An organoselenium composition for inhibiting tumor cells and its preparation method

Abstract

This invention belongs to the field of biomedical technology, specifically relating to an organic selenium composition for inhibiting tumor cells and its preparation method. The organic selenium composition comprises the following components in parts by weight: 7-12 parts selenobenzonitrile-glycyrrhizinamide, 2-5 parts trazastilbene, and 0.5-1 part selenized carrageenan. This invention utilizes the reaction of the carbonyl α,β-unsaturated olefin bond in methyl glycyrrhizinic acid with 4-mercaptobenzonitrile to introduce a benzonitrile structure to enhance targeting while retaining the core active structure of methyl glycyrrhizinic acid. Simultaneously, a selenium compound is introduced onto the carboxyl group of methyl glycyrrhizinic acid, thereby preparing selenobenzonitrile-glycyrrhizinamide. When selenobenzonitrile-glycyrrhizinamide is used in combination with trazastilbene and selenized carrageenan, the prepared organic selenium composition exhibits significant inhibitory effects on the proliferation of various tumor cells, while showing no significant toxicity to normal human liver cells (L02).

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to an organic selenium composition for inhibiting tumor cells and its preparation method. Background Technology

[0002] Cancer is a global health problem and one of the leading causes of death. Cancer occurs when normal cells in the body transform into abnormal cells, and these abnormal cells divide and proliferate uncontrollably, forming tumor tissue that obtains nutrients from the host and damages the host's body.

[0003] Currently, the main treatments for cancer include surgery, chemotherapy, and radiation therapy. Surgery is more effective in the early stages of cancer, removing the tumor, but sometimes it may require the removal of the entire organ, leading to side effects and the risk of cancer cell spread. Radiation therapy is effective in treating cancers in specific organs, but it exposes patients to the risk of other cancers and cannot prevent the spread of cancer cells to other organs; patients also experience significant pain during treatment. Chemotherapy uses anticancer drugs, but these drugs not only act on cancer cells but also damage normal cells, leading to severe side effects. Although existing organoselenium compounds have some applications in anticancer treatment, their anticancer efficacy still needs improvement, their anticancer spectrum is limited, and the types of compound structures are insufficient to meet the needs of cancer prevention and treatment.

[0004] Glycyrrhetinic acid can inhibit the proliferation and invasion of tumor cells, induce apoptosis, and exhibit strong cytotoxic activity against tumor cells. It has inhibitory effects on various malignant tumors, including gastric cancer and liver cancer. However, after absorption, glycyrrhetinic acid is concentrated in the liver tissue within a short period, with relatively low concentrations in other organs and the blood. Therefore, to achieve the desired therapeutic effect, the dosage must be increased, which also increases the risk of drug use. Further research is needed to develop new anticancer drugs with higher selectivity for cancer cells and lower toxicity, especially organoselenium compositions that can effectively inhibit tumor cells, which is crucial for cancer prevention and treatment. Summary of the Invention

[0005] To overcome the shortcomings of the prior art, the primary objective of this invention is to provide an organic selenium composition that inhibits tumor cells.

[0006] Another object of the present invention is to provide a method for preparing the above-mentioned organoselenium composition for inhibiting tumor cells.

[0007] The objective of this invention is achieved through the following technical solution:

[0008] An organic selenium composition for inhibiting tumor cells comprises the following components in parts by weight: 2-5 parts trazastilbene, 7-12 parts selenobenzonitrile-glycyrrhizinamide, and 0.5-1 part selenized carrageenan.

[0009] The structural formula of the selenobenzonitrile-glycyrrhizinamide is:

[0010] .

[0011] Furthermore, the selenobenzonitrile-glycyrrhizamide is prepared by the following process:

[0012] (1) Methylated glycyrrhetinic acid was added to N,N-dimethylformamide, followed by 1-hydroxybenzotriazole and N,N'-dicyclohexylcarbodiimide. After reacting for 20-30 min, allylamine was added to continue the reaction. After the reaction was completed, glycyrrhetinic acid was obtained by purification.

[0013] The structural formula of the methylated glycyrrhetinic acid is:

[0014] ;

[0015] The structural formula of the glycyrrhetinic acid is:

[0016] ;

[0017] (2) At -5 to 0°C, the glycyrrhizamide was added to tetrahydrofuran, and then an aqueous solution of benzyltrimethylammonium hydroxide and a decane solution of tert-butyl hydroperoxide were added to react. After the reaction was completed, the glycyrrhizamide was obtained by purification.

