Triazole derivatives with inhibitory activity on hepatoma cells, preparation method and application thereof

Abstract

The present application discloses a triazole derivative with liver cancer cell activity, a preparation method and application, and a structure of the derivative is as follows: wherein n is 0, 1, 2 or 3; R 1 , R 2 , R 3 , R 4 and R 5 are H, halogen, trifluoromethyl, ethyl, phenyl, hydroxyl, cyano, nitro, an aromatic ring or the like. The preparation method of the above compound can improve the purity of the product, the triazole derivative with liver cancer cell activity is applied to preparation of an anti-alcoholic liver fibrosis drug or an anti-tumor drug, and the tumor is a liver cancer cell HepG2, HUH7, a lung cancer cell line NCI-H1299, NCI-H1437 or A549.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical synthesis technology, specifically to triazole derivatives that inhibit the activity of liver cancer cells, their preparation methods, and applications. Background Technology

[0002] Heterocyclic organic chemistry is one of the most important and well-studied branches of medicinal chemistry. A key characteristic of heterocyclic bioactive compounds is their diverse constituent heteroatoms, including nitrogen, sulfur, and oxygen. These heteroatoms directly influence the reactivity of the target skeleton, the compound's activity (or toxicology), the interaction between the target drug and different target inhibitors, and can affect metabolism and pharmacokinetics. The most promising heterocyclic compounds are azoles, which are five-membered nitrogen heterocycles. The nitrogen-containing azole skeleton provides the advantage of two nitrogen heteroatoms in the ring, which has a significant impact on structural modification and biological interactions. The chemistry and biology of azole derivatives have become very hot topics in the past decade. Using each azole (“imidazolium,” “pyrazole,” “triazole,” “tetraazole,” and “pentaazole,” etc.) as keywords in database searches, we found that the triazole motif is the most frequently studied, especially the 1,2,3-triazole group. The 1,2,3-triazole core provides a variety of pharmacophore properties, and when hybrids contain or are fused with 1,2,3-triazole rings, they are most often considered “lead compounds.”

[0003] The 1,2,3-triazole group can be prepared via azide-alkyne cycloaddition reactions catalyzed by copper (I) in click chemistry. This is a common structural feature in new drug development, representing bioisosteres of amides, esters, carboxylic acids, and other heterocycles. It readily interacts with various proteins, enzymes, and receptors in organisms through weak bond interactions (e.g., hydrophobic interactions, hydrogen bonds, and van der Waals forces). 1,2,3-triazole derivatives possess significant biological and pharmacological properties, including anti-Alzheimer's disease, anticancer, antimalarial, antituberculosis, antiviral, antibacterial, antiacetylcholinesterase, antioxidant, antidiabetic, and anti-HIV activities. Therefore, 1,2,3-triazole derivatives serve as a unique scaffold for new drug development. Furthermore, some triazole-based drugs have already been used clinically to treat various diseases, demonstrating promising drug profiles.

[0004] The 1,2,3-triazole ring holds undeniable importance in medicinal chemistry due to its unique properties, including metabolic stability, high dipole moment, and ability to form hydrogen bonds. Copper (I)-catalyzed 1,3-dipolar cycloaddition between organozynes and terminal alkynes, commonly known as CuAAC or click chemistry, has been identified as one of the most successful, versatile, reliable, and modular strategies for the rapid and regioselective construction of 1,4-disubstituted 1,2,3-triazoles as multifunctional molecules. This reaction is readily induced, modular, and efficient, with a broad substrate range and high to excellent yields (often quantitative), and is easily scaled up, providing excellent regioselectivity and regiospecificity with few byproducts. The reaction is typically carried out in environmentally friendly solvents or easily removable solvent systems, and the substrates (alkynes / azides) and catalysts are readily available. Therefore, the CuAAC reaction can be considered an ideal click reaction and is one of the most thoroughly studied reactions in the last two decades. Thus, this application chooses to begin with click chemistry reactions to synthesize a series of 1,2,3-triazole derivatives for anticancer activity studies. Summary of the Invention

[0005] To address the aforementioned problems, this invention provides a method for preparing triazole derivatives with inhibitory activity against liver cancer cells and their applications. A batch of novel compounds containing 1,2,3-triazole were designed, and their antitumor activity tests showed good results. Furthermore, the preparation method can improve the purity of the products.

[0006] This invention is achieved through the following technical solution:

[0007] Triazole derivatives that inhibit the activity of liver cancer cells have the following structural formula:

[0008]

[0009]

[0010] Where n is 0, 1, 2, or 3; R 1 R 2 R 3 R 4 and R 5 It can be one or more of the following: H, halogen, trifluoromethyl, ethyl, phenyl, hydroxyl, cyano, nitro, aromatic ring, etc.

[0011] The above-mentioned method for preparing triazole derivatives with inhibitory activity against liver cancer cells, wherein the derivative structural formula is The preparation method includes the following steps:

[0012] S1. Add 2-hydroxy-4-Boc amino-aniline and potassium carbonate to a dichloromethane solution. Set the reaction system to 0°C. Then, slowly add a dichloromethane solution containing chloroacetyl chloride to the system, keeping the temperature stable. After the addition is complete, continue stirring the reaction. Detect the reaction is complete by TLC. Cool to room temperature and transfer the reaction solution in batches to a separatory funnel. Wash and extract with saturated brine, collect the organic phase, add anhydrous sodium sulfate, stir thoroughly, let stand, filter to remove anhydrous sodium sulfate, and remove the organic solvent DCM by vacuum distillation using a rotary evaporator to obtain 7-Boc amino-2H-1,4-benzoxazine-3(4H)-one.

[0013] S2. Add 7-Boc amino-2H-1,4-benzoxazine-3(4H)-one to dichloromethane, stir until completely dissolved, then add trifluoroacetic acid, stir at room temperature, adjust the pH of the reaction system to 6 with saturated sodium carbonate solution, separate the organic phase, concentrate and obtain 7-amino-2H-1,4-benzoxazine-3(4H)-one.

