Boron-containing compound and preparation method thereof

Abstract

The invention discloses a boron-containing compound and a preparation method thereof. The invention provides a compound as shown in a formula 5. The compound can be used for synthesizing the compound shown in the formula I. The compound shown in the formula I is low in toxicity to tumor cells and can be combined with boron neutron capture therapy for targeted cancer treatment. # imgabs0 #

Description

[0001] This application is a divisional of a Chinese invention patent with an application date of August 2, 2022, application number 2022109235473, and name “A kind of olefin-containing compound and its application”. Technical Field

[0002] The invention relates to a boron-containing compound and a preparation method thereof. Background Art

[0003] The latest statistics show that deaths from malignant tumors account for 23.91% of all deaths among residents. Cancer has become the leading cause of death among urban and rural residents in my country. In 2015, there were approximately 3.928 million cases of malignant tumors and approximately 2.338 million deaths. On average, more than 10,000 people are diagnosed with cancer every day. Poverty caused by cancer and people returning to poverty due to cancer have become serious economic and social problems in my country.

[0004] Boron Neutron Capture Therapy (BNCT) is a new binary radiotherapy targeted cancer treatment technology. Its principle is to inject a neutron-containing substance with specific affinity to the tumor into the 10 Drug B is injected into the patient's body, and the boron-containing drug is specifically concentrated in the tumor. Then, the tumor site is irradiated with an epithermal neutron beam. 10 The B nucleus captures a thermal neutron to form an unstable composite nucleus. 11 B, 11 B then spontaneously splits into an alpha particle with a kinetic energy of 1.78 MeV and a 7 Li recoil nucleus (reaction cross section 6.3%); or an alpha particle with a kinetic energy of 1.47 MeV and a 7 Li recoils the nucleus and emits a photon with an energy of 0.48 MeV (reaction cross section is 93.7%). Because the released particles have high energy, high linear energy conversion (LET) and low oxygen enhancement ratio, they can achieve high selectivity and high intensity to kill tumor cells.

[0005] 7 The range of Li recoil nuclei and α particles in biological tissues is about 5 μm and 9 μm, which is smaller than the diameter of tumor cells (10 μm). 7 The killing effect of Li and α particles is limited to ingestion 10 B cells and their immediate neighbors. 10 B(n,α) 7 The Li reaction cross section reaches 3840 barns, which is much larger than the reaction cross section between neutrons and nuclides in normal human tissues and blood. 10The radiation damage effect of normal tissues of B can be controlled at a safe dose level. Therefore, this treatment is known as cellular level radiotherapy, which combines the excellent characteristics of external radiotherapy and internal radiotherapy, and effectively avoids the side effects of conventional external radiotherapy and internal radiotherapy.

[0006] Compared with other tumor treatment technologies, BNCT has special advantages:

[0007] 1) Targetedness of BNCT therapy

[0008] Boron-containing drugs have targeting properties. Boron-containing drugs have a high affinity with tumor cells and are mainly absorbed by cancerous tissues. 10 The content of B is greater than that in normal cells 10 B content, so the therapeutic dose for tumor cells is much greater than that for normal cells.

[0009] 2) High Linear Energy Transfer (LET) characteristics

[0010] Traditional radiotherapy x-rays and gamma rays are of low LET nature, that is, the relative biological effect (RBE) on the body is low, and oxygen is needed to enhance the biological radiation effect. However, due to the rapid and invasive growth of malignant tumors, tumor tissues are often insufficiently supplied with blood, resulting in local hypoxia and relatively poor treatment effects. 7 Li particles are high LET charged particles that can kill both oxygen-rich and oxygen-deficient tumor cells. Even for heavy ions such as protons and carbon ions, the RBE of alpha particles produced by BNCT nuclear reactions is still the highest.

[0011] 3) The therapeutic effect does not depend on the state of the cancer cells

[0012] Chemotherapy, X-knife, gamma knife and conventional radiotherapy generally have effects on tumor cells in the proliferation phase (G1, S, G2 and M phases), but are insensitive to tumor cells in the quiescent phase (G0 phase). The continued growth of tumor cells in the G0 phase is the root cause of tumor recurrence. 7 The killing effect of Li particles on tumor cells does not depend on the cell growth cycle, and can also kill tumor cells in the dormant phase. Hypoxic tumors become more resistant to conventional radiotherapy such as low-LET gamma rays and electron beams, while BNCT-based treatment can kill hypoxic cancer cells as long as there is enough boron entering.

[0013] 4) Fine treatment scale

[0014] Among cancer therapies to date, neutron capture therapy has the only other method that has a treatment range as fine as the micron level. Minimally invasive surgery, the most precise form of surgery, has a treatment range limited to the millimeter level, and the Bragg peak focus dose depth in heavy ion therapy is about 2.5 to 3.0 cm. The finer the treatment size, the smaller the side effects.