[0018] The structural formula of the epoxy glycyrrhizinamide is:

[0019] ;

[0020] (3) Add the epoxy glycyrrhizinamide and 4-mercaptobenzonitrile to water, adjust the pH to 8.5-9 and carry out the reaction; after the reaction is completed, benzonitrile-glycyrrhizinamide is obtained by purification.

[0021] The structural formula of the benzonitrile-glycyrrhizamide is:

[0022] ;

[0023] (4) The benzonitrile-glycyrrhizamide was added to 1,2-dichloroethane, and then selenophenol was added and stirred evenly. The reaction was carried out under ultraviolet light and heating conditions. After the reaction was completed, the selenobenzonitrile-glycyrrhizamide was obtained by purification treatment.

[0024] Further, in step (1), the molar ratio of methylated glycyrrhetinic acid, 1-hydroxybenzotriazole, N,N'-dicyclohexylcarbodiimide, and allylamine is 1:(1.2-1.5):(1.2-1.5):(1.2-1.5); and the reaction continues for 16-24 h.

[0025] Further, in step (2), the molar ratio of glycyrrhetinic acid, benzyltrimethylammonium hydroxide and tert-butylhydrogen peroxide is 1:(2.5-3):(10-12); the reaction temperature is 0-5℃ and the time is 1.5-3 h.

[0026] Further, in step (3), the molar ratio of epoxy glycyrrhizinamide and 4-mercaptobenzonitrile is 1:(1-1.2); the reaction time is 3-5 h.

[0027] Further, in step (4), the molar ratio of benzonitrile-glycyrrhetinic acid and selenophenol is 1:(1.5-2); the heating temperature is 40-55°C, and the reaction time is 10-16 h.

[0028] Furthermore, the organoselenium composition that inhibits tumor cells also includes pharmaceutically acceptable excipients.

[0029] Furthermore, the dosage form of the organoselenium composition that inhibits tumor cells is an injection, tablet, capsule, powder, pill, granule, or oral liquid. This invention does not impose any particular limitation on the preparation method of the dosage form of the organoselenium composition that inhibits tumor cells; conventional methods are acceptable. Preferably, sucrose is added to the organoselenium composition that inhibits tumor cells, and granules are prepared using conventional methods; sucrose and sodium benzoate are added to the organoselenium composition that inhibits tumor cells, and an oral liquid is prepared using conventional methods; the organoselenium composition that inhibits tumor cells is then filled into capsules as capsule powder to obtain capsules.

[0030] The preparation method of the above-mentioned organic selenium composition for inhibiting tumor cells includes the following steps: mixing 2-5 parts of trazastilbene, 7-12 parts of selenobenzonitrile-glycyrrhizamide, and 0.5-1 part of selenized carrageenan evenly to obtain the final product.

[0031] The present invention has the following advantages over the prior art:

[0032] This invention utilizes the reaction of the carbonyl α,β-unsaturated olefin bond in methylated glycyrrhetinic acid with 4-mercaptobenzonitrile to introduce a benzonitrile structure to enhance targeting while retaining the core active structure of methylated glycyrrhetinic acid. Simultaneously, a selenium compound is introduced onto the carboxyl group of methylated glycyrrhetinic acid, thereby preparing selenobenzonitrile-glycyrrhetinamide. When selenobenzonitrile-glycyrrhetinamide is used in combination with trazastilbene and selenized carrageenan, the resulting organoselenium composition exhibits significant inhibitory effects on various human tumor cells and shows no significant toxicity to normal human liver cells (L02). Furthermore, compared to formulations using glycyrrhetinic acid or lacking any of these components, the composition of this invention requires a relatively lower dosage to achieve the same inhibitory effect, thus helping to reduce the risk of drug use. Detailed Implementation

[0033] The technical solution of the present invention will be further described below with reference to specific embodiments. However, those skilled in the art should understand that the following embodiments are only for illustrating the present invention and should not be regarded as limiting the present invention. Specific conditions not specified in the embodiments are performed according to conventional conditions or conditions recommended by the manufacturer. Unless otherwise specified, the reagents or instruments used are all conventional products obtained through commercial channels.

