Synthesis and application of chrysin-1, 3, 5-triazine derivative

Abstract

The invention relates to a synthetic method of a chrysin-1, 3, 5-triazine derivative and an antitumor activity research of the chrysin-1, 3, 5-triazine derivative. According to the invention, a series of chrysin-1, 3, 5-triazine derivatives are synthesized by modifying chrysin and a 1, 3, 5-triazine compound. The synthesis method is simple, convenient and efficient, and has better selectivity and yield. The synthesized derivatives show remarkable anti-tumor activity through in-vitro cell experiments and in-vivo animal experiments, and especially in the aspect of inhibiting growth of breast cancer (MDA-MB-231), gastric cancer (7901), pancreatic cancer (PANC-1) and lung cancer (A549) cells, part of the derivatives show a low nanomole-level IC50 value. Researches show that these chrysin-1, 3, 5-triazine derivatives may play an anti-tumor role by inhibiting tumor cell proliferation, inducing apoptosis and other mechanisms, and have the potential to become novel anti-tumor drugs. The synthesis method of the compound, the antitumor activity and the mechanism research of the compound provide effective thinking and basis for developing new antitumor drugs, and the compound has a wide application prospect.

Description

Technical Field

[0001] The present invention relates to the technical field of drug synthesis, and particularly to a synthesis method and application of a series of chrysin-1,3,5-triazine derivatives. Background Art

[0002] Malignant tumors are one of the diseases that seriously endanger human health. Their pathogenesis is quite complex, and there are quite many factors inducing their occurrence and development. Currently, the relatively clear factors can be roughly divided into two major categories: exogenous factors and endogenous factors. Exogenous factors include environmental, occupational pollution, living habits, iatrogenic trauma, irritation and other factors. The latter include factors such as the body's own immunity, endocrine, and genetics. Currently, most of the chemotherapy drugs used in clinical cancer treatment achieve the treatment purpose by blocking cell chromosome division or inhibiting the replication of the cell genetic material DNA. However, most traditional chemotherapy drugs do not have specificity and targeting. They can not only inhibit the growth of cancer cells but also damage the growth of normal cells, and have many toxic side effects.

[0003] Natural products have always been one of the important sources of lead compounds for tumor treatment. Chrysin, as a natural flavonoid compound, has attracted the attention of scholars at home and abroad. Such compounds are extracted from low-toxic or non-toxic plants and have a variety of biological activities, such as anti-tumor, anti-inflammatory, antibacterial, anti-anxiety, antioxidant and other pharmacological activities. However, during the in vivo metabolism process, the C-5 or C-7 hydroxyl groups of chrysin are rapidly glycosylated, resulting in a decrease in activity, which limits its clinical application. Therefore, it is of great significance to modify the structure of its nuclear parent to improve its lipophilicity and drug activity. Due to the stability and strong reactivity of 1,3,5-triazine in biological media, it has been explored in drug discovery programs. In fact, many studies have shown that 1,3,5-triazine derivatives have anti-cancer activity, and the 1,3,5-triazine skeleton has been applied to the design of anti-cancer drugs. Therefore, adding 1,3,5-triazine to the structure of chrysin can effectively increase its anti-tumor activity. Summary of the Invention

[0004] The present invention aims to provide a chrysin-1,3,5-triazine derivative, and provide an efficient and legal method for the new compound, as well as the application of the new compound in the treatment of tumors.

