A kind of mitomycin C prodrug and its preparation method and application

Abstract

The present invention provides a prodrug of mitomycin C and its preparation method and application. The prodrug of mitomycin C of the present invention uses a 4-azido-2,3,5,6-tetrafluorobenzyl alcohol structure as a response group. Compared with the original drug of mitomycin C, the prodrug can obtain semi-active targeting with the guidance of X-rays in the combination of radiotherapy and chemotherapy, and can reduce the damage to normal cells throughout the body without sacrificing the therapeutic effect, and reduce the adverse reactions of patients; compared with the antibody-coupled drug with the same targeting, the semi-active targeting of the prodrug of mitomycin C of the present invention comes from the X-rays emitted by the radiotherapy equipment, can be directly controlled artificially, does not require cancer cells to have specific antigenic properties, and has a wider range of applications; at the same time, the prodrug of mitomycin C of the present invention is a relatively simple small molecule compound with stable chemical properties, and has significant advantages over antibody-coupled drugs in terms of production, storage and transportation costs.

Description

Technical Field

[0001] The present invention relates to the technical field of anticancer drugs, and in particular to a mitomycin C prodrug and a preparation method and application thereof. Background Art

[0002] Cancer is a general term for a series of diseases involving abnormal cell growth that may invade or spread to other parts of the body. As life expectancy increases, the incidence of cancer is also increasing. Chemotherapy and radiotherapy are the two most commonly used cancer treatments. Chemotherapy uses chemical drugs to kill cancer cells or inhibit their growth. Most chemotherapy drugs have systemic effects. While killing cancer cells, the drugs also cause damage to normal cells throughout the body; radiotherapy uses high doses of ionizing radiation to irradiate the tumor area, producing substances such as oxygen free radicals that damage the DNA of cancer cells, thereby killing cancer cells or inhibiting their growth. Commonly used external beam radiotherapy equipment can now achieve precise control of the radiation dose around the tumor. However, since the tumor microenvironment is often hypoxic, substances such as oxygen free radicals produced by radiotherapy are easily reduced by reducing substances in the tumor microenvironment, resulting in some types of cancer cells being insensitive to radiotherapy.

[0003] Mitomycin C is a cell cycle non-specific drug with a broad anti-tumor spectrum and rapid action, but a low therapeutic index and high systemic toxicity. Based on the defects of mitomycin C, it is necessary to improve it. Summary of the invention

[0004] In view of this, the present invention provides a prodrug of mitomycin C and a preparation method and application thereof, so as to solve or at least partially solve the problems existing in the prior art.

[0005] In a first aspect, the present invention provides a prodrug of mitomycin C, the structural formula of which is as follows:

[0006]

[0007] In a second aspect, the present invention also provides a method for preparing the mitomycin C prodrug, comprising the following steps:

[0008] Utilizing the reaction of 4-azido-2,3,5,6-tetrafluorobenzyl alcohol and 4-nitrophenyl chloroformate to generate (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate;

[0009] The prodrug of mitomycin C is prepared by reacting (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate with the methylene group of mitomycin C.

[0010] Preferably, the preparation method of the mitomycin C prodrug, the preparation method of 4-azido-2,3,5,6-tetrafluorobenzyl alcohol is:

[0011] Reducing 4-amino-2,3,5,6-tetrafluorobenzoic acid to 4-amino-2,3,5,6-tetrafluorobenzyl alcohol;

[0012] Reacting 4-amino-2,3,5,6-tetrafluorobenzyl alcohol with sodium azide or potassium azide to generate 4-azido-2,3,5,6-tetrafluorobenzyl alcohol;

[0013] Or, the preparation method of the 4-azido-2,3,5,6-tetrafluorobenzyl alcohol is:

[0014] Reducing 2,3,4,5,6-pentafluorobenzaldehyde to obtain 2,3,4,5,6-pentafluorobenzyl alcohol;

[0015] 2,3,4,5,6-Pentafluorobenzyl alcohol is reacted with trimethylsilyl azide to give 4-azido-2,3,5,6-tetrafluorobenzyl alcohol.

