A benzopyridine ring-containing compound and a preparation method thereof

Abstract

The application discloses a benzopyridine ring-containing compound and a preparation method thereof. The application provides a compound shown in formula 4. The compound can be used for synthesizing a compound shown in formula I. The compound shown in formula I has low toxicity to tumor cells and can be used for targeted cancer treatment in combination with boron neutron capture therapy.

Description

[0001] This application is a divisional application of the Chinese Invention Patent with the application number 2022109235473 and the name "A compound containing alkenyl and its application", the filing date of which is August 2, 2022. TECHNICAL FIELD

[0002] The present application relates to a kind of compounds containing benzopyridine ring and preparation method thereof. BACKGROUND

[0003] The latest statistics show that malignant tumor death accounts for 23.91% of all causes of death of residents, and cancer has become the first cause of death of urban and rural residents in China. In 2015, about 3.928 million people were diagnosed with cancer, and about 2.338 million people died, with an average of more than 10,000 people diagnosed with cancer every day.

[0004] Boron Neutron Capture Therapy (BNCT) is a new technology of binary radiotherapy targeted cancer treatment. Its principle is to inject a specific affinity B-containing drug into the tumor, and then irradiate the tumor site with an ultrahot neutron beam, 10 The B-containing drug is injected into the patient's body, and the B-containing drug is specifically concentrated in the tumor. Then, an ultrahot neutron beam is used to irradiate the tumor site, 10 The B atom nucleus captures a hot neutron to form an unstable compound nucleus 11 B, 11 B spontaneously splits into an α particle with a kinetic energy of 1.78 MeV and a 7 Li recoil nucleus (reaction cross section is 6.3%); or an α particle with a kinetic energy of 1.47 MeV and a 7 Li recoil nucleus and emits a photon with an energy of 0.48 MeV (reaction cross section is 93.7%). Because the energy of the released particles is high, it has the characteristics of high linear energy transfer (LET) and low oxygen enhancement ratio, so it can achieve the effect of high selectivity and high intensity killing tumor cells.

[0005] 7 The range of Li recoil nucleus and α particle in biological tissue is about 5 μm and 9 μm, which is less than the diameter of tumor cell 10 μm, so 7 The killing effect of Li and α particle is limited to cells that have taken up 10 B and their immediately adjacent cells. On the other hand, 10 B (n, α) 7 The reaction cross section of Li reaches 3840 target barns, which is much larger than the reaction cross section of neutron and normal tissue and blood composition nuclei. Therefore, the neutron is highly selective for cells that have taken up 10The radiation damage effect of normal tissue of B can be controlled at a safe dose level. Thus the treatment is praised as cell level radiotherapy, combining the excellent features of external radiotherapy and internal radiotherapy, and effectively avoiding the side effects of conventional external radiotherapy and internal radiotherapy.

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

[0007] 1) Targeting of BNCT therapy

[0008] The boron-containing drug has targeting properties. The boron-containing drug has high affinity with tumor cells, is mainly absorbed by cancerous tissues, and is mainly accumulated in tumor cells 10 The content of B in tumor cells is much higher than that in normal cells 10 The content of B in tumor cells is much higher than that in normal cells, and thus the therapeutic dose of tumor cells is much higher than that of normal cells.

[0009] 2) High energy transfer line density (LET) property

[0010] Traditional radiotherapy χ-rays, γ-rays, etc. belong to low LET property, i.e. the relative biological effect (RBE) of the body is low, and oxygen is needed to enhance the biological radiation effect. However, due to the rapid invasive growth of malignant tumors, the tumor tissue is often hypoxic, and the treatment effect is relatively poor. α particles and 7 Li particles are high LET charged particles, which can kill tumor cells with or without oxygen. Even for protons, carbon ions and other heavy ions, the RBE of α particles generated by BNCT nuclear reaction is still the highest.

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

[0012] Chemotherapy, X-knife, γ-knife and ordinary radiotherapy generally have effects on tumor cells in the proliferation phase (G1, S, G2 and M phase), but are not sensitive to tumor cells in the resting phase (G0 phase). G0 phase tumor cells can still grow, which is the root cause of tumor recurrence. BNCT based on high LET, α particles and 7 Li particles kill tumor cells regardless of the cell growth cycle, and can also kill tumor cells in the resting phase. Hypoxic tumor cells become more resistant to low LET γ-rays, electron beams and other conventional radiotherapy, and BNCT-based treatment can kill hypoxic cancer cells as long as sufficient boron enters.

