Amino acid amine-borane compound, preparation method therefor and use thereof

Abstract

The present invention belongs to the technical field of drugs for treating tumors. Disclosed in the present invention are an amino acid amine-borane compound, and a preparation method therefor and the use thereof. An amino acid amine-borane compound as represented by Formula (I) is obtained by means of the protection of functional groups in an amino acid, borane complexation, and deprotection reactions. Cell experimental results show that tumor cells exhibit a much higher uptake capacity for such compounds than for clinically used drug 4-dihydroxyboryl-L-phenylalanine (BPA) for boron neutron capture therapy, with no signs of toxicity, indicating that the compound has significant potential as a new boron-delivery agent for boron neutron capture therapy.

Description

Amino acid amine-borane compounds, their preparation methods and applications Technical Field

[0001] This invention belongs to the field of tumor therapeutic drug technology, specifically relating to amino acid amine-borane compounds, their preparation methods, and applications. Background Technology

[0002] Malignant tumors are among the most serious diseases threatening human life and health. The development of efficient diagnostic and treatment technologies for cancer is in high demand, and significant progress has been made in the treatment of malignant tumors, including surgery, drug therapy, radiotherapy, and immunotherapy. Among these, approximately 70% of cancer patients choose radiotherapy each year. With the development of modern medical technology, radiotherapy techniques are constantly evolving, progressing from non-selective radiotherapy to proton and heavy ion radiotherapy with a certain degree of longitudinal energy selectivity. Despite this, high-grade gliomas and other malignant tumors still exhibit strong resistance to current treatments. As a cell-level precise binary targeted tumor therapy technology, boron neutron capture therapy (BNCT) combines biological targeting with the effect of heavy ions to selectively and precisely kill cancer cells at the cellular level, requiring only 1-2 irradiations. It has significant advantages for treating advanced malignant tumors that are difficult to operate on and cannot be treated with traditional radiotherapy and chemotherapy. BNCT will carry... 10 B-targeted drugs are injected into the patient's body. 10 B will specifically accumulate in tumor tissue, and the affected area will be irradiated with hyperthermic neutrons. The neutrons and 10 B occurred 10 B(n,α) 7 Li nuclear reactions form its isotopes. 11 B. Subsequently. 11 B-cell fission produces high energy. 7Li ions and alpha particles, with a range of only about one cell diameter, precisely kill cancer cells because their DNA undergoes irreversible breakage under the influence of heavy ions, while normal cells remain largely undamaged. Currently, boron-targeted neurotransmitter therapy (BNCT) is experiencing explosive growth, with neutron sources shifting from reactors to more economical, safe, and reliable accelerator neutron sources, achieving a key technological breakthrough in neutron beam acquisition. However, the development of boron-targeted BNCT drugs still faces challenges. Currently, only 4-dihydroxyboron-L-phenylalanine (BPA) and sodium undecylmercaptododecoboride (BSH) are used clinically, with BPA being the only marketed clinical treatment globally. However, BPA suffers from insufficient tumor specificity, short blood half-life and retention time, and extremely poor solubility, resulting in low boron atom enrichment in tumor tissue. Clinically, BPA must be excipiented with fructose to improve water solubility and is administered via large-volume intravenous injection, causing significant nephrotoxicity and severely impacting the efficacy of BNCT. Therefore, developing a class of small-molecule organoboron compounds with economical and efficient synthesis processes, readily available raw materials, good water solubility, and potentially broad tumor targeting as candidate drug molecules is crucial for improving the clinical treatment efficacy of BNCT. Summary of the Invention

[0003] To overcome the shortcomings of the prior art, the present invention aims to provide amino acid amine-borane compounds, their preparation methods, and applications, thereby solving the technical problems of existing BNCT drugs, such as difficult synthesis, poor water solubility, and insufficient tumor targeting. To achieve the above objective, the present invention employs the following technical solution:

[0004] The first aspect of the present invention discloses amino acid amine-borane compounds having compounds of formula (I) or pharmaceutically acceptable salts thereof, as well as stereoisomers, isotopic products and derivatives of said compound (I) or pharmaceutically acceptable salts thereof:

[0005]

[0006] Wherein, R is a hydrogen atom, alkyl, aryl, heteroaryl, alkyl containing a substituent, aryl containing a substituent, or heteroaryl containing a substituent; R′ is a hydrogen atom, alkyl, or alkyl containing a substituent.