[0015] Ideal boron-containing targeted drugs for BNCT treatment should meet the following requirements:

[0016] It is non-toxic to humans at clinical doses;

[0017] 10 Drug B has a high affinity for tumor tissue, and the concentration ratio of tumor to normal tissue can reach 4:1~3:1;

[0018] Per gram of tumor tissue 10 B drug concentration reaches 20~35μg;

[0019] During the treatment period, a certain therapeutic concentration can be maintained in the tumor tissue.

[0020] At present, the development of BNCT is relatively slow, which is closely related to the relative lag in the research and development of boron-containing drugs. Up to now, the only boron-containing drugs that have entered clinical use are disodium thiododecaborane (BSH) and para-dicarboxyborylphenylalanine (BPA). Among them, BPA was approved for marketing in Japan in 2020, making it the world's first BNCT boron-containing drug approved for marketing. As a key breakthrough in BNCT technology, the effects of BSH and BPA are far from the requirements, and there are problems such as insufficient tumor targeting specificity and insufficient affinity with tumor cells. After in-depth research, it was found that the ratio of boron concentration in tumor to normal tissue measured by BSH for human glioma samples is generally <1 (average 0.6), and it is considered to be a non-specific boron carrier. However, the boron concentration ratio produced by BPA is generally not greater than 2.4, and it cannot penetrate into the internal components of tumor cells, so it is only retained in the cells for a short time. In addition, there are groups of some tumor cells that do not take up BPA. In clinical practice, BNCT is mainly used to treat brain tumors, gliomas, melanomas and other cancers, which is mainly limited by the small number of boron-containing drugs currently available in clinical practice and their limited indications. Therefore, it is imperative to develop boron-targeted drugs with more indications, low toxicity and good targeting.

[0021] Fibroblast activation protein (FAP) is one of the specific markers of tumor-associated fibroblasts (TAF), with special biological characteristics, stable genome, abundant and specific expression in tumor stroma. Existing FAP inhibitors are mainly used for tumor treatment. However, there is currently no FAP targeted drug that can be used in boron neutron capture therapy. Summary of the invention

[0022] The technical problem to be solved by the present invention is that the existing compounds suitable for BNCT have a single structure. To this end, the present invention provides a boron-containing compound and a preparation method thereof, which can be used to synthesize the compound shown in formula I. The compound shown in formula I has low toxicity to tumor cells and can be combined with boron neutron capture therapy for targeted cancer treatment.

[0023] The present invention provides a compound as shown in Formula I or a pharmaceutically acceptable salt thereof:

[0024]

[0025] Among them, R 1 For single bond, C1~C 10 Straight chain alkylene, C2~C 10 Straight chain alkenylene, C2~C 10 Branched chain alkylene, C5~C8 cycloalkylene, C6~C 10 Arylene-C1~C4 alkylene, C1~C4 alkylene-C6~C 10 Arylene-C1~C4 alkylene, or C6~C 10 arylene;

[0026] R 2 Hydroxyl, C1~C5 straight chain alkyl, C3~C6 branched chain alkyl, C5~C8 cycloalkyl, C6~C 10 Aryl-C1~C4 alkyl, C1~C4 alkyl-C6~C 10 Aryl-C1~C4 alkyl, or C6~C 10 Aryl;

[0027] R 3 Hydroxyl, C1~C5 straight chain alkyl, C3~C6 branched chain alkyl, C5~C8 cycloalkyl, C6~C 10 Aryl-C1~C4 alkyl, C1~C4 alkyl-C6~C 10 Aryl-C1~C4 alkyl, or C6~C 10 Aryl;

[0028] R 4 is H, F or Cl;

[0029] R 5 is H, F or Cl;

[0030] The above n1, n2 and n3 are independently 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0031] In a certain embodiment, in the compound of formula I or a pharmaceutically acceptable salt thereof, some of the groups are defined as follows, and the remaining groups are defined as described in any other embodiment (hereinafter referred to as "in a certain embodiment"): 1 Can be a single bond.

[0032] In one embodiment, the R 2 It may be a hydroxyl group.

[0033] In one embodiment, the R 3 It may be a hydroxyl group.

[0034] In one embodiment, the R 4 It can be F or Cl.

[0035] In one embodiment, the R 4 Can be F.

[0036] In one embodiment, the R 5 It can be F or Cl.

[0037] In one embodiment, the R 5 Can be F.

[0038] In one embodiment, the compound represented by formula I may be compound I-1:

[0039] .

[0040] The present invention also provides a compound as shown in Formula 2, Formula 3, Formula 4, Formula 5 or Formula 6:

[0041] , , , or

[0042] Among them, R 51 is a C1~C4 alkyl group;

[0043] In the compound shown in Formula 3, R 32 is C1~C3 alkyl; R 33 is C1~C3 alkyl; R 34 is a C1~C3 alkyl group;

[0044] In the compound represented by Formula 5, R52 is C1~C3 alkyl; R 53 is C1~C3 alkyl; R 54 is C1~C3 alkyl; R 55 is a C1~C3 alkyl group;

[0045] R 1 , R 2 , R 3 The definitions are as above.

[0046] In one embodiment, the compound represented by Formula 2 is compound 2-1:

[0047] .