[0013] 4) Fine treatment scale

[0014] To date, only neutron capture therapy (NCT) has achieved a therapeutic range down to the micron level. Minimally invasive surgery, the most precise form of surgery, operates on a scale limited to the millimeter level. The Bragg peak focal dose depth in heavy ion therapy is approximately 2.5–3.0 cm. The finer the treatment area, 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 20-35 μg;

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

[0020] Currently, the development of BNCT is relatively slow, closely related to the relatively slow development of boron-containing drugs. To date, the only boron-containing drugs in clinical use are disodium thiododecaborane (BSH) and para-dicarboxyborylphenylalanine (BPA). BPA was approved for marketing in Japan in 2020, becoming the world's first approved boron-containing BNCT drug. While boron-containing drugs are a key breakthrough in BNCT technology, the effectiveness of BSH and BPA remains far from satisfactory, with issues such as insufficient tumor targeting specificity and insufficient affinity for tumor cells. In-depth research has found that the boron concentration ratio in tumor to normal tissue measured by BSH in human glioma samples is generally <1 (average 0.6), suggesting that it is a nonspecific boron carrier. BPA, on the other hand, generally produces a boron concentration ratio no greater than 2.4, preventing it from penetrating the internal components of tumor cells and resulting in only transient intracellular retention. Furthermore, some tumor cells do not take up BPA. Clinically, BNCT is primarily used to treat cancers such as brain tumors, gliomas, and melanomas. This is primarily due to the limited availability of boron-containing drugs and their limited indications. Therefore, the development of boron-targeted drugs with broader indications, lower toxicity, and better targeting is imperative.

[0021] Fibroblast activation protein (FAP) is one of the specific markers of tumor-associated fibroblasts (TAF), which has special biological characteristics, genome stability, and rich and specific expression in tumor stroma. Existing FAP inhibitors are mainly used for tumor treatment. However, there is no FAP-targeted drug that can be applied to boron neutron capture therapy. SUMMARY

[0022] The technical problem to be solved by the present application is that the existing compounds suitable for BNCT have single structure, and therefore the present application provides a benzopyridine ring compound and a preparation method thereof, which can be used for synthesizing the compound shown in formula I, the compound shown in formula I has low toxicity to tumor cells, and can be used for targeted cancer treatment in boron neutron capture therapy.

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

[0024]

[0025] wherein, R 1 is a single bond, C1~C 10 straight-chain alkylene, C2~C 10 straight-chain alkenylene, C2~C 10 branched alkylene, C5~C8cycloalkylene, C6~C 10 arylene-C1~C4alkylene, C1~C4alkylene-C6~C 10 arylene-C1~C4alkylene, or C6~C 10 arylene;

[0026] R 2 is hydroxyl, C1~C5straight-chain alkyl, C3~C6branched alkyl, C5~C8cycloalkyl, C6~C 10 aryl-C1~C4alkyl, C1~C4alkyl-C6~C 10 aryl-C1~C4alkyl, or C6~C 10 aryl;

[0027] R 3 is hydroxyl, C1~C5straight-chain alkyl, C3~C6branched alkyl, C5~C8cycloalkyl, C6~C 10 aryl-C1~C4alkyl, C1~C4alkyl-C6~C 10 aryl-C1~C4alkyl, or C6~C 10 aryl;

[0028] R 4H, F or Cl;

[0029] R 5 H, F or Cl;

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

[0031] In a certain embodiment, the compound of Formula I, or a pharmaceutically acceptable salt thereof, is described in which the definition of the moiety is as described below and the definition of the remaining moieties is as described in any other embodiment (hereinafter referred to as "in a certain embodiment"): R 1 may be a single bond.

[0032] In a certain embodiment, the compound of Formula I, or a pharmaceutically acceptable salt thereof, is described in which the definition of the moiety is as described below and the definition of the remaining moieties is as described in any other embodiment (hereinafter referred to as "in a certain embodiment"): R 2 may be a hydroxyl group.

[0033] In a certain embodiment, the compound of Formula I, or a pharmaceutically acceptable salt thereof, is described in which the definition of the moiety is as described below and the definition of the remaining moieties is as described in any other embodiment (hereinafter referred to as "in a certain embodiment"): R 3 may be a hydroxyl group.