[0007] In a second aspect, the present invention discloses a method for preparing the above-mentioned amino acid amine-borane compound, comprising the following steps: 1) dissolving an amino acid and N,O-bis(trimethylsilyl)acetamide in an organic solvent and reacting to obtain a silicon-protected amino acid; 2) adding a borane complex to the reaction mixture obtained in step 1 (i.e., the silicon-protected amino acid) and reacting to obtain the amino acid amine-borane compound.

[0008] A third aspect of the present invention discloses the use of the above-mentioned amino acid amine-borane compound in the preparation of a pharmaceutical formulation for BNCT. Preferably, the BNCT pharmaceutical formulation is a drug or formulation for treating tumors, or a drug or formulation for inhibiting tumor progression. More preferably, the drug or formulation for inhibiting tumor progression is a drug for inhibiting the progression of malignant tumors or metastatic tumors.

[0009] Compared with existing technologies, the present invention has the following beneficial effects: The amino acid amine-borane compound provided by the present invention has the structure shown in formula (I). In this structure, the amino group of the amino acid has a lone pair of electrons, and the boron atom on the borane has an empty orbital. The lone pair of electrons on the amino group readily coordinates with the boron atom to form an amine-borane product. By protecting the carboxyl functional group in the amino acid, a silicon-based protecting intermediate is obtained to reduce the polarity of the amino acid molecule. Then, by coordinating the amino group with the borane, an amine-borane derivative is obtained, and then the amino acid amine-borane product is successfully obtained after deprotection. Since this structure is similar to that of the amino acid, it can be transported into tumor cells through amino acid transport proteins on the cell membrane surface. The preparation method of the entire compound is simple and low in cost, and is suitable for the synthesis of various amino acids and amino acid structural analogs. Cell experiments show that the uptake capacity of this compound by tumor cells is much higher than that of the boron neutron capture clinical therapeutic drug BPA, and no signs of toxicity have been observed, indicating that it has important potential as a novel boron delivery agent for boron neutron capture therapy. Attached Figure Description

[0010] Figure 1 shows the phenylalanine amine-borane of Example 1. 1 1H NMR spectrum; Figure 2 shows the phenylalanine amine-borane of Example 1. 13 C NMR spectrum; Figure 3 shows the phenylalanine amine-borane spectrum of Example 1. 11 B NMR spectrum; Figure 4 shows the tryptophan amine-borane spectrum from Example 2. 1 1H NMR spectrum; Figure 5 shows the tryptophan amine-borane spectrum from Example 2. 13 C NMR spectrum; Figure 6 shows the tryptophan amine-borane spectrum from Example 2. 11 B NMR spectrum; Figure 7 shows the methionine amine-borane of Example 3. 1 1H NMR spectrum; Figure 8 shows the methionine amine-borane of Example 3. 13 C NMR spectrum; Figure 9 shows the methionine amine-borane of Example 3. 11 B NMR spectrum; Figure 10 shows the threonine amine-borane of Example 4. 1 1H NMR spectrum; Figure 11 shows the threonine amine-borane of Example 4. 13 C10 NMR spectrum; Figure 12 shows the threonine amine-borane of Example 4. 11B NMR spectrum; Figure 13 shows isoleucine amine-borane from Example 5. 1 1H NMR spectrum; Figure 14 shows isoleucine amine-borane from Example 5. 13 C NMR spectrum; Figure 15 shows isoleucine amine-borane from Example 5. 11 B NMR spectrum; Figure 16 shows the cytotoxicity test results of compound 1 of the present invention; Figure 17 shows the cytotoxicity test results of compound 2 of the present invention; Figure 18 shows the cytotoxicity test results of compound 3 of the present invention; Figure 19 shows the cytotoxicity test results of compound 4 of the present invention; Figure 20 shows the cytotoxicity test results of compound 5 of the present invention; Figure 21 shows the cytotoxicity test results of compound 6 of the present invention; Figure 22 shows the cytotoxicity test results of compound 7 of the present invention; Figure 23 shows the cytotoxicity test results of compound 8 of the present invention; Figure 24 shows the boron atom uptake test results of compounds 1-7 of the present invention in 4T1 cells. Embodiments of the present invention