[0005] In order to achieve the above object, the present invention specifically adopts the following technical solutions:

[0006] A new chrysin-1,3,5-triazine derivative:

[0007] Dissolve 2,4,6-trichloro-1,3,5-triazine (1.0 mmol, 1.0 equiv.) and anhydrous K2CO3 (1.1 mmol, 1.1 equiv.) in an appropriate amount of acetone and cool the solution to -10 °C. Then, slowly add a secondary amine (1.1 mmol, 1.1 equiv.) dissolved in an appropriate amount of acetone dropwise. After the reaction is completed (indicated by TLC), evaporate the solvent under vacuum, and then extract the mixture with ethyl acetate and saturated brine multiple times to remove anhydrous K2CO3 and the remaining 2,4,6-trichloro-1,3,5-triazine. Dry the organic layer over anhydrous Na2SO4. Remove the dried organic solvent by rotary evaporation to obtain the compound. Dissolve chrysin (1.0 equiv.), anhydrous K2CO3 (1.1 equiv.) and compound 2a-2i (1.0 equiv.) in an appropriate amount of anhydrous dimethyl sulfoxide and heat the solution at 45 °C for 8 hours. After determining the total conversion to the product by thin layer chromatography (TLC) analysis, purify the reaction mixture by column chromatography (petroleum ether:ethyl acetate = 10:1) to obtain compounds 4a-4i.

[0008]

[0009] The present invention provides a series of novel chrysin-1,3,5-triazine derivatives.

[0010] The present invention provides a method for synthesizing a novel chrysin-1,3,5-triazine derivative, which has the advantages of simple operation, safety, few reaction by-products, high yield, and easy separation and purification of the product.

[0011] The chrysin-1,3,5-triazine derivatives of the present invention have strong inhibitory activities against a variety of tumor cells. The compound with the strongest inhibitory effect has an IC50 value of 15.36 μM against breast cancer, showing broad application prospects in the treatment of various cancers. Brief Description of the Drawings

[0012] Figure 1 1H-NMR spectrum of the final product in Example 1 1 ; Figure 2 13C-NMR spectrum of the final product in Example 1 13 ;

[0013] Figure 3 1H-NMR spectrum of the final product in Example 2 1 ; Figure 4 13C-NMR spectrum of the final product in Example 2 13 ;

[0014] Figure 5 1H-NMR spectrum of the final product in Example 3 1 ;Figure 6 13C-NMR spectrum of the final product of Example 3; 13 C-NMR spectrum;

[0015] Figure 7 1H-NMR spectrum of the final product of Example 4; 1 H-NMR spectrum; Figure 8 13C-NMR spectrum of the final product of Example 4; 13 C-NMR spectrum;

[0016] Figure 9 1H-NMR spectrum of the final product of Example 5; 1 H-NMR spectrum; Figure 10 13C-NMR spectrum of the final product of Example 5; 13 C-NMR spectrum;

[0017] Figure 11 1H-NMR spectrum of the final product of Example 6; 1 H-NMR spectrum; Figure 12 13C-NMR spectrum of the final product of Example 6; 13 C-NMR spectrum;

[0018] Figure 13 1H-NMR spectrum of the final product of Example 7; 1 H-NMR spectrum; Figure 14 13C-NMR spectrum of the final product of Example 7; 13 C-NMR spectrum;

[0019] Figure 15 1H-NMR spectrum of the final product of Example 8; 1 H-NMR spectrum; Figure 16 13C-NMR spectrum of the final product of Example 8; 13 C-NMR spectrum;

[0020] Figure 17 1H-NMR spectrum of the final product of Example 9; 1 H-NMR spectrum; Figure 18 13C-NMR spectrum of the final product of Example 9; 13 C-NMR spectrum;

[0021] Figure 19 1H-NMR spectrum of the final product of Example 10; 1 H-NMR spectrum; Figure 20 13C-NMR spectrum of the final product of Example 10; 13 C-NMR spectrum;

[0022] Figure 21 1H-NMR spectrum of the final product of Example 11; 1 H-NMR spectrum; Figure 22 13C-NMR spectrum of the final product of Example 11; 13 C-NMR spectrum;

[0023] Figure 23 1H-NMR spectrum of the final product of Example 12; 1H-NMR spectrum; Figure 24 of the final product of Example 12 13 C-NMR spectrum. Detailed implementation manner

[0024] The technical solutions in the specific embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. The following description of at least one exemplary embodiment is actually only illustrative and in no way limits the present invention and its application or use. Based on the embodiments of the present invention, other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the scope of protection of the present invention.