[0034] In a certain embodiment, the compound of Formula I, or a pharmaceutically acceptable salt thereof, is described in which the definition of the moiety is as described below and the definition of the remaining moieties is as described in any other embodiment (hereinafter referred to as "in a certain embodiment"): R 4 may be F or Cl.

[0035] In a certain embodiment, the compound of Formula I, or a pharmaceutically acceptable salt thereof, is described in which the definition of the moiety is as described below and the definition of the remaining moieties is as described in any other embodiment (hereinafter referred to as "in a certain embodiment"): R 4 may be F.

[0036] In a certain embodiment, the compound of Formula I, or a pharmaceutically acceptable salt thereof, is described in which the definition of the moiety is as described below and the definition of the remaining moieties is as described in any other embodiment (hereinafter referred to as "in a certain embodiment"): R 5 may be F or Cl.

[0037] In a certain embodiment, the compound of Formula I, or a pharmaceutically acceptable salt thereof, is described in which the definition of the moiety is as described below and the definition of the remaining moieties is as described in any other embodiment (hereinafter referred to as "in a certain embodiment"): R 5 may be F.

[0038] In a certain embodiment, the compound of Formula I is Compound I-1:

[0039] .

[0040] The present application also provides a compound of Formula 2, Formula 3, Formula 4, Formula 5 or Formula 6:

[0041] , , , or

[0042] wherein R 51 is C1-C4 alkyl;

[0043] In the compound of Formula 3, R 32 is C1-C3 alkyl; R 33 is C1-C3 alkyl; R 34 is C1-C3 alkyl;

[0044] R is C1-C3 alkyl; R 52 is C1-C3 alkyl; R 53 is C1-C3 alkyl; R 54 is C1-C3 alkyl; R 55 is C1-C3 alkyl;

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

[0046] In one aspect, the compound of Formula 2 is compound 2-1:

[0047] .

[0048] In one aspect, the compound of Formula 3 is compound 3-1:

[0049] .

[0050] In one aspect, the compound of Formula 4 is compound 4-1:

[0051] .

[0052] In one aspect, the compound of Formula 5 is compound 5-1:

[0053] .

[0054] In one aspect, the compound of Formula 6 is compound 6-1:

[0055] .

[0056] The present application also provides a method for preparing the compound of Formula I, comprising the following steps: reacting a compound of Formula 6 with a compound of Formula a in a solvent in the presence of an amidation reagent to produce a compound of Formula I, i.e.

[0057] .

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

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

[0060] In one aspect, the amidation reagent can be a combination of N,N,N',N'-tetramethylchloroformamidinium hexafluorophosphate (TCFH) and 1-methylimidazole.

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

[0062] In an embodiment, the solvent can be acetonitrile.

[0063] In an embodiment, the compound of formula 6 can be compound 6-1 described above.

[0064] In an embodiment, the molar volume ratio of the compound of formula 6 to the solvent can be 0.121 mol / L.

[0065] In an embodiment, the molar ratio of the compound of formula 6 to N,N,N',N'-tetramethylchloroformamidinium hexafluorophosphate (TCFH) in the amidation reagent can be 0.845.

[0066] In an embodiment, the molar volume ratio of the compound of formula 6 to 1-methylimidazole in the amidation reagent can be 2.417 mol / L.

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

[0068] In an embodiment, the reaction condition can be stirring for 2 hours.

[0069] In an embodiment, the preparation method can further comprise isolation of the reaction product, which can be acidification, extraction, washing, drying, column chromatography in sequence.

[0070] In an embodiment, the acidification can be acidification using trifluoroacetic acid. Preferably, 1% trifluoroacetic acid acidification.

[0071] In an embodiment, the extraction can be ethyl acetate extraction. Preferably, equal volume ethyl acetate extraction.

[0072] In an embodiment, the washing can be sodium bicarbonate solution washing. Preferably, equal volume sodium bicarbonate solution washing.

[0073] In an embodiment, the drying can be anhydrous sodium sulfate drying.

[0074] In an embodiment, the developing agent used in the column chromatography can be petroleum ether, ethyl acetate.

[0075] In an embodiment, the acidification can be acidification using 1% trifluoroacetic acid; the extraction can be equal volume ethyl acetate extraction for 3 times; the washing can be equal volume sodium bicarbonate solution washing for 2 times; the drying can be anhydrous sodium sulfate drying; the developing agent used in the column chromatography can be petroleum ether, ethyl acetate.