[0034] The selenium content in the selenized carrageenan of this invention is 19-22 mg / g.

[0035] Preparation example: Preparation of methylated glycyrrhetinic acid

[0036]

[0037] Under a nitrogen atmosphere, glycyrrhetinic acid (5 mmol) was added to 20 mL of tetrahydrofuran. After cooling to -78 °C, bis(trimethylsilylamine) lithium (LiHMDS, 15 mmol) was added and the reaction was carried out for 1 h. Then, iodomethane (15 mmol) was added and the reaction was continued at room temperature for another 1 h. The reaction mixture was quenched with saturated ammonium chloride aqueous solution, and the entire mixture was extracted with ethyl acetate. The combined organic layers were washed with water and saturated brine, dried with anhydrous sodium sulfate, filtered, and evaporated under reduced pressure to obtain the intermediate product, which was directly used in the next reaction.

[0038] Under a nitrogen atmosphere, the obtained intermediate was added to 20 mL of tetrahydrofuran, cooled to -78 °C, and then lithium diisopropylamino (LDA, 12 mmol) was added and reacted for 1 h. Iodomethane (15 mmol) was then added and the mixture was heated to room temperature and reacted for another 1 h. The reaction mixture was quenched with saturated ammonium chloride aqueous solution, and the entire mixture was extracted with ethyl acetate. The combined organic layers were adjusted to pH 6.5 with 1 M hydrochloric acid, washed with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (n-heptane / ethyl acetate = 5:1, v / v) to obtain methylated glycyrrhetinic acid. 1 H NMR (C 32 H 50 O4, 400 MHz, DMSO): δ 12.10 (s, 1H), 5.78 (s, 1H), 4.75(s, 1H), 3.38 (t, 1H), 1.90-1.25 (m, 16H), 1.23 (s, 9H), 1.20 (s, 3H), 1.05-0.95 (m, 3H), 0.90 (s, 15H); HRMS(ESI + ): [M+H] + The calculation yields 499.37, and the search yields 499.38. Example 1

[0039] An organic selenium composition for inhibiting tumor cells comprises the following components in parts by weight: 3 parts trazastilbene, 10 parts selenobenzonitrile-glycyrrhizamide, and 0.8 parts selenized carrageenan.

[0040] The preparation process of the selenobenzonitrile-glycyrrhizinamide is as follows:

[0041]

[0042] (1) Methylated glycyrrhetinic acid (10 mmol) was added to anhydrous N,N-dimethylformamide (60 mL), followed by 1-hydroxybenzotriazole (HOBT, 13 mmol) and N,N'-dicyclohexylcarbodiimide (DCC, 13 mmol). After reacting for 25 min, allylamine (CAS: 107-11-9, 14 mmol) was added and reacted for 20 h. The reaction was quenched with an aqueous solution, then extracted with ethyl acetate. The ethyl acetate phase was washed successively with saturated sodium bicarbonate solution and saturated brine, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (eluent: n-hexane / ethyl acetate = 90 / 10, v / v) to obtain glycyrrhetinic acid.

[0043] glycyrrhizamide 1 H NMR (C35 H 55 NO3, 400 MHz, DMSO): δ 9.35 (s, 1H), 5.85 -5.76 (m, 2H), 5.20 - 5.17 (m, 1H), 5.07 - 5.05 (m, 1H), 4.65 (s, 1H), 3.88(d, 2H), 3.54 (t, 1H), 1.90-1.25 (m, 19H), 1.23 (s, 9H), 1.05-0.95 (m, 3H),0.90 (s, 15H); HRMS(ESI + ): [M+H] + The calculation yields 538.42, and the value is found to be 538.42.

[0044] (2) At 0°C, glycyrrhizinamide (10 mmol) was added to tetrahydrofuran (60 mL), followed by 40 w% benzyltrimethylammonium hydroxide aqueous solution (28 mmol) and tert-butyl hydroperoxide (TBHP, 5.5 mol / L, 110 mmol) in n-decane solution. The reaction was carried out at 5°C for 2 h. The reaction solution was quenched with saturated ammonium chloride aqueous solution, then extracted with ethyl acetate. The ethyl acetate phase was washed successively with saturated sodium bicarbonate solution and saturated brine, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (eluent, n-hexane / ethyl acetate = 92 / 8, v / v) to obtain epoxy glycyrrhizinamide.