[0014] S3. Add 7-amino-2H-1,4-benzoxazine-3(4H)-one, 3-ethynylaniline, HATU and DIPEA sequentially to a round-bottom flask. Then add N,N-dimethylformamide to the system. After stirring at room temperature, add water to the reaction system. During stirring, a solid precipitates out. Filter and dry to obtain the terminal alkyne compound.

[0015] S4. Obtain the target compound through terminal alkyne compounds.

[0016] Furthermore, the molar ratio of 2-hydroxy-4-Boc amino-aniline to chloroacetyl chloride in S1 is 1:1;

[0017] The molar ratio of 7-amino-2H-1,4-benzoxazine-3(4H)-one, 3-ethynylaniline, HATU and DIPEA in S3 is 1:1.5:1.5:3.

[0018] Furthermore, the specific preparation method of the target compound in S4 is as follows: Terminal alkyne compound, azide compound, L-ascorbic acid sodium, and anhydrous CuSO4 are added to a round-bottom flask. Then, a mixed solvent of water:tert-butanol:THF = 1:1:1 is added to the flask. The mixture is stirred at room temperature until completely dissolved. After the reaction is complete as detected by TLC, diatomaceous earth is placed in a funnel, and the mixture is filtered under reduced pressure to remove solid impurities. The filtrate is collected and transferred to a separatory funnel. The mixture is extracted multiple times with dichloromethane, and the lower organic phase is collected. Then, it is back-extracted multiple times with saturated brine, and the lower organic phase is collected. Anhydrous sodium sulfate is added to the mixture, and after thorough stirring and standing, the anhydrous sodium sulfate is removed by filtration. The DCM organic solvent is removed by vacuum distillation at 35°C to obtain the crude product. The crude product is then separated and purified by column chromatography to finally obtain the target compound.

[0019] Furthermore, the specific preparation method of the target compound in S4 is as follows: take the terminal alkyne compound, azide compound, copper carbonate, and sodium ascorbate, add them to a flask equipped with a stir bar, then add a mixed solvent of water and tert-butanol in a ratio of 1:1 to the flask, heat and reflux the mixture and stir to react, add water and ethyl acetate, extract and separate the emulsion layer, centrifuge the emulsion layer to remove the liquid, retain the solid and dry it to obtain the target compound.

[0020] A method for preparing triazole derivatives with inhibitory activity against liver cancer cells, wherein the derivative has the following structural formula: The preparation method includes the following steps:

[0021] Step 1: Methyl 4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate was dissolved in acetic acid solution. The system was cooled to below 10°C, and a mixture of nitric acid and sulfuric acid was added dropwise. After the addition was complete, the system was gradually raised to room temperature. The system was reacted at room temperature for 24 hours, and the reaction was monitored by TLC. After the reaction was completed, the system was quenched with water, extracted with ethyl acetate, dried over anhydrous sodium sulfate, and the crude product was purified by column chromatography to obtain methyl 2-nitro-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate.

[0022] Step 2: Weigh methyl 2-nitro-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate, dissolve it in anhydrous methanol, add Pd / C for hydrogenation reaction, and let the system react overnight at room temperature. Monitor the reaction by TLC until the starting material disappears. Filter the system directly to remove palladium on carbon, evaporate excess solvent, and purify by column chromatography to obtain methyl 2-amino-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate.

[0023] Step 3: Methyl 2-amino-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate and ammonium formate were dissolved in DMF. The system was heated to 160-165℃ and reacted for 5 hours under nitrogen atmosphere. The reaction was monitored by TLC. After the reaction was completed, the system was poured into water and extracted with ethyl acetate to separate the organic phase. The organic phase was then dried with anhydrous magnesium sulfate, filtered and evaporated under reduced pressure to obtain 7-methoxy-6-(3-morpholin-4-ylpropoxy)quinazolin-4(3H)-one.

[0024] Step 4: Dissolve 7-methoxy-6-(3-morpholin-4-ylpropoxy)quinazoline-4(3H)-one in phosphorus oxychloride, heat the system to reflux, and continue to monitor the reaction by TLC after 20 hours. When the reaction is complete, pour the system into ice water, adjust the pH of the aqueous phase to neutral with saturated sodium bicarbonate, extract with dichloromethane, separate the organic phase, dry with anhydrous magnesium sulfate, filter and evaporate under reduced pressure to obtain 4-chloro-6-methoxy-7-[3-(4-morpholin)propoxy]quinazoline;

[0025] Step 5: Dissolve 4-chloro-6-methoxy-7-[3-(4-morpholino)propoxy]quinazoline in isopropanol, add m-alkynyl aniline, heat the system to reflux, and react for 10 hours. Monitor the reaction by TLC. When the reaction is complete, a solid product precipitates from the system. Filter to obtain a terminal alkyne intermediate.

[0026] Step 6: Obtain the product through terminal alkyne intermediates.

[0027] Furthermore, the molar ratio of methyl 4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate to nitric acid and sulfuric acid in step 1 is 1:1 to 1.3:1 to 1.3;

[0028] In step 2, the mass ratio of methyl 2-nitro-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate to Pd / C is 10:1 to 0.5.

[0029] In step 3, the molar ratio of methyl 2-amino-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate to ammonium formate is 1:3-4.

[0030] Furthermore, the specific preparation method of the product in step 6 is as follows: In a reaction flask, a terminal alkyne intermediate, an azide compound, tert-butanol, water, tetrahydrofuran, copper sulfate pentahydrate, and sodium ascorbate are added sequentially. The mixture is reacted at 90°C for 12 hours. After the raw materials have completely reacted and converted into the product, excess tetrahydrofuran and tert-butanol are evaporated from the system. Secondary water is added, and the mixture is placed in an ultrasonic instrument for sonication. After standing for solid formation, the mixture is centrifuged, the supernatant is removed, and the mixture is recrystallized with a small amount of methanol. The product is then obtained by filtration.

