Boric acid ester compound as well as preparation method and application thereof

Abstract

The invention relates to the technical field of medicines, in particular to a borate compound as well as a preparation method and application thereof. Wherein the structural formula of the boric acid ester compound is as shown in I. According to the invention, through molecular structure design, 3-carboxyl is utilized for modification, water solubility and intramolecular hydrogen bonds are enhanced, hydrolysis is inhibited, and stability is improved; a uracil structure is utilized, thymidine / deoxyuridine is simulated, the thymidine / deoxyuridine is actively taken by TK1 high-expression cancer cells and is enriched near DNA in a targeted manner, the BNCT efficiency is improved, and a novel boron drug with high water solubility, stability and targeting property is successfully developed. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical technology, and particularly relates to a borate compound, a preparation method thereof, and an application thereof. Background Art

[0002] Radiotherapy is a common means for treating tumors. However, for tumors in some special locations (such as glioblastoma, which grows deep in the body), the effect of conventional radiotherapy is not good. As a new radiotherapy method, Boron Neutron Capture Therapy (BNCT) has significant advantages.

[0003] Boron Neutron Capture Therapy (BNCT) is a targeted particle radiation therapy based on 10 the nuclear capture reaction of ¹⁰B. Its principle is to enrich the boron agent containing 10 ¹⁰B into tumor cells, and then irradiate with an external thermal neutron beam to trigger 10 the ¹⁰B(n,α) 7 ⁷Li nuclear reaction, releasing high-energy α particles and lithium ions. These particles have a very short range (only about 5 - 9 microns), and the energy is almost completely concentrated within the range of a single cell, which can accurately damage the DNA of tumor cells, thereby killing tumor cells, while causing less damage to surrounding normal tissues.

[0004] The core problem of existing boron agents lies in their "functional singularity", that is, most boron agents only rely on the passive uptake mechanism of the tumor microenvironment (such as the enhanced permeability and retention effect), lacking active targeting function. This leads to unstable selective adsorption efficiency of tumor cells for 10 ¹⁰B, and it is difficult to simultaneously meet the comprehensive requirements of high enrichment, long-term retention, and low toxicity. For example, commonly used BPA (boron phenylalanine) and BSH (sodium thioborate) can enrich in tumor tissues to a certain extent, but due to insufficient targeting, it is necessary to increase the dosage during treatment, which may exacerbate the risk of systemic toxicity. This limitation not only affects the treatment effect but also restricts the safety and wide promotion of Boron Neutron Capture Therapy (BNCT) in clinical applications. Summary of the Invention

[0005] The main object of the present application is to provide a borate compound, a preparation method thereof, and an application thereof, aiming to improve the water solubility and chemical stability of the boron agent.

[0006] To achieve the above object, a borate compound proposed in the present application has a structural formula as shown in Formula I:

[0007]

[0008] In some embodiments, the solubility of the borate ester compound in water is ≥50 mmol / L at 10°C to 30°C.

[0009] In some embodiments, the hydrolysis rate of the borate ester compound in water is <10% in 24 hours at 10°C to 30°C.

[0010] On the other hand, the present application also provides a preparation method of a borate ester compound, comprising the following steps: mixing uridine with 3-carboxyphenylboronic acid, performing an esterification reaction in a polar solvent, performing a cyclic reaction 2 to 5 times to obtain a crude product, and purifying the crude product to obtain the borate ester compound.

[0011] In some embodiments, the molar ratio of the uridine to the 3-carboxyphenylboronic acid is 3:1 to 1:1; and / or, the ratio of the total mass of the uridine and the 3-carboxyphenylboronic acid to the volume of the polar solvent is (0.5 - 4) g: 5 mL.

[0012] In some embodiments, the temperature of the esterification reaction is 120°C to 200°C.

[0013] In some embodiments, the organic solvent includes at least one of dimethyl sulfoxide, N,N-dimethylformamide, and water.

[0014] In some embodiments, the purification treatment includes a recrystallization step, and the recrystallization step is performed in ethanol.

[0015] In some embodiments, after the recrystallization step, the recrystallized product is dissolved in a water-methanol mixed mobile phase, and is subjected to high-pressure liquid preparation separation through a C18 reverse-phase chromatography column, and the target eluate is collected to obtain the borate ester compound.

