Phenyl thiazole derivative with anti-liver cancer and anti-inflammatory activity as well as preparation method and application thereof

Abstract

The invention relates to a phenyl thiazole derivative with anti-liver cancer and anti-inflammatory activity as well as a preparation method and application thereof, and belongs to the technical field of synthesis of medicines. According to the invention, 2-phenylthiazole is used as a mother nucleus structure, terminal alkyne is introduced to the structure, and then the 2-phenylthiazole reacts with an azide compound through a click reaction to obtain a plurality of compounds with novel structures, and the compounds have good anti-tumor, anti-inflammatory and anti-oxidation activities, and provide certain application value for research and development of innovative drugs.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of synthesis with drugs, and particularly relates to a phenylthiazole derivative with anti-liver cancer and anti-inflammatory activities, a preparation method and application thereof. BACKGROUND

[0002] Cancer cells have the characteristics of unlimited proliferation, infiltration and diffusion to normal tissues or organs, which makes cancer very difficult, seriously affects people's health, and its prevention and treatment has become the core content of global health strategy. Thiazole derivatives are a class of very important heterocyclic compounds with wide biological activities. Many important compounds contain thiazole ring structure. Thiazole derivatives are often used to synthesize compounds with high efficiency, low toxicity and good drug metabolism characteristics. Thiazole ring is the basic component of some antitumor drugs such as dasatinib, dalaferine and ixabepilone. In recent years, some new high-efficiency low-toxicity antitumor thiazole derivatives have been synthesized. For example, 2-thiazole carboxamide compounds related to 2-phenylthiazole have obvious inhibitory effect on Akt, and can inhibit the phosphorylation of downstream MDM2 and GSK3β proteins. Mahapatra et al. designed to synthesize melanin-schiff base derivatives by changing the thiazole substituent, and found that the antitumor activity of the derivatives was generated by the cytotoxicity of EGFR inhibition mechanism. Salem et al. synthesized a series of new 1,3-thiazole compounds, which showed good antitumor activity. It was found that the mechanism of antitumor activity of compound 2-[5-(4-chlorophenyl)-1,3-thiazol-2-yl)]-1-acetylhydrazine was its obvious inhibitory activity on vascular endothelial growth factor receptor-2 (VEGFR-2), which could induce cell arrest in G1 phase and induce mitochondrial depolarization. Sabry et al. synthesized a series of new imidazothiazole derivatives, among which compounds N-acetyl-5-(4-bromophenyl) imidazothiazole-3-formylhydrazine and 5-(4-methoxyphenyl)-6-(morpholinomethyl) imidazothiazole-3-formylhydrazine had double EGFR / HER2 kinase inhibition effect, which induced cell arrest in G1 / S phase and G1 phase, and the modeling research results showed that the thiazole part had antitumor activity.

[0003] 1,2,3-triazole is an important drug molecule building block, which has a planar rigid structure that can be inserted into the DNA of tumor cells, interact with DNA and cause DNA damage in tumor cells. Such structures have similar amide, ester, carboxylic acid, etc. The electronic isosteric effect makes it have the ability to coordinate with metal ions, can act on metal ion-containing proteases, inhibit their activity and affect the growth of tumor cells; at the same time, 1,2,3-triazole ring can link different pharmacological molecules into a new molecule, and often such derivatives have low toxicity and high activity characteristics. These characteristics of triazoles make them widely used in the modification of natural products, active azole compounds or quinone compounds, and widely used in the development of innovative drugs. Our team cooperates with Jinan Aishiyangke Technology Co., Ltd. to carry out the synthesis and activity test of phenylthiazole derivatives based on 1,2,3-triazole modification. Jinan Aishiyangke Technology Co., Ltd. is responsible for the synthesis of compounds, and our team is responsible for the activity test. SUMMARY

[0004] In view of the above problems, the purpose of the present application is to provide a phenylthiazole derivative with anti-hepatoma and anti-inflammatory activity and a preparation method and application thereof. The present application introduces a terminal alkyne on the structure of 2-phenylthiazole, and then reacts with azide compounds through click reaction to obtain a variety of novel compounds with good anti-tumor, anti-inflammatory and antioxidant activities.

[0005] In order to achieve the above purpose, the specific scheme adopted by the present application is:

[0006] In the first aspect, the present application provides a phenylthiazole derivative with anti-hepatoma and anti-inflammatory activity, which has the structure of: In the formula, R 1 , R 2 , R 3 , R 4 and R 5 are one or more of H, halogen, trifluoromethyl, ethyl, phenyl, hydroxyl, cyano, nitro, aromatic ring, etc.