[0076] In an embodiment, the compound of formula I is a compound of formula I-2: .

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

[0078] (2) reacting the compound of formula 2 with a compound of formula B in a solvent to obtain a compound of formula 3;

[0079] (3) reacting the compound of formula 3 in a solvent to obtain a compound of formula 4;

[0080] (4) reacting the compound of formula 4 with a compound of formula C in a solvent to obtain a compound of formula 5-2;

[0081] (5) reacting the compound of formula 5-2 in a solvent to obtain a compound of formula 6-2;

[0082] (6) amidating a compound of formula 6-1 with a compound of formula A in the presence of an amidation reagent in a solvent to obtain a compound of formula I-2, and

[0083] .

[0084] The solvent is a solvent commonly used in the art for such reactions.

[0085] In an embodiment, in step (1), the reaction can be carried out at normal pressure; the reaction can be carried out in the presence of oxygen; the solvent can be methanol; the compound of formula D can be methanol; the reaction can be carried out in the presence of dichlorosulfoxide; 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 further comprise isolation of the reaction product, which can be carried out by sequentially adjusting the pH of the reaction solution and filtering.

[0086] In an embodiment, in step (1), the molar volume ratio of the compound of formula 1 to the solvent can be 0.8 mol / L; the molar volume ratio of the compound of formula 1 to the compound of formula D can be 0.8 mol / L; the molar volume ratio of the compound of formula 1 to dichlorosulfoxide can be 6.6 mol / L; the pH of the reaction solution can be adjusted using sodium bicarbonate.

[0087] In an embodiment, in step (2), the reaction can be carried out under normal pressure; the reaction can be carried out under inert atmosphere; the solvent can be tetrahydrofuran; the compound of formula 2 can be compound 2-1 described above; the compound of formula B can be trimethylsilyl ethynyl; the reaction can 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 further comprise isolation and purification of the reaction product, which can be filtration of the reaction solution, dilution, extraction, washing, drying, filtration, concentration and purification in sequence.

[0088] In an embodiment, in step (2), the molar volume ratio of the compound of formula 2 to the solvent can be 0.278 mol / L; the molar ratio of the compound of formula 2 to the compound of formula B can be 0.5; the molar ratio of the compound of formula 2 to trimethylsilyl ethynyl can be 20; the molar ratio of the compound of formula 2 to cuprous iodide can be 20; the molar ratio of the compound of formula 2 to triethylamine can be 0.67; the extraction in the isolation and purification can be ethyl acetate extraction; the purification in the isolation and purification can be column chromatography purification.

[0089] In an embodiment, in step (2), in the column chromatography purification in the isolation and purification, the developing agent can be petroleum ether and ethyl acetate.

[0090] In an embodiment, in step (3), the reaction can be carried out under normal pressure; the reaction can be carried out under inert atmosphere; the compound of formula 3 can be compound 3-1 described above; 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 further comprise isolation and purification of the reaction product, which can be filtration of the reaction solution, dilution, extraction, washing, drying, filtration, concentration, slurry, filtration and drying in sequence.

[0091] In an embodiment, in step (3), the molar volume ratio of the compound of formula 3 to the solvent can be 0.45 mol / L; the molar ratio of the compound of formula 3 to potassium fluoride can be 0.33; the extraction in the isolation and purification can be ethyl acetate extraction; the slurry in the isolation and purification can be slurry with petroleum ether and ethyl acetate.

[0092] In a certain embodiment of the step (4), the reaction can be carried out under normal pressure; the reaction can be carried out under inert atmosphere; the solvent can be toluene; the compound of formula 4 can be compound 4-1 described above; the compound of formula C can be pinacol borane; the reaction can be carried out in the presence of carbonylchlorohydrogen tris(triphenylphosphine) ruthenium (II); the reaction temperature can be 50℃; the reaction time can be 14 hours; and the reaction can further comprise isolation and purification of the reaction product, which can be filtration of the reaction solution, dilution, extraction, washing, drying, filtration, concentration and purification in sequence.

[0093] In a certain embodiment of the step (4), the molar volume ratio of the compound of formula 4 to the solvent can be 0.47 mol / L; the molar ratio of the compound of formula 4 to the compound of formula C can be 0.26; the molar ratio of the compound of formula 4 to carbonylchlorohydrogen tris(triphenylphosphine) ruthenium (II) can be 71; the extraction in the isolation and purification can be ethyl acetate extraction; and the purification in the isolation and purification can be column chromatography purification.