[0016] On the other hand, the present application also provides an application of a borate ester compound in the preparation of a boron neutron capture therapy drug, and the borate ester compound is the above-mentioned borate ester compound or the borate ester compound prepared by the above-mentioned preparation method.

[0017] The present application combines uridine with 3-carboxyphenylboronic acid to form a borate ester compound with both water solubility, stability and targeting properties. Utilizing the affinity of thymidine kinase (TK1) highly expressed in cancer cells for nucleoside substances, the enrichment of boron drugs in the tumor cell nucleus or sensitive targets is achieved, and the selective killing effect of BNCT is enhanced. The problems of poor water solubility, insufficient stability, weak targeting and toxicity of existing BNCT boron agents are solved. Description of the Drawings

[0018] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for the description of the embodiments or the prior art. Obviously, the drawings in the following description are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can be obtained based on the structures shown in these drawings.

[0019] Figure 1 is the infrared spectrum of the borate compound (3-carboxyphenylboronic acid uridine monoester) in Example 1 of the present application;

[0020] Figure 2 is the nuclear magnetic H 1 spectrum of the borate compound (3-carboxyphenylboronic acid uridine monoester) in Example 1 of the present application;

[0021] Figure 3 is the mass spectrum (negative ion) of the aqueous solution of 3-carboxyphenylboronic acid uridine diester in Comparative Example 1 of the present application;

[0022] Figure 4 is the mass spectrum (negative ion) of the aqueous solution of 3-carboxyphenylboronic acid uridine monoester in Example 1 of the present application;

[0023] Figure 5 is the comparison chart of the cell survival rates of 3-carboxyphenylboronic acid uridine monoester (UB) in Example 1 of the present application and BPA in Comparative Example 2.

[0024] The realization, functional characteristics, and advantages of the object of the present invention will be further described with reference to the embodiments and the accompanying drawings. Detailed Embodiments

[0025] The following will clearly and completely describe the technical solutions in the embodiments of the present invention with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, rather than all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts belong to the scope of protection of the present invention.

[0026] Hereinafter, the non-aqueous anode sodium metal battery, electrolyte, its preparation method, and electrical device of the present application will be specifically disclosed in detail with appropriate reference to the accompanying drawings. However, there may be cases where unnecessary detailed descriptions are omitted. For example, there are cases where detailed descriptions of well-known matters and repeated descriptions of actually identical structures are omitted. This is to avoid the following description from becoming unnecessarily lengthy and to facilitate the understanding of those skilled in the art. In addition, the accompanying drawings and the following description are provided for those skilled in the art to fully understand the present application and are not intended to limit the subject matter described in the claims.

[0027] The "ranges" disclosed in this application are defined in the form of a lower limit and an upper limit. A given range is defined by selecting a lower limit and an upper limit, and the selected lower limit and upper limit define the boundaries of a particular range. The ranges defined in this way can include the end values or not include the end values, and can be combined arbitrarily, that is, any lower limit can be combined with any upper limit to form a range. For example, if ranges of 60 - 120 and 80 - 110 are listed for a specific parameter, ranges of 60 - 110 and 80 - 120 are also contemplated. In addition, if the minimum range values 1 and 2 are listed, and if the maximum range values 3, 4, and 5 are listed, then the following ranges are all contemplated: 1 - 3, 1 - 4, 1 - 5, 2 - 3, 2 - 4, and 2 - 5. In this application, unless otherwise specified, the numerical range "a - b" represents an abbreviated representation of any real number combination between a and b, where a and b are both real numbers. For example, the numerical range "0 - 5" means that all real numbers between "0 - 5" are fully listed herein, and "0 - 5" is only an abbreviated representation of these numerical combinations. Additionally, when stating that a certain parameter is an integer ≥2, it is equivalent to disclosing that the parameter is, for example, the integers 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, etc.

[0028] If there is no special instruction, all embodiments and optional embodiments of this application can be combined with each other to form new technical solutions.

[0029] If there is no special instruction, all technical features and optional technical features of this application can be combined with each other to form new technical solutions.

[0030] If there is no special instruction, all steps of this application can be carried out in sequence or randomly, and preferably in sequence. For example, the method includes steps (a) and (b), which means that the method can include steps (a) and (b) carried out in sequence, or can also include steps (b) and (a) carried out in sequence. For example, it is mentioned that the method may further include step (c), which means that step (c) can be added to the method in any order. For example, the method can include steps (a), (b), and (c), or can also include steps (a), (c), and (b), or can also include steps (c), (a), and (b), etc.