[0031] Furthermore, the specific preparation method of the product in step 6 is as follows: take the terminal alkyne compound, azide compound, copper carbonate, and sodium ascorbate, add them to a flask equipped with a stir bar, then add a mixed solvent of water and tert-butanol in a ratio of 1:1 to the flask, stir at room temperature, add water and ethyl acetate, extract and separate the emulsion layer, remove the liquid after centrifugation of the emulsion layer, retain the solid and dry it to obtain the target compound.

[0032] The above-mentioned triazole derivatives with the ability to inhibit the activity of liver cancer cells are used in the preparation of anti-alcoholic liver fibrosis drugs or anti-tumor drugs.

[0033] Furthermore, the tumor is a liver cancer cell line HepG2, HUH7, or a lung cancer cell line NCI-H1299, NCI-H1437, or A549.

[0034] The beneficial effects of this invention are as follows:

[0035] (1) The present invention has prepared a series of novel 1,2,3-triazole derivatives, some of which target tumor cells by inducing DNA damage in tumor cells and have an inhibitory effect on HUH7 liver cancer cells, but no effect on A549 lung cancer cells, and can be used to prepare anti-alcoholic liver fibrosis drugs.

[0036] (2) At the same time, some compounds have an inhibitory effect on lung cancer cells, and some compounds inhibit tumor cells by targeting MMP9, and can be used as MMP9 inhibitors.

[0037] (3) The preparation method of the compound in this invention is quite unique because triazole exhibits acidity and can coordinate with metals under acidic or neutral conditions. The product coordinates with copper ions in the catalyst, which will affect the purity. This problem can be avoided under alkaline conditions, which can improve the purity and yield of the product. Attached Figure Description

[0038] Figure 1 This is a schematic diagram illustrating the ability to minimize the molecular structure of compound 3;

[0039] Figure 2 for Schematic diagram of energy minimization in molecular structure;

[0040] Figure 3 The hydrogen NMR spectrum of compound 3;

[0041] Figure 4 This is the hydrogen NMR spectrum of compound 18. Detailed Implementation

[0042] The preparation method includes the following steps:

[0043] S1. A certain amount of 2-hydroxy-4-Boc amino-aniline and potassium carbonate were added to a dichloromethane solution. The reaction system was kept at 0°C. Then, a dichloromethane solution containing chloroacetyl chloride was slowly added dropwise to the system while maintaining a stable and constant temperature. After the addition was complete, the mixture was stirred for a period of time. The reaction was detected by TLC and then cooled to room temperature. The reaction solution was transferred in batches to a separatory funnel and extracted with saturated brine. The upper organic phase was collected and anhydrous sodium sulfate was added to it. After stirring thoroughly, the mixture was allowed to stand and filtered to remove the anhydrous sodium sulfate. The organic solvent was removed by vacuum distillation using a rotary evaporator at 35°C. The crude product was then separated and purified by column chromatography to finally obtain 7-Boc amino-2H-1,4-benzoxazine-3(4H)-one. The molar ratio of 2-hydroxy-4-Boc amino-aniline to chloroacetyl chloride was 1:1.

[0044] S2. A certain amount of 7-Boc amino-2H-1,4-benzoxazine-3(4H)-one was added to dichloromethane and stirred until completely dissolved. Then trifluoroacetic acid was added and stirred at room temperature for a period of time. The pH of the reaction system was adjusted to 6 using saturated sodium carbonate solution. The organic phase was separated and concentrated to obtain 7-amino-2H-1,4-benzoxazine-3(4H)-one.

[0045] S3. 7-Amino-2H-1,4-benzoxazine-3(4H)-one, 3-ethynylaniline, HATU, and DIPEA are added sequentially to a round-bottom flask. Then, N,N-dimethylformamide is added to the system. The mixture is stirred at room temperature for a period of time. Water is added to the reaction system. A large amount of solid precipitates during stirring. The solid is filtered and dried to obtain the terminal alkyne compound. The molar ratio of 7-amino-2H-1,4-benzoxazine-3(4H)-one to 3-ethynylaniline to HATU to DIPEA is 1:1.5:1.5:3.

[0046] S4. A certain amount of terminal alkyne compound, azide compound, L-ascorbic acid sodium, and anhydrous CuSO4 were added to a round-bottom flask. The azide compound was 2,6-dichlorobenzyl azide or 3-chlorobenzyl azide. Then, a mixed solvent of water:tert-butanol:THF = 1:1:1 (v / v / v) was added to the flask. The mixture was stirred at room temperature until completely dissolved. After the reaction was completed by TLC, diatomaceous earth was placed in a funnel, and the mixture was filtered under reduced pressure to remove solid impurities. The filtrate was collected and transferred to a separatory funnel. The mixture was extracted multiple times with dichloromethane, and the lower organic phase was collected. The mixture was then back-extracted multiple times with saturated brine, and the lower organic phase was collected. Anhydrous sodium sulfate was added to the mixture, and after thorough stirring and standing, the anhydrous sodium sulfate was removed by filtration. The mixture was then distilled under reduced pressure at 35°C to remove organic solvents such as DCM to obtain the crude product. The crude product was then separated and purified by column chromatography to finally obtain the target compound.

[0047] S4 can also be replaced by: taking a terminal alkyne compound, an azide compound, copper carbonate, and sodium ascorbate. The azide compound can be 3-trifluoromethylphenyl azide or 2-fluorophenyl azide. Add these to a flask equipped with a stir bar, then add a mixed solvent of water and tert-butanol in a 1:1 ratio to the flask. Heat and reflux the mixture while stirring. Add water and ethyl acetate, extract the mixture, and separate the emulsion layer. After centrifugation, remove the liquid from the emulsion layer, retain the solid, and dry it to obtain the target compound.

[0048] The preparation method includes the following steps:

[0049] Step 1: A certain amount of methyl 4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate was dissolved in acetic acid solution. The system was cooled to below 10°C, and a mixture of nitric acid and sulfuric acid was added dropwise. After the addition was complete, the system was gradually raised to room temperature. The reaction was monitored by TLC at room temperature. After the reaction was completed, the system was quenched with water, extracted with ethyl acetate, dried over anhydrous sodium sulfate, and the crude product was purified by column chromatography to obtain methyl 2-nitro-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate. The molar ratio of methyl 4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate to nitric acid and sulfuric acid was 1:1 to 1.3:1 to 1.3.