[0094] In a certain embodiment of the step (4), in the column chromatography purification in the isolation and purification, the developing agent can be petroleum ether and ethyl acetate.

[0095] In a certain embodiment of 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 of formula 5-2 can be compound 5-1 described above; the reaction can be carried out in the presence of hydrochloric acid; the reaction temperature can be 85℃; the reaction time can be 12 hours; and the reaction can further comprise isolation and purification of the reaction product, which can be extraction of the reaction solution, liquid separation, drying, beating, filtration and drying in sequence.

[0096] In a certain embodiment of the step (5), the molar volume ratio of the compound of formula 5-2 to the solvent can be 0.59 mol / L; the concentration of the hydrochloric acid can be 6 mol / L; the molar volume ratio of the compound of formula 5-2 to the hydrochloric acid can be 0.39 mol / L; the extraction in the isolation and purification can be ethyl acetate extraction; and the beating in the isolation and purification can be beating with methyl tert-butyl ether.

[0097] In a certain embodiment of the step (6), the reaction conditions can refer to the reaction conditions in the preparation method of the compound of formula I described above.

[0098] The present application also provides a pharmaceutical composition comprising substance X and a pharmaceutically acceptable excipient; the substance X is the compound of formula I described above or a pharmaceutically acceptable salt thereof.

[0099] The application also provides a use of a substance X in the preparation of a medicament for treating tumors; the substance X is the compound of formula I or a pharmaceutically acceptable salt thereof; the medicament is a medicament for treating tumors.

[0100] In some embodiments, the medicament for treating tumors is a radiotherapy targeting medicament.

[0101] In some embodiments, the radiotherapy targeting medicament is a boron neutron capture therapy medicament.

[0102] Unless otherwise specified, the terms used in the present application have the following meanings:

[0103] The term "alkyl" refers to a straight chain or branched alkyl group having the specified number of carbon atoms (e.g., C1-C4). Alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, i-propyl, n-butyl, t-butyl, i-butyl, s-butyl, and the like.

[0104] The term "alkylene" refers to a straight chain or branched divalent alkyl group having the specified number of carbon atoms (e.g., C1-C4) that connects to other groups at two sites. Alkylene groups include, but are not limited to, , , , and the like.

[0105] The term "alkenylene" refers to a straight chain or branched divalent alkenyl group having the specified number of carbon atoms (e.g., C2-C4) that connects to other groups at two sites. Alkenylene groups include, but are not limited to, , , and the like.

[0106] The term "cycloalkyl" refers to a saturated monocyclic ring group consisting only of carbon atoms having the specified number of carbon atoms (e.g., C5-C8). Cycloalkyl groups include, but are not limited to, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, and the like.

[0107] The term "aryl" refers to a cyclic group consisting only of carbon atoms having the specified number of carbon atoms (e.g., C6-C 10 ), which is monocyclic or polycyclic, and each ring is aromatic (complying with Huckel's rule). Aryl groups include, but are not limited to, phenyl, naphthyl, and the like.

[0108] The term "arylene" refers to a divalent cyclic group consisting only of carbon atoms having the specified number of carbon atoms (e.g., C6-C 10 ), which is monocyclic or polycyclic, and each ring is aromatic (complying with Huckel's rule), that connects to other groups at two sites. Arylene groups include, but are not limited to, , and the like.

[0109] The term "pharmaceutically acceptable salt" refers to a salt of a compound that is produced by reaction of the compound with a pharmaceutically acceptable (relatively non-toxic, safe, suitable for patient use) 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 the 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 the pharmaceutically acceptable acid in a suitable inert solvent. Pharmaceutically acceptable acid addition salts include, but are not limited to, hydrochloride salts, sulfate salts, methanesulfonate salts, and the like. See Handbook of Pharmaceutical Salts: Properties, Selection, and Use (P. Heinrich Stahl, 2002) for details.

[0110] On the basis of common general knowledge in the art, the above-mentioned preferred conditions can be combined in any manner, thereby obtaining preferred embodiments of the present application.

[0111] The reagents and raw materials used in the present application are commercially available.

[0112] The positive progress effect of the present application is that the compound has low toxicity to tumor cells and can target cancer treatment by boron neutron capture therapy. DETAILED DESCRIPTION

[0113] The present application is further illustrated by the following examples, but the present application is not limited to the scope of the examples. The experimental methods in the following examples, for which no specific conditions are indicated, are selected according to conventional methods and conditions, or according to the instructions of the commercial products.