[0016] Preferably, the method for preparing the mitomycin C prodrug, reducing 4-amino-2,3,5,6-tetrafluorobenzoic acid to 4-amino-2,3,5,6-tetrafluorobenzyl alcohol specifically comprises:

[0017] Lithium aluminum hydride is added to the first organic solvent, and then 4-amino-2,3,5,6-tetrafluorobenzoic acid is added to obtain 4-amino-2,3,5,6-tetrafluorobenzyl alcohol through reaction.

[0018] Preferably, the method for preparing the mitomycin C prodrug is to react 4-amino-2,3,5,6-tetrafluorobenzyl alcohol with sodium azide or potassium azide to generate 4-azido-2,3,5,6-tetrafluorobenzyl alcohol as follows:

[0019] Dissolve 4-amino-2,3,5,6-tetrafluorobenzyl alcohol and p-toluenesulfonic acid monohydrate in water, then cool to -5 to 0°C, add sodium azide or potassium azide, continue stirring and then add sodium nitrite, and after reaction, 4-azido-2,3,5,6-tetrafluorobenzyl alcohol is obtained.

[0020] Preferably, the preparation method of the mitomycin C prodrug is to react 4-azido-2,3,5,6-tetrafluorobenzyl alcohol with 4-nitrophenyl chloroformate to generate (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate as follows:

[0021] Add 4-azido-2,3,5,6-tetrafluorobenzyl alcohol to the second organic solvent, then add p-nitrophenyl chloroformate, cool to -5 to 0°C, add pyridine, continue stirring, and react at 20 to 30°C to obtain (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate;

[0022] The mitomycin C prodrug is generated by reacting (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate with the methylene group of mitomycin C:

[0023] Mitomycin C is added to a third organic solvent, and then (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate is added, and triethylamine is added after stirring, and the mixture is reacted at 20-30° C. to obtain a mitomycin C prodrug.

[0024] Preferably, in the method for preparing the mitomycin C prodrug, the mass volume ratio of lithium aluminum hydride, 4-amino-2,3,5,6-tetrafluorobenzoic acid and the first organic solvent is (0.1-0.3) g: (0.8-1.2) g: (15-25) mL;

[0025] The first organic solvent is tetrahydrofuran.

[0026] Preferably, in the method for preparing the mitomycin C prodrug, the mass volume ratio of the 4-amino-2,3,5,6-tetrafluorobenzyl alcohol, p-toluenesulfonic acid monohydrate, sodium azide or potassium azide, sodium nitrite and water is (0.8-1.2) g: (8-9) g: (0.3-0.7) g: (0.8-1.2) g: (15-25) mL.

[0027] Preferably, in the method for preparing the mitomycin C prodrug, the mass volume ratio of the 4-azido-2,3,5,6-tetrafluorobenzyl alcohol, p-nitrophenyl chloroformate, pyridine, and the second organic solvent is (0.8-1.2) g: (1.5-2.0) g: (3-4) g: (15-25) mL;

[0028] The mass volume ratio of the mitomycin C, (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate, triethylamine, and the third organic solvent is (0.8-1.2) g: (1-2) g: (1-2) mL: (40-60) mL;

[0029] The second organic solvent is tetrahydrofuran;

[0030] The third organic solvent is N,N-dimethylformamide.

[0031] In a third aspect, the present invention further provides a use of the mitomycin C prodrug or the mitomycin C prodrug prepared by the preparation method in the preparation of anticancer drugs.

