyl;

[0012] Y and Z are each independently selected from N or C(R). 10 );

[0013] Q is selected from N or C(R). 11 );

[0014] M is selected from -O- or -C(R) 12 ,R 13 )-;

[0015] R1, R2, R3, R4, R5, R6, R7, R9, R 10 R 11 R 12 and R 13 Each is independently selected from hydrogen, halogen, trifluoromethyl, difluoromethyl, C1-C6 alkyl, C3-C8 cycloalkyl, cyano, nitro, methoxy, hydroxy or amino.

[0016] Preferably, X is selected from -O- or -NH-;

[0017] R1, R2, R3, R4, R5, R6, R7, R9 and R 10 Each is independently selected from hydrogen, halogen, trifluoromethyl, difluoromethyl, C1-C3 alkyl, C3-C6 cycloalkyl, cyano or methoxy;

[0018] R8, R 11 R 12 and R 13 Each is independently selected from hydrogen or C1-C3 alkyl groups;

[0019] Y and Z are each independently selected from -N- or -C(R) 10 )-.

[0020] Preferably, R1, R2, R3, R4, R5, R6, R7, and R9 are each independently selected from hydrogen, fluorine, chlorine, trifluoromethyl, difluoromethyl, methyl, cyano, or methoxy.

[0021] R8 is selected from methyl;

[0022] R 10 R 11 R 12 and R13 Each is independently selected from hydrogen.

[0023] Preferably, the compound is:

[0024]

[0025] Compound S1, as shown above;

[0026]

[0027] Compound S2, as shown above;

[0028]

[0029] Compound S3, as shown above;

[0030]

[0031] Compound S4, as shown above.

[0032] The use of the compound as described in any of the above, or a pharmaceutically acceptable salt, deuterated compound, solvate, racemic mixture, enantiomer, diastereomer, and tautomer thereof, in the preparation of an AD therapeutic agent.

[0033] The use of the compound as described in any of the above, or a pharmaceutically acceptable salt, deuterated compound, solvate, racemic mixture, enantiomer, diastereomer, and tautomer thereof, in the preparation of a drug for enhancing microglial cell activity.

[0034] A pharmaceutical composition comprising any one of the compounds described above or a pharmaceutically acceptable salt thereof, a deuterated compound, a solvate, a racemic mixture, an enantiomer, a diastereomer, a tautomer, and a pharmaceutically acceptable carrier.

[0035] Beneficial effects:

[0036] This invention relates to a class of pyrazole-derived compounds characterized by their ability to promote the binding of TREM2 to DAP12, which can be used to prepare drugs for the treatment of Alzheimer's disease (AD). Detailed Implementation

[0037] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0038] Example 1: Synthesis of compound S1

[0039] Synthesis route:

[0040]

[0041] Synthesis process:

[0042] first step:

[0043] 1,7-Naphthoglycoside-8(7H)-1 (4.5 g, 30.79 mmol, 1 eq) was dissolved in DMF (45 mL) and NBS (5.5 g, 30.79 mmol, 1 eq) was added. The reaction mixture was stirred at room temperature for 3 hours. After the reaction was complete, the mixture was filtered, the filter cake was washed with H2O, and dried under vacuum to give Al (5.7 g, yield 82.66%) as a yellow solid.

[0044] Step Two:

[0045] A1 (5.7 g, 25.33 mmol, 1 eq) was dissolved in THF (53 mL), and K2CO3 (7 g, 50.66 mmol, 2 eq) and MeI (7.2 g, 50.66 mmol, 2 eq) were added. The reaction mixture was stirred overnight at 40 °C. After the reaction was complete, it was diluted with H2O (300 mL), extracted with EA (100 mL × 3), washed with saturated brine, dried over anhydrous Na2SO4, filtered and concentrated to give A2 (3.5 g, yield 58.06%) as a yellow solid.