[0011] To enable those skilled in the art to understand the features and effects of the present invention, the following is a general description and definition of the terms and expressions mentioned in the specification and claims. Unless otherwise specified, all technical and scientific terms used herein have the ordinary meaning understood by those skilled in the art regarding the present invention, and in case of conflict, the definitions in this specification shall prevail. The theories or mechanisms described and disclosed herein, whether right or wrong, shall not in any way limit the scope of the invention; that is, the present invention can be practiced without being limited by any particular theory or mechanism. In this document, the term "boron neutron capture therapy" (BNCT) refers to a tumor treatment method comprising the steps of administering a boron-containing compound to a subject requiring treatment and irradiating said subject with thermal neutrons. The term "tumor" refers to the uncontrolled and progressive growth of tissue with cells proliferating. Uncontrolled proliferation is a state different from normal cell proliferation, such as a state of significantly increased cell proliferation rate. The term "progressive" means strong progression or increase. The term "tumor cell" refers to cells in histology. The term "tumor treatment" refers to the treatment of diseases caused by or related to tumors. Unless otherwise specified, “→” in equation (Ⅰ) represents a coordinate bond.

[0012] In a first aspect, the present invention provides an amino acid amine-borane compound with the structure shown in formula (I):

[0013]

[0014] Wherein, R is a hydrogen atom, alkyl, aryl, heteroaryl, alkyl containing a substituent, aryl containing a substituent, or heteroaryl containing a substituent; R′ is a hydrogen atom, alkyl, or alkyl containing a substituent; alkyl or alkyl containing a substituent includes, but is not limited to, hydrogen atoms, alkyl, or alkyl containing a substituent. , , , , , , , , , , , , or ; aryl or aryl containing substituents include, but are not limited to, , , , or Heteroaryl groups or heteroaryl groups containing substituents include, but are not limited to, those containing... or .

[0015] Preferably, the pharmaceutically acceptable salt of the amino acid amine-borane compound is its ammonium, lithium, sodium, potassium, rubidium, cesium, magnesium, or calcium salt. Preferably, it comprises all enantiomers of the amino acid amine-borane compound or its pharmaceutically acceptable salt, and mixtures of enantiomers in any proportion. Preferably, the isotopic product of the enantiomers of the amino acid amine-borane compound or its pharmaceutically acceptable salt contains... 10 B 11 B 12 C 13 C 18 O or 2 H atom.

[0016] A second aspect of the present invention provides a method for preparing the above-mentioned amino acid amine-borane compound, comprising the following steps:

[0017]

[0018] (1) Dissolve natural or non-natural amino acids or N-substituted amino acids (1.0 equivalent) and N,O-bis(trimethylsilyl)acetamide (BSA; 1.0~5.0 equivalent) in an organic solvent and react to obtain a silicon-protected amino acid derivative; wherein, N,O-bis(trimethylsilyl)acetamide is used as a carboxyl group protecting agent.

[0019]

[0020] (2) Add borane complex (1.0~5.0 equivalents) to a solution of silicon-protected amino acid derivative, and remove the silicon-protecting group after the reaction is complete to obtain an amino acid amine-borane compound.