[0050] Step 2: Weigh a certain amount of methyl 2-nitro-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate, dissolve it in 10 mL of anhydrous methanol, add Pd / C for hydrogenation reaction, and let the system react overnight at room temperature. Monitor the reaction by TLC until the starting material disappears. Filter the system directly to remove palladium on carbon, evaporate excess solvent, and purify by column chromatography to obtain methyl 2-amino-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate. The mass ratio of methyl 2-nitro-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate to Pd / C is 10:1 to 0.5.

[0051] Step 3: A certain amount of methyl 2-amino-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate and ammonium formate were dissolved in DMF. The system was heated to 160-165℃ and reacted for 5 hours under nitrogen atmosphere. The reaction was monitored by TLC. After the reaction was completed, the system was poured into water and extracted with ethyl acetate to separate the organic phase. The organic phase was then dried with anhydrous magnesium sulfate, filtered, and evaporated under reduced pressure to obtain 7-methoxy-6-(3-morpholin-4-ylpropoxy)quinazolin-4(3H)-one. The molar ratio of methyl 2-amino-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate to ammonium formate was 1:3-4.

[0052] Step 4: Dissolve 2.6 g (7.8 mmol) of 7-methoxy-6-(3-morpholin-4-ylpropoxy)quinazoline-4(3H)-one in 10 mL of phosphorus oxychloride. Heat the system to reflux and continue to monitor the reaction by TLC for 20 hours. Once the reaction is complete, pour the system into ice water, adjust the pH of the aqueous phase to neutral with saturated sodium bicarbonate, extract with dichloromethane, separate the organic phase, dry with anhydrous magnesium sulfate, filter and evaporate under reduced pressure to obtain 4-chloro-6-methoxy-7-[3-(4-morpholin)propoxy]quinazoline;

[0053] Step 5: Dissolve 1.5 g (4.5 mmol) of 4-chloro-6-methoxy-7-[3-(4-morpholino)propoxy]quinazoline in 40 mL of isopropanol, add 1.56 g (13.4 mmol) of m-alkynyl aniline, heat the system to reflux, and react for 10 hours. Monitor the reaction by TLC. When the reaction is complete, a large amount of solid product precipitates. Filter the product to obtain 1.6 g of terminal alkyne intermediate, with a yield of 65%.

[0054] Step 6: In the reaction flask, add the terminal alkyne intermediate (250 mg, 0.56 mmol), azide compound (0.7 mmol), 10 mL of tert-butanol / 10 mL of water / 10 mL of tetrahydrofuran, copper sulfate pentahydrate (14 mg, 0.1 mmol), and sodium ascorbate (23 mg, 0.1 mmol) in sequence. The azide compound is 3-fluorobenzyl azide or 4-chlorophenyl azide.

[0055] The reaction was carried out at 90℃ for 12 hours. After the raw materials were completely reacted and converted into the product, excess tetrahydrofuran and tert-butanol were removed from the system by evaporation. 50 mL of deionized water was added, and the system was placed in an ultrasonic instrument for ultrasonication for 20 min. After standing for 1 hour, the solid was generated. After centrifugation for 10 min, the supernatant was removed, and the product was obtained by recrystallization with a small amount of methanol and then filtered.

[0056] Step 6 can also be replaced by: taking the terminal alkyne compound, the azide compound, copper carbonate, and sodium ascorbate (the azide compound can be pomalidomide azide), adding them to a flask equipped with a stir bar, then adding a mixed solvent of water and tert-butanol in a 1:1 ratio to the flask, stirring at room temperature for 30 minutes, adding water and ethyl acetate, extracting, separating the emulsion layer, centrifuging the emulsion layer to remove the liquid, retaining the solid, drying it, and obtaining the target compound.

[0057] The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings.

[0058] Example 1

[0059]

[0060] The preparation process involves the following steps:

[0061] S1、

[0062] 2-hydroxy-4-Boc amino-aniline (0.10 mol) and potassium carbonate (0.10 mol) were added to 280 mL of dichloromethane solution in a clean 500 mL round-bottom flask equipped with a stir bar. The reaction system was kept at 0 °C. Then, 70 mL of dichloromethane solution containing chloroacetyl chloride (0.10 mol) was slowly added dropwise to the system, keeping the temperature stable. After the addition was complete, the mixture was stirred for 30 min. After the reaction was completed by TLC, the mixture was cooled to room temperature. The reaction solution was transferred in batches to a separatory funnel and washed and extracted with saturated saline (200 mL × 3). The organic phase was collected, and 20 g of anhydrous sodium sulfate was added to it. After stirring thoroughly, the mixture was allowed to stand for 30 min. The anhydrous sodium sulfate was removed by filtration, and the organic solvent DCM was removed by vacuum distillation using a rotary evaporator to obtain 25.3 g of 7-Boc amino-2H-1,4-benzoxazine-3(4H)-one.

[0063] S2,

[0064] 0.1 mol of 7-Boc amino-2H-1,4-benzoxazine-3(4H)-one was added to 200 mL of dichloromethane and stirred until completely dissolved. Then, 20 mL of trifluoroacetic acid was added and stirred at room temperature for 2 h. The pH of the reaction system was adjusted to 6 using saturated sodium carbonate solution. The organic phase was separated and concentrated to obtain 15.5 g of 7-amino-2H-1,4-benzoxazine-3(4H)-one.