[0046] Step 3:

[0047] A2 (3.64 g, 15.23 mmol, 1 eq) was dissolved in 1,4-Dioxane (60 mL) and H2O (6 mL), and Pd(dppf)Cl2 (2.2 g, 3.05 mmol, 0.2 eq), (4-chloro-2-fluorophenyl)boronic acid (4.5 g, 25.89 mmol, 1.7 eq), and K2CO3 (6.3 g, 45.69 mmol, 3 eq) were added. The reaction mixture was stirred at 80 °C for 1 hour. After the reaction was complete, the reaction mixture was concentrated, diluted with H2O (200 mL), extracted with EA (60 mL × 3), washed with saturated brine, dried over anhydrous Na2SO4, filtered, concentrated, and purified by pre-hplc (mobile phase: water and acetonitrile) to give A3 (2.49 g, yield 56.77%) as a yellow solid.

[0048] Step 4:

[0049] A3 (2.49 g, 8.62 mmol, 1 eq) was dissolved in DCM (40 mL), and UHP (2.4 g, 25.86 mmol, 3 eq) was added. The mixture was stirred at room temperature for 5 min. Then TFFA (7.2 g, 34.48 mmol, 4 eq) was added, and the mixture was stirred at room temperature for 30 min. After the reaction was complete, the mixture was diluted with H2O (200 mL), extracted with DCM (60 mL × 3), washed with saturated brine, dried over anhydrous Na2SO4, filtered, and concentrated to give A4 (1.6 g, yield 61.06%) as a yellow solid.

[0050] Step 5:

[0051] A4 (1.6 g, 5.28 mmol, 1 eq) was dissolved in DCM (30 mL), and (COCl)2 (871 mg, 6.86 mmol, 1.3 eq) was added. The mixture was stirred at room temperature for 30 min. After the reaction was complete, the mixture was diluted with H2O (100 mL), extracted with DCM (60 mL × 3), dried over anhydrous Na2SO4, filtered, concentrated, and purified by silica gel column chromatography (PE / EA) to give A5 (500 mg, yield 29.41%) as a yellow solid.

[0052] Step 6:

[0053] A5 (150 mg, 0.46 mmol, 1 eq) was dissolved in toluene (5 mL), and 2-(1-methyl-1H-pyrazole-4-yl)-ethanol (58 mg, 0.46 mmol, 1 eq), Pd(OAc)2 (15 mg, 0.037 mmol, 0.08 eq), BINAP (30 mg, 0.048 mmol, 0.1 eq) and Cs2CO3 (450 mg, 1.38 mmol, 3 eq) were added, followed by nitrogen purging. The reaction mixture was stirred overnight at 100 °C. After the reaction was complete, the mixture was concentrated, diluted with H2O (50 mL), extracted with EA (20 mL × 3), washed with saturated brine, dried over anhydrous Na2SO4, and purified by pre-hplc (mobile phase: water and acetonitrile) to give S1 (45 mg, yield 23.74%) as a white solid.

[0054] ESI-MS: 413.0 [M+H]+;

[0055] 1 H NMR (400MHz, DMSO-d6): δ7.63-7.67(m,4H),7.52-7.46(m,2H),7.35(s,1H),7.16(d, J=9.2Hz,1H),4.51(t,J=6.8Hz,2H),3.78(s,3H),3.57(s,3H),2.90(t,J=6.8Hz,2H).

[0056] Example 2: Synthesis of compound S2

[0057] Synthesis route:

[0058]

[0059] Synthesis process:

[0060] first step:

[0061] 1,7-Naphthoglycoside-8(7H)-1 (4.5 g, 30.79 mmol, 1 eq) was dissolved in DMF (45 mL) and NBS (5.5 g, 30.79 mmol, 1 eq) was added. The reaction mixture was stirred at room temperature for 3 hours. After the reaction was complete, the mixture was filtered, the filter cake was washed with H2O, and dried under vacuum to give Al (5.7 g, yield 82.66%) as a yellow solid.