[0007] In a second aspect, the present application provides a preparation method of the phenylthiazole derivative, which comprises the following steps: adding 2-(3-nitrile-4-isobutoxyphenyl)-4-methyl-N-(prop-2-yn-1-yl)thiazole-5-formamide, an azide compound, sodium L-ascorbate and anhydrous CuSO4 into a round-bottom flask with a stirrer, then adding a mixed solvent of water, t-butyl alcohol and THF in a volume ratio of 1:1:1 into the flask, stirring at room temperature to make them all dissolved; stirring at room temperature overnight; after the reaction is completed by TLC detection, extracting with dichloromethane, collecting the lower organic phase; re-extracting with saturated brine, collecting the lower organic phase, adding anhydrous sodium sulfate thereto, stirring sufficiently, standing for 30 min, and then removing the anhydrous sodium sulfate by filtration; removing dichloromethane at 35°C under reduced pressure to obtain a crude product, then separating and purifying the obtained crude product by column chromatography, and finally obtaining a solid, which is the phenylthiazole derivative; The azide compound is any one of 2-nitrile benzyl azide, 4-bromobenzyl azide, 2-chloro-5-fluorobenzyl azide, 3-fluorobenzyl azide, 2-bromobenzyl azide, zidovudine, 2-chlorophenyl azide, 2-fluorophenyl azide, 3-chlorophenyl azide, 4-chlorophenyl azide, 4-fluorophenyl azide, 2-trifluoromethylbenzyl azide, 2-trifluoromethoxyphenyl azide, 3-trifluoromethylbenzyl azide, 2-chloro-6-fluorobenzyl azide, 4-trifluoromethylbenzyl azide and 4-fluorobenzyl azide.

[0008] Further, the present application also relates to a preparation process of a plurality of raw materials.

[0009] In a third aspect, the present application provides an application of the phenylthiazole derivative in preparing an antitumor drug.

[0010] Beneficial effects: the present application introduces a terminal alkyne into a 2-phenylthiazole as a mother nucleus structure, and then reacts with an azide compound through a click reaction to obtain a plurality of novel compounds with good antitumor, anti-inflammatory and antioxidant activities, which provides great application value for the research and development of innovative antitumor drugs. BRIEF DESCRIPTION OF DRAWINGS

[0011] Figure 1 is a nuclear magnetic hydrogen spectrum of compound 7.2.

[0012] Figure 2 is a nuclear magnetic hydrogen spectrum of compound 7.3.

[0013] Figure 3 is a nuclear magnetic hydrogen spectrum of compound 7.8.

[0014] Figure 4 is a molecular docking structure schematic diagram of compound 7.2.

[0015] Figure 5is a schematic diagram of a molecular docking structure of compound 7.3.

[0016] Figure 6 is a schematic diagram of a molecular docking structure of compound 7.8. DETAILED DESCRIPTION

[0017] The present application takes 2-phenylthiazole as a mother nucleus structure, introduces a terminal alkyne on the structure, then reacts with an azide compound through a click reaction to obtain more than ten novel compounds, and tests the antitumor, anti-inflammatory and antioxidant activities.

[0018] The above content of the present application is further described in detail through the following examples, but this should not be understood as the scope of the above subject matter of the present application being limited to the following examples only, and any technology realized based on the above content of the present application belongs to the scope of the present application.

[0019] Example 1

[0020]

[0021] In a reaction kettle with a temperature regulating device, 27 g of polyphosphoric acid was added, 3 g of water was slowly added dropwise under stirring, and after the addition was completed, stirring was continued for 10 min, and the temperature of the reaction system was controlled at 40℃; 26.3 g of ethyl 2-(4-hydroxyphenyl)-4-methylthiazole-5-carboxylate was added, stirring was continued for 10 min, the temperature of the reaction liquid was controlled, 15.4 g of urotropine was slowly added to the reaction system, the temperature was maintained at 40-50℃ during the addition process, and after the addition was completed, stirring was continued for 20 min, the reaction liquid was slowly heated, the temperature of the reaction liquid was raised to 90-95℃, and the reaction was maintained for about 2 h. After the reaction was completed, the temperature was lowered to below 40℃, 48 g of water was slowly added dropwise, and after the addition was completed, stirring was continued at room temperature for 2 h, and then the temperature was lowered to 5℃, stirring was continued for 1 h, and then centrifugation was performed, the filter cake was dried, 70 g of ethanol was added, and then heating was continued until complete dissolution, and then the temperature was lowered to 5℃, stirring was continued for 1 h, and then centrifugation was performed, the filter cake was washed once with cold ethanol, and then centrifugation was performed to obtain about 25.7 g of ethyl 2-(3-formyl-4-hydroxyphenyl)-4-methylthiazole-5-carboxylate. 1 H NMR (400 MHZ, DMSO-d6): 11.41 (s, 1H), 10.31 (s, 1H), 8.22 (d, J = 4.0 Hz, 1H), 8.09 (dd, J1 = 4.0 Hz, J2 = 4.0 Hz, 1H), 7.11 (d, J = 8.0 Hz, 1H), 4.29 (dd, J1 = 8.0 Hz, J2 = 4.0 Hz, 2H), 2.66 (s, 3H), 1.03 (t, J = 8.0 Hz, 3H).