[0065] S3

[0066] 0.02 mol of 7-amino-2H-1,4-benzoxazine-3(4H)-one, 0.03 mol of 3-ethynylaniline, 0.03 mol of HATU, and 0.06 mol of DIPEA were added sequentially to a clean 500 mL round-bottom flask equipped with a stir bar. Then, 300 mL of N,N-dimethylformamide was added to the system, and the mixture was stirred at room temperature for 1 h. 200 mL of water was then added to the reaction system. During stirring, a large amount of solid precipitated out. The solid was filtered and dried to obtain the terminal alkyne compound. 1H NMR (400MHz, DMSO-d6): 10.76 (s, 1H), 10.26 (s, 1H), 8.04 (s, 1H), 7.96 (d, J=8.0Hz, 1H), 7.68 (d, J= 8.0Hz,1H),7.54-7.52(m,2H),7.25-7.22(m,1H),6.93(d,J=8.0Hz,1H),4.54(s,2H),4.30(s,1H);

[0067] S4

[0068]

[0069] 1 mmol of the terminal alkyne compound, 1.2 mmol of 2,6-dichlorobenzyl azide, 2 mmol of sodium L-ascorbate, and 1 mmol of anhydrous CuSO4 were added to a 100 mL round-bottom flask equipped with a stir bar. Then, 50 mL of a mixed solvent of water, tert-butanol, and THF (v / v / v) was added to the flask, and the mixture was stirred at room temperature for 10 min until completely dissolved. The reaction was allowed to proceed overnight at room temperature with stirring. After the reaction was complete as detected by TLC, the mixture was extracted three times with DCM, and the lower organic phase was collected. The mixture was then back-extracted three times with saturated brine, and the lower organic phase was collected. Anhydrous sodium sulfate was added to this phase, and the mixture was stirred thoroughly and allowed to stand for 30 min. The anhydrous sodium sulfate was then removed by filtration. The crude product was obtained by vacuum distillation at 35°C to remove organic solvents such as DCM. The crude product was then separated and purified by column chromatography (DCM:MeOH = 500:1 v / v, column packed with neutral Al2O3) to finally obtain white solid b1 (yield 77.1%).

[0070] Example 2

[0071]

[0072] The synthesis of compound b2 was similar to that of compound b1, except that 3-chlorobenzyl azide was used instead of 2,6-dichlorobenzyl azide. The crude product was purified by column chromatography to obtain a white solid b2 (yield 81.7%).

[0073] Example 3

[0074]

[0075] The synthesis method of compound b3 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b3 (yield 76.2%).

[0076] Example 4

[0077]

[0078] The synthesis method of compound b4 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b4 (yield 72.4%).

[0079] Example 5

[0080]

[0081] The synthesis method of compound b5 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b5 (yield 73.9%).

[0082] Example 6

[0083]

[0084] The synthesis method of compound b6 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b6 (yield 76.3%).

[0085] Example 7

[0086]

[0087] The synthesis method of compound b7 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b7 (yield 63.8%).

[0088] Example 8

[0089]

[0090] The synthesis method of compound b8 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b8 (yield 66.4%).

[0091] Example 9

[0092]

[0093] The synthesis method of compound b9 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b9 (yield 71.7%).

[0094] Example 10

[0095]

[0096] The synthesis method of compound b10 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b10 (yield 74.6%).

[0097] Example 11

[0098]

[0099] The synthesis method of compound b11 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b11 (yield 55.8%).

[0100] Example 12

[0101]

[0102] The synthesis method of compound b12 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b12 (yield 68.3%).

[0103] Example 13

[0104]

[0105] The synthesis method of compound b13 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b13 (yield 75.9%).

[0106] Example 14

[0107]

[0108] The synthesis method of compound b14 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b14 (yield 66.4%).

[0109] Example 15

[0110]

[0111] The synthesis method of compound b15 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b15 (yield 52.7%).

[0112] Example 16

[0113]

[0114] The synthesis method of compound b16 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b16 (yield 63.3%).

[0115] Example 17

[0116]

[0117] The synthesis method of compound b17 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b17 (yield 65.7%).

[0118] Example 18

[0119]

[0120] The synthesis method of compound b18 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b18 (yield 54.3%).

[0121] Example 19

[0122]

[0123] The synthesis method of compound b19 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b19 (yield 61.6%).

[0124] Example 20

[0125]

[0126] The synthesis method of compound b20 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b20 (yield 73.7%).

[0127] Example 21

[0128]

[0129] The synthesis method of compound b21 is similar to that of compound b1. The crude product was purified by column chromatography to obtain white solid b21 (yield 69.2%).

[0130] Example 22

[0131]

[0132] Unlike S4 in Example 1, S4 in this Example is as follows: 1 mmol of terminal alkyne compound 6, 1.0 mmol of 3-trifluoromethylphenyl azide, 0.05 mmol of copper carbonate, and 0.1 mmol of sodium ascorbate were added to a 100 mL round-bottom flask equipped with a stir bar. Then, 50 mL of a mixed solvent of water and tert-butanol in a ratio of 1:1 (v / v) was added to the flask. The mixture was heated under reflux and stirred for 10 min. 10 mL of water and 5 mL of ethyl acetate were added. After extraction, a large amount of emulsion layer appeared in the middle. The emulsion layer was centrifuged to remove the liquid, and the solid was dried to obtain b22 (yield 94.1%).

[0133] Example 23

[0134]

[0135] Unlike the azide compounds used in Example 22, the following steps were taken: 1 mmol of terminal alkyne compound 6, 1.0 mmol of 2-fluorophenyl azide, 0.01 mmol of copper carbonate, and 0.02 mmol of sodium ascorbate were added to a 100 mL round-bottom flask equipped with a stir bar. Then, 50 mL of a mixed solvent of water and tert-butanol in a ratio of 1:1 (v / v) was added to the flask, and the mixture was heated under reflux and stirred for 20 min. 10 mL of water and 5 mL of ethyl acetate were added. After extraction, a large amount of emulsion layer appeared in the middle. The emulsion layer was centrifuged to remove the liquid, and the solid was dried to obtain b23 (yield 92.5%).

[0136] Example 24

[0137]

[0138] Unlike S4 in Example 1, S4 in this Example is as follows: 1 mmol of terminal alkyne compound, 1.2 mmol of 4-((2-(2-(2-(2-azido)ethoxy)ethoxy)ethyl)amino)-pomalidomide, and 0.01 mmol of cuprous iodide were added to a 100 mL round-bottom flask equipped with a stir bar. Then, 50 mL of a mixed solvent of water and tert-butanol in a ratio of 1:1 (v / v) was added to the flask. The mixture was stirred at room temperature for 10 min, and then stirred for 2 h at room temperature. 10 mL of water and 5 mL of ethyl acetate were added. After extraction, the intermediate emulsion layer was separated. The emulsion layer was centrifuged to remove the liquid, and the solid was dried to obtain b24 (yield 51.8%).