[0045] Epoxy glycyrrhizinamide 1 H NMR (C 35 H 55 NO4, 400 MHz, DMSO): δ 9.35 (s, 1H),5.85 - 5.82 (m, 1H), 5.20 - 5.17 (m, 1H), 5.07 - 5.05 (m, 1H), 4.65 (s, 1H),3.88 (d, 2H), 3.54 (t, 1H), 3.20 (s, 1H), 1.90-1.27 (m, 21H), 1.20 (s, 3H), 0.95-0.90 (m, 19H), 0.83 (s, 3H); HRMS(ESI + ): [M+H] + The calculation yields 554.41, and the result is 554.43.

[0046] (3) Add epoxy glycyrrhizinamide (10 mmol) and 4-mercaptobenzonitrile (11 mmol) to deionized water (100 mL), then adjust the pH to 8.5 with 1 mol / L sodium hydroxide aqueous solution, and react for 4 h; extract the reaction solution with ethyl acetate, dry the ethyl acetate phase with anhydrous sodium sulfate, concentrate, and purify by column chromatography (eluent, n-hexane / ethyl acetate = 85 / 15, v / v) to obtain benzonitrile-glycyrrhizinamide;

[0047] Benzonitrile-Glycyrrhizamide 1 H NMR (C 42 H 58 N2O3S, 400 MHz, DMSO): δ 9.35 (s, 1H), 7.72 (d, 2H), 7.62 (d, 2H), 5.85 - 5.76 (m, 1H), 5.20 - 5.17 (m, 1H), 5.07 -5.05 (m, 1H), 4.65 (s, 1H), 3.88 (d, 2H), 3.54 (t, 1H), 1.90-1.25 (m, 19H), 1.23 (s, 9H), 1.05-0.95 (m, 3H), 0.90 (s, 15H); HRMS(ESI + ): [M+H] + The calculation yields 671.42, and the value is found to be 671.42.

[0048] (4) Add benzonitrile-glycyrrhizinamide (20 mmol) to 1,2-dichloroethane (200 mL), then add selenophenol (35 mmol) and stir evenly at room temperature. Place the reaction apparatus under the light of an ultraviolet lamp and control the temperature at 45°C for 12 h. Add n-heptane (150 mL) to the reaction solution and stir for 10 min. Filter the solution and beat the filter cake with a mixed solution (n-heptane / acetonitrile / acetone = 10:1.5:1, v / v / v) for 30 min. Filter to obtain selenobenzonitrile-glycyrrhizinamide.

[0049] Selenobenzonitrile-glycyrrhizamide 1 H NMR (C 48 H 64N2O3SSe, 400 MHz, CDCl3): δ 8.03(s, 1H), 7.72 (d, 2H), 7.62 (d, 2H), 7.37-7.33 (m, 5H), 4.65 (s, 1H), 3.54(t, 1H), 3.19 (t, 2H), 1.90-1.26 (m, 23H), 1.23 (s, 9H), 1.05-0.95 (m, 3H),0.90 (s, 15H); HRMS(ESI + ): [M+H] + The calculation yields 829.38, and the value is found to be 829.38.

[0050] Example 1 also provides a method for preparing the above-mentioned organoselenium composition for inhibiting tumor cells, comprising the following steps:

[0051] According to the stated weight proportions, trazastilbene, selenobenzonitrile-glycyrrhetinamide, and selenized carrageenan are mixed evenly to obtain the organic selenium composition that inhibits tumor cells. Example 2

[0052] An organic selenium composition for inhibiting tumor cells comprises the following components in parts by weight: 2 parts trazastilbene, 7 parts selenobenzonitrile-glycyrrhizamide, and 0.5 parts selenized carrageenan.