[0022] Example 2

[0023]

[0024] In a 1000L reactor, add 263kg of polyphosphoric acid, slowly add 27kg of drinking water under stirring, continue stirring for 10min after the addition, then open the cooling water valve to cool the mixture to 40°C. Add 40kg of ethyl 2-(4-hydroxyphenyl)-4-methylthiazole-5-carboxylate, open the circulating water valve to control the temperature of the reaction liquid, slowly add 235.5kg of urotropine, keep the temperature at 40-50°C during the addition. After the addition, open the steam valve to slowly heat the reaction liquid, raise the temperature of the reaction liquid to 90-95°C, keep the reaction for about 1.0h, take a sample for testing, HPLC control, when the content of ethyl 2-(4-hydroxyphenyl)-4-methylthiazole-5-carboxylate is less than 0.5%, the reaction is completed, if not, take a sample every 0.5h. After the reaction is completed, close the circulating water, cool the jacket with drinking water. When the internal temperature drops to below 40°C, slowly add drinking water, stir for 2h after the addition, cool to 5±5°C, stir for 2h for crystallization, centrifuge. Wash the filter cake with drinking water (3-4 times the amount each time) for 2 times, elute once (1 times the amount), wash until the pH of the filtrate is close to neutral (the number of washing times is determined according to the actual effect of centrifugation), spin dry (try to extend the time as much as possible to make the moisture content of the product less than 50%), after centrifugation, take a sample for testing, if the wet product meets the related substances and quality standards, then directly proceed to drying. Crush the filter cake and transfer it to a forced air drying oven, dry at 55-65°C for more than 6h, then transfer the filter cake to another clean reaction kettle containing ethanol (the mass of the filter cake*(1-water content)*3), add water to make the ethanol content 75%, then heat to complete dissolution, then cool to 5±5°C, stir for 1h for crystallization, centrifuge, elute the filter cake with cold ethanol (75%, 5-10°C) once, spin dry. Crush the filter cake and transfer it to a forced air drying oven, dry at 55-65°C for more than 6h, to obtain 202.1g of ethyl 2-(3-formyl-4-hydroxyphenyl)-4-methylthiazole-5-carboxylate

[0025] Example 3

[0026]

[0027] In a reaction kettle with temperature regulating device, 2-(3-formyl-4-hydroxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester 29 g, bromoiso-butane 20 g and anhydrous potassium carbonate 20 g were added into N,N-dimethylformamide 500 g, stirred uniformly, heated to 90°C, and kept for 5 h. Sampling detection was performed by HPLC. When the content of 2-(3-formyl-4-hydroxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester was less than 0.5%, the reaction was completed. If the reaction was not completed, sampling was performed every 1 h. After the reaction was completed, the temperature was slowly decreased to 20-30°C. Drinking water 150 g was slowly added. The reaction solution was cooled to 5-10°C during stirring. Stirring and crystallization were performed for 1 h. Centrifugation was performed. The filter cake was dried and transferred to another clean reaction kettle containing isopropanol 130 g. Slurry was prepared at room temperature for 1 h. The temperature was decreased to 5-10°C. Stirring and crystallization were performed for 1 h. Centrifugation was performed. The filter cake was washed with isopropanol (0.5 times the amount). After suction filtration, 2-(3-formyl-4-isobutoxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester 31.4 g was obtained by drying. 1 H NMR (400 MHz, DMSO-d6): 10.42 (s, 1H), 8.25 (s, 2H), 7.38 (d, J = 8.0 Hz, 1H), 4.33-4.28 (m, 2H), 4.01 (d, J = 4.0 Hz, 2H), 2.69 (s, 3H), 2.18-2.06 (m, 1H), 1.31 (t, J = 8.0 Hz, 3H), 1.05-1.03 (m, 6H).

[0028] Example 4

[0029]

[0030] In a reaction kettle with temperature regulating device, add 300 g of anhydrous formic acid, then add 35 g of 2-(3-formyl-4-isobutoxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester, 7.7 g of hydroxylamine hydrochloride and 12.5 g of sodium formate dihydrate in sequence, heat to reflux for 2 h, take sample for detection, HPLC control, when 2-(3-formyl-4-isobutoxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester is less than 0.5%, the reaction is completed (the color of the reaction solution changes from light yellow to yellow during the reaction process), if not completely reacted, take sample every 0.5 h, during heating, the reaction solution will quickly solidify, the stirring speed needs to be accelerated to make it uniform, after the reaction is completed, the reaction solution is cooled to 20-30°C, which will solidify, the stirring speed needs to be accelerated to make it uniform, then drink water is slowly added, after the addition, the reaction solution is cooled to 10-20°C, stirred for crystallization for 1 h, centrifuged, the filter cake is soaked and washed with drink water (2 times the amount) once, washed with drink water (1 times the amount) once, washed with ethanol water (1:1, v:v, 1 times the amount) once, finally the washing liquid is close to neutral, after drying, 140 kg of ethanol is added, heated to reflux for 1 h, then cooled to 5-10°C, stirred for crystallization for 1 h, centrifuged. The filter cake is washed with ethanol (0.5 times the amount), and about 32.2 g of 2-(3-nitrile-4-isobutoxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester is obtained after drying and spinning. 1 H NMR (400 MHZ, DMSO-d6): 8.32 (s, 1H), 8.25 (dd, J1= 4.0 Hz, J2= 4.0 Hz, 1H), 7.38 (d, J = 8.0 Hz, 1H), 4.33-4.28 (m, 2H), 4.01 (d, J = 4.0 Hz, 2H), 2.68 (s, 3H), 2.13-2.05 (m, 1H), 1.31 (t, J = 8.0 Hz, 3H), 1.03 (s, 3H), 1.02 (s, 3H).