[0062] Step Two:

[0063] A1 (5.7 g, 25.33 mmol, 1 eq) was dissolved in THF (53 mL), and K2CO3 (7 g, 50.66 mmol, 2 eq) and MeI (7.2 g, 50.66 mmol, 2 eq) were added. The reaction mixture was stirred overnight at 40 °C. After the reaction was complete, it was diluted with H2O (300 mL), extracted with EA (100 mL × 3), washed with saturated brine, dried over anhydrous Na2SO4, filtered and concentrated to give A2 (3.5 g, yield 58.06%) as a yellow solid.

[0064] Step 3:

[0065] A2 (3.64 g, 15.23 mmol, 1 eq) was dissolved in 1,4-Dioxane (60 mL) and H2O (6 mL), and Pd(dppf)Cl2 (2.2 g, 3.05 mmol, 0.2 eq), (4-chloro-2-fluorophenyl)boronic acid (4.5 g, 25.89 mmol, 1.7 eq), and K2CO3 (6.3 g, 45.69 mmol, 3 eq) were added. The reaction mixture was stirred at 80 °C for 1 hour. After the reaction was complete, the reaction mixture was concentrated, diluted with H2O (200 mL), extracted with EA (60 mL × 3), washed with saturated brine, dried over anhydrous Na2SO4, filtered, concentrated, and purified by pre-hplc (mobile phase: water and acetonitrile) to give A3 (2.49 g, yield 56.77%) as a yellow solid.

[0066] Step 4:

[0067] A3 (2.49 g, 8.62 mmol, 1 eq) was dissolved in DCM (40 mL), and UHP (2.4 g, 25.86 mmol, 3 eq) was added. The mixture was stirred at room temperature for 5 min. Then TFFA (7.2 g, 34.48 mmol, 4 eq) was added, and the mixture was stirred at room temperature for 30 min. After the reaction was complete, the mixture was diluted with H2O (200 mL), extracted with DCM (60 mL × 3), washed with saturated brine, dried over anhydrous Na2SO4, filtered, and concentrated to give A4 (1.6 g, yield 61.06%) as a yellow solid.

[0068] Step 5:

[0069] A4 (1.6 g, 5.28 mmol, 1 eq) was dissolved in DCM (30 mL), and (COCl)2 (871 mg, 6.86 mmol, 1.3 eq) was added. The mixture was stirred at room temperature for 30 min. After the reaction was complete, the mixture was diluted with H2O (100 mL), extracted with DCM (60 mL × 3), dried over anhydrous Na2SO4, filtered, concentrated, and purified by silica gel column chromatography (PE / EA) to give A5 (500 mg, yield 29.41%) as a yellow solid.

[0070] Step 6:

[0071] A4 (250 mg, 0.78 mmol, 1 eq) was dissolved in dimethyl sulfoxide (3 mL), and 2-(1-methyl-1h-pyrazol-4-yl)ethylenediamine (146 mg, 1.16 mmol, 1.48 eq) and DIEA (501 mg, 3.88 mmol, 5 eq) were added. The reaction mixture was stirred overnight at 130 °C. The solution was diluted with H2O (30 mL), extracted with EA (15 mL × 3), washed with brine, dried over anhydrous Na2SO4, filtered, concentrated, and purified by pre-hplc (mobile phase: water and acetonitrile) to give S3 (48.15 mg, yield 15.10%) as a yellow solid.

[0072] ESI-MS: 412.3 [M+H]+;

[0073] 1 H NMR (400MHz, DMSO-d6): δ7.60-7.55(m,2H),7.48-7.41(m,2H),7.33(s,1H),7.31(s,1H),7.2 5-7.18(m,2H),6.84(d,J=8.8Hz,1H),3.771(s,3H),3.54-3.49(m,5H),2.69(t,J=7.2Hz,2H).

[0074] Example 3: Synthesis of compound S3

[0075] Synthesis route:

[0076]

[0077] Synthesis process:

[0078] first step:

[0079] 2-Amino-6-methoxypyridine (13.0 g, 90.00 mmol, 2.25 eq) was dissolved in trimethyl orthoformate (55 mL) and stirred at 75 °C for 2 h. Michaelis acid (5.0 g, 40.00 mmol, 1 eq) was added and the mixture was stirred at 75 °C for another 30 min. After the reaction was completed, the mixture was cooled to room temperature, filtered, and the filter cake was washed with EA to obtain Al (9 g, yield 35%) as a white solid.