[0053] The reaction formula for the selenobenzonitrile-glycyrrhizamide is the same as in Example 1, and specifically includes the following steps:

[0054] (1) Methylated glycyrrhetinic acid (10 mmol) was added to anhydrous N,N-dimethylformamide (70 mL), followed by HOBT (12 mmol) and DCC (12 mmol). After reacting for 20 min, allylamine (12 mmol) was added and the reaction was allowed to proceed for 16 h. The reaction was quenched with an aqueous solution, then extracted with ethyl acetate. The ethyl acetate phase was washed successively with saturated sodium bicarbonate solution and saturated brine, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (eluent: n-hexane / ethyl acetate = 90 / 10, v / v) to obtain glycyrrhetinic acid. 1 H NMR and HRMS (ESI) + (Same as Example 1)

[0055] (2) At -5°C, glycyrrhizinamide (10 mmol) was added to tetrahydrofuran (60 mL), followed by 40 wt% benzyltrimethylammonium hydroxide aqueous solution (25 mmol) and tert-butyl hydroperoxide solution in n-decane (5.5 mol / L, 100 mmol). The reaction was carried out at 0°C for 1.5 h. The reaction solution was quenched with saturated ammonium chloride aqueous solution, then extracted with ethyl acetate. The ethyl acetate phase was washed successively with saturated sodium bicarbonate solution and saturated brine, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (eluent: n-hexane / ethyl acetate = 92 / 8, v / v) to obtain epoxy-based glycyrrhizinamide. 1 H NMR and HRMS (ESI) + (Same as Example 1)

[0056] (3) Epoxy glycyrrhizinamide (10 mmol) and 4-mercaptobenzonitrile (10 mmol) were added to deionized water (110 mL), and the pH was adjusted to 8.5 with 1 mol / L sodium hydroxide aqueous solution. The reaction was allowed to proceed for 3 h. The reaction solution was extracted with ethyl acetate, and the ethyl acetate phase was dried with anhydrous sodium sulfate. The solution was concentrated and purified by column chromatography (eluent: n-hexane / ethyl acetate = 85 / 15, v / v) to obtain benzonitrile-glycyrrhizinamide. 1 H NMR and HRMS (ESI) + (Same as Example 1)

[0057] (4) Add benzonitrile-glycyrrhizinamide (20 mmol) to 1,2-dichloroethane (200 mL), then add selenophenol (30 mmol) and stir evenly at room temperature. Place the reaction apparatus under ultraviolet light and control the temperature at 55°C for 10 h. Add n-heptane (150 mL) to the reaction solution, stir for 10 min, and filter. Pulverize the filter cake with a mixed solution (n-heptane / acetonitrile / acetone = 10:1.5:1, v / v / v) for 30 min, and filter to obtain selenobenzonitrile-glycyrrhizinamide; the selenobenzonitrile-glycyrrhizinamide 1 H NMR and HRMS (ESI) + (Same as Example 1)

[0058] Example 2 also provides a method for preparing the above-mentioned organoselenium composition for inhibiting tumor cells, the same as in Example 1. Example 3

[0059] An organic selenium composition for inhibiting tumor cells comprises the following components in parts by weight: 5 parts trazastilbene, 12 parts selenobenzonitrile-glycyrrhizamide, and 1 part selenized carrageenan.

[0060] The reaction formula for the selenobenzonitrile-glycyrrhizamide is the same as in Example 1, and specifically includes the following steps:

[0061] (1) Methylated glycyrrhetinic acid (10 mmol) was added to anhydrous N,N-dimethylformamide (80 mL), followed by HOBT (15 mmol) and DCC (15 mmol). After reacting for 30 min, allylamine (15 mmol) was added and the reaction was allowed to proceed for 24 h. The reaction was quenched with an aqueous solution, then extracted with ethyl acetate. The ethyl acetate phase was washed successively with saturated sodium bicarbonate solution and saturated brine, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (eluent: n-hexane / ethyl acetate = 90 / 10, v / v) to obtain glycyrrhetinic acid. 1 H NMR and HRMS (ESI) + (Same as Example 1)

[0062] (2) At 0°C, glycyrrhizinamide (10 mmol) was added to 60 mL of tetrahydrofuran, followed by 40 wt% aqueous solution of benzyltrimethylammonium hydroxide (30 mmol) and a decane solution of tert-butyl hydroperoxide (5.5 mol / L, 120 mmol). The reaction was carried out at 0°C for 3 h. The reaction solution was quenched with saturated ammonium chloride aqueous solution, then extracted with ethyl acetate. The ethyl acetate phase was washed successively with saturated sodium bicarbonate solution and saturated brine, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (eluent: hexane / ethyl acetate = 92 / 8, v / v) to obtain epoxy-based glycyrrhizinamide. 1 H NMR and HRMS (ESI) + (Same as Example 1)