[0048] In one embodiment, the compound represented by formula 3 is compound 3-1:

[0049] .

[0050] In one embodiment, the compound represented by formula 4 is compound 4-1:

[0051] .

[0052] In one embodiment, the compound represented by Formula 5 is compound 5-1:

[0053] .

[0054] In one embodiment, the compound represented by Formula 6 is Compound 6-1:

[0055] .

[0056] The present invention also provides a method for preparing the compound of formula I, which comprises the following steps: in a solvent, subjecting the compound of formula 6 to an amidation reaction with the compound of formula a in the presence of an amidating agent as shown below to obtain the compound of formula I, that is,

[0057] .

[0058] In one embodiment, the amidation reaction can be carried out under normal pressure.

[0059] In one embodiment, the amidation reaction can be carried out in the presence of oxygen.

[0060] In a certain embodiment, the amidation agent may be a combination of N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (TCFH) and 1-methylimidazole.

[0061] Wherein, the solvent is a commonly used solvent for such reactions in the art.

[0062] In one embodiment, the solvent may be acetonitrile.

[0063] In one embodiment, the compound represented by Formula 6 may be the above-mentioned compound 6-1.

[0064] In one embodiment, the molar volume ratio of the compound represented by Formula 6 to the solvent may be 0.121 mol / L.

[0065] In one embodiment, the molar ratio of the compound represented by Formula 6 to the N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (TCFH) in the amidation reagent may be 0.845.

[0066] In one embodiment, the molar volume ratio of the compound represented by Formula 6 to the 1-methylimidazole in the amidation reagent may be 2.417 mol / L.

[0067] In one embodiment, the reaction temperature can be 25°C.

[0068] In one embodiment, the reaction condition may be stirring the reaction for 2 hours.

[0069] In a certain embodiment, the preparation method may further include separation of the reaction product, and the separation may be acidification, extraction, washing, drying, and column chromatography in sequence.

[0070] In a certain embodiment, the acidification may be acidification using trifluoroacetic acid, preferably 1% trifluoroacetic acid acidification.

[0071] In a certain embodiment, the extraction can be ethyl acetate extraction, preferably an equal volume of ethyl acetate extraction.

[0072] In a certain embodiment, the washing can be washing with a sodium bicarbonate solution, preferably washing with an equal volume of sodium bicarbonate solution.

[0073] In a certain embodiment, the drying may be drying with anhydrous sodium sulfate.

[0074] In a certain embodiment, the developing solvent used in the column chromatography can be petroleum ether or ethyl acetate.

[0075] In a certain embodiment, the acidification can be performed by using 1% trifluoroacetic acid; the extraction can be performed by using an equal volume of ethyl acetate for 3 times; the washing can be performed by using an equal volume of sodium bicarbonate solution for 2 times; the drying can be performed by drying over anhydrous sodium sulfate; and the developing solvent used for the column chromatography can be petroleum ether or ethyl acetate.

[0076] In one embodiment, the compound represented by formula I may be a compound represented by formula I-2: .

[0077] The present invention also provides a method for preparing the compound of formula I-2, comprising the following steps: (1) reacting the compound of formula 1 with the compound of formula D in a solvent as shown below to obtain a compound of formula 2;

[0078] (2) In a solvent, reacting the compound represented by Formula 2 with the compound represented by Formula B as shown below to obtain a compound represented by Formula 3;

[0079] (3) In a solvent, the compound represented by Formula 3 is subjected to the following reaction to obtain a compound represented by Formula 4;

[0080] (4) In a solvent, reacting the compound represented by Formula 4 with the compound represented by Formula C as shown below to obtain a compound represented by Formula 5-2;

[0081] (5) In a solvent, the compound represented by Formula 5-2 is subjected to the following reaction to obtain a compound represented by Formula 6-2;

[0082] (6) In a solvent, the compound represented by Formula 6-1 and the compound represented by Formula A are subjected to an amidation reaction as shown below in the presence of an amidating agent to obtain a compound represented by Formula I-2. .

[0083] Wherein, the solvent is a commonly used solvent for such reactions in the art.

[0084] In a certain embodiment, in the step (1), the reaction can be carried out under normal pressure; the reaction can be carried out in the presence of oxygen; the solvent can be methanol; the compound represented by formula D can be methanol; the reaction can be carried out in the presence of dichlorothionyl; the reaction temperature can be 80°C; the reaction time can be 4 hours; the reaction can be quenched by adding water; the reaction can also include separation of the reaction product, and the separation can be sequentially adjusting the pH of the reaction solution and filtering.

[0085] In a certain embodiment, in the step (1), the molar volume ratio of the compound represented by formula 1 to the solvent may be 0.8 mol / L; the molar volume ratio of the compound represented by formula 1 to the compound represented by formula D may be 0.8 mol / L; the molar volume ratio of the compound represented by formula 1 to dichloride may be 6.6 mol / L; and the pH of the reaction solution may be adjusted by sodium bicarbonate.