[0031] Existing BNCT drugs (such as BPA, BSH) have low solubility (requiring high - dose administration) and insufficient in - vivo stability (rapid hydrolysis), resulting in insufficient boron concentration at the tumor site, making it difficult to trigger an effective neutron capture reaction and limiting the therapeutic effect.

[0032] A borate ester compound proposed in this application has a structural formula as shown in Formula Ⅰ:

[0033]

[0034] The molecular structure design of this compound is based on the synergistic effect of two major functional modules: the construction of a targeting skeleton and the design of a hydrophilic stabilizing unit. Its targeting skeleton constructs a six-membered heterocyclic structure with a uracil ring as the core, and the distribution characteristics of its nitrogen and oxygen atoms are highly similar to those of thymidine / deoxyuridine. This structure simulation strategy enables the compound to be actively recognized and taken up by thymidine kinase (TK1) highly expressed in cancer cells, achieving precise enrichment in the tumor cell nucleus and providing a positioning basis for subsequent boron neutron capture therapy (BNCT). Its hydrophilic stabilizing unit is achieved by introducing the following modifying groups into the phenylborate group: (1) The carboxyl group (-COOH) at the 3-position of the benzene ring ionizes to carboxylate (-COO - ) at physiological pH, significantly enhancing the molecular polarity; (2) The hydroxyl groups (-OH) and amino groups (-NH-) distributed near the uracil ring and glycosidic bond form a three-dimensional hydrogen bond network with water molecules, further expanding the hydrophilic interface and breaking through the hydrophobic barrier of traditional boron agents; (3) A monoester structure is used to connect the phenylborate and uracil (instead of the easily hydrolyzed diester bond), and combined with the stabilizing effect of the π-electron conjugation system of the benzene ring, the hydrolysis rate of the compound is reduced; (4) The hydroxyl group forms an intramolecular hydrogen bond with the adjacent carbonyl / amino group, locking the conformation of the borate group; at the same time, the steric structures of the uracil ring and benzene ring form steric hindrance to shield the attack of external nucleophiles, and the double protection significantly improves the chemical stability.

[0035] In summary, through the synergistic effect of multiple hydrophilic groups (carboxylate, hydroxyl, amino), the solubility of this compound is improved compared with traditional drugs; the anti-hydrolysis property of the monoester bond combined with the intramolecular stabilization mechanism prolongs the in vivo circulation time; the characteristics of the uracil analog drive TK1-mediated tumor-selective enrichment and optimize the targeting property.

[0036] In some embodiments, the solubility of the borate compound in water is ≥50 mmol / L at 10°C to 30°C.

[0037] Traditional boron agents (such as the solubility of BPA is only 5 mmol / L) have poor water solubility, require high-concentration administration or rely on complex formulations (such as amino acid complexes), resulting in limited clinical use and increased patient burden. The borate compound of the embodiments of this application has a high solubility higher than 50 mmol / L. The high solubility allows for higher-dose administration, enabling more boron atoms to accumulate at the tumor site. At the same time, the high solubility can also avoid the use of cosolvents or carriers, reducing the risk of allergies or metabolic burden.

[0038] In some embodiments, the hydrolysis rate of the borate compound in water is <10% in 24 hours at 10°C to 30°C.

[0039] Traditional boron agents have the problem of easy hydrolysis, resulting in a short in vivo circulation time and low tumor enrichment efficiency; or the prematurely released boric acid loses its targeting ability, causing damage to normal tissues. The borate compounds of the embodiments of the present application inhibit hydrolysis through the design of a monoester bond and intramolecular hydrogen bond, enabling the compounds to remain intact in the blood, extending the tumor targeting window, and reducing off-target toxicity. The stability at 10 - 30 °C covers both normal temperature storage and in vivo physiological temperature, ensuring the stability of the drug throughout the process from storage to use.

[0040] On the other hand, the present application also provides a preparation method of a borate compound, comprising the following steps: mixing uridine with 3-carboxyphenylboronic acid, performing an esterification reaction in an organic solvent, circulating the reaction 2 - 5 times to obtain a crude product, and purifying the crude product to obtain the borate compound.