[0139] Example 25

[0140] Unlike Example 24S4, Example S4 was prepared as follows: 1 mmol of terminal alkyne compound, 1.2 mmol of 4-((2-(2-(2-(2-azidoethoxy)ethoxy)ethoxy)ethyl)amino)-pomalidomide, 0.02 mmol of copper carbonate, and 0.05 mmol of sodium ascorbate were added to a 100 mL round-bottom flask equipped with a stir bar. Then, 50 mL of a mixed solvent of water and tert-butanol (1:1 v / v) was added to the flask. The mixture was stirred at room temperature for 10 min, and then stirred for 2 h at room temperature. 10 mL of water and 5 mL of ethyl acetate were added. After extraction, a large emulsion layer was separated. The emulsion layer was centrifuged to remove the liquid, and the solid was dried to obtain b24 (yield 90.9%).

[0141] Example 26

[0142] The preparation of [the substance] involves the following steps:

[0143] Step 1

[0144]

[0145] 50 mmol of methyl 4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate was dissolved in 100 mL of acetic acid. The system was cooled to below 10 °C, and a mixture of 62 mmol of nitric acid and 62 mmol of sulfuric acid was added dropwise. After the addition was complete, the system was gradually raised to room temperature. The system was reacted at room temperature for 30 hours, and the reaction was monitored by TLC. After the reaction was completed, the system was quenched with water, extracted with ethyl acetate, dried over anhydrous sodium sulfate, and the crude product was purified by column chromatography to obtain 12.7 g of methyl 2-nitro-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate.

[0146] Step 2

[0147]

[0148] 1 g of methyl 2-nitro-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate was weighed and dissolved in 12 mL of anhydrous methanol. Pd / C (10%, 0.1 g) was added for hydrogenation. The reaction was carried out overnight at room temperature and monitored by TLC until the starting material disappeared. The system was directly filtered to remove palladium on carbon. After removing excess solvent by evaporation, the system was purified by column chromatography to obtain 0.71 g of methyl 2-amino-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate.

[0149] Step 3

[0150]

[0151] Methyl 2-amino-4-methoxy-3-(3-(morpholin-4-yl)propoxy)benzoate (10 mmol) and ammonium formate (32 mmol) were dissolved in 30 mL of DMF. The system was heated to 160–165 °C for 5 hours under nitrogen atmosphere and the reaction was monitored by TLC. After the reaction was completed, the system was poured into water and extracted multiple times with 50 mL of ethyl acetate to separate the organic phase. The organic phase was then dried with anhydrous magnesium sulfate, filtered, and evaporated under reduced pressure to obtain 2.87 g of 7-methoxy-6-(3-morpholin-4-ylpropoxy)quinazolin-4(3H)-one.

[0152] Step 4

[0153]

[0154] 3.2 g of 7-methoxy-6-(3-morpholin-4-ylpropoxy)quinazoline-4(3H)-one was dissolved in 15 mL of phosphorus oxychloride. The system was heated to reflux and the reaction was monitored by TLC for 20 hours. After the reaction was completed, the system was poured into ice water. The pH of the aqueous phase was adjusted to neutral with saturated sodium bicarbonate. The organic phase was extracted with dichloromethane and dried with anhydrous magnesium sulfate. The solution was filtered and evaporated under reduced pressure to obtain 2.51 g of 4-chloro-6-methoxy-7-[3-(4-morpholin)propoxy]quinazoline.

[0155] Step 5

[0156]

[0157] 4-Chloro-6-methoxy-7-[3-(4-morpholino)propoxy]quinazoline (1.5 g) was dissolved in 40 mL of isopropanol, and m-alkynyl aniline (1 g) was added. The system was heated to reflux and reacted for 15 hours. The reaction was monitored by TLC. After the reaction was completed, 10 mL of isopropanol was concentrated and a large amount of solid product precipitated. The product was filtered to obtain 1.6 g of terminal alkyne intermediate.

[0158] Step 6

[0159]

[0160] In a reaction flask, a terminal alkyne intermediate (0.56 mmol), 3-fluorobenzyl azide (0.7 mmol), 10 mL tert-butanol / 10 mL water / 10 mL tetrahydrofuran, copper sulfate pentahydrate (0.1 mmol), and sodium ascorbate (0.1 mmol) were added sequentially. The reaction was carried out at 90 °C for 12 h. After the reactants were completely converted into the product, excess tetrahydrofuran and tert-butanol were evaporated from the system. 50 mL of deionized water was added, and the mixture was ultrasonicated for 20 min. After standing for 1 h, the solid was generated. After centrifugation for 10 min, the supernatant was removed, and the product (4c) was recrystallized from a small amount of methanol and filtered to obtain the product (4c). The yield was 81.9%.

[0161] Example 27

[0162]

[0163] The difference from Example 26 is that the azide compound in step 6 is different. Specifically, in the reaction flask, a terminal alkyne intermediate (0.56 mmol), 4-chlorophenyl azide (0.7 mmol), 10 mL of tert-butanol / 10 mL of water / 10 mL of tetrahydrofuran, copper sulfate pentahydrate (0.1 mmol), and sodium ascorbate (0.1 mmol) were added sequentially. The reaction was carried out at 90°C for 12 h. After the raw materials were completely reacted and converted into the product, excess tetrahydrofuran and tert-butanol were evaporated from the system. 50 mL of deionized water was added, and the system was ultrasonicated for 20 min. After standing for 1 h, the solid was generated. After centrifugation for 10 min, the supernatant was removed, and the product was recrystallized with a small amount of methanol and filtered to obtain the product (4d). The yield was 91.4%.

[0164] Example 28

[0165]

[0166] The synthesis method of compound 4e is similar to that of compound 4d. The crude product was purified by column chromatography to obtain white solid 4e (yield 88.6%).