[0086] In a certain embodiment, in the step (2), the reaction can be carried out under normal pressure; the reaction can be carried out under an inert atmosphere; the solvent can be tetrahydrofuran; the compound represented by formula 2 can be the above-mentioned compound 2-1; the compound represented by formula B can be trimethylethynylsilane; the reaction can also be carried out in the presence of bis(triphenylphosphine)palladium(II) chloride, cuprous iodide and triethylamine; the reaction temperature can be 50°C; the reaction time can be 3 hours; the reaction can also include separation and purification of the reaction product, and the separation and purification can be filtering the reaction solution, diluting, extracting, washing, drying, filtering, concentrating and purifying in sequence.

[0087] In a certain embodiment, in the step (2), the molar volume ratio of the compound represented by formula 2 to the solvent may be 0.278 mol / L; the molar ratio of the compound represented by formula 2 to the compound represented by formula B may be 0.5; the molar ratio of the compound represented by formula 2 to trimethylethynyl silicon may be 20; the molar ratio of the compound represented by formula 2 to cuprous iodide may be 20; the molar ratio of the compound represented by formula 2 to triethylamine may be 0.67; the extraction in the separation and purification is ethyl acetate extraction; and the purification in the separation and purification is column chromatography purification.

[0088] In one embodiment, in the step (2), the developing solvents in the column chromatography purification in the separation and purification are petroleum ether and ethyl acetate.

[0089] In a certain embodiment, in the step (3), the reaction can be carried out under normal pressure; the reaction can be carried out under an inert atmosphere; the compound represented by formula 3 can be the above-mentioned compound 3-1; the solvent can be a mixture of methanol and dichloromethane; the reaction can be carried out in the presence of potassium fluoride; the reaction temperature can be room temperature; the reaction time can be 12 hours; the reaction can also include separation and purification of the reaction product, and the separation and purification can be filtering the reaction solution, diluting, extracting, washing, drying, filtering, concentrating, beating, filtering and drying in sequence.

[0090] In a certain embodiment, in the step (3), the molar volume ratio of the compound represented by formula 3 to the solvent may be 0.45 mol / L; the molar ratio of the compound represented by formula 3 to potassium fluoride may be 0.33; the extraction in the separation and purification is ethyl acetate extraction; and the pulping in the separation and purification is pulping with petroleum ether and ethyl acetate.

[0091] In a certain embodiment, in the step (4), the reaction can be carried out under normal pressure; the reaction can be carried out under an inert atmosphere; the solvent can be toluene; the compound represented by formula 4 can be the above-mentioned compound 4-1; the compound represented by formula C can be pinacol borane; the reaction can be carried out in the presence of carbonyl chloride tris(triphenylphosphine)ruthenium (II); the reaction temperature can be 50°C; the reaction time can be 14 hours; the reaction can also include separation and purification of the reaction product, and the separation and purification can be filtering the reaction solution, diluting, extracting, washing, drying, filtering, concentrating and purifying in sequence.

[0092] In a certain embodiment, in the step (4), the molar volume ratio of the compound represented by formula 4 to the solvent may be 0.47 mol / L; the molar ratio of the compound represented by formula 4 to the compound represented by formula C may be 0.26; the molar ratio of the compound represented by formula 4 to carbonylchlorohydrotris(triphenylphosphine)ruthenium (II) may be 71; the extraction in the separation and purification is ethyl acetate extraction; and the purification in the separation and purification is column chromatography purification.

[0093] In one embodiment, in the step (4), the developing solvents in the column chromatography purification in the separation and purification are petroleum ether and ethyl acetate.

[0094] In a certain embodiment, in the step (5), the reaction can be carried out under normal pressure; the reaction can be carried out in the presence of oxygen; the solvent can be 1,4-dioxane; the compound represented by formula 5-2 can be the above-mentioned compound 5-1; the reaction can be carried out in the presence of hydrochloric acid; the reaction temperature can be 85°C; the reaction time can be 12 hours; the reaction can also include separation and purification of the reaction product, and the separation and purification can be sequentially extracting the reaction liquid, separating the liquid, drying, pulping, filtering and drying.

[0095] In a certain embodiment, in the step (5), the molar volume ratio of the compound represented by formula 5-2 to the solvent may be 0.59 mol / L; the concentration of hydrochloric acid is 6 mol / L; the molar volume ratio of the compound represented by formula 5-2 to the hydrochloric acid may be 0.39 mol / L; the extraction in the separation and purification is ethyl acetate extraction; and the pulping in the separation and purification is pulping with methyl tert-butyl ether.

[0096] In one embodiment, the reaction conditions in step (6) can refer to the reaction conditions in the preparation method of the compound represented by formula I above.

[0097] The present invention also provides a pharmaceutical composition, which comprises a substance X and a pharmaceutical excipient; the substance X is the compound shown in Formula I or a pharmaceutically acceptable salt thereof.

[0098] The present invention also provides an application of a substance X in the preparation of a drug; the substance X is the compound shown in Formula I or a pharmaceutically acceptable salt thereof; the drug is a drug for treating tumors.

[0099] In one embodiment, the drug used to treat tumors is a radiotherapy targeted drug.