[0032] The mitomycin C prodrug and the preparation method thereof of the present invention have the following technical effects compared with the prior art:

[0033] The mitomycin C prodrug of the present invention uses a 4-azido-2,3,5,6-tetrafluorobenzyl alcohol structure as a response group. Compared with the mitomycin C original drug, the prodrug can obtain semi-active targeting under the guidance of X-rays in the combination of radiotherapy and chemotherapy, and can reduce damage to normal cells throughout the body without sacrificing the treatment effect, thereby reducing the adverse reactions of patients. Compared with antibody-coupled drugs with the same targeting, the semi-active targeting of the X-ray-activated mitomycin C prodrug of the present invention comes from the X-rays emitted by the radiotherapy equipment, can be directly controlled artificially, does not require cancer cells to have specific antigenic properties, and has a wider range of applications. At the same time, since the mitomycin C prodrug of the present application is a relatively simple small molecule compound, the chemical properties are quite stable, and the dry powder can even be stored at room temperature, it has significant advantages over antibody-coupled drugs in terms of production, storage and transportation costs. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0035] Figure 1 is the hydrogen nuclear magnetic resonance spectrum of the mitomycin C prodrug prepared in Example 1 of the present invention;

[0036] Figure 2 is the mass spectrum of the mitomycin C prodrug prepared in Example 1 of the present invention;

[0037] Figure 3 is a high performance liquid chromatogram of the mitomycin C prodrug prepared in Example 1 of the present invention;

[0038] Figure 4 This is a diagram of the mitomycin C prodrug prepared in Example 1 of the present invention placed in phosphate buffered saline (PBS) and irradiated with 60Gy X-rays by a linear accelerator;

[0039] Figure 5The cytotoxicity comparison chart of the mitomycin C prodrug (MMC-P) and mitomycin C original drug (MMC) prepared in Example 1 is shown. DETAILED DESCRIPTION

[0040] In order to make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Generally, the components of the embodiments of the present invention described and shown in the drawings here can be arranged and designed in various different configurations.

[0041] Therefore, the following detailed description of the embodiments of the present invention provided in the accompanying drawings is not intended to limit the scope of the invention claimed for protection, but merely represents selected embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0042] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0043] The present application provides a prodrug of mitomycin C, the structural formula of which is shown below:

[0044]

[0045] The mitomycin C prodrug of the present application uses a 4-azido-2,3,5,6-tetrafluorobenzyl alcohol structure as a response group. Compared with the original mitomycin C drug, the prodrug can obtain semi-active targeting with the guidance of X-rays in the combination of radiotherapy and chemotherapy, and can reduce damage to normal cells throughout the body without sacrificing the therapeutic effect, and reduce the adverse reactions of patients; the mitomycin C prodrug of the present application can reduce the cytotoxicity of mitomycin C before irradiation, decompose after irradiation to produce mitomycin C, restore the original toxicity, and the killing effect on cancer cells is the same as that of untreated mitomycin C; compared with antibody-coupled drugs with the same targeting, the semi-active targeting of the X-ray-activated mitomycin C prodrug of the present application comes from the X-rays emitted by the radiotherapy equipment, can be directly controlled artificially, does not require cancer cells to have specific antigenic properties, and has a wider range of applications; at the same time, since the mitomycin C prodrug of the present application is a relatively simple small molecule compound, the chemical properties are quite stable, and the dry powder can even be stored at room temperature, it has significant advantages over antibody-coupled drugs in terms of production, storage and transportation costs.

[0046] Based on the same inventive concept, the present application also provides a method for preparing the above-mentioned mitomycin C prodrug, comprising the following steps:

[0047] S1, using 4-azido-2,3,5,6-tetrafluorobenzyl alcohol to react with 4-nitrophenyl chloroformate to generate (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate;

[0048] S2. Using (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate to react with the methylene group of mitomycin C to generate a mitomycin C prodrug.

[0049] In some embodiments, the preparation method of 4-azido-2,3,5,6-tetrafluorobenzyl alcohol is:

[0050] Reducing 4-amino-2,3,5,6-tetrafluorobenzoic acid to 4-amino-2,3,5,6-tetrafluorobenzyl alcohol;

[0051] Reacting 4-amino-2,3,5,6-tetrafluorobenzyl alcohol with sodium azide or potassium azide to generate 4-azido-2,3,5,6-tetrafluorobenzyl alcohol;

[0052] Or, the preparation method of 4-azido-2,3,5,6-tetrafluorobenzyl alcohol is:

[0053] Reducing 2,3,4,5,6-pentafluorobenzaldehyde to obtain 2,3,4,5,6-pentafluorobenzyl alcohol;

[0054] 2,3,4,5,6-Pentafluorobenzyl alcohol is reacted with trimethylsilyl azide to give 4-azido-2,3,5,6-tetrafluorobenzyl alcohol.