[0041] In the embodiments of the present application, the borate compound is synthesized through an esterification reaction. Due to the large difference in the reaction activity between boric acid and hydroxyl groups, problems such as the generation of diesters and the residue of unreacted raw materials are likely to occur. In this preparation method, through 2 - 5 cycles of reaction, the unreacted 3-carboxyphenylboronic acid is gradually consumed, promoting the progress of the esterification reaction. By means of a purification process, the monoester product is separated from the residual raw materials and diester impurities, improving the purity of the product.

[0042] In some embodiments, the molar ratio of uridine to 3-carboxyphenylboronic acid is from 3:1 to 1:1.

[0043] Uridine contains multiple hydroxyl groups (such as the 2', 3', and 5'-hydroxyl groups of ribose). If 3-carboxyphenylboronic acid is in excess (such as a molar ratio of 1:3), it is likely to react with multiple hydroxyl groups to form diester or polyester by-products (with unstable structures and poor targeting properties); when uridine is in excess (such as a molar ratio of 3:1), it can preferentially occupy the reaction sites of phenylboronic acid, forcing the reaction to proceed in the direction of monoester. When the molar ratio of the two is within the above range, the excess degree of uridine is controllable, which can not only drive the main monoester reaction (such as the preferential esterification of the 5'-hydroxyl group of ribose) but also avoid excessive waste. Through the optimization of the molar ratio, the proportion of the monoester product in the crude product increases. Combining with subsequent purification, a small amount of unreacted raw materials can be efficiently removed, reducing the production cost.

[0044] In some embodiments, the ratio of the total mass of uridine and 3-carboxyphenylboronic acid to the volume of the polar solvent is (0.5 - 4) g: 5 mL. As an example, it can be typical but non-limiting values such as 0.5 g: 5 mL, 1 g: 5 mL, 2 g: 5 mL, 3 g: 5 mL, 4 g: 5 mL, 5 g: 5 mL, etc.

[0045] In some embodiments, the temperature of the esterification reaction is 120 °C - 200 °C.

[0046] The temperature of the esterification reaction ranges from 120°C to 200°C, which can significantly reduce the reaction activation energy and shorten the reaction time. In this temperature range, the preferential esterification of the 5'-hydroxyl group of uridine can be promoted (due to less steric hindrance), the participation of the 2'- and 3'-hydroxyl groups in the reaction can be inhibited, the formation of poly-ester by-products can be reduced, and the yield of mono-ester can be increased. In summary, this temperature range for the esterification reaction can not only avoid the thermal decomposition of the uracil ring or phenylboronic acid group, but also enable the esterification reaction to proceed fully.

[0047] In some embodiments, the polar solvent includes at least one of dimethyl sulfoxide, N,N-dimethylformamide, and water.

[0048] As high-boiling polar aprotic solvents, dimethyl sulfoxide and N,N-dimethylformamide can fully dissolve uridine and 3-carboxyphenylboronic acid, promoting intermolecular collisions and increasing the yield of mono-ester. At the same time, the high-boiling characteristics of the solvent match the esterification reaction temperature (120°C - 200°C), maintaining the stability of the liquid-phase reaction environment and avoiding concentration fluctuations or pressure risks caused by solvent volatilization. Water can be used as a co-solvent to reduce the amount of organic solvent, which is environmentally friendly. Moreover, the presence of water can stabilize ionic intermediates (such as borates) and inhibit the hydrolysis of ester bonds.

[0049] In some embodiments, the purification treatment includes a recrystallization step, which is carried out in ethanol.

[0050] The crude product after the esterification reaction usually contains unreacted uridine, 3-carboxyphenylboronic acid, di-ester by-products, and solvent residues. Ethanol has a moderate polarity and can dissolve the target mono-ester compound at high temperatures, while the poorly soluble di-ester by-products (due to stronger hydrophobicity) and unreacted 3-carboxyphenylboronic acid (with too high polarity) preferentially precipitate out upon cooling, enabling selective separation. Moreover, the aprotic nature and weak acidity (pH≈7.5) of ethanol can inhibit the hydrolysis of borate ester bonds, maintaining the chemical stability of the product during the purification process.