[0080] Step Two:

[0081] A1 (9.0 g, 32.37 mmol, 1 eq) was dissolved in diphenyl ether (15 mL), heated to 150 °C, stirred for 3 h, and after complete conversion was monitored by TLC, the reaction solution was cooled to room temperature. After the addition of petroleum ether, a large amount of solid precipitated out. The solution was filtered to obtain pure A2 (3.9 g, yield 75%), which was a white solid.

[0082] Step 3:

[0083] A2 (1 g, 5.68 mmol, 1 eq) was dissolved in POCl3 (10 mL), heated to 100 °C, and stirred at 100 °C for 1 h. The conversion was monitored by TLC to ensure complete conversion. After the reaction solution cooled to room temperature, it was slowly poured into ice water (50 mL) and stirred continuously until no significant exothermic reaction occurred. Then, EA (20 mL x 3) was added for extraction. The mixture was dried over anhydrous sodium sulfate, filtered, and evaporated to dryness to obtain a relatively pure crude product A3 (1.1 g, 85% yield), which was a yellow solid.

[0084] Step 4:

[0085] A3 (1.1 g, 5.67 mmol, 1 eq), 4-chloro-2-fluorophenylboronic acid pinacol ester (1.08 g, 6.23 mmol, 1.1 eq), Pd(dppf)Cl2 (0.23 g, 1.13 mmol, 0.05 eq), and Na2CO3 (1.2 g, 10.73 mmol, 2 eq) were dissolved in 1,4-dioxane (10 mL) and H2O (2.5 mL). After purging with nitrogen three times, the mixture was refluxed at 110 °C for 5 h under nitrogen protection. The conversion was monitored by TLC until complete. 50 mL of water was added, and the mixture was extracted with EA (20 mL x 2). The solution was dried over anhydrous sodium sulfate, filtered, and evaporated to dryness. The solution was then subjected to silica gel column chromatography (PE / EA) to give A4 (1.2 g, 45% yield) as a yellow solid.

[0086] Step 5:

[0087] A4 (1.2 g, 41.67 mmol, 1 eq) was dissolved in an aqueous solution of HBr (10 mL), stirred at 100 °C for 4 h, and after complete conversion by TLC monitoring, it was directly concentrated and evaporated to dryness to obtain a relatively pure crude product A5 (1 g, yield 90%), which was a yellow solid.

[0088] Step 6:

[0089] A5 (500.0 mg, 1.82 mmol, 1 eq) was dissolved in THF (5 mL), and (Tf)₂O (1.1 g, 3.64 mmol, 2 eq) and DIEA (0.47 g, 3.64 mmol, 2 eq) were added under ice bath conditions. The mixture was stirred at room temperature for 3 h, and the conversion was monitored by TLC until complete. Water (30 mL) was added to the reaction solution, and the mixture was extracted with EA (20 mL x 2). The solution was dried over anhydrous sodium sulfate, filtered, concentrated, and then purified by silica gel column chromatography (PE / EA) to obtain A6 (190 mg, yield 26%) as a pale yellow solid.

[0090] Step 7:

[0091] A6 (190.0 mg, 0.47 mmol, 1 eq), 2-(1-methyl-1H-pyrazol-4-yl)morpholine (93 mg, 0.56 mmol, 1.2 eq), and K2CO3 (130 mg, 0.94 mmol, 2 eq) were dissolved in DMF (2 mL) and stirred at room temperature for 1 h. After the reaction was completed, water (50 mL) was added to the reaction solution, and the mixture was extracted with EA (20 mL x 2). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated, and purified by prep-TLC (mobile phase: water and acetonitrile) to give S3 (70 mg, yield 40%) as a light brown solid.