[0021] A third aspect of this invention provides the application of the aforementioned amino acid amine-borane compound in the preparation of pharmaceutical formulations for BNCT, which can be used to treat various types of tumors or inhibit tumor progression. The boron neutron capture therapy drug is a drug for treating tumors or inhibiting tumor progression; the tumors include, but are not limited to, malignant tumors; the malignant tumors include, but are not limited to, glioblastoma multiforme, malignant meningioma, intramedullary spinal glioma, advanced or recurrent head and neck cancer, thyroid cancer, malignant melanoma, recurrent breast cancer, metastatic liver cancer, malignant brain tumors, osteosarcoma, lung cancer, squamous cell carcinoma of the skin, or nasopharyngeal carcinoma. The amino acid amine-borane compound can be directly dissolved in physiological saline, phosphate buffer, sterile water, or glucose solution to prepare a boron drug formulation for infusion. The amino acid amine-borane compound requires no excipients, its water solubility is more than 35 times that of commercially available BPA boron drugs, and the boron atom uptake by tumor cells is 5 to 26 times that of commercially available BPA boron drugs.

[0022] The following examples use instruments and equipment conventional in the art. Experimental methods in the following examples, unless otherwise specified, are generally performed under standard conditions or as recommended by the manufacturer. All raw materials used in the following examples are conventional commercially available products with specifications in the art, unless otherwise stated.

[0023] I. Preparation of Amino Amine-Boronane Compounds

[0024] Example 1: Preparation of phenylalanine amine-borane

[0025]

[0026] 165.2 mg of phenylalanine and 306.6 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 2 mL of 1,4-dioxane, and the mixture was stirred at 10 °C for 24 hours to obtain a reaction mixture. A 1 M (1 mol / L) and 1 mL volume of a borane dimethyl sulfide complex (i.e., 1.0 mmol of borane dimethyl sulfide complex) was added to the reaction mixture, and the mixture was stirred for another 24 hours. After the reaction was complete, 2 mL of water was added to quench the reaction, and then phenylalanine amine-borane (denoted as compound 1) was obtained. The NMR spectra of compound 1 are shown in Figures 1-3. 1H NMR (400 MHz, D2O) δ 7.42 – 7.23 (m, 5H), 3.38 (t,J= 6.5 Hz, 1H), 3.05 – 2.99 (m, 2H), 1.43 (s, 3H). 13 C NMR (101 MHz, DO) δ 177.9, 139.0, 133.3, 133.1, 131.6, 69.0, 40.3. 11 B NMR (128 MHz, D2O) δ -21.3.

[0027] Example 2: Preparation of tryptophanamine-borane

[0028]

[0029] 204.2 mg of tryptophan and 613.2 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of chloroform and stirred at 20 °C for 36 hours to obtain a reaction mixture. A 1 M, 2 mL solution of boranetetrahydrofuran complex was added to the reaction mixture, and stirring was continued for 24 hours. After the reaction was complete, 2 mL of methanol was added to quench the reaction, and then tryptophan amine-borane (compound 2) was obtained. The NMR spectra of compound 2 are shown in Figures 4-6. 1 H NMR (400 MHz, D2O) δ 7.71 (d,J= 8.0 Hz, 1H), 7.49 (d,J= 8.2 Hz, 1H), 7.31 – 7.20 (m, 2H), 7.16 (q,J= 7.5 Hz, 1H), 3.46 (t,J= 6.2 Hz, 1H), 3.36 – 3.28 (m, 1H), 3.17 (dd,J= 14.9, 7.0 Hz, 1H), 1.26 (s, 3H). 13 C NMR (101 MHz, DO) δ 178.5, 140.3, 130.6, 129.0, 126.1, 123.4, 122.4, 115.9, 111.55, 59.0, 30.35. 11 B NMR (128 MHz, D2O) δ -22.2.