[0051] The specific process of the recrystallization step can be as follows: Mix the crude product with an excess of ethanol, heat it to 70°C - 80°C to dissolve it, and then filter it while hot to remove the undissolved di-esters and raw material impurities; after the filtrate cools naturally to room temperature, further cool it in an ice bath (0 - 4°C) to promote the slow precipitation of the target mono-ester crystals, collect the crystals by vacuum filtration, and wash them 2 - 3 times with pre-cooled ethanol to remove the surface residual impurities; finally, dry them under vacuum at 40 - 50°C for 4 - 6 hours to obtain a high-purity mono-ester product.

[0052] In some embodiments, after the recrystallization step, the recrystallized product is dissolved in a water-methanol mixed mobile phase and subjected to high-pressure liquid chromatography separation using a C18 reverse-phase column to collect the target eluate and obtain the borate ester compound.

[0053] Although recrystallization improved the purity of the crude product, trace amounts of structurally similar substances (such as isomers of monoesters, residual diesters, etc.) were still present. Based on the hydrophobicity differences of compounds, the C18 reverse-phase chromatographic column used gradient elution with a water-methanol mixed mobile phase to precisely separate the monoesters from impurities with similar polarities (such as diesters), enabling the purity of the final product to meet the pharmaceutical-grade purity requirements (>99%).

[0054] On the other hand, the present application also provides the use of a borate compound in the preparation of a boron neutron capture therapy drug, and the borate compound is the above-mentioned borate compound or the borate compound prepared by the above preparation method.

[0055] Through the uracil-TK1 targeting mechanism, this compound achieves highly selective enrichment near the tumor cell nucleus. The α particles released after neutron irradiation precisely kill cancer cells, avoiding damage to normal tissues. High solubility (≥50 mM) allows for a lower dosage, still meeting the tumor boron concentration threshold and reducing the metabolic burden. The anti-hydrolysis property of the monoesters structure (hydrolysis rate <10% in 24 hours) extends the in vivo circulation time, enabling the boron concentration in the tumor to be at its peak during neutron irradiation (animal experiments show an extended irradiation window period).

[0056] The following is illustrated with specific examples.

[0057] Example 1

[0058] This example provides a borate compound, and the synthetic route of this borate compound is as follows:

[0059]

[0060] The preparation steps of the borate compound in this example are as follows: Weigh 1.5 g of raw materials of uridine and 3-carboxyphenylboronic acid in a molar ratio of 2:1, add them to 5 mL of a 0.5% dimethyl sulfoxide aqueous solution to form a mixed solution, and conduct cyclic reactions at 160 °C for 4 times; Recrystallize the crude product with ethanol until the yield reaches 80%, and perform high-pressure liquid-phase preparation and separation on a C18 column with a water-methanol mobile phase to remove the unstable 3-carboxyphenylboronic acid uridine diester, obtaining 3-carboxyphenylboronic acid uridine monoester, which is the borate compound in this example.

[0061] Both uridine and 3-carboxyphenylboronic acid were purchased from Shanghai Macklin Biochemical Co., Ltd.

[0062] Example 2

[0063] The difference between Example 2 and Example 1 is that the molar ratio of uridine and 3-carboxyphenylboronic acid is 1:1.

[0064] Example 3

[0065] Example 3 is different from Example 1 in that the molar ratio of uridine to 3-carboxyphenylboronic acid is 3:1.

[0066] Comparative Example 1

[0067] Example 3 is different from Example 1 in that the molar ratio of uridine to 3-carboxyphenylboronic acid is 1:2. The borate ester compound obtained is uridine 3-carboxyphenylboronic acid diester.

[0068] Comparative Example 2

[0069] The boron drug uses BPA (boronophenylalanine).

[0070] Performance Test

[0071] 1. Infrared Detection:

[0072] Use a Nicolet 6700 Fourier transform infrared spectrometer and detect the infrared spectrum of the borate ester compound by the pressing tablet method.

[0073] Figure 1 This is the infrared spectrum of the borate ester compound of Example 1 of this application (the uridine phenylborate in the figure refers to uridine 3-carboxyphenylboronic acid monoester). The successful synthesis of the borate ester structure can be verified by the characteristic peak at 900 cm -1 −1.

[0074] 2. NMR Detection:

[0075] Use an AVANCE III 600 MHz superconducting Fourier nuclear magnetic resonance spectrometer to detect the nuclear magnetic resonance 1H spectrum of the uridine 3-carboxyphenylboronic acid monoester product. The solvent is deuterated water, and the chemical shift of the element is expressed in ppm.