[0063] (3) Epoxy glycyrrhizinamide (10 mmol) and 4-mercaptobenzonitrile (12 mmol) were added to deionized water (120 mL), and the pH was adjusted to 9 with 1 mol / L sodium hydroxide aqueous solution. The reaction was allowed to proceed for 5 h. The reaction solution was extracted with ethyl acetate, and the ethyl acetate phase was dried with anhydrous sodium sulfate. The solution was concentrated and purified by column chromatography (eluent: n-hexane / ethyl acetate = 85 / 15, v / v) to obtain benzonitrile-glycyrrhizinamide. 1 H NMR and HRMS (ESI) + (Same as Example 1)

[0064] (4) Add benzonitrile-glycyrrhizinamide (20 mmol) to 1,2-dichloroethane (200 mL), then add selenophenol (40 mmol) and stir evenly at room temperature. Place the reaction apparatus under ultraviolet light and control the temperature at 40°C for 10 h. Add n-heptane (150 mL) to the reaction solution, stir for 10 min, and filter. Pulverize the filter cake with a mixed solution (n-heptane / acetonitrile / acetone = 10:1.5:1, v / v / v) for 30 min, and filter to obtain selenobenzonitrile-glycyrrhizinamide; selenobenzonitrile-glycyrrhizinamide 1 H NMR and HRMS (ESI) + (Same as Example 1)

[0065] Example 3 also provides a method for preparing the above-mentioned organoselenium composition for inhibiting tumor cells, the same as in Example 1.

[0066] Comparative Example 1

[0067] Comparative Example 1 is basically the same as Example 1, except that the selenobenzonitrile-glycyrrhetinic acid in Example 1 is replaced with glycyrrhetinic acid.

[0068] Comparative Example 2

[0069] Comparative Example 2 is basically the same as Example 1, except that trazastilbene in Example 1 is replaced with selenized carrageenan.

[0070] Experimental Example 1

[0071] Antitumor activity test of the selenobenzonitrile-glycyrrhizinamide prepared in this invention

[0072] 1 mg of the selenobenzonitrile-glycyrrhizinamide prepared in Example 1 was added to 200 μL of DMSO to prepare a stock solution of 5 mg / mL. This stock solution was then serially diluted with fresh DMEM culture medium (containing 800,000 units of penicillin, 1.0 g of streptomycin, and 10% inactivated fetal bovine serum per 1000 mL of medium) to prepare test sample solutions with selenobenzonitrile-glycyrrhizinamide concentrations of 0.5, 1, 2, 4, 8, 16, 32, 64, 128, and 256 μg / mL. Gefitinib sample solutions with concentrations of 0.5, 1, 2, 4, 8, 16, 32, 64, 128, and 256 μg / mL were prepared using the same method.

[0073] Human cervical cancer HeLa cells, human liver cancer HepG2 cells, human gastric cancer BGC-823 cells, human lung cancer A549 cells, and normal human hepatocytes L02 were cultured in DMEM medium containing 10% inactivated fetal bovine serum and incubated at 37°C in a 5% CO2 incubator. The cells grew in an adherent state. HeLa cells, HepG2 cells, BGC-823 cells, A549 cells, and L02 cells in logarithmic growth phase were seeded at 1×10⁻⁶ cells per cell line. 4 Cells were cultured in 96-well plates and, after 24 h of growth, were treated according to the following groups: Experimental groups received 100 μL of different concentrations of selenobenzonitrile-glycyrrhizinamide or gefitinib sample solution, with three replicates for each concentration; the negative control group received 100 μL of DMEM culture medium without selenobenzonitrile-glycyrrhizinamide or gefitinib; the blank control group received only DMEM culture medium without cells; and the solvent control group received 100 μL of DMEM culture medium containing the same volume of DMSO as the experimental group (256 μg / mL). After 72 h of incubation, the supernatant was discarded, and 100 μL of serum-free MTT culture medium containing 0.5 mg / mL was added to each well. The cells were cultured for another 4 h, centrifuged after incubation, and the supernatant was carefully discarded. 150 μL of dimethyl sulfoxide was added to each well. The absorbance (A570) of each well at 570 nm was measured using an ELISA reader. The average value of each group was taken, and the inhibition rate was calculated as follows: Inhibition rate = [1 - (A570 of each experimental group - A570 of the blank control group) / (A570 of the solvent control group - A570 of the blank control group)] × 100%. The half-maximal inhibitory concentration (IC50) of the sample for tumor cell growth was calculated using the logit method. 50 The data are expressed as mean ± standard deviation (Mean ± SD, n=3), and the specific results are shown in Table 1.