[0092] ESI-MS: 424.3 [M+H]+;

[0093] 1 H NMR (400MHz, DMSO-d6) δ8.78(s,1H),7.68-7.64(m,3H),7.54-7.45(m,3H),7.33(d,J=8,1H),7.18(d,J=4,1 H),4.57-4.48(m,2H),4.37(d,J=1H),3.99(d,J=12,1H),3.79(s,3H),3.65(t,J=20,1H),3.18-3.04(m,2H).

[0094] Example 4: Synthesis of compound S4

[0095] Synthesis route:

[0096]

[0097] Synthesis process:

[0098] first step:

[0099] Add 12.4 g of 4-amino-2-methoxypyridine (1 eq, 0.1 mol) and 125 mL of acetonitrile to a 500 mL single-necked flask and cool in an ice bath. Add NBS (17.8 g, 1 eq, 0.1 mol) in portions and react for 2 h. The reaction mixture was allowed to react completely by TLC. Add the reaction solution to 200 mL of saturated saline solution and extract the product by EA (100 mL x 3). Combine the organic phases, dry and concentrate, and purify by silica gel column chromatography (PE / EA) to obtain product A1 (16 g, 90% yield), which is a white solid.

[0100] Step Two:

[0101] Triethyl orthoformate (80 mL) and Michaelis-Menten acid (25.2 g, 2 eq, 0.157 mol) were added to a 250 mL single-necked flask. The mixture was heated to 100 °C and reacted for 1.5 h. After cooling to room temperature, A1 (16 g, 1 eq, 0.078 mol) was added, and the mixture was heated to 90 °C and reacted for 2 h. The reaction was completed by TLC. The reaction solution was cooled to room temperature, and a large amount of solid precipitated. The mixture was filtered, and the filter cake was washed twice with ethanol and dried to obtain A2 (12 g, 80% yield), which was a white solid.

[0102] Step 3:

[0103] Add 55 mL of diphenyl ether and 12 g (1 eq, 33.6 mmol) of A2 to a 100 mL single-necked flask. Heat to 180 °C and react for 2 h. The reaction is complete by TLC. Cool to room temperature, and a large amount of solid precipitates. Filter the mixture, wash the filter cake twice with EA, and dry to obtain A3 (7 g, 75% yield), which is a yellow solid.

[0104] Step 4:

[0105] Add A3 (8g, 1eq, 31.3mmol), methanol (160mL), ammonium formate (20g, 10eq, 0.313mol), and Pd / C (1.6g) to a 250mL single-necked flask. Heat to 60℃ and react for 2 hours. Monitor the reaction by TLC until complete. Cool to room temperature, filter the reaction solution through diatomaceous earth, concentrate the filtrate to dryness, and purify by silica gel column chromatography with PE / EA to obtain 5.5g of crude product. Pulverize the crude product with 100mL (PE:EA = 1:1) to obtain A4 (4.8g, yield 56%), which is a yellow solid.

[0106] Step 5:

[0107] Add A4 (2 g, 1 eq, 11.3 mmol) to a 100 mL single-necked flask, then add acetonitrile (20 mL), phosphorus oxychloride (8.6 g, 5 eq, 56.5 mmol), and DIPEA (4.37 g, 3 eq, 33.9 mmol). Heat to 70 °C and react for 2 h. Monitor the reaction by TLC until complete. Concentrate the reaction solution to dryness, add 100 mL of water, adjust the pH to 5-6 with saturated sodium carbonate solution, and extract the product with EA (50 mL x 2). Combine the organic phases, dry and concentrate, and purify by silica gel column chromatography (PE / EA) to obtain A5 (1.5 g, 65% yield) as a white solid.

[0108] Step 6:

[0109] Add A5 (1.5 g, 1 eq, 7.7 mmol), dioxane (30 mL), 2N sodium carbonate solution (6 mL), and 2-fluoro-4-chlorophenylboronic acid (1.34 g, 1 eq, 7.7 mmol) to a 100 mL single-necked flask. Add Pd(dppf)Cl2 (200 mg) under nitrogen protection. Heat the mixture to 80 °C for 2 h under nitrogen protection. TLC showed that the starting material reacted completely. Concentrate the reaction solution to dryness and purify by column chromatography to obtain A6 (1.8 g, 60% yield), which is a white solid.