[0055] In some embodiments, reducing 4-amino-2,3,5,6-tetrafluorobenzoic acid to 4-amino-2,3,5,6-tetrafluorobenzyl alcohol specifically comprises:

[0056] Lithium aluminum hydride is added to the first organic solvent, and then 4-amino-2,3,5,6-tetrafluorobenzoic acid is added to obtain 4-amino-2,3,5,6-tetrafluorobenzyl alcohol through reaction.

[0057] In some embodiments, 4-amino-2,3,5,6-tetrafluorobenzyl alcohol is reacted with sodium azide or potassium azide to generate 4-azido-2,3,5,6-tetrafluorobenzyl alcohol:

[0058] Dissolve 4-amino-2,3,5,6-tetrafluorobenzyl alcohol and p-toluenesulfonic acid monohydrate in water, then cool to -5 to 0°C, add sodium azide or potassium azide, continue stirring and then add sodium nitrite, and after reaction, 4-azido-2,3,5,6-tetrafluorobenzyl alcohol is obtained.

[0059] In some embodiments, the reaction of 4-azido-2,3,5,6-tetrafluorobenzyl alcohol with 4-nitrophenyl chloroformate to generate (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate is specifically:

[0060] Add 4-azido-2,3,5,6-tetrafluorobenzyl alcohol to the second organic solvent, then add p-nitrophenyl chloroformate, cool to -5 to 0°C, add pyridine, continue stirring, and react at 20 to 30°C to obtain (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate;

[0061] The mitomycin C prodrug is generated by reacting (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate with the methylene group of mitomycin C:

[0062] Mitomycin C is added to a third organic solvent, and then (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate is added, and triethylamine is added after stirring, and the mixture is reacted at 20-30° C. to obtain a mitomycin C prodrug.

[0063] In some embodiments, the mass volume ratio of lithium aluminum hydride, 4-amino-2,3,5,6-tetrafluorobenzoic acid and the first organic solvent is (0.1-0.3) g: (0.8-1.2) g: (15-25) mL;

[0064] The first organic solvent is tetrahydrofuran.

[0065] In some embodiments, the mass volume ratio of 4-amino-2,3,5,6-tetrafluorobenzyl alcohol, p-toluenesulfonic acid monohydrate, sodium azide or potassium azide, sodium nitrite and water is (0.8-1.2) g:(8-9) g:(0.3-0.7) g:(0.8-1.2) g:(15-25) mL.

[0066] In some embodiments, the mass volume ratio of 4-azido-2,3,5,6-tetrafluorobenzyl alcohol, p-nitrophenyl chloroformate, pyridine, and the second organic solvent is (0.8-1.2) g: (1.5-2.0) g: (3-4) g: (15-25) mL;

[0067] The mass volume ratio of mitomycin C, (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate, triethylamine, and the third organic solvent is (0.8-1.2) g: (1-2) g: (1-2) mL: (40-60) mL;

[0068] The second organic solvent is tetrahydrofuran;

[0069] The third organic solvent is N,N-dimethylformamide.

[0070] Specifically, in some embodiments, the synthesis method of 4-amino-2,3,5,6-tetrafluorobenzyl alcohol comprises the following steps:

[0071] Add 0.1-0.3 g of lithium aluminum hydride to 10-20 mL of tetrahydrofuran and stir for 5-15 minutes to obtain a first solution;

[0072] Add 0.8-1.2 g of 4-amino-2,3,5,6-tetrafluorobenzoic acid to 3-7 mL of tetrahydrofuran and stir to obtain a second solution;

[0073] The second solution is added to the first solution, and heated under reflux for 10 to 15 hours at a reflux temperature of 66 to 70°C;

[0074] After heating to reflux, tetrahydrofuran (recoverable) is removed by rotary evaporation, and then 3-7 mL of water is added to quench the excess lithium aluminum hydride, and then 15-25 mL of ethyl acetate is added to dissolve the product, filtered, and the precipitate is washed twice with 15-25 mL of ethyl acetate, the organic phases are combined, and the organic phases are washed twice with 15-25 mL of saturated sodium bicarbonate aqueous solution, and dried with anhydrous sodium sulfate, and ethyl acetate is removed by rotary evaporation at 45-55° C. After cooling, the product naturally crystallizes, which is 4-amino-2,3,5,6-tetrafluorobenzyl alcohol.