[0074] Table 1

[0075]

[0076] As can be seen from Table 1, the selenobenzonitrile-glycyrrhizinamide prepared in this invention has a strong inhibitory effect on the proliferation of human cervical cancer HeLa cells, human liver cancer HepG2 cells, human gastric cancer BGC-823 cells, and human lung cancer A549 cells, while its inhibitory effect on normal human cells L02 is relatively small, showing obvious selectivity.

[0077] Test Example 2: Antitumor Activity Test of the Organoselenium Composition Prepared in This Invention

[0078] 1 mg of each of the compositions prepared in Examples 1-3 and Comparative Examples 1-2 was added to 200 μL of DMSO to prepare a stock solution of 5 mg / mL. Before the test, the stock solution was continuously diluted with fresh DMEM culture medium (containing 800,000 units of penicillin, 1.0 g of streptomycin, and 10% inactivated fetal bovine serum per 1000 mL of culture medium) to prepare test sample solutions with composition concentrations of 0.5, 1, 2, 4, 8, 16, 32, 64, 128, and 256 μg / mL.

[0079] The tumor cell growth inhibition rate of each group of compositions was detected using the method described in Example 1. The half-maximal inhibitory concentration (IC50) of the samples for tumor cell growth was calculated using the logit method. 50 The data are expressed as mean ± standard deviation (Mean ± SD, n=3), and the results are shown in Table 2.

[0080] Table 2

[0081]

[0082] As can be seen from Table 2, the organic selenium compositions prepared in Examples 1-3 of this invention have significant inhibitory activity against the proliferation of various human tumor cells, and have a relatively small inhibitory effect on human normal cells LO2.

[0083] Compared to Example 1, Comparative Example 1 replaced selenobenzonitrile-glycyrrhetinic acid with glycyrrhetinic acid, and Comparative Example 2 replaced trazastilbene in Example 1 with selenized carrageenan. 50 The values ​​are all higher than those in Example 1, which indicates that the selenobenzonitrile-glycyrrhetinic acid, trazastilbene, and selenized carrageenan in the organic selenium composition of the present invention work synergistically. On the one hand, it enhances the inhibitory effect on tumor cells to a certain extent, and on the other hand, its dosage is reduced compared to the formulation using glycyrrhetinic acid or lacking any of the components, thereby helping to reduce the risk of medication.

[0084] This invention utilizes the reaction of the carbonyl α,β-unsaturated olefin bond in methylated glycyrrhetinic acid with 4-mercaptobenzonitrile to introduce a benzonitrile structure to enhance targeting while retaining the core active structure of methylated glycyrrhetinic acid. Simultaneously, a selenium compound is introduced onto the carboxyl group of methylated glycyrrhetinic acid, thereby preparing selenobenzonitrile-glycyrrhetinamide. When selenobenzonitrile-glycyrrhetinamide is used in combination with trazastilbene and selenized carrageenan, the resulting organic selenium composition significantly reduces the IC50 of various tumor cells. 50 It has no significant toxicity to normal hepatocytes L02, and compared to formulations using glycyrrhetinic acid or lacking any of the components, the composition of the present invention requires a reduced dosage to achieve the same inhibitory effect, which helps to reduce the risk of medication.

[0085] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them. The basic principles and main features of the present invention have been described above with specific implementation schemes. Based on the present invention, some modifications or substitutions can be made, but these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of protection claimed by the present invention.

Claims

1. An organoselenium composition for inhibiting tumor cells, characterized in that, It is composed of the following components by weight: 7-12 parts of seleno-benzonitrile-glycyrrhetamide, 2-5 parts of trizygocannabinoid and 0.5-1 part of selenized carrageenan; The structural formula of the seleno-benzonitrile-glycyrrhetamide is: ； The tumor cells are Hela cells, HepG2 cells, BGC-823 cells or A549 cells.