[0031] Example 5

[0032] The ethanol was vacuum extracted into a dried reaction kettle, stirring was started, 2-(3-cyano-4-isobutoxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester 30 g was added, heated to 55±5°C, 20% sodium hydroxide 25 g was slowly added dropwise, the temperature was maintained at 55±5°C during the dropwise addition, after the dropwise addition was completed, the reaction was continued for 1 h with stirring, a sample was taken for testing, HPLC control, when the content of 2-(3-cyano-4-isobutoxyphenyl)-4-methylthiazole-5-carboxylic acid ethyl ester was less than 0.1%, the reaction was completed, if not completely reacted, a sample was taken every 0.5 h. After the reaction was completed, the reaction liquid was cooled to below 20°C, 2 mol / L dilute hydrochloric acid was slowly added dropwise, the pH was adjusted to 2-3, the temperature of the reaction liquid was controlled to be less than 30°C during the acidification process, and a large amount of white solid was precipitated. After acidification, the reaction was stirred at 10-20°C for 1 h, centrifuged, the filter cake was soaked and washed with drinking water (2 times the amount) once, eluted once, and finally the washing liquid was close to neutral, eluted with ethanol (90%, 0.5 times the amount), and dried. The filter cake was transferred to a forced air drying oven and dried at 60-70°C for more than 6 h, the moisture content was less than or equal to 2.0%, and 2-(3-cyano-4-isobutoxyphenyl)-4-methylthiazole-5-carboxylic acid 24.8 g was obtained.

[0033] Example 6

[0034]

[0035] In a reaction kettle with temperature regulating device, 2-(3-cyano-4-isobutoxyphenyl)-4-methylthiazole-5-carboxylic acid 31.6 g was added, then N,N-dimethylformamide 300 mL was added, then propargylamine 11 g, HATU 38 g and DIPEA 13 g were added, stirring was started at room temperature for 6 h, 500 mL of water was added to the reaction system, a large amount of solid was precipitated during stirring, and 2-(3-nitrile-4-isobutoxyphenyl)-4-methyl-N-(prop-2-yn-1-yl)thiazole-5-carboxamide 33.6 g was obtained by filtration and drying.

[0036] Example 7

[0037] To a 100 mL round bottom flask equipped with a stir bar, was added 2-(3-cyano-4- isobutoxyphenyl)-4-methyl-N-(prop-2-yn-1-yl)thiazole-5-carboxamide (1 mmol), 2- cyano benzyl azide (1.2 mmol), sodium L-ascorbate (2 mmol), anhydrous CuSO4 (1 mmol), and then 50 mL of a mixture of water:t-butanol:THF = 1:1:1 (v / v / v) was added to the flask, and stirred at room temperature for 10 min to make it completely dissolved. The reaction was stirred at room temperature overnight for 2 h. After the reaction was completed by TLC detection, it was extracted with dichloromethane 3 times, and the lower organic phase was collected. It was back-extracted with saturated brine 3 times, and the lower organic phase was collected, and anhydrous sodium sulfate was added to it, stirred well, and then left to stand for 30 min, and then anhydrous sodium sulfate was filtered out. The dichloromethane was removed by distillation under reduced pressure at 35°C to obtain a crude product, and then the obtained crude product was separated and purified by column chromatography to obtain a solid 7.1; 1 H NMR (400 MHz, DMSO-d6): 8.82 (t, J = 8.0 Hz, 1H), 8.24 (d, J = 4.0 Hz, 1H), 8.17 (dd, J1= 4.0 Hz, J2= 4.0 Hz, 1H), 8.11 (s, 1H), 7.92 (d, J = 8.0 Hz, 1H), 7.73 (t, J = 8.0 Hz, 1H), 7.56 (t, J = 12.0 Hz, 1H), 7.38 (d, J = 8.0 Hz, 2H), 5.80 (s, 2H), 4.50 (d, J = 4.0 Hz, 2H), 4.01 (d, J = 8.0 Hz, 2H), 2.60 (s, 3H), 1.03 (s, 3H), 1.01 (s, 3H).

[0038] Example 8

[0039]

[0040] The synthesis method of compound 7.2 is similar to that of compound 7.1, using 4- bromobenzyl azide instead of 2-cyano benzyl azide, and the crude product is separated and purified by column chromatography to obtain a solid 7.2; 1H NMR (400 MHZ, DMSO-d6): 8.82 (t, J = 8.0 Hz, 1H), 8.24 (d, J = 4.0 Hz, 1H), 8.17 (dd, J1= 4.0 Hz, J2= 4.0 Hz, 1H), 8.09 (s, 1H), 7.58 (dd, J1= 4.0 Hz, J2= 4.0 Hz, 1H), 7.38 (d, J = 12.0 Hz, 1H), 7.32-7.27 (m, 1H), 7.09 (dd, J1= 4.0 Hz, J2= 4.0 Hz, 1H), 5.69 (s, 2H), 4.50 (d, J = 8.0 Hz, 2H), 4.01 (d, J = 8.0 Hz, 2H), 2.59 (s, 3H), 1.03 (s, 3H), 1.02 (s, 3H).