[0100] In one embodiment, the radiotherapy targeted drug is a boron neutron capture therapy drug.

[0101] Unless otherwise specified, the terms used in the present invention have the following meanings:

[0102] The term "alkyl" refers to a straight or branched chain alkyl group with a specified number of carbon atoms (e.g., C1-C4). Alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl, sec-butyl, etc.

[0103] The term "alkylene" refers to a divalent alkyl group having a specified number of carbon atoms (e.g., C1-C4) and having a straight or branched chain and connected to other groups at two positions. Alkylene includes but is not limited to , , , wait.

[0104] The term "alkenylene" refers to a straight or branched divalent alkenyl group having a specified number of carbon atoms (e.g., C2-C4) and connected to other groups at two positions. Alkenylene includes but is not limited to , , wait.

[0105] The term "cycloalkyl" refers to a saturated monocyclic ring group consisting only of carbon atoms with a specified number of carbon atoms (e.g., C5-C8). Cycloalkyl includes, but is not limited to, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, etc.

[0106] The term "aryl" refers to a group having a specified number of carbon atoms (e.g. C6~C 10 ) is a cyclic group consisting only of carbon atoms, which is monocyclic or polycyclic, and each ring is aromatic (in accordance with Huckel's rule). Aryl includes but is not limited to phenyl, naphthyl, etc.

[0107] The term "arylene" refers to a group having a specified number of carbon atoms (e.g. C6~C 10 ) is a divalent cyclic group consisting only of carbon atoms connected to other groups through two sites, which is a monocyclic or polycyclic group, and each ring is aromatic (in accordance with Huckel's rule). Arylene includes but is not limited to , wait.

[0108] The term "pharmaceutically acceptable salt" refers to a salt obtained by reacting a compound with a pharmaceutically acceptable (relatively non-toxic, safe, and suitable for use by patients) acid or base. When the compound contains a relatively acidic functional group, a base addition salt can be obtained by contacting the free form of the compound with a sufficient amount of a pharmaceutically acceptable base in a suitable inert solvent. Pharmaceutically acceptable base addition salts include, but are not limited to, sodium salts, potassium salts, calcium salts, aluminum salts, magnesium salts, bismuth salts, ammonium salts, and the like. When the compound contains a relatively basic functional group, an acid addition salt can be obtained by contacting the free form of the compound with a sufficient amount of a pharmaceutically acceptable acid in a suitable inert solvent. Pharmaceutically acceptable acid addition salts include, but are not limited to, hydrochlorides, sulfates, mesylate salts, and the like. For details, see Handbook of Pharmaceutical Salts: Properties, Selection, and Use (P. Heinrich Stahl, 2002).

[0109] Without violating the common sense in the art, the above-mentioned preferred conditions can be arbitrarily combined to obtain the preferred embodiments of the present invention.

[0110] The reagents and raw materials used in the present invention are commercially available.

[0111] The positive and progressive effects of the present invention are that the compound has low toxicity to tumor cells and can be combined with boron neutron capture therapy to target cancer. DETAILED DESCRIPTION

[0112] The present invention is further described below by way of examples, but the present invention is not limited to the scope of the examples. The experimental methods in the following examples without specifying specific conditions are carried out according to conventional methods and conditions, or selected according to the product specifications.

[0113] The A549 cells, HepG2 cells, U87MG cells, SCC9 cells, and A375 cells in the following examples are all from the Cell Bank of the Chinese Academy of Sciences in Shanghai;

[0114] In the following embodiments

[0115] The manufacturer of the CCK-8 cell viability assay kit is Shanghai Donghuan Biotechnology Co., Ltd.;

[0116] The manufacturer of PBS is Shanghai Shuanglu Biotechnology Co., Ltd.;

[0117] The manufacturer of DMEM high-glucose culture medium is Shanghai Donghuan Biotechnology Co., Ltd.

[0118] Example 1 Preparation of Compound I-1 of the Present Invention

[0119]

[0120] 1. Preparation of Compound I-1

[0121] The preparation route of I-1 is as follows:

[0122]

[0123] Step 1: Compound 1 (50 g, 198 mmol) was dissolved in methanol (250 ml), and thionyl chloride (30 ml) was slowly added at room temperature. The reaction solution was reacted at 80°C for 4 hours. The reaction solution was quenched by adding water (50 ml), and the pH was adjusted to 7 with sodium bicarbonate solution. The solid was filtered and collected. The white compound 2-1 was obtained with a yield of 93%.

[0124] The identification data of compound 2-1 are as follows: LC-MS: M (C 11 H8BrNO2) = 266.09 (m / z), [M+H] + 267.2

[0125] Step 2: Compound 2-1 (37.0 g, 139 mmol) and trimethylethynylsilane (b) (27.3 g, 278 mmol) were dissolved in tetrahydrofuran (500 ml), and then Pd(PPh3)2Cl2 (4.88 g, 6.95 mmol), CuI (1.32 g, 6.95 mmol), and Et3N (208 mmol, 29.0 mL) were slowly added under nitrogen protection. The reaction solution was reacted at 50 °C for 3 hours under nitrogen protection. The reaction solution was filtered and collected, diluted with water (100 ml), and extracted with ethyl acetate twice (250 ml / time). The organic phases were combined and washed twice with saturated brine (100 ml / time), the organic phases were dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (developing solvent: petroleum ether, ethyl acetate). Compound 3-1 was obtained as a yellow solid with a yield of 96%.