[0075] Specifically, the synthesis reaction formula of 4-amino-2,3,5,6-tetrafluorobenzyl alcohol is as follows:

[0076]

[0077] Specifically, in some embodiments, the synthesis method of 4-azido-2,3,5,6-tetrafluorobenzyl alcohol comprises the following steps:

[0078] Dissolve 0.8-1.2 g of 4-amino-2,3,5,6-tetrafluorobenzyl alcohol and 8-9 g of p-toluenesulfonic acid monohydrate (or p-toluenesulfonic acid) in 10-20 mL of water, cool to 0° C. after dissolution, add 0.5 g of sodium azide, and stir for 10-20 min in an ice bath (i.e., at 0° C.) to obtain a third solution;

[0079] Dissolve 0.8-1.2 g of sodium nitrite in 3-7 mL of water to obtain a fourth solution;

[0080] The fourth solution was added dropwise to the third solution within 10 minutes, and the reaction was continued under stirring in an ice bath for 15 to 25 minutes;

[0081] After the reaction is completed, 5 to 15 mL of saturated sodium carbonate aqueous solution is added to the reaction system, the ice bath is removed, and solid sodium carbonate is continued to be added until the pH is 8, and 40 to 50 mL of ethyl acetate is used for three extractions, the organic phases are combined, and the organic phases are washed with 15 to 25 mL of saturated sodium bicarbonate aqueous solution, and dried with anhydrous sodium sulfate. The ethyl acetate is removed by rotary evaporation at 25 to 35° C., and the solvent and water are pumped out with an oil pump to obtain a solid product, namely 4-azido-2,3,5,6-tetrafluorobenzyl alcohol.

[0082] Specifically, the synthesis reaction formula of 4-azido-2,3,5,6-tetrafluorobenzyl alcohol is as follows:

[0083]

[0084] Specifically, in some embodiments, the synthesis method of (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate is:

[0085] Dissolve 0.8-1.2 g of 4-azido-2,3,5,6-tetrafluorobenzyl alcohol in 5-15 mL of tetrahydrofuran to obtain a fifth solution;

[0086] Add 1.5-2.0 g of p-nitrophenyl chloroformate to 5-15 mL of tetrahydrofuran to obtain a sixth solution;

[0087] The sixth solution was added to the fifth solution, mixed and cooled to 0°C, 3-4 g of pyridine was added, stirred for 5-15 min, and then heated to 20-30°C for reaction for 24 h;

[0088] After the reaction is completed, tetrahydrofuran is removed by rotary evaporation at 25 to 35° C., 45 to 55 mL of water is added, and 70 to 80 mL of ethyl acetate is used for three extractions, the organic phases are combined, and the organic phases are washed four times with 50 to 150 mL of saturated sodium carbonate aqueous solution, dried over anhydrous sodium sulfate, and ethyl acetate is removed by rotary evaporation at 25 to 35° C. The product is purified by column chromatography, namely (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate.