[0167] Example 29

[0168]

[0169] The synthesis method of compound 4f is similar to that of compound 4d. The crude product was purified by column chromatography to obtain white solid 4f (yield 91.4%).

[0170] Example 30

[0171]

[0172] The synthesis method of compound 4g is similar to that of compound 4d. The crude product was purified by column chromatography to obtain 4g of white solid (yield 92.7%).

[0173] Example 31

[0174]

[0175] The synthesis method of compound 4h is similar to that of compound 4d. The crude product was purified by column chromatography to obtain white solid 4h (yield 88.6%).

[0176] Example 32

[0177]

[0178] The synthesis method of compound 4i is similar to that of compound 4d. The crude product was purified by column chromatography to obtain white solid 4i (yield 94.6%).

[0179] Example 33

[0180]

[0181] Unlike step 6 in Example 26, step 6 in this example is as follows: Take terminal alkyne compound 6 (1 mmol), pomalidomide azide (1.2 mmol), copper carbonate (0.01 mmol), and sodium ascorbate (0.02 mmol) and add them to a 100 mL round-bottom flask equipped with a stir bar. Then add 50 mL of a mixed solvent of water and tert-butanol = 1:1 (v / v) to the flask, stir at room temperature for 30 min, add 10 mL of water and 5 mL of ethyl acetate, extract, and separate the emulsion layer that appears in the middle. After centrifugation, remove the liquid from the emulsion layer, retain the solid, dry it, and obtain 4j (yield 96.2%).

[0182] Example 34

[0183]

[0184] The synthesis of compound 4k was similar to that of compound 4d. The crude product was purified by column chromatography to obtain a white solid 4i (yield 90.5%).

[0185] Example 35

[0186]

[0187] The synthesis method of compound 7 is similar to that of compound 4d. The crude product was purified by column chromatography to obtain white solid 7 (yield 94.7%).

[0188] Example 36

[0189] Anti-alcoholic liver fibrosis activity assay: Mouse hepatic stellate cells (HSC-T6) were seeded into sterile 6cm culture dishes and cultured in DMEM medium. After cell attachment, the culture medium was discarded, and DMEM medium containing 100mM ethanol was added. The dishes were then incubated at 37℃ in a CO2 incubator for 48 hours to activate HSC-T6 cells into HSC+ cells. 5000 HSC+ cells / well were seeded into 96-well plates and placed in a cell culture incubator. After complete attachment (approximately 24 hours), the waste medium in the wells was aspirated, and drug concentrations of 0μM, 2μM, 4μM, 8μM, 16μM, and 32μM were added sequentially, with 5 replicates per group. After drug administration, the plates were incubated for another 48 hours. Compounds B10, B11, and B12 showed better performance, with IC50 values... 50 The values ​​were 69.16, 94.03, and 16.04 μM, respectively.

[0190] Example 37

[0191] Antitumor activity assay: Cells were cultured in DMEM or RPMI 1640 medium containing 10% fetal bovine serum (FBS) and 1% penicillin / streptomycin at 37°C and 5% CO2. CCK-8 assays were performed on several human cancer cell lines, including Huh7, MCF-7, and A549, according to the CCK-8 manufacturer's instructions. Cells were plated at a density of 3000–5000 cells per well in 96-well plates. After 24 hours of incubation, cells were treated with a 50 μM compound for 72 hours, and 10 μL of CCK-8 reagent was added to each well. The CCK-8 reagent was dissolved in the cell culture medium for 1 hour, and cell viability was measured (Table 1). Compounds B3 and B18 can induce DNA damage in HUH7 cells. For example, compound B18 at 20 μM can increase the expression level of the DNA damage protein H2AX by more than 14 times compared to the blank control, indicating that these compounds target the induction of DNA damage in tumor cells. Our IDO1 inhibition experiment, induced by activation of HeLa cells, revealed that compounds 3 and 18 had no inhibitory effect on IDO1 protein, and the IC50 value was [not specified]. 50 Greater than 10 μM; compound 3 ratio It exhibits a strong inhibitory effect on Huh7. Through molecular structure energy minimization analysis, it was found that the 1,4-benzoxazinone and 2-fluorobenzyl structures, compared to the phenyltriazole structure, exhibit an expanded structure, facilitating the insertion of the phenyltriazole structure into tumor cell DNA. The 1,4-benzoxazinone structure in the molecular structure folds towards the phenyltriazole structure, increasing steric hindrance and easily hindering insertion into DNA. B22, B23, and B24, at 40 μM, inhibited HUH7 cells by 17.71%, 13.25%, and 9.83%, respectively. Specific test results are shown in Table 1 below.

[0192] Table 1

[0193]

[0194] Example 38

[0195] Human lung cancer cell lines NCI-H1299, NCI-H1437, A549, and HepG2 were used. NCI-H1299 and NCI-H1437 cells were cultured in complete medium containing 10% fetal bovine serum and 1% RPMI-1640; A549 cells were cultured in complete medium containing 10% fetal bovine serum and 1% Ham's F-12K. Cell lines were aseptically cultured at 37°C in a humidified incubator with 5% CO2. The compounds and gefitinib were dissolved in DMSO to prepare stock solutions with a final concentration of 12800 μM. The compounds were then diluted with the corresponding cell culture medium to different concentrations for subsequent experiments (0, 2, 4, 8, 16, 32 μmol / L, where the final concentration of DMSO did not exceed 0.1% in all experiments).