[0126] The identification data of compound 3-1 are as follows: LC-MS: M (C 16 H 17 NO2Si) =283.40 (m / z), [M+H] + 284.2

[0127] Step 3: Compound 3-1 (38 g, 134 mmol) was dissolved in methanol (150 ml) and dichloromethane (150 ml), and potassium fluoride (23.4 g, 402 mmol) was slowly added under nitrogen protection. The reaction solution was stirred at room temperature for 12 hours. The reaction solution was filtered and collected, diluted with water (100 ml), and extracted with ethyl acetate twice (250 ml / time). The organic phases were combined and washed twice with saturated brine (100 ml / time), the organic phases were dried with anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and the crude product was slurried with petroleum ether and ethyl acetate (1:3, 30 ml) at room temperature for 30 minutes. Compound 4-1 was obtained by filtration and drying as a yellow solid with a yield of 53%.

[0128] The identification data of compound 4-1 are as follows: LC-MS: M (C 13 H9NO2) =211.22 (m / z), [M+H] + 212.0

[0129] Step 4: Compound 4-1 (15g, 71mmol) was dissolved in toluene (150ml), and carbonyl chloride tris(triphenylphosphine)ruthenium (II) (16971-33-8) (1g, 1mmol) and pinacol borane (c) (35.3g, 275mmol) were slowly added under nitrogen protection. The reaction liquid was stirred at 50°C for 14 hours under nitrogen protection. The reaction liquid was filtered and collected, diluted with water (50ml), and extracted with ethyl acetate twice (100ml / time). The organic phases were combined and washed twice with saturated brine (50ml / time), the organic phases were dried with anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (developing solvent: petroleum ether, ethyl acetate). Compound 5-1 was obtained as a yellow solid with a yield of 85%.

[0130] The identification data of compound 5-1 are as follows: LC-MS: M (C 19 H 22 BNO4) =339.2 (m / z), [M+H] + 340.0

[0131] Step 5: Compound 5-1 (20 g, 59 mmol) was dissolved in 1,4-dioxane (100 ml), and hydrochloric acid (6 mol / L, 150 ml) was slowly added. The reaction solution was stirred at 85 °C for 12 hours. The reaction solution was extracted with ethyl acetate three times (80 ml / times). The aqueous phase was collected and freeze-dried, and the crude product was slurried with methyl tert-butyl ether at room temperature for 30 minutes. After filtration and drying, compound 6-1 was obtained as a yellow solid with a yield of 91%.

[0132] The structural identification data of compound 6-1 are as follows: LC-MS: M (C 12 H10 BNO4) =243.03 (m / z), [M+H] + 244.1

[0133] Step 6: Compound 6-1 (8 g, 29 mmol) and compound a (18.9 g, 15.7 mmol) were dissolved in acetonitrile (240 ml), and N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (TCFH) (9.6 g, 34.3 mmol) and 1-methylimidazole (12 ml, 12.4 g) were slowly added. The reaction solution was stirred at 25 ° C for 2 hours. The pH of the reaction solution was adjusted to 5 with 1% trifluoroacetic acid, and the reaction solution was extracted with ethyl acetate three times (500 ml / time). The organic phases were combined and washed twice with sodium bicarbonate solution (200 ml / time), and the organic phases were dried with anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography (developing solvent: petroleum ether, ethyl acetate). The product I-1 was obtained as a yellow solid with a yield of 46%.

[0134] 2. Structural identification

[0135] The LC-MS and NMR identification data of compound I-1 are as follows:

[0136] LC-MS: M (C 19 H 17 BF2N4O4) =414.18 (m / z), [M+H]+ 415.2

[0137] 1 H NMR: (400 MHz, DMSO-d6) δ 9.18-9.15 (m, 1H), 8.98 (d, J = 4.4 Hz,1H), 8.29 (s, 1H), 8.09-7.60 (m, 5H), 7.45-7.40 (m, 1H), 6.36-6.32 (m, 1H), 5.22 (d, J = 9.6 Hz, 1H), 4.34-4.16 (m, 4H), 2.97-2.83 (m, 2H).

[0138] F NMR: (400 MHz, DMSO-d6) δ -95.054, -103.651.

[0139] Example 2: Cytotoxicity of Compound I-1 to A549 cells

[0140] A549 cells in the logarithmic growth phase were seeded in 96-well plates, with 1*10 4Cells were cultured overnight in DMEM high-glucose medium, the medium was removed, and the cells were washed twice with PBS (0.01 mol / L). Compound I-1 was added to the fresh medium to prepare a series of solutions containing compound I-1 with a concentration gradient. The concentrations of compound I-1 were 0, 100, 250, 500, and 750 µg / ml, respectively. 100 µl was added to the test cell plate. After the cells were treated with drugs for 48 hours, the cell culture medium was removed with a pipette, the cells were washed twice with PBS (0.01 mol / L), 100uL of CCK8 working solution was added, and the culture plate was incubated in the incubator for 2 hours; the absorbance at 450 nm was measured with an ELISA reader to calculate the effect of the drug on cell viability.