[0110] Step 7:

[0111] Add A6 (1.8 g, 1 eq, 6.25 mmol) to a 50 mL single-necked flask, add 48% hydrobromic acid (20 mL), heat to 100 °C and react for 2 h, cool to room temperature, filter, and dry to obtain A7 (1.5 g, 70% yield), which is a white solid.

[0112] Step 8:

[0113] Add A7 (500 mg, 1 eq, 1.82 mmol), DCM (10 mL), TEA (460 mg, 2.5 eq, 4.56 mmol), and bis(trifluoromethanesulfonylaniline) (975 mg, 1.5 eq, 2.74 mmol) to a 25 mL single-necked flask. React overnight for 16 h. Monitor the reaction of the starting materials by TLC until complete. Concentrate the reaction solution to dryness and purify PE / EA by silica gel column chromatography to obtain A8 (350 mg).

[0114] Step 9:

[0115] Add A8 (300 mg, 1 eq, 0.74 mmol), THF (10 mL), DIPEA (150 mg, 1.5 eq, 1.1 mmol), and 2-(1-methyl-1H-pyrazol-4-yl)morpholine (150 mg, 1.2 eq, 0.88 mmol) to a 10 mL single-necked flask. Heat at 60 °C for 2 days under nitrogen protection. TLC and HPLC showed that the reactant had reacted to about 40%. Concentrate the reaction solution to dryness and purify PE / EA by silica gel column chromatography to obtain S4 (65 mg, yield 15%), which is a white solid.

[0116] ESI-MS: 424.3 [M+H]+;

[0117] 1 H NMR(400MHz,Chloroform-d)δ8.94-8.93(d,1H),8.81-8.80(d,,1H),7.58(s,1H),7.47(s,1H),7.40-7.32(m,3H), 7.12-7.11(d,2H),4.72-4.69(m,1H),4.49-4.45(d,1H),4.19-4.15(m,2H),3.95-3.89(m,4H),3.26-3.11(m,2H).

[0118] Example 5: Measurement of pSYK levels in THP-1 cells induced by the compound

[0119] 1. Reagents and Consumables

[0120] Table 1 Reagents and Consumables

[0121]

[0122] 2. Experimental Apparatus

[0123] Table 2 Experimental Instruments

[0124]

[0125] 3 cells

[0126] Table 3 Cells

[0127]

[0128] 4 Experimental Steps

[0129] 4.1 Cell Culture

[0130] Transfer cells from the culture flask to cell culture medium and count the number of cells. Dilute the cells to the desired density with medium (RPMI + 10% FBS + 1% PS + 30 nM PMA) and add 100 μL of cell suspension to each well of a 96-well cell culture plate as specified. The final seeding density is 100,000 cells / well. Cap the plate and incubate overnight at 37°C in a 5% CO2 incubator. Remove the PMA-containing medium and add 100 μL of fresh, complete PMA-free medium (RPMI + 10% FBS + 1% PS) to each well. Cap the plate and incubate for 24 hours at 37°C in a 5% CO2 incubator to recover.

[0131] 4.2 Compound Treatment

[0132] Wash with 100 μL / well HBSS at the end of culture and before assay. Remove HBSS and add 40 μL / well HBSS. Transfer 10 μL of diluted compound (200×) to a cell plate. After treating the compound at 37°C for 7 min, perform Alphalisa assays as described in the "Detection" section.

[0133] 4.3 Testing

[0134] Remove the processing solution, add 25 μL of lysis buffer 1x (diluted in H2O), and incubate on a plate shaker at 450 rpm for 10 minutes. Transfer 10 μL / well of lysate to an optiplate-384. Add 5 μL of acceptor mixture to each well and incubate at room temperature for 1 hour. Add 5 μL of donor mixture to each well and incubate at room temperature in the dark for 1 hour. Read the plate using a standard Alphalisa setup on a BMG plate.