[0076] Figure 2 This is the NMR H 1 spectrum of the borate ester compound (uridine 3-carboxyphenylboronic acid monoester) of Example 1 of this application. By comparison, it is found that the peak of uridine at 5.8 ppm (H at the 2 and 3 positions) is twice the peak at 7.5 ppm (at the 5 position) and has similar intensity in the uridine 3-carboxyphenylboronic acid monoester, indicating that the borate esterification occurs at the 2 or 3 position of uridine. In addition, the H peaks of 3-carboxyphenylboronic acid at 7.8 ppm and 7.9 ppm on the boronic acid itself do not change relatively in the uridine 3-carboxyphenylboronic acid monoester, indicating that the esterification randomly occurs at one of the two boronic acids, but the peak at 8.2 ppm relatively decreases in the uridine 3-carboxyphenylboronic acid monoester, indicating esterification.

[0077] 3. Mass Spectrometry Detection:

[0078] Analyze using a Thermo Fisher liquid chromatography-mass spectrometry instrument (Q Exactive).

[0079] Figure 3 This is the mass spectrum (negative ion) of the aqueous solution of 3-carboxyphenylboronic acid uridine diester for Comparative Example 1 of this application. As can be seen from Figure 3 it, the aqueous solution of 3-carboxyphenylboronic acid uridine diester is unstable and decomposes into uridine and 3-carboxyphenylboronic acid at room temperature, with a 9% change in 10 hours.

[0080] Figure 4 This is the mass spectrum (negative ion) of the aqueous solution of 3-carboxyphenylboronic acid uridine monoester for Example 1 of this application. As can be seen from Figure 4 it, the prepared aqueous solution of 3-carboxyphenylboronic acid uridine monoester is relatively stable, and the decomposition product 165 increases by less than 1% after 24 hours. It is very likely that the hydrogen bond formed between 3-carboxyl and uridine residue leads to the increased stability.

[0081] 4. Effects of BPA (borophenylalanine) and 3-carboxyphenylboronic acid uridine monoester on 3 kinds of cancer cells

[0082] (1) Cell culture and cell viability experiment

[0083] Using A549 lung cancer cell line, Hela cervical cancer cell line, and F98 glioma cell line as the research objects, the cells were purchased from Shenzhen Top Biological Technology Co., Ltd., and DMEM-HG complete medium (containing 10% FBS and 1% double antibody) was used as the cell culture medium.

[0084] The effects of two boron agents, BPA and 3-carboxyphenylboronic acid uridine monoester (UB), on the viability of cancer cells were detected using a cell apoptosis detection kit (CCK-8, cell counting kit-8). The cell lines were cultured in RPMI-1640 cell culture medium. The cell morphology on each culture dish (15 cm, about 3x10 7 cells) was observed under a fluorescence microscope, and 4 boxes with similar cell states were grouped together. 3 ml of each of the two boron agents (3-carboxyphenylboronic acid uridine monoester, BPA) with a molar concentration of 5 mM (adjusted to neutral pH) and an actual concentration of 1 mM were added to 15 ml of the medium respectively. At the same time, a positive control group (3 ml of 30% H2O2 was added) and a negative control group (3 ml of PBS was added) were set up, and the cells were cultured at 37°C. After incubation for 3 hours, CCK-8 reagent was added dropwise to the culture dish at a ratio of 10% of the total volume, and after thorough mixing, the cells were further cultured at 37°C for 3 h. The absorbance of the mixed solution was detected using a 754NPC type ultraviolet-visible spectrophotometer, and the cell viability was compared (cell viability = (absorbance of the cell group solution after exposure to the toxicant / absorbance of the control without the toxicant molecule). The results are as Figure 5 shown. The results show that these two boron drugs have no obvious effect on the viability of cancer cells at a concentration of 1 mM.

[0085] (2) Comparative experiments on the uptake of BPA and 3-carboxylphenylboronic acid uridine monophosphate (hereinafter referred to as UB) by three types of cancer cells

[0086] Using A549 lung cancer cell line, Hela cervical cancer cell line, and F98 glioma cell line as the research objects, the cells were purchased from Shenzhen Top Biological Technology Co., Ltd., and DMEM-HG complete medium (containing 10% FBS and 1% double antibody) was used as the cell culture medium.