[0114] The A549 cells, HepG2 cells, U87MG cells, SCC9 cells, and A375 cells in the following examples are from the Shanghai Cell Bank of the Chinese Academy of Sciences;

[0115] In the following examples

[0116] The CCK-8 cell viability detection kit is produced by Shanghai Donguan Biological Technology Co., Ltd.

[0117] The PBS is produced by Shanghai Shuangmo Biological Technology Co., Ltd.

[0118] The DMEM high-sugar culture solution is produced by Shanghai Donguan Biological Technology Co., Ltd.

[0119] Example 1 Preparation of the compound I-1 of the present application

[0120]

[0121] 1. Preparation of compound I-1

[0122] The preparation route of I-1 is as follows:

[0123]

[0124] First step: Compound 1 (50 g, 198 mmol) was dissolved in methanol (250 ml), and dichlorosulfoxide (30 ml) was slowly added at room temperature. The reaction solution was reacted at 80°C for 4 hours. The reaction solution was quenched with water (50 ml), and the pH was adjusted to 7 with sodium bicarbonate solution. Filtration was performed, and the solid was collected. White compound 2-1 was obtained with a yield of 93%.

[0125] The identification data of compound 2-1 are as follows: LC-MS: M (C 11 H8BrNO2) = 266.09 (m / z), [M+H] + 267.2

[0126] Second step: Compound 2-1 (37.0 g, 139 mmol), trimethylacetylenylsilane (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 twice with ethyl acetate (250 ml / time). The organic phase was combined, washed twice with saturated brine (100 ml / time), and dried with anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure, and column chromatography purification was performed (developing agent: petroleum ether, ethyl acetate). Compound 3-1 was obtained as a yellow solid with a yield of 96%.

[0127] 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

[0128] Third step: 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 phase was combined and washed with saturated brine twice (100 ml / time). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. 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 as a yellow solid after filtration and drying, with a yield of 53%.

[0129] The identification data of compound 4-1 are as follows: LC-MS: M (C 13 H9NO2) =211.22 (m / z), [M+H] + 212.0

[0130] Fourth step: compound 4-1 (15 g, 71 mmol) was dissolved in toluene (150 ml), and carbonyl chloro hydrogen tris (triphenylphosphine) ruthenium (II) (16971-33-8) (1 g, 1 mmol) and pinacol borane (c) (35.3 g, 275 mmol) were slowly added under nitrogen protection. The reaction solution was stirred at 50°C for 14 hours under nitrogen protection. The reaction solution was filtered and collected, diluted with water (50 ml), and extracted with ethyl acetate twice (100 ml / time). The organic phase was combined and washed with saturated brine twice (50 ml / time). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. Column chromatography was used for purification (developing agent: petroleum ether, ethyl acetate). Compound 5-1 was obtained as a yellow solid, with a yield of 85%.

[0131] 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

[0132] Fifth step: 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 / time). The aqueous phase was collected and freeze-dried. 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%.

[0133] The identification data of compound 6-1 are as follows: LC-MS: M (C 12 H10 BNO4) =243.03 (m / z), [M+H] + 244.1

[0134] Step 6: Compound 6-1 (8 g, 29 mmol), compound a (18.9 g, 15.7 mmol) were dissolved in acetonitrile (240 ml), N,N,N',N'-tetramethylchloroformamidium hexafluorophosphate (TCFH) (9.6 g, 34.3 mmol), 1-methylimidazole (12 ml, 12.4 g) were added slowly. 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 phase was combined and washed with sodium bicarbonate solution twice (200 ml / time), and the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and then purified by column chromatography (developing agent: petroleum ether, ethyl acetate). The product I-1 was obtained as a yellow solid in a yield of 46%.

[0135] 2. Structure identification

[0136] The LC-MS, NMR identification data of compound I-1 are as follows:

[0137] LC-MS: M (C 17 H 1 BF2N4O4) =414.18 (m / z), [M+H]+ 415.2

[0138] 4 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).

[0139] F NMR: (400 MHz, DMSO-d6) δ -95.054, -103.651.