[0135] The fold difference in pSYK levels between the compound group and the DMSO blank control group was calculated using the following formula:

[0136] Vehicle(%)=100X ReadSample / DMSOave

[0137] pSYK levels in THP-1 cells under the influence of 5 compounds.

[0138] The results showed that the compound had the activity of enhancing SYK phosphorylation.

[0139] Table 4. Effects of compounds on pSYK levels in THP-1 cells.

[0140]

[0141] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above, and that the invention can be implemented in other specific forms without departing from its spirit or essential characteristics. Therefore, the embodiments should be considered in all respects as exemplary and non-limiting, and the scope of the invention is defined by the appended claims rather than the foregoing description. Thus, all variations falling within the meaning and scope of equivalents of the claims are intended to be included within the present invention. No markings in the claims should be construed as limiting the scope of the claims.

Claims

1. A class of compounds of formula I and II, or pharmaceutically acceptable salts, deuterated compounds, solvates, racemic mixtures, enantiomers, diastereomers, and tautomers thereof, characterized in that, As shown in Equations I and II below, in, X is selected from -O-, -S-, or -NH-; R8 is selected from hydrogen or C1-C6 alkyl; Y and Z are each independently selected from N or C(R). 10 ); Q is selected from N or C(R). 11 ); M is selected from -O- or -C(R) 12 ,R 13 )-; R1, R2, R3, R4, R5, R6, R7, R9, R 10 R 11 R 12 and R 13 Each is independently selected from hydrogen, halogen, trifluoromethyl, difluoromethyl, C1-C6 alkyl, C3-C8 cycloalkyl, cyano, nitro, methoxy, hydroxy or amino.

2. The compound according to claim 1, or a pharmaceutically acceptable salt, deuterated compound, solvate, racemic mixture, enantiomer, diastereomer, and tautomer thereof, characterized in that, in, X is selected from -O- or -NH-; R1, R2, R3, R4, R5, R6, R7, R9 and R 10 Each is independently selected from hydrogen, halogen, trifluoromethyl, difluoromethyl, C1-C3 alkyl, C3-C6 cycloalkyl, cyano or methoxy; R8, R 11 R 12 and R 13 Each is independently selected from hydrogen or C1-C3 alkyl groups; Y and Z are each independently selected from -N- or -C(R) 10 )-.

3. The compound according to claim 1, or a pharmaceutically acceptable salt, deuterated compound, solvate, racemic mixture, enantiomer, diastereomer, and tautomer thereof, characterized in that, in, R1, R2, R3, R4, R5, R6, R7, and R9 are each independently selected from hydrogen, fluorine, chlorine, trifluoromethyl, difluoromethyl, methyl, cyano, or methoxy. R8 is selected from methyl; R 10 R 11 R 12 and R 13 Each is independently selected from hydrogen.

4. The compound according to claim 1, or a pharmaceutically acceptable salt, deuterated compound, solvate, racemic mixture, enantiomer, diastereomer, and tautomer thereof, characterized in that, The compound is: Compound S1, as shown above; Compound S2, as shown above; Compound S3, as shown above; Compound S4, as shown above.

5. The compound according to any one of claims 1 to 4, or a pharmaceutically acceptable salt, deuterated compound, solvate, racemic mixture, enantiomer, diastereomer, and tautomer thereof, characterized in that, The application of the compound in the preparation of AD treatment drugs.

6. The compound according to any one of claims 1 to 4, or a pharmaceutically acceptable salt, deuterated compound, solvate, racemic mixture, enantiomer, diastereomer, and tautomer thereof, characterized in that, The compound is used in the preparation of drugs that enhance microglial cell activity.

7. A pharmaceutical composition, characterized in that, It includes the compound of any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof, a deuterated compound, a solvate, a racemic mixture, an enantiomer, a diastereomer, a tautomer, and a pharmaceutically acceptable carrier.

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