[0141] The test results are shown in Table 1. The low cytotoxicity of compound I-1 is an important indicator of BNCT, which can ensure the maximum boron concentration in tumor cells. The compound I-1 of the present invention shows low toxicity to tumor cells. The IC 50 It is 458.1 µg / ml, which meets the requirement of low toxicity of boron-containing drugs in boron neutron capture therapy.

[0142] Table 1 Inhibitory effect of different concentrations of compounds on A549 cells

[0143]

[0144] Example 3: Cytotoxicity of Compound I-1 on HepG2 cells

[0145] HepG2 cells in the logarithmic growth phase were seeded in 96-well plates, with 1*10 4 Cells were cultured overnight in DMEM high-glucose medium, the medium was removed, and the cells were washed twice with PBS (0.01mol / L). Compound I-1 was added to fresh culture medium to prepare a series of solutions containing compound I-1 with a concentration gradient. The concentrations of compound I-1 were 0, 100, 250, 500, and 750 µg / ml, respectively. 100 µl was added to the test cell plate. After the cells were treated with drugs for 48 hours, the cell culture medium was removed with a pipette, the cells were washed twice with PBS (0.01mol / L), 100uL of CCK8 working solution was added, and the culture plate was incubated in the incubator for 2 hours; the absorbance at 450 nm was measured with an enzyme-labeled instrument to calculate the effect of the drug on cell viability.

[0146] The test results are shown in Table 2. The low cytotoxicity of compound I-1 is an important indicator of BNCT, which can ensure the maximum boron concentration in tumor cells. The compound I-1 of the present invention shows low toxicity to tumor cells, and the IC 50It is 340.1 µg / ml, which meets the requirement of low toxicity of boron-containing drugs in boron neutron capture therapy.

[0147] Table 2 Inhibitory rate of HepG2 cells at different concentrations of compounds

[0148]

[0149] Example 4: Cytotoxicity of Compound I-1 to U87MG cells

[0150] U87MG cells in the logarithmic growth phase were seeded in 96-well plates, with 1*10 4 Cells were cultured overnight in DMEM high-glucose medium, the medium was removed, and the cells were washed twice with PBS (0.01 mol / L). Compound I-1 was added to the fresh medium to prepare a series of solutions containing compound I-1 with a concentration gradient. The concentrations of compound I-1 were 0, 100, 250, 500, and 750 µg / ml, respectively. 100 µl was added to the test cell plate. After the cells were treated with drugs for 48 hours, the cell culture medium was removed with a pipette, the cells were washed twice with PBS (0.01 mol / L), 100uL of CCK8 working solution was added, and the culture plate was incubated in the incubator for 2 hours; the absorbance at 450 nm was measured with an ELISA reader to calculate the effect of the drug on cell viability.

[0151] The test results are shown in Table 3. The low cytotoxicity of compound I-1 is an important indicator of BNCT, which can ensure the maximum boron concentration in tumor cells. The compound I-1 of the present invention shows low toxicity to tumor cells, and the IC 50 It is 411.8 µg / ml, which meets the requirement of low toxicity of boron-containing drugs in boron neutron capture therapy.

[0152] Table 3 Inhibitory effect of different concentrations of compounds on U87MG cells

[0153]

[0154] Example 5: Cytotoxicity of Compound I-1 on SCC9 cells

[0155] SCC9 cells in the logarithmic growth phase were seeded in 96-well plates, with 1*10 4Cells were cultured overnight in DMEM high-glucose medium, the medium was removed, and the cells were washed twice with PBS (0.01mol / L). Compound I-1 was added to fresh culture medium to prepare a series of solutions containing compound I-1 with a concentration gradient. The concentrations of compound I-1 were 0, 100, 250, 500, and 750 µg / ml, respectively. 100 µl was added to the test cell plate. After the cells were treated with drugs for 48 hours, the cell culture medium was removed with a pipette, the cells were washed twice with PBS (0.01mol / L), 100uL of CCK8 working solution was added, and the culture plate was incubated in the incubator for 2 hours; the absorbance at 450 nm was measured with an enzyme-labeled instrument to calculate the effect of the drug on cell viability.

[0156] The test results are shown in Table 4. The low cytotoxicity of compound I-1 is an important indicator of BNCT, which can ensure the maximum boron concentration in tumor cells. The compound I-1 of the present invention shows low toxicity to tumor cells, and the IC 50 It is 289.1 µg / ml, which meets the requirement of low toxicity of boron-containing drugs in boron neutron capture therapy.