[0140] Example 2: Cytotoxicity of compound I-1 on A549 cells

[0141] A549 cells in the logarithmic growth phase were inoculated on a 96-well plate at 1*10 50Cells were cultured overnight in DMEM high-glucose medium. The medium was aspirated and the cells were washed twice with PBS (0.01 mol / L). Compound I-1 was added to fresh culture medium to prepare a series of solutions containing compound I-1 at concentrations of 0, 100, 250, 500, and 750 µg / ml. 100 µl of the solution was added to the test cell wells. After 48 hours of drug treatment, the cell culture medium was aspirated with a pipette, the cells were washed twice with PBS (0.01 mol / L), and 100 µL of CCK8 working solution was added. The culture plates were incubated in an incubator for 2 hours. The absorbance at 450 nm was measured using a microplate reader to calculate the effect of the drug on cell viability.

[0142] 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, and its IC 50 It is 458.1 μg / ml, which meets the low toxicity requirement of boron neutron capture therapy for boron-containing drugs.

[0143] Table 1 Inhibitory rate of A549 cells by different concentrations of compounds

[0144]

[0145] Example 3: Cytotoxicity of Compound I-1 on HepG2 cells

[0146] HepG2 cells in the logarithmic growth phase were seeded in 96-well plates, with 1*10 cells per well. 4 Cells were cultured overnight in DMEM high-glucose medium. The medium was aspirated and the cells were washed twice with PBS (0.01 mol / L). Compound I-1 was added to fresh culture medium to prepare a series of solutions containing compound I-1 at concentrations of 0, 100, 250, 500, and 750 µg / ml. 100 µl of the solution was added to the test cell wells. After 48 hours of drug treatment, the cell culture medium was aspirated with a pipette, the cells were washed twice with PBS (0.01 mol / L), and 100 µL of CCK8 working solution was added. The culture plates were incubated in an incubator for 2 hours. The absorbance at 450 nm was measured using a microplate reader to calculate the effect of the drug on cell viability.

[0147] 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 its IC 50340.1 μg / ml, which meets the requirement of low toxicity of boron-containing drugs for boron neutron capture therapy.

[0148] Table 2 Inhibition rate of different concentrations of the compound on HepG2 cells

[0149]

[0150] Example 4: Cytotoxicity of compound I-1 on U87MG cells

[0151] U87MG cells in the logarithmic growth phase were inoculated on a 96-well plate at 1*10 4 cells per well, and were cultured in DMEM high-glucose medium overnight. The medium was aspirated, and the cells were washed twice with PBS (0.01 mol / L). Compound I-1 was added to fresh medium to prepare a series of concentration gradients of the compound I-1 solution, and the concentration of compound I-1 was 0, 100, 250, 500, and 750 μg / ml, respectively. 100 μl was taken and added to the cell well plate to be tested. After the cells were treated with the drug for 48 hours, the cell culture medium was aspirated, the cells were washed twice with PBS (0.01 mol / L), 100 uL of CCK8 working solution was added, and the culture plate was incubated in an incubator for 2 hours. The absorbance at 450 nm was determined by an enzyme marker, and the effect of the drug on cell viability was calculated.

[0152] The test results are shown in Table 3. The low cytotoxicity of compound I-1 is an important indicator of BNCT, which can maximize the boron concentration in tumor cells. The compound I-1 described in the present application exhibits low toxicity to tumor cells, and the IC 50 411.8 μg / ml, which meets the requirement of low toxicity of boron-containing drugs for boron neutron capture therapy.

[0153] Table 3 Inhibition rate of different concentrations of the compound on U87MG cells

[0154]

[0155] Example 5: Cytotoxicity of compound I-1 on SCC9 cells

[0156] SCC9 cells in the logarithmic growth phase were inoculated on a 96-well plate at 1*10 4Cells were cultured overnight in DMEM high-glucose medium. The medium was aspirated and the cells were washed twice with PBS (0.01 mol / L). Compound I-1 was added to fresh culture medium to prepare a series of solutions containing compound I-1 at concentrations of 0, 100, 250, 500, and 750 µg / ml. 100 µl of the solution was added to the test cell wells. After 48 hours of drug treatment, the cell culture medium was aspirated with a pipette, the cells were washed twice with PBS (0.01 mol / L), and 100 µL of CCK8 working solution was added. The culture plates were incubated in an incubator for 2 hours. The absorbance at 450 nm was measured using a microplate reader to calculate the effect of the drug on cell viability.

[0157] 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 its IC 50 It is 289.1 μg / ml, which meets the low toxicity requirement of boron neutron capture therapy for boron-containing drugs.