[0196] Lung cancer cells in the logarithmic growth phase were divided into groups of 5 × 10⁶ cells per well. 3 Cells were seeded at a density of 1000 μL in 96-well plates and cultured at 37°C in a 5% CO2 incubator. After cell attachment, the cells were divided into a normal control group, a blank control group, and different drug concentration groups (0, 2, 4, 8, 16, 32 μmol / L). Except for the normal control group, which received no intervention, all other groups were treated with the corresponding drugs and cultured at 37°C in a 5% CO2 incubator for 48 h, with three replicates per group. 10 μL of CCK8 reagent was added to each well, and the cells were incubated in the dark at 37°C for 1-4 h. The absorbance (OD) was measured at 450 nm using a Bio-Tek microplate reader. The inhibition rate of cell growth in each group was calculated as follows: Inhibition rate (%) = [(OD value of experimental group - OD value of blank group) / (OD value of control group - OD value of blank group)] × 100%. The half-maximal inhibitory concentration (IC50) of the drug was then calculated. 50 The experiment was repeated three times. The IC50 values ​​of compounds 4c, 4i, 4k, and 7 against the HepG2 cell line were... 50 The concentrations were approximately 3.64, 3.83, 8.19, and 0.81 μM. Through an IDO1 inhibition assay induced by HeLa cell activation, we found that compound 4c had no inhibitory effect on IDO1 protein, with an IC50 of approximately 3.64, 3.83, 8.19, and 0.81 μM. 50The concentration was greater than 30 μM, and it did not inhibit EGFR protein as shown in the EGFR inhibition assay. However, protein immunoblotting experiments revealed that at a concentration of 4 μM, MMP9 protein expression was reduced to approximately 75%, and at 16 μM, it was reduced to approximately 20%. Compound 7, at 16 μM, reduced MMP9 protein expression to approximately 17%. In contrast, the positive control compound (the click reaction product of icotinib and benzyl azide) had little effect on MMP9 reduction, showing an 81% reduction at 16 μM. Therefore, it can be determined that the target of this type of compound in inhibiting tumor cells is MMP9, and it can be used as an MMP9 inhibitor. Specific test results are shown in Table 2 below:

[0197] Table 2

[0198]

[0199] The foregoing has shown and described the basic principles, main features, and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The embodiments and descriptions in the specification are merely illustrative of the principles of the present invention. Various changes and modifications can be made to the present invention without departing from its spirit and scope. All such changes and modifications fall within the scope of the present invention as claimed, which is defined by the appended claims and their equivalents.

Claims

1. A triazole derivative having the ability to inhibit the activity of liver cancer cells, characterized in that: Its structural formula is: ; Where n is 1; R 1 R 2 R 3 R 4 and R 5 Each is independently selected from H, halogen, trifluoromethyl, ethyl, hydroxy, cyano or nitro.

2. The method for preparing the triazole derivative with inhibitory activity against liver cancer cells according to claim 1, characterized in that: Includes the following steps: S1. Add 2-hydroxy-4-Boc amino-aniline and potassium carbonate to a dichloromethane solution. Set the reaction system to 0°C. Then, slowly add a dichloromethane solution containing chloroacetyl chloride to the system, keeping the temperature stable. After the addition is complete, continue stirring and detect the reaction by TLC. After the reaction is complete, cool to room temperature. Transfer the reaction solution in batches to a separatory funnel, extract with saturated brine, collect the upper organic phase, add anhydrous sodium sulfate, stir thoroughly, let stand, filter to remove anhydrous sodium sulfate, remove the organic solvent by vacuum distillation using a rotary evaporator at 35°C, and then separate and purify the crude product by column chromatography to finally obtain 7-Boc amino-2H-1,4-benzoxazine-3(4H)-one. S2. Add 7-Boc amino-2H-1,4-benzoxazine-3(4H)-one to dichloromethane, stir until completely dissolved, then add trifluoroacetic acid, stir at room temperature, adjust the pH of the reaction system to 6 with saturated sodium carbonate solution, separate the organic phase, concentrate and obtain 7-amino-2H-1,4-benzoxazine-3(4H)-one. S3. Add 7-amino-2H-1,4-benzoxazine-3(4H)-one, 3-ethynylaniline, HATU and DIPEA sequentially to a round-bottom flask. Then add N,N-dimethylformamide to the system. After stirring at room temperature, add water to the reaction system. During stirring, a solid precipitates out. Filter and dry to obtain the terminal alkyne compound. S4. The target compound is obtained by using terminal alkyne compounds.

3. The method for preparing the triazole derivative with inhibitory activity against liver cancer cells according to claim 2, characterized in that: The molar ratio of 2-hydroxy-4-Boc amino-aniline to chloroacetyl chloride in S1 is 1:1; The molar ratio of 7-amino-2H-1,4-benzoxazine-3(4H)-one, 3-ethynylaniline, HATU and DIPEA in S3 is 1:1.5:1.5:

3.

4. The method for preparing the triazole derivative with inhibitory activity against liver cancer cells according to claim 2, characterized in that: The specific preparation method of the target compound in S4 is as follows: Terminal alkyne compound, azide compound, L-ascorbic acid sodium, and anhydrous CuSO4 are added to a round-bottom flask. Then, a mixed solvent of water, tert-butanol, and THF (1:1:1) is added to the flask. The mixture is stirred at room temperature until completely dissolved. After the reaction is complete as detected by TLC, diatomaceous earth is placed in a funnel, and the mixture is filtered under reduced pressure to remove solid impurities. The filtrate is collected and transferred to a separatory funnel. The mixture is extracted multiple times with dichloromethane, and the lower organic phase is collected. Then, it is back-extracted multiple times with saturated brine, and the lower organic phase is collected. Anhydrous sodium sulfate is added to the mixture, and after thorough stirring and standing, the anhydrous sodium sulfate is removed by filtration. The DCM organic solvent is removed by vacuum distillation at 35°C to obtain the crude product. The crude product is then separated and purified by column chromatography to finally obtain the target compound.

5. The method for preparing the triazole derivative with inhibitory activity against liver cancer cells according to claim 2, characterized in that: The specific preparation method of the target compound in S4 is as follows: Take the terminal alkyne compound, azide compound, copper carbonate, and sodium ascorbate, add them to a flask equipped with a stir bar, then add a mixed solvent of water and tert-butanol = 1:1 to the flask, heat and reflux and stir the reaction, add water and ethyl acetate, extract and separate the emulsion layer, remove the liquid after centrifugation of the emulsion layer, retain the solid and dry it to obtain the target compound.

6. The use of the triazole derivative of claim 1, which has the activity of inhibiting liver cancer cells, in the preparation of an antitumor drug, wherein the tumor is liver cancer cell HUH7.

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