[0087] The cell morphology on each culture dish (15 cm, approximately 3x10 7 cells) was observed under a fluorescence microscope, and four boxes with similar cell states were grouped together. 2 ml - 6 ml of two boron agents, BPA with a molar concentration of 5 mM (adjusted to neutral pH), and 5 - 15 mM 3-carboxylphenylboronic acid uridine monophosphate were added to 15 ml of the culture medium respectively. At the same time, a negative control group (2 ml of PBS was added) was set up and cultured at 37°C. After incubating for 3 hours, the culture medium was poured out, and the cells were rinsed 3 times with 3 ml of PBS and then drained. The cells were digested with spectroscopic grade nitric acid + 30% hydrogen peroxide in a graphite digestion instrument and fixed to 10 ml, and the content of B element was detected by ICP-MS (Agilent inductively coupled plasma mass spectrometer model 7900). The results are shown in Table 1.

[0088] Table 1. Comparative experiments on the uptake of BPA and 3-carboxylphenylboronic acid uridine monophosphate (UB) by three types of cancer cells

[0089] Amount of B added (μg) B content in cells (μg) Proportion of B element entering cells (%) A549 control 0 (2 ml PBS) 0.05+0.03 UB - A549 110 (5 mM 2 ml) 0.23+0.03 0.20 UB - A549 220 (10 mM 2 ml) 0.47+0.05 0.21 UB - A549 330 (15 mM 2 ml) 1.35+0.06 0.41 BPA - A549 110 (5 mM 2 ml) 0.81+0.06 0.74 BPA - A549 330 (5 mM 6 ml) 1.72+0.08 0.52 Hela control 0 (2 ml PBS) 0.04+0.03 UB - Hela 110 (5 mM 2 ml) 0.32+0.04 0.29 UB - Hela 330 (15 mM 2 ml) 0.58+0.05 0.18 BPA - Hela 110 (5 mM 2 ml) 0.60+0.03 0.55 F98 control 0 (2 ml PBS) 0.05+0.02 UB - F98 110 (5 mM 2 ml) 0.18+0.05 0.16 UB - F98 330 (15 mM 2 ml) 0.41+0.03 0.12 BPA - F98 110 (5 mM 2 ml) 0.39+0.04 0.35

[0090] (3) Analysis of the entry of BPA and 3-carboxylphenylboronic acid uridine monophosphate (UB) into Hela cells

[0091] Since cytoplasmic thymidine kinase (TK1) is widely and highly expressed in Hela cancer cells, its substrates thymidine or deoxyuridine have targeted enrichment. Therefore, the nucleoside boron drug - 3-carboxylphenylboronic acid uridine monophosphate (UB) may be enriched in the nucleus (solid phase), so the B content in the supernatant and precipitate was compared.

[0092] Experimental procedure: Select four culture dishes with a diameter of 15 cm of Hela cells with similar cell states in each group. Add 2 ml of 5 mmol of the drug (3-carboxylphenylboronic acid uridine monophosphate (UB)) and BPA to 15 ml of the culture medium. After poisoning for 1 - 3 hours, wash with PBS 3 times. Add 1 ml of 80% acetonitrile-aqueous solution to each dish, fix, then scrape off, and break the cells by ultrasonic wave at 50% power. After centrifuging at 13000 r / min for 5 minutes, a small amount of the supernatant was spotted for HLCP-MS mass spectrometry, and most of the remaining approximately 2.5 ml of the supernatant was directly measured by ICP mass spectrometry for the B content. The centrifuged precipitate of approximately 0.1 ml was digested with 5 ml of nitric acid-hydrogen peroxide for ICP-MS measurement. The results are shown in Table 2.

[0093] Table 2. Analysis of the state of BPA and 3-carboxylphenylboronic acid uridine monoester (UB) entering Hela cells

[0094]

[0095]

[0096] As can be seen from Table 2, about 1 / 3 of 3-carboxylphenylboronic acid uridine monoester (UB) enters the solid phase when entering Hela cells, thus entering the relatively sensitive target points of the cells, and has a more sensitizing effect on BNCT treatment.