[0030] Example 3: Preparation of methionine amine-borane

[0031]

[0032] 149.2 mg of methionine and 306.6 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of tetrahydrofuran and stirred at 50 °C for 36 hours to obtain a reaction mixture. A 1 M, 1 mL solution of borane dimethyl sulfide complex was added to the reaction mixture, and stirring continued for 24 hours. After the reaction was complete, 2 mL of methanol was added to quench the reaction, and then the methionine amine-borane (denoted as compound 3) was obtained. The NMR spectra of compound 3 are shown in Figures 7-9. 1 H NMR (400 MHz, D2O) δ 3.15 (t,J= 6.2 Hz, 1H), 2.44 (td,J= 7.5, 4.1 Hz, 2H), 1.99 – 1.83 (m, 5H), 1.60 – 0.73 (m, 3H). 13 C NMR (101 MHz, DO) δ 177.4, 61.7, 30.6, 28.9, 14.0. 11 B NMR (128 MHz, D2O) δ -21.3.

[0033] Example 4: Preparation of threonine amine-borane

[0034]

[0035] 119.1 mg of threonine and 1.02 g of N,O-bis(trimethylsilyl)acetamide were dissolved in 2 mL of diethyl ether and stirred at 60 °C for 48 hours to obtain a reaction mixture. A 1 M, 5 mL solution of boranetetrahydrofuran complex was added to the reaction mixture, and stirring was continued for 36 hours. After the reaction was complete, 2 mL of water was added to quench the reaction, and then threonine amine-borane (denoted as compound 4) was obtained. The NMR spectra of compound 4 are shown in Figures 10-12. 1 H NMR (400 MHz, D2O) δ 3.80 (dq,J= 8.1, 6.4 Hz, 1H), 2.92 (d,J= 7.9 Hz, 1H), 1.77 – 1.34 (m, 3H), 1.20 (d,J= 6.5 Hz, 3H). 13 C NMR (101 MHz, DO) δ 172.8, 65.9, 60.4, 19.4. 11 B NMR (128 MHz, D2O) δ -21.2.

[0036] Example 5: Preparation of isoleucine amine-borane

[0037]

[0038] 131.2 mg of isoleucine and 408.8 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of acetonitrile and stirred at 25 °C for 24 hours to obtain a reaction mixture. A 1 M, 1.5 mL solution of borane dimethyl sulfide complex was added to the reaction mixture, and stirring was continued for 36 hours. After the reaction was complete, 2 mL of ethanol was added to quench the reaction, and then isoleucine amine-borane (denoted as compound 5) was obtained. The NMR spectra of compound 5 are shown in Figures 13-15. 1 H NMR (400 MHz, D2O) δ 3.01 – 2.90 (m, 1H), 1.67 (ddd,J= 11.9, 9.4, 5.0 Hz, 1H), 1.57 – 1.02 (m, 5H), 0.90 – 0.74 (m, 6H). 13 C NMR (101 MHz, DO) δ 174.1, 59.5, 35.8, 24.4, 14.6, 11.0. 11 B NMR (128 MHz, D2O) δ -21.9.

[0039] Example 6: Preparation of leucine amine-borane

[0040]

[0041] 131.2 mg of leucine and 408.8 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of dichloromethane and stirred at 0 °C for 48 hours to obtain a reaction mixture. A 1 M, 1 mL solution of boranetetrahydrofuran complex was added to the reaction mixture, and stirring was continued for another 48 hours. After the reaction was complete, 2 mL of methanol was added to quench the reaction, and then leucine amine-borane (denoted as compound 6) was obtained. The NMR spectrum of compound 6 is shown below: 1 H NMR (400 MHz, D2O) δ 3.08 (t,J= 7.1 Hz, 1H), 1.68 – 1.48 (m, 2H), 1.47 (t,J= 7.0 Hz, 4H), 0.83 (dd,J= 6.5, 5.4 Hz, 6H). 13 C NMR (101 MHz, DO) δ 178.7, 61.7, 41.6, 24.4, 21.8, 21.7. 11 B NMR (128 MHz, D2O) δ -20.4.