[0089] Specifically, the synthesis reaction formula of (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate is as follows:

[0090]

[0091] Specifically, in some embodiments, the synthesis method of mitomycin C prodrug is:

[0092] Dissolve 0.8-1.2 g of mitomycin C in 25-35 mL of N,N-dimethylformamide to obtain a seventh solution;

[0093] Add 1-2 g (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate to 5-15 mL N,N-dimethylformamide to obtain an eighth solution;

[0094] Add 1-2 mL of triethylamine to 5-15 mL of N,N-dimethylformamide, and stir to obtain a ninth solution;

[0095] Add the eighth solution to the seventh solution, stir for 5 to 15 minutes, then add the ninth solution, and react at 20 to 30° C. for 60 to 80 hours;

[0096] After the reaction is completed, 90-110 mL of saturated sodium carbonate aqueous solution is added for dilution, and 140-160 mL of ethyl acetate is used for three extractions. The organic phases are combined, washed twice with 40-60 mL of saturated sodium carbonate aqueous solution, dried over anhydrous sodium sulfate, and ethyl acetate is removed by rotary evaporation. The product is purified by column chromatography, which is the prodrug of mitomycin C.

[0097] Specifically, the synthesis reaction formula of mitomycin C prodrug is as follows:

[0098]

[0099] The following further illustrates the preparation method of the mitomycin C prodrug of the present application with specific examples. This section further illustrates the content of the present invention in conjunction with specific examples, but should not be construed as limiting the present invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0100] Example 1

[0101] The present invention provides a method for preparing a prodrug of mitomycin C, comprising the following steps:

[0102] S1. The synthesis of 4-amino-2,3,5,6-tetrafluorobenzyl alcohol specifically comprises the following steps:

[0103] S11, adding 0.22 g of lithium aluminum hydride to 15 mL of tetrahydrofuran and stirring for 10 min to obtain a first solution;

[0104] S12, adding 1 g of 4-amino-2,3,5,6-tetrafluorobenzoic acid to 5 mL of tetrahydrofuran, stirring to obtain a second solution;

[0105] S13, adding the second solution to the first solution, heating under reflux at 66° C. for 12 h;

[0106] After heating to reflux, tetrahydrofuran (recyclable) was removed by rotary evaporation, and then 5 mL of water was added to quench the excess lithium aluminum hydride, and then 20 mL of ethyl acetate was added to dissolve the product, filtered, and the precipitate was washed twice with 20 mL of ethyl acetate, the organic phases were combined, and the organic phases were washed twice with 20 mL of saturated sodium bicarbonate aqueous solution, dried with anhydrous sodium sulfate, and ethyl acetate was removed by rotary evaporation at 50°C. After cooling, the product naturally crystallized, which was 4-amino-2,3,5,6-tetrafluorobenzyl alcohol;

[0107] S2, the synthesis of 4-azido-2,3,5,6-tetrafluorobenzyl alcohol specifically comprises the following steps:

[0108] S21, dissolving 1 g of 4-amino-2,3,5,6-tetrafluorobenzyl alcohol and 8.77 g of p-toluenesulfonic acid monohydrate in 15 mL of water, cooling to 0° C. after dissolution, adding 0.5 g of sodium azide, and stirring for 10 min in an ice bath (i.e., at 0° C.) to obtain a third solution;

[0109] S22, dissolving 1.06 g of sodium nitrite in 5 mL of water to obtain a fourth solution;

[0110] S23, dropping the fourth solution into the third solution within 10 minutes, and continuing to stir and react in an ice bath for 20 minutes;

[0111] S24, after the reaction is completed, add 10 mL of saturated sodium carbonate aqueous solution to the reaction system, remove the ice bath, continue to add sodium carbonate solid until pH = 8, extract with 45 mL of ethyl acetate three times, combine the organic phases, wash the organic phases with 20 mL of saturated sodium bicarbonate aqueous solution, dry with anhydrous sodium sulfate, remove ethyl acetate by rotary evaporation at 30°C, and use an oil pump to drain the solvent and water to obtain a solid product, namely 4-azido-2,3,5,6-tetrafluorobenzyl alcohol;

[0112] S3, the synthesis of (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate specifically comprises the following steps:

[0113] S31, dissolving 1 g of 4-azido-2,3,5,6-tetrafluorobenzyl alcohol in 10 mL of tetrahydrofuran to obtain a fifth solution;

[0114] S32, adding 1.82 g of p-nitrophenyl chloroformate to 10 mL of tetrahydrofuran to obtain a sixth solution;