2. The organic selenium composition for inhibiting tumor cells according to claim 1, wherein The seleno-benzonitrile-glycyrrhetamide is prepared by the following preparation process: (1) methylated glycyrrhetinic acid is added into N,N-dimethylformamide, then 1-hydroxybenzotriazole and N,N'-dicyclohexyl carbodiimide are added, after 20-30 min of reaction, allylamine is added for further reaction; after the reaction is completed, glycyrrhetamide is obtained by purification treatment; The structural formula of the methylated glycyrrhetinic acid is: ； The structural formula of the glycyrrhetamide is: ； The purification treatment steps are: adding an aqueous solution to quench the reaction, then extracting with ethyl acetate, then washing the ethyl acetate phase with saturated sodium bicarbonate solution and saturated brine, drying with anhydrous sodium sulfate, concentrating, and purifying by column chromatography; (2) at-5-0℃, the glycyrrhetamide is added into tetrahydrofuran, then an aqueous solution of benzyltrimethylammonium hydroxide and a solution of tert-butyl hydroperoxide in n-decane are added for reaction; after the reaction is completed, epoxy glycyrrhetamide is obtained by purification treatment; The structural formula of the epoxy glycyrrhetamide is: ； The purification treatment steps are: quenching the reaction liquid with saturated ammonium chloride aqueous solution, then extracting with ethyl acetate, then washing the ethyl acetate phase with saturated sodium bicarbonate solution and saturated brine, drying with anhydrous sodium sulfate, concentrating, and purifying by column chromatography; (3) the epoxy glycyrrhetamide and 4-mercaptobenzonitrile are added into water, and the pH is adjusted to 8.5-9 for reaction; after the reaction is completed, benzonitrile-glycyrrhetamide is obtained by purification treatment; The structural formula of the benzonitrile-glycyrrhetamide is: ； The purification treatment steps are: extracting the reaction liquid with ethyl acetate, drying the ethyl acetate phase with anhydrous sodium sulfate, concentrating, and purifying by column chromatography; (4) the benzonitrile-glycyrrhetamide is added into 1,2-dichloroethane, then phenyl selenol is added and stirred uniformly, and the reaction is carried out under the conditions of ultraviolet light irradiation and heating; after the reaction is completed, the seleno-benzonitrile-glycyrrhetamide is obtained by purification treatment; The purification treatment steps are: adding n-heptane to the reaction liquid and stirring, then filtering, and then beating the filter cake with a mixed solution of n-heptane / acetonitrile / acetone, and filtering.

3. The organic selenium composition for inhibiting tumor cells according to claim 2, wherein In step (1), the molar ratio of the methylated glycyrrhetinic acid, 1-hydroxybenzotriazole, N,N'-dicyclohexyl carbodiimide and allylamine is 1:(1.2-1.5):(1.2-1.5):(1.2-1.5); the time for further reaction is 16-24 h.

4. The organic selenium composition for inhibiting tumor cells according to claim 2, wherein In step (2), the molar ratio of the glycyrrhetamide, benzyltrimethylammonium hydroxide and tert-butyl hydroperoxide is 1:(2.5-3):(10-12); the reaction temperature is 0-5℃, and the reaction time is 1.5-3 h.

5. The organic selenium composition for inhibiting tumor cells according to claim 2, wherein In step (3), the molar ratio of the epoxy glycyrrhetamide and 4-mercaptobenzonitrile is 1:(1-1.2); the reaction time is 3-5 h.

6. The organic selenium composition for inhibiting tumor cells according to claim 2, wherein The molar ratio of the benzonitrile-glycyrrhetamide and phenylselenol in step (4) is 1:(1.5-2); the heating temperature is 40-55℃, and the reaction time is 10-16 h.

7. The organic selenium composition of claim 1, wherein the organic selenium composition is administered in an amount of 0.1 to 10 mg / kg per day. The organic selenium composition for inhibiting tumor cells also comprises a pharmaceutically acceptable adjuvant.

8. The organic selenium composition of claim 1, wherein, The dosage form of the organic selenium composition for inhibiting tumor cells is injection, tablet, capsule, powder, pill, granule or oral liquid.

9. A method for preparing the organic selenium composition for inhibiting tumor cells according to any one of claims 1 to 8, characterized by, The following steps are included: uniformly mixing 2-5 parts of kuazhaquinoside, 7-12 parts of selenobenzonitrile-glycyrrhetamide and 0.5-1 part of selenized carrageenan, to obtain the product.

Images