[0158] Table 4 Inhibitory rate of SCC9 cells at different concentrations of compounds

[0159]

[0160] Example 6: Cytotoxicity of Compound I-1 on A375 cells

[0161] 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 aspirated and the cells were washed twice with PBS (0.01 mol / L). Compound I-1 was added to fresh culture medium to prepare a series of solutions containing compound I-1 at concentrations of 0, 100, 250, 500, and 750 µg / ml. 100 µl of the solution was added to the test cell wells. After 48 hours of drug treatment, the cell culture medium was aspirated with a pipette, the cells were washed twice with PBS (0.01 mol / L), and 100 µL of CCK8 working solution was added. The culture plates were incubated in an incubator for 2 hours. The absorbance at 450 nm was measured using a microplate reader to calculate the effect of the drug on cell viability.

[0162] 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, and its IC 50The concentration of the compound is 586.5 μg / ml, which meets the requirement of low toxicity of boron-containing drugs for boron neutron capture therapy.

[0163] Table 5 Inhibition rate of different concentrations of the compound on A375 cells

[0164] .

Claims

1. A compound as shown in Formula 4: in, R 51 It is a C1~C4 alkyl group.

2. The compound as described in claim 1, wherein The compound represented by Formula 4 is compound 4-1:

3. A method for preparing a compound as shown in formula 4, characterized in that: In a solvent, in the presence of potassium fluoride, the compound represented by formula 3 is subjected to the following reaction to obtain the compound represented by formula 4. R 51 is a C1-C4 alkyl group; Among them, in the compound shown in Formula 3, R 32 is a C1-C3 alkyl group; R 33 is a C1-C3 alkyl group; R 34 It is a C1~C3 alkyl group.

4. The preparation method according to claim 3, wherein 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 compound represented by formula 3 is compound 3-1: (4) The solvent is a mixture of methanol and dichloromethane; (5) The reaction time is 12 hours; (6) The reaction also includes separation and purification of the reaction product, and the separation and purification is sequentially filtering the reaction solution, diluting, extracting, washing, drying, filtering, concentrating, beating, filtering and drying.

5. The preparation method according to claim 4, wherein: It meets one or more of the following conditions: (1) The molar volume ratio of the compound shown in 3 to the solvent is 0.45 mol / L; (2) the molar ratio of the compound represented by formula 3 to potassium fluoride is 0.33; (3) The extraction in the separation and purification is ethyl acetate extraction; (4) The pulping in the separation and purification is performed by using petroleum ether and ethyl acetate.

6. A compound as shown in Formula 3: in, R 32 is a C1-C3 alkyl group; R 33 is a C1-C3 alkyl group; R 34 is a C1-C3 alkyl group; R 51 is a C1-C4 alkyl group; the compound shown in Formula 3 is not 7. The compound as shown in formula 3 according to claim 6, characterized in that The compound represented by Formula 3 is Compound 3-1:

8. A method for preparing a compound as shown in formula 3, characterized in that: In a solvent, in the presence of bis(triphenylphosphine)palladium(II) chloride, cuprous iodide and triethylamine, the compound represented by Formula 2 and the compound represented by Formula B are reacted as shown below to obtain the compound represented by Formula 3. Among them, R 32 is a C1-C3 alkyl group; R 33 is a C1-C3 alkyl group; R 34 is a C1-C3 alkyl group; R 51 It is a C1~C4 alkyl group.

9. The preparation method according to claim 8, wherein 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 tetrahydrofuran; (4) The compound represented by formula 2 is compound 2-1: (5) The compound represented by formula B is trimethylethynylsilane; (6) The reaction temperature is 50°C; (7) The reaction also includes separation and purification of the reaction product, which is carried out in sequence by filtering the reaction solution, diluting, extracting, washing, drying, filtering, concentrating and purifying.

10. The preparation method according to claim 9, characterized in that: It meets one or more of the following conditions: (1) The molar volume ratio of the compound represented by Formula 2 to the solvent is 0.278 mol / L; (2) The molar ratio of the compound represented by Formula 2 to the compound represented by Formula B is 0.5; (3) The molar ratio of the compound represented by Formula 2 to trimethylethynylsilane is 20; (4) The molar ratio of the compound represented by Formula 2 to cuprous iodide is 20; (5) The molar ratio of the compound represented by Formula 2 to triethylamine is 0.67; (6) 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 solvents in the column chromatography purification in the separation and purification are petroleum ether and ethyl acetate.

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