[0115] S33, adding the sixth solution to the fifth solution, mixing, cooling to 0°C, adding 3.58 g of pyridine, stirring for 10 min, heating to 25°C and reacting for 24 h;

[0116] S34, after the reaction is completed, tetrahydrofuran is removed by rotary evaporation at 30°C, 50 mL of water is added, and the mixture is extracted three times with 75 mL of ethyl acetate, the organic phases are combined, and the organic phases are washed four times with 10 mL of saturated sodium carbonate aqueous solution, dried over anhydrous sodium sulfate, and the ethyl acetate is removed by rotary evaporation at 30°C. The product is purified by column chromatography, namely (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate;

[0117] S4, the synthesis of mitomycin C prodrug, specifically comprising the following steps:

[0118] S41, dissolving 1 g of mitomycin C in 30 mL of N,N-dimethylformamide to obtain a seventh solution;

[0119] S42, adding 1.39 g (4-azido-2,3,5,6-tetrafluorophenyl)methyl-nitrophenyl carbonate to 10 mL N,N-dimethylformamide to obtain an eighth solution;

[0120] S43, adding 1.5 mL of triethylamine to 10 mL of N,N-dimethylformamide, stirring to obtain a ninth solution;

[0121] S44, adding the eighth solution to the seventh solution, stirring for 10 min, then adding the ninth solution, and reacting at 25° C. for 72 h;

[0122] S45. After the reaction is completed, add 100 mL of saturated sodium carbonate aqueous solution to dilute, extract with 150 mL of ethyl acetate three times, combine the organic phases, wash twice with 50 mL of saturated sodium carbonate aqueous solution, dry with anhydrous sodium sulfate, remove ethyl acetate by rotary evaporation, and purify by column chromatography to obtain the product, which is the prodrug of mitomycin C.

[0123] Performance Testing

[0124] The nuclear magnetic resonance hydrogen spectrum of the mitomycin C prodrug prepared in Example 1 is as follows: Figure 1 The mass spectrum of the mitomycin C prodrug prepared in Example 1 is shown in Figure 2 As shown; the HPLC chromatogram of the mitomycin C prodrug prepared in Example 1 is as shown Figure 3 shown.

[0125] The mitomycin C prodrug prepared in Example 1 was placed in phosphate buffered saline (PBS) and irradiated with 60 Gy X-rays by a linear accelerator. Figure 4 shown. Figure 4 In the figure, a represents 60 Gy X-ray irradiation, and b represents 0 Gy X-ray irradiation.

[0126] from Figure 4 It can be seen that the prodrug of mitomycin C is responsive to X-rays, and the prodrug of mitomycin C is greatly reduced after 60Gy X-ray irradiation.

[0127] Figure 5 The cytotoxicity comparison diagram of the mitomycin C prodrug (MMC-P) and the mitomycin C original drug (MMC) prepared in Example 1 is as follows: the drug (MMC-P or MMC) is dissolved in DMSO (dimethyl sulfoxide) to prepare three stock solutions of 10 mmol / L, 1 mmol / L and 0.1 mmol / L; the stock solution is then diluted to the desired final concentration (i.e., Figure 5 0.1μM~50μM); 10000 cells (CFPAC-1 human pancreatic cancer cells) were inoculated into each well 24h in advance, and the medium was aspirated to 96-well plates, 100μL per well, incubated for 48h, the medium containing drugs was aspirated, and serum-free medium containing 10% CCK-8 solution was added, and the absorbance at 450nm was measured after incubation for 1h to calculate the cell survival ratio. Figure 5 Control is the control group without drug.

[0128] from Figure 5It can be seen that at the same drug concentration, the cell survival rate of mitomycin C prodrug (MMC-P) was significantly higher than that of mitomycin C, which indicates that the toxicity of mitomycin C original drug (MMC) is significantly lower than that of mitomycin C.

[0129] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. Use of a prodrug of mitomycin C in the preparation of anticancer drugs; The cancer is human pancreatic cancer; The mitomycin C prodrug is responsive to X-rays; The structural formula of the mitomycin C prodrug is as follows: 。

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