[0042] Example 7: Preparation of valine amine-borane

[0043]

[0044] 117.1 mg of valine and 817.6 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of acetone and stirred at 45 °C for 36 hours to obtain a reaction mixture. A 1 M, 2 mL solution of a borane dimethyl sulfide complex was added to the reaction mixture, and stirring was continued for 18 hours. After the reaction was complete, 2 mL of tert-butanol was added to quench the reaction, and then the valine amine-borane (denoted as compound 7) was isolated. The NMR spectrum of compound 7 is shown below: 1 H NMR (400 MHz, D2O) δ 2.96 (d,J= 5.3 Hz, 1H), 2.07 – 1.93 (m, 2H), 1.79 – 1.09 (m, 1H), 0.94 (s, 4H), 0.93 (s, 3H). 13 C NMR (101 MHz, DO) δ 176.1, 59.5, 31.1, 19.2, 17.8.

[0045] Example 8: Preparation of lysine amine-borane

[0046]

[0047] 146.2 mg of lysine and 204.4 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of tetrahydrofuran, and the mixture was stirred at 20 °C for 24 hours to obtain a reaction mixture. A 1 M, 1.5 mL solution of a borane-tetrahydrofuran complex was added to the reaction mixture, and the mixture was stirred for another 36 hours. After the reaction was complete, 2 mL of water was added to quench the reaction, and the lysine amine-borane (designated as compound 8) was then isolated. The NMR spectrum of compound 8 is shown below: 1 H NMR (400 MHz, D2O) δ 3.67 (t,J= 6.1 Hz, 1H), 2.65 (t,J= 7.5 Hz, 2H), 1.83 (dq,J= 10.1, 6.2 Hz, 2H), 1.66 – 1.55 (m, 2H), 1.47 – 0.88 (m, 5H). 13 C NMR (101 MHz, DO) δ 175.4, 58.5, 40.1, 30.8, 27.2, 25.3.

[0048] Example 9: Preparation of glutamate-borane

[0049]

[0050] 147.1 mg of glutamic acid and 408.8 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of 1,4-dioxane, and the mixture was stirred at 30 °C for 36 hours to obtain a reaction mixture. A 1 M, 1.5 mL solution of boranetetrahydrofuran complex was added to the reaction mixture, and the mixture was stirred for another 48 hours. After the reaction was complete, 2 mL of ethanol was added to quench the reaction, and the glutamic acid-borane mixture (designated as compound 9) was then isolated. The NMR spectrum of compound 9 is shown below: 1 H NMR (400 MHz, D2O) δ 4.19 – 4.05 (m, 1H), 2.31 (ddd,J= 8.9, 7.3, 4.4 Hz, 2H), 2.15 – 2.02 (m, 2H), 1.69 – 0.88 (m, 3H). 13 C NMR (101 MHz, DO) δ 181.9, 175.9, 56.7, 34.0, 25.2.

[0051] Example 10: Preparation of tyrosine amine-borane

[0052]

[0053] 181.2 mg of tyrosine and 204.4 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of acetonitrile and stirred at 25 °C for 40 hours to obtain a reaction mixture. A 1 M, 3.5 mL solution of a borane dimethyl sulfide complex was added to the reaction mixture, and stirring was continued for 36 hours. After the reaction was complete, 2 mL of isopropanol was added to quench the reaction, and then the tyrosine amine-borane (denoted as compound 10) was obtained. The NMR spectrum of compound 10 is shown below: 1 H NMR (400 MHz, D2O) δ 7.07 (d,J= 8.6 Hz, 2H), 6.75 (d,J= 8.6 Hz, 2H), 3.23 (d,J= 13.1 Hz, 1H), 2.89 (dd,J= 14.8, 4.8 Hz, 1H), 2.81 (dd,J= 14.2, 8.6 Hz, 1H), 1