[0041] NMR hydrogen spectrum as shown in Figure 1 , molecular docking as shown in Figure 4 .

[0042] Example 9

[0043]

[0044] The synthesis method of compound 7.3 is similar to that of compound 7.1, using 2-chloro-5-fluorobenzyl azide to replace 2-nitrile benzyl azide, and the crude product is separated and purified by column chromatography to obtain solid 7.3; 1 H NMR (400 MHZ, DMSO-d6): 8.82 (t, J = 8.0 Hz, 1H), 8.24 (d, J = 4.0 Hz, 1H), 8.17 (dd, J1= 4.0 Hz, J2= 4.0 Hz, 1H), 8.09 (s, 1H), 7.58 (dd, J1= 4.0 Hz, J2= 4.0 Hz, 1H), 7.38 (d, J = 12.0 Hz, 1H), 7.32-7.27 (m, 1H), 7.09 (dd, J1= 4.0 Hz, J2= 4.0 Hz, 1H), 5.69 (s, 2H), 4.50 (d, J = 8.0 Hz, 2H), 4.01 (d, J = 8.0 Hz, 2H), 2.59 (s, 3H), 1.03 (s, 3H), 1.02 (s, 3H).

[0045] NMR hydrogen spectrum as shown in Figure 2 , molecular docking as shown in Figure 5 .

[0046] Example 10

[0047]

[0048] The synthesis method of compound 7.4 is similar to that of compound 7.1, using 3-fluorobenzyl azide to replace 2-nitrile benzyl azide, and the crude product is separated and purified by column chromatography to obtain solid 7.4; 1H NMR (400 MHz, DMSO-d6): 8.80 (t, J = 8.0 Hz, 1H), 8.24 (d, J = 4.0 Hz, 1H), 8.17 (dd, J1= 4.0 Hz, J2= 4.0 Hz, 1H), 8.10 (s, 1H), 7.46-7.37 (m, 2H), 7.20-7.15 (m, 3H), 5.62 (s, 2H), 4.49 (d, J = 8.0 Hz, 2H), 4.01 (d, J = 8.0 Hz, 2H), 2.60 (s, 3H), 1.03 (s, 3H), 1.02 (s, 3H).

[0049] Example 11

[0050]

[0051] The synthesis method of compound 7.5 is similar to compound 7.1, using 2-bromobenzyl azide to replace 2-cyano benzyl azide, and the crude product is separated and purified by column chromatography to obtain solid 7.5; 1 H NMR (400 MHz, DMSO-d6): 8.80 (t, J = 8.0 Hz, 1H), 8.24 (d, J = 4.0 Hz, 1H), 8.17 (dd, J1= 4.0 Hz, J2= 4.0 Hz, 1H), 8.10 (s, 1H), 7.46-7.37 (m, 2H), 7.20-7.15 (m, 3H), 5.62 (s, 2H), 4.49 (d, J = 8.0 Hz, 2H), 4.01 (d, J = 8.0 Hz, 2H), 2.60 (s, 3H), 1.03 (s, 3H), 1.02 (s, 3H).

[0052] Example 12

[0053] The synthesis method of compound 7.6 is similar to compound 7.1, using zidovudine to replace 2-cyano benzyl azide, and the crude product is separated and purified by column chromatography to obtain solid 7.6; 1H NMR (400 MHz, DMSO-d6): 8.89 (t, J = 4.0 Hz, 1H), 8.48 (s, 1H), 8.24 (d, J = 4.0 Hz, 1H), 8.19-8.16 (m, 1H), 7.84 (d, J = 8.0 Hz, 1H), 7.64-7.54 (m, 2H), 7.47-7.37 (m, 2H), 4.61 (d, J = 8.0 Hz, 2H), 4.01 (d, J = 8.0 Hz, 2H), 2.63 (s, 3H), 1.03 (s, 3H), 1.01 (s, 3H).

[0054] Example 13

[0055]

[0056] The synthesis of compound 7.7 was similar to compound 7.1, using 2-chlorophenyl azide to replace 2-nitrilebenzyl azide, and the crude product was separated and purified by column chromatography to obtain solid 7.7.

[0057] Example 14

[0058]

[0059] The synthesis of compound 7.8 was similar to compound 7.1, using 2-fluorophenyl azide to replace 2-nitrilebenzyl azide, and the crude product was separated and purified by column chromatography to obtain solid 7.8; 1 H NMR (400 MHz, DMSO-d6): 8.89 (t, J = 4.0 Hz, 1H), 8.48 (s, 1H), 8.24 (d, J = 4.0 Hz, 1H), 8.19-8.16 (m, 1H), 7.84 (d, J = 8.0 Hz, 1H), 7.64-7.54 (m, 2H), 7.47-7.37 (m, 2H), 4.61 (d, J = 8.0 Hz, 2H), 4.01 (d, J = 8.0 Hz, 2H), 2.63 (s, 3H), 1.03 (s, 3H), 1.01 (s, 3H).