[0025] Example 1: Preparation of 7-((4-chloro-6-(dicyclohexylamino)-1,3,5-triazin-2-yl)oxy)-5-hydroxy-2-phenyl-4H-benzofuran-4-one

[0026]

[0027] Dissolve 2,4,6-trichloro-1,3,5-triazine (1.0 mmol, 1.0 equiv.) and anhydrous K2CO3 (1.1 mmol, 1.1 equiv.) in an appropriate amount of acetone (Dissolve) and cool it to -10 °C, then dropwise add N-cyclohexyl-N-methylcyclohexylamine (1.1 mmol, 1.1 equiv.) dissolved in an appropriate amount of acetone. After the reaction is completed (shown by TLC), evaporate the solvent under vacuum, and then extract with ethyl acetate and saturated brine for multiple times to remove anhydrous K2CO3 and the remaining 2,4,6-trichloro-1,3,5-triazine. Dry the organic layer over anhydrous Na2SO4. The dried organic solvent is removed by rotary evaporation to obtain the compound. Dissolve chrysin (1.0 equiv.), anhydrous K2CO3 (1.1 equiv.) and compound 2a-2i (1.0 equiv.) in an appropriate amount of anhydrous dimethyl sulfoxide (Dissolve), and heat at 45 °C for 8 hours. After determining the total conversion to the product by thin-layer chromatography (TLC) analysis, purify the reaction mixture by column chromatography technique (petroleum ether: ethyl acetate = 10:1) to obtain 250 mg of compound 4a, white solid, yield 91%, 1H NMR (500 MHz, CDCl3) δ12.73 (s, 1H), 7.89 (d, J = 7.3 Hz, 2H), 7.55 (dq, J = 14.6, 6.9 Hz, 3H), 6.86 (d, J = 2.3 Hz, 1H), 6.75 (s, 1H), 6.66 (d, J = 2.3 Hz, 1H), 1.80 (d, J = 13.3 Hz, 4H), 1.72 - 1.50 (m, 7H), 1.49 - 1.24 (m, 6H), 1.11 (t, J = 12.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 182.8, 169.3, 164.8, 164.7, 161.9, 157.5, 156.8, 132.3, 130.9, 129.2, 126.4, 108.8, 106.3, 106.1, 101.4, 56.5, 56.3, 29.9, 29.3, 26.1, 25.8, 25.3, 24.8.

[0028] Example 2: Preparation of 7-(4-(benzyl(tert-butyl)amino)-6-chloro-1,3,5-triazin-2-yl)oxy)-5-hydroxy-2-phenyl-4H-benzofuran-4-one

[0029]

[0030] The preparation method refers to Example 1. A white solid was obtained with a yield of 90.5%. 1H NMR (500 MHz, CDCl3) δ 12.89 - 12.55 (m, 1H), 7.96 - 7.81 (m, 2H), 7.64 - 7.54 (m, 3H), 7.45 - 6.87 (m, 6H), 6.82 - 6.50 (m, 3H), 5.38 - 4.62 (m, 2H), 2.15 - 0.91 (m, 12H). 13C NMR (126 MHz, CDCl3) δ 183.2, 132.6, 131.3, 128.9, 126.80, 59.7.