[0157] Table 4 Inhibitory effect of different concentrations of compounds on SCC9 cells

[0158]

[0159] Example 6: Cytotoxicity of Compound I-1 to A375 cells

[0160] A375 cells in the logarithmic growth phase were seeded in 96-well plates, with 1*10 4 Cells were cultured overnight in DMEM high-glucose medium, the medium was removed, and the cells were washed twice with PBS (0.01mol / L). Compound I-1 was added to fresh culture medium to prepare a series of solutions containing compound I-1 with a concentration gradient. The concentrations of compound I-1 were 0, 100, 250, 500, and 750 µg / ml, respectively. 100 µl was added to the test cell plate. After the cells were treated with drugs for 48 hours, the cell culture medium was removed with a pipette, the cells were washed twice with PBS (0.01mol / L), 100uL of CCK8 working solution was added, and the culture plate was incubated in the incubator for 2 hours; the absorbance at 450 nm was measured with an enzyme-labeled instrument to calculate the effect of the drug on cell viability.

[0161] The test results are shown in Table 5. The low cytotoxicity of compound I-1 is an important indicator of BNCT, which can ensure the maximum boron concentration in tumor cells. The compound I-1 of the present invention shows low toxicity to tumor cells. The IC 50It is 586.5 µg / ml, which meets the requirement of low toxicity of boron-containing drugs in boron neutron capture therapy.

[0162] Table 5 Inhibition rate of A375 cells by compounds at different concentrations

[0163] .

Claims

1. A compound as shown in Formula 5: ， in, R 1 For single bond, C1~C 10 Straight chain alkylene, C2~C 10 Straight chain alkenylene, C2~C 10 Branched chain alkylene, C5~C8 cycloalkylene, C6~C 10 Arylene-C1~C4 alkylene, C1~C4 alkylene-C6~C 10 Arylene-C1~C4 alkylene, or C6~C 10 arylene; R 51 is a C1~C4 alkyl group; R 52 is a C1~C3 alkyl group; R 53 is a C1~C3 alkyl group; R 54 is a C1~C3 alkyl group; R 55 It is a C1~C3 alkyl group.

2. The compound as shown in formula 5 according to claim 1, characterized in that The R 1 For a single key.

3. The compound as shown in formula 5 according to claim 1, characterized in that The compound represented by Formula 5 is Compound 5-1: .

4. A method for preparing a compound as shown in formula 5-2, characterized in that: In a solvent, the compound represented by Formula 4 and the compound represented by Formula C are reacted as shown below to obtain a compound represented by Formula 5-2. ， Among them, R 51 is C1~C4 alkyl; R 52 is C1~C3 alkyl; R 53 is C1~C3 alkyl; R 54 is C1~C3 alkyl; R 55 It is a C1~C3 alkyl group.

5. The preparation method according to claim 4, characterized in that: It meets one or more of the following conditions: (1) The reaction is carried out under normal pressure; (2) The reaction is carried out under an inert atmosphere; (3) The solvent is toluene; (4) The compound represented by formula 4 is compound 4-1: ; (5) The compound represented by formula C is pinacol borane; (6) The reaction is carried out in the presence of tris(triphenylphosphine)ruthenium(II)carbonyl chloride; (7) The reaction temperature is 50°C; (8) The reaction further includes separation and purification of the reaction product, and the separation and purification sequentially comprises filtering the reaction solution, diluting, extracting, washing, drying, filtering, concentrating and purifying.

6. The preparation method according to claim 5, characterized in that: It meets one or more of the following conditions: (1) The molar volume ratio of the compound represented by formula 4 to the solvent is 0.47 mol / L; (2) The molar ratio of the compound represented by Formula 4 to the compound represented by Formula C is 0.26; (3) The molar ratio of the compound represented by formula 4 to carbonyl chlorohydrotris(triphenylphosphine)ruthenium(II) is 7:1; (4) The extraction in the separation and purification is ethyl acetate extraction; the purification in the separation and purification is column chromatography purification, wherein the developing solvent in the column chromatography purification in the separation and purification is petroleum ether and ethyl acetate.

7. The preparation method according to claim 4, characterized in that: The method comprises a preparation method of a compound represented by formula 4, comprising the following steps: (1) In a solvent, reacting the compound represented by Formula 1 with the compound represented by Formula D as shown below to obtain the compound represented by Formula 2; (2) In a solvent, reacting the compound represented by Formula 2 with the compound represented by Formula B as shown below to obtain a compound represented by Formula 3; (3) In a solvent, the compound represented by formula 3 is subjected to the following reaction to obtain a compound represented by formula 4. ， Among them, R 51 is a C1~C4 alkyl group; In the compound shown in Formula 3, R 32 is C1~C3 alkyl; R 33 is C1~C3 alkyl; R 34 It is a C1~C3 alkyl group.

8. A method for preparing a compound of formula I-2, characterized in that: It includes the following steps: A method for preparing a compound as shown in formula 5-2 as claimed in any one of claims 4 to 7; and, a method for preparing a compound of formula I from a compound of formula 5-2, ， Among them, R 32 , R 33 , R 34 and R 51 As claimed in claim 7, R 4 is H, F or Cl; R 5 is H, F or Cl.

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