[0060] The nuclear magnetic hydrogen spectrum is as shown in Figure 3 The molecular docking is as shown in Figure 6

[0061] Example 15

[0062]

[0063] ​The synthesis of compound 7.9 was similar to compound 7.1 using 3-chlorophenyl azide to replace 2-cyano benzyl azide, the crude product was purified by column chromatography to give solid 7.9.

[0064] Example 16

[0065]

[0066] The synthesis of compound 7.10 was similar to compound 7.1 using 4-chlorophenyl azide to replace 2-cyano benzyl azide, the crude product was purified by column chromatography to give solid 7.10.

[0067] Example 17

[0068]

[0069] The synthesis of compound 7.11 was similar to compound 7.1 using 4-fluorophenyl azide to replace 2-cyano benzyl azide, the crude product was purified by column chromatography to give solid 7.11.

[0070] Example 18

[0071]

[0072] The synthesis of compound 7.12 was similar to compound 7.1 using 2-trifluoromethyl benzyl azide to replace 2-cyano benzyl azide, the crude product was purified by column chromatography to give solid 7.12.

[0073] Example 19

[0074]

[0075] The synthesis of compound 7.13 was similar to compound 7.1 using 2-trifluoromethoxy phenyl azide to replace 2-cyano benzyl azide, the crude product was purified by column chromatography to give solid 7.13.

[0076] Example 20

[0077]

[0078] The synthesis of compound 7.14 was similar to compound 7.1 using 3-trifluoromethyl benzyl azide to replace 2-cyano benzyl azide, the crude product was purified by column chromatography to give solid 7.14.

[0079] Example 21

[0080]

[0081] The synthesis of compound 7.15 was similar to compound 7.1, using 2-chloro-6- fluorobenzyl azide to replace 2-cyano benzyl azide, the crude product was purified by column chromatography to give solid 7.15.

[0082] Example 22

[0083]

[0084] The synthesis of compound 7.16 was similar to compound 7.1, using 4- trifluoromethylbenzyl azide to replace 2-cyano benzyl azide, the crude product was purified by column chromatography to give solid 7.16.

[0085] Example 23

[0086]

[0087] The synthesis of compound 7.17 was similar to compound 7.1, using 4- fluorobenzyl azide to replace 2-cyano benzyl azide, the crude product was purified by column chromatography to give solid 7.17; 1 H NMR (400 MHZ, DMSO-d6): 8.79 (t, J = 8.0 Hz, 1H), 8.23 (d, J = 4.0 Hz, 1H), 8.17 (dd, J1= 4.0 Hz, J2= 4.0 Hz, 1H), 8.05 (s, 1H), 7.42-7.37 (m, 3H), 7.21 (t, J = 8.0 Hz, 2H), 5.57 (s, 2H), 4.48 (d, J = 4.0 Hz, 2H), 4.01 (d, J = 4.0 Hz, 2H), 2.59 (s, 3H), 1.03 (s, 3H), 1.01 (s, 3H).

[0088] Example 24

[0089] HepG2 cells were plated in 96-well plates at about 2000-5000 cells per well. Cells were treated with compounds at a concentration of 40 μM, respectively, and four replicates were set for each concentration group. After incubation in the cell incubator for 72 hours, CCK8 reagent was added to each well according to the proportion of 10 ul CCK8 reagent per 100 ul, and the cells were incubated in the cell incubator for 1 hour. The absorbance value at 450 nm of the cells was determined by an enzyme-labeled instrument. The experiment was set in triplicate. According to the absorbance value, the activity of the cells at different concentrations was obtained, and the cell survival rate at different concentrations was calculated. At the same time, the Hela cell-based IDO1 enzyme activity determination method was used to study the inhibitory activity of the compounds with better inhibitory activity on HepG2. Under the condition of 10 μM, it was found that some compounds could inhibit the activity of IDO1 by 47.19%. Through molecular docking with the IDO1 target, it was found that the triazole in the structure of the compound could interact with the key active site of IDO1, and we found that if the methyl group connecting the amide and the triazole was replaced by a phenyl group, due to the increase in molecular structure, it could not enter the active pocket of the IDO1 target.

[0090] Example 25

[0091] HUH-7, A549, PC-9, H460, H1299 and L02 cells were plated in 96-well plates at about 2000 cells per well. Cells were treated with compounds at concentrations of 0 μM, 5 μM, 10 μM, 20 μM, 40 μM and 80 μM, respectively, and four replicates were set for each concentration group. After incubation in the cell incubator for 72 hours, CCK8 reagent was added to each well according to the proportion of 10 ul CCK8 reagent per 100 ul, and the cells were incubated in the cell incubator for 30 minutes. The absorbance value at 450 nm of the cells was determined by an enzyme-labeled instrument. The experiment was set in triplicate. According to the absorbance value, the activity of the cells at different concentrations was obtained, and the cell inhibition rate at different concentrations was calculated. F compound HUH-7 A549 PC-9 H460 H1299 LO2 7.1 86.48±2.80 86.97±0.78 64.61±2.68 96.12±1.42 50.65±4.51 108.58±0.65 7.2 84.41±2.99 82.89±1.78 71.32±2.91 88.39±0.77 115.13±7.23 112.03±0.75 7.3 88.14±3.15 96.76±0.65 47.15±0.22 95.65±0.22 119.43±5.73 114.42±1.07 7.4 92.53±2.86 106.56±1.48 58.08±2.16 68.53±3.79 138.85±8.17 68.56±0.28 7.5 86.85±3.02 111.97±2.67 92.87±1.56 86.19±3.29 123.52±13.79 71.64±2.21 7.6 107.31±1.58 104.70±2.35 91.40±3.70 98.90±0.67 87.46±8.66 117.98±2.96 7.7 64.12±1.60 87.54±0.42 60.17±2.27 89.67±5.11 89.98±4.86 103.67±0.78 7.8 65.40±3.77 74.20±1.88 47.66±0.95 92.64±1.34 125.97±1.64 103.82±0.85 7.9 77.25±1.62 82.45±1.13 91.99±1.80 66.89±0.94 127.13±1.60 102.02±2.20 7.10 77.89±3.29 63.34±0.54 94.43±0.72 104.68±1.96 118.06±1.95 107.56±2.08 7.11 79.70±3.47 97.87±0.83 101.86±0.96 102.44±1.91 146.28±3.24 107.92±1.83 7.12 70.44±2.67 107.31±0.53 72.08±2.01 106.49±2.19 134.76±3.01 119.99±1.51 7.13 70.20±1.39 101.33±1.01 69.59±0.62 103.41±1.77 146.22±3.46 112.55±0.83 7.14 64.12±1.43 95.08±1.54 77.98±3.09 80.27±0.93 85.69±8.45 87.12±0.59 7.15 70.04±4.21 79.17±1.33 55.80±0.91 102.81±2.07 77.71±7.08 105.80±0.69 7.16 95.39±4.15 88.16±1.53 90.43±1.23 50.67±8.88 118.47±5.52 91.09±1.49 7.17 107.53±8.96 70.83±1.39 79.16±1.98 102.68±3.56 116.84±4.29 103.71±2.24

[0092] Example 26

[0093] Lipopolysaccharide (LPS) induced BV2 cell inflammation model: BV2 cells were seeded in 96-well plates at 2 x 10 4Detection plates were seeded per well on 96-well plates and incubated in a 37°C incubator containing 5% CO2. After 24 hours of incubation, the corresponding concentration of the test compound was added to the drug-treated group and incubated for 2 hours. Subsequently, LPS was added to both the drug-treated group and the LPS model group at a final concentration of 100 ng / ml. After 24 hours of incubation, the supernatant from each group was reacted with an equal volume of Griess buffer, and the OD value of each group was measured at 540 nm using a BioTek microplate reader.

[0094] Table 1. Results of the compound's effect on improving LPS-induced inflammation in BV2 cells In the NO generation rate detection, the NO generation rate of the LPS model group was set to 100% (n=3). The data in the table are expressed as mean ± SEM. * P<0.05, ** P<0.01 relative to the LPS group.

[0095] Example 27

[0096] Cell viability assay: BV2 cells were used at a concentration of 2 × 10⁻⁶. 4 Cells were seeded per well in a 96-well plate and incubated at 37°C in a 5% CO2 incubator. After 24 hours of incubation, the corresponding concentration of the test compound was added to the drug-treated group and incubated for 2 hours. Subsequently, LPS was added to both the drug-treated group and the LPS model group at a final concentration of 100 ng / mL. After another 24 hours of incubation, MTT was added to each well at a final concentration of 0.5 mg / mL for live cell staining. After incubation for 1 hour, the culture medium was discarded, and 100 μL of DMSO was added to each well. The solution was shaken on a shaker to dissolve the solution completely, and the OD value of each group was measured at 490 nm using a Biotek microplate reader.

[0097] Table 2. Toxicity of compounds on BV2 cells

[0098] In the cell viability assay, the cell viability of the LPS model group was set to 100% (n=3). The data in the table are expressed as mean ± SEM. * P<0.05, *** P<0.001 relative to the LPS group.

[0099] The above examples describe the basic principles, main features and advantages of the present application, and those skilled in the art should understand that the present application is not limited to the above examples, and the above examples and descriptions in the specification are only to illustrate the principles of the present application, and various changes and improvements can be made without departing from the principles of the present application, and such changes and improvements fall within the scope of the present application.

Claims

1. A phenylthiazole derivative with anti-hepatocellular carcinoma and anti-inflammatory activities, characterized in that: Its structure is as follows: In the formula, 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.

2. The method for preparing the phenylthiazole derivative according to claim 1, characterized in that: 2-(3-Cryptyl-4-isobutoxyphenyl)-4-methyl-N-(prop-2-yn-1-yl)thiazolyl-5-carboxamide, an azide compound, sodium L-ascorbate, and anhydrous CuSO4 were added to a flask. A mixed solvent of water, tert-butanol, and THF was then added to the flask, and the mixture was stirred at room temperature until completely dissolved. The reaction was carried out overnight at room temperature with stirring. After the reaction was complete, the mixture was extracted with dichloromethane, and the lower organic phase was collected. The mixture was then back-extracted with saturated brine, and the lower organic phase was collected again. Anhydrous sodium sulfate was added to the mixture, and after thorough stirring and standing for a period of time, the anhydrous sodium sulfate was removed by filtration. Dichloromethane was removed by vacuum distillation at 35°C to obtain the crude product. The crude product was then separated and purified by column chromatography to obtain a solid, which was the phenylthiazolyl derivative. The azide compound is any one of 2-acrylonitrile benzyl azide, 4-bromobenzyl azide, 2-chloro-5-fluorobenzyl azide, 3-fluorobenzyl azide, 2-bromobenzyl azide, zidovudine, 2-chlorophenyl azide, 2-fluorophenyl azide, 3-chlorophenyl azide, 4-chlorophenyl azide, 4-fluorophenyl azide, 2-trifluoromethylbenzyl azide, 2-trifluoromethoxyphenyl azide, 3-trifluoromethylbenzyl azide, 2-chloro-6-fluorobenzyl azide, 4-trifluoromethylbenzyl azide, and 4-fluorobenzyl azide.

3. The preparation method according to claim 2, characterized in that: The preparation method of 2-(3-acrylonitrile-4-isobutoxyphenyl)-4-methyl-N-(prop-2-yn-1-yl)thiazolyl-5-carboxamide, the raw material used in the preparation of phenylthiazole derivatives, includes the following steps: Step 1: Add polyphosphoric acid to the reaction vessel, slowly add water dropwise while stirring, and control the reaction system temperature at 40℃; add ethyl 2-(4-hydroxyphenyl)-4-methylthiazolium-5-carboxylate, stir, control the reaction solution temperature, slowly add hexamethylenetetramine to the reaction system, slowly heat the reaction solution to 90-95℃ and maintain the temperature for the reaction. After the reaction is complete, cool down to below 40℃, slowly add water dropwise, stir at room temperature, cool down to allow crystallization, centrifuge, dry the filter cake, add ethanol, heat until completely dissolved, then cool down to allow crystallization, centrifuge, wash the filter cake with ethanol, and spin dry to obtain ethyl 2-(3-aldehyde-4-hydroxyphenyl)-4-methylthiazolium-5-carboxylate; the reaction formula is as follows: Step 2: Add ethyl 2-(3-aldehyde-4-hydroxyphenyl)-4-methylthiazol-5-carboxylate, isobutane bromide, and anhydrous potassium carbonate to the reaction vessel, then add N,N-dimethylformamide. After stirring evenly, heat to 90℃ and maintain the temperature for reaction. After the reaction is complete, slowly cool to 20-30℃ and slowly add drinking water. Cool to allow crystals to precipitate, centrifuge, dry the filter cake, and transfer it to another clean reaction vessel containing isopropanol. Pulverize at room temperature for 1 hour, then cool to allow crystals to precipitate, centrifuge, wash the filter cake with isopropanol, filter, and dry to obtain ethyl 2-(3-aldehyde-4-isobutoxyphenyl)-4-methylthiazol-5-carboxylate; the reaction formula is as follows: Step 3: Add anhydrous formic acid to the reaction vessel, then add ethyl 2-(3-aldehyde-4-isobutoxyphenyl)-4-methylthiazol-5-carboxylate, hydroxylamine hydrochloride, and sodium formate dihydrate sequentially. Heat to reflux and react. After the reaction is complete, cool to 20-30°C, and slowly add drinking water. After the addition is complete, cool the reaction solution and stir to induce crystallization. Centrifuge, soak, wash, and rinse the filter cake, adjust the pH to neutral, dry, add ethanol, heat to reflux and slurry, then cool and stir to induce crystallization. Centrifuge, rinse the filter cake with ethanol, and spin dry to obtain ethyl 2-(3-cyano-4-isobutoxyphenyl)-4-methylthiazol-5-carboxylate; the reaction formula is as follows: Step 4: Vacuum extract ethanol into a dried reaction vessel, start stirring, add ethyl 2-(3-acrylonitrile-4-isobutoxyphenyl)-4-methylthiazol-5-carboxylic acid, heat to 55±5℃, slowly add sodium hydroxide solution dropwise. After the addition is complete, continue stirring. After the reaction is complete, cool the reaction solution to below 20℃, slowly add dilute hydrochloric acid to adjust the pH to 2-3, acidify, and then keep at 10-20℃ with stirring to precipitate crystals. Centrifuge, and soak, wash, rinse, and dry the filter cake to obtain 2-(3-acrylonitrile-4-isobutoxyphenyl)-4-methylthiazol-5-carboxylic acid; the reaction formula is as follows: Step 5: Add 2-(3-acrylonitrile-4-isobutoxyphenyl)-4-methylthiazolyl-5-carboxylic acid and N,N-dimethylformamide sequentially to the reaction vessel, followed by propargylamine, HATU, and DIPEA. Stir at room temperature, add water to the reaction system, and a solid precipitates during stirring. Filter and dry to obtain 2-(3-acrylonitrile-4-isobutoxyphenyl)-4-methyl-N-(prop-2-yn-1-yl)thiazolyl-5-carboxamide; the reaction formula is as follows:

4. The use of the phenylthiazole derivative according to claim 1 in the preparation of antitumor drugs.

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