[0031] Example 3: Preparation of 7-(4-chloro-6-(cyclohexyl(methyl)amino)-1,3,5-triazin-2-yl)oxy)-5-hydroxy-2-phenyl-4H-benzofuran-4-one

[0032]

[0033] The preparation method refers to Example 1. A white solid was obtained with a yield of 89.1%. 1H NMR (500 MHz, CDCl3) δ 12.76 (s, 1H), 12.73 (s, 1H), 7.89 (d, J = 6.9 Hz, 4H), 7.63 - 7.49 (m, 7H), 6.89 (dd, J = 15.0, 2.1 Hz, 2H), 6.75 (d, J = 2.6 Hz, 2H), 6.69 (dd, J = 8.5, 2.1 Hz, 2H), 3.07 (s, 3H), 2.92 (s, 3H), 1.87 - 1.54 (m, 15H), 1.46 (d, J = 8.2 Hz, 8H). 13C NMR (126 MHz, CDCl3) δ 183.3, 165.2, 162.2, 157.6, 157.2, 132.6, 131.4, 129.6, 126.8, 109.2, 106.6, 106.2, 101.3, 56.0, 55.4, 30.2, 30.0, 29.8, 29.5, 25.9.

[0034] Example 4: Determination of the acetylcholinesterase inhibitory activity of the 13 2-cinnamoylaminothiazole-4-carboxamide derivatives described above in this patent.

[0035] The present invention tested the antitumor activities of 12 chrysin-1,3,5-triazine derivatives against four types of tumor cells.

[0036] The specific test method is as follows: Select MDA-MB-231 (human breast cancer cells), 7901 (human gastric cancer cells), PANC-1 (human pancreatic cancer cells), and A549 (human lung cancer cells) as experimental cell lines. Dissolve 12 chrysin-1,3,5-triazine derivatives (C1-C12) using anhydrous DMSO, and prepare a stock solution of 100 mM for each compound. Further dilute the stock solution using RPMI-1640 medium (containing 0.1% DMSO) to prepare working solutions of different concentrations (e.g., 1 μM, 5 μM, 10 μM, 50 μM, 100 μM, 200 μM). Set at least 3 replicate wells for each concentration of the compound to ensure the accuracy of the data. Add the prepared working solutions into a 96-well plate, and add compounds of different concentrations to each well. Set a negative control group (add the same volume of DMSO solution), and the positive control group can be treated with common anti-tumor drugs (such as cisplatin or docetaxel). After 48 hours of compound treatment, perform MTT assay. Add 20 μL of MTT reagent (5 mg / mL, dissolved in PBS) to each well, and continue to incubate in a 37 °C, 5% CO2 incubator for 4 hours. Aspirate the medium in each well, and add 150 μL of DMSO solution to dissolve the formed purple formazan crystals. Measure the absorbance (OD value) of each well at a wavelength of 570 nm using an ELISA Reader. After dissolution, the OD value is proportional to the number of viable cells. The cell survival rate (%) is calculated by the following formula: Inhibition rate (%) = [(A control - A sample) / A control] × 100%. For each compound, calculate the cell survival rate at different concentrations. Based on the cell survival rate data of chrysin-1,3,5-triazine derivatives at different concentrations, plot a curve of compound concentration vs. cell survival rate. Use software (such as GraphPad Prism) for non-linear regression analysis to fit the concentration-survival curve. The IC50 value refers to the concentration at which the compound concentration reduces the cell survival rate by 50%. Through curve fitting, obtain the IC50 value of each compound. Calculate the IC50 values in the MDA-MB-231, 7901, PANC-1, and A549 cell lines, and judge their anti-tumor activities based on the IC50 values. Among them, the IC50 of compound 4i against breast cancer cells is the highest, which is 15.36 μM.

[0037] The above-described embodiments merely represent several implementation manners of the present invention. The description is relatively specific and detailed, but it should not be construed as a limitation on the scope of the patent of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the concept of the present invention, several modifications and improvements can still be made, and these all belong to the protection scope of the present invention.

Claims

1. Synthesis and application of a series of chrysin-1,3,5-triazine derivatives, the structure is shown as follows:

2. Application of the chrysin-1,3,5-triazine derivative according to claim 1 in the preparation of anti-tumor agents, characterized in that the chrysin-1,3,5-triazine derivative has broad-spectrum anti-tumor inhibitory activity.

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