[0097] (4) Experiment of 3-carboxylphenylboronic acid uridine monoester (UB) entering A549 and F98 tumor-bearing mice

[0098] The tumor-bearing nude mice were purchased from Shenzhen Top-Bio Technology Co., Ltd. Tumors with a volume of 1000 mm 3 , about 20 g in weight for each of the 3 male mice in a group. A total of 2 drugs (UB and BPA) were injected. The administration route was intravenous injection via the tail vein. The administration dose was 5 mM, 0.1 ml, injected once every 3 hours for a total of 2 times. PBS was injected as a control. After 6 hours, the mice were sacrificed, and 0.2 g of tumor tissue and 0.2 g of surrounding tissue were taken. After digestion with spectroscopic-grade nitric acid + 30% hydrogen peroxide in a graphite digestion instrument, the volume was fixed at 10 ml, and the content of B element was detected by ICP-MS (Agilent inductively coupled plasma mass spectrometer model 7900). The results are shown in Table 3. A solvent control group was set up.

[0099] Table 3. Comparative analysis of BPA and 3-carboxylphenylboronic acid uridine monoester (UB) entering the tumors of tumor-bearing mice

[0100] Amount of B added per box (μg) B content in tumor mass (μg) B content in peripheral substances (μg) Control 0 0.01+0.01 0.01+0.01 UB - F98 11.0 0.04+0.02 0.02+0.01 BPA - F98 11.0 0.06+0.02 0.01+0.01 UB - A549 11.0 0.04+0.01 0.02+0.01 BPA - A549 11.0 0.03+0.01 0.01+0.01

[0101] As can be seen from Table 3, after intravenous injection of 3-carboxylphenylboronic acid uridine monoester (UB) for 6 hours, the tumor can enrich UB, and it can be enriched in the tumor mass at a concentration of 5 mM. The average distribution of B is 0.2 g / 20 g = 1%, while the B content distribution in the tumor mass is only greater than 0.3%. Therefore, most of it is metabolized through the blood, similar to BPA. However, the B content in the tumor mass is significantly higher than that in the surrounding tissues. Therefore, this boron drug has a certain tumor enrichment ability, similar to BPA. Therefore, 3-carboxylphenylboronic acid uridine monoester (UB) can be used as a boron drug in BNCT treatment.

[0102] The above are only the preferred embodiments of this application, and do not limit the patent scope of this application. Any equivalent structural transformation made under the application concept of this application, or direct / indirect application in other related technical fields, is included in the patent protection scope of this application.

Claims

1. A borate compound, characterized in that, Its structural formula is shown in Formula I:

2. The borate compound according to claim 1, characterized in that, The solubility of the borate ester compound in water is ≥ 50 mmol / L at 10°C to 30°C.

3. The borate compound according to claim 1, wherein The hydrolysis rate of the borate ester compound in water within 24 hours is < 10% at 10°C to 30°C.

4. A method for preparing a borate compound as described in any one of claims 1 to 3, characterized in that, It includes the following steps: Uridine is mixed with 3-carboxyphenylboronic acid, and an esterification reaction is carried out in a polar solvent. The reaction is cycled 2 to 5 times to obtain a crude product. After purifying the crude product, the borate ester compound is obtained.

5. The preparation method of the borate compound according to claim 4, characterized in that, The molar ratio of uridine to 3-carboxyphenylboronic acid is 3:1 to 1:1; and / or, The ratio of the total mass of uridine and 3-carboxyphenylboronic acid to the volume of the polar solvent is (0.5 - 4) g : 5 mL.

6. The preparation method of the borate compound according to claim 4, characterized in that, The temperature of the esterification reaction is 120°C to 200°C.

7. The preparation method of the borate compound according to claim 4, wherein, The polar solvent includes at least one of dimethyl sulfoxide, N,N-dimethylformamide, and water.

8. The preparation method of the borate compound according to claim 4, characterized in that, The purification treatment includes a recrystallization step, and the recrystallization step is carried out in ethanol.

9. The preparation method of the borate compound according to claim 8, characterized in that, After the recrystallization step, the recrystallized product is dissolved in a water-methanol mixed mobile phase, and high-pressure liquid-phase preparative separation is carried out through a C18 reverse-phase chromatography column. The target eluate is collected to obtain the borate ester compound.

10. Use of a borate ester compound in the preparation of a boron neutron capture therapy drug, characterized in that, The borate ester compound is the borate ester compound described in any one of Claims 1 to 3, or the borate ester compound prepared by the preparation method described in any one of Claims 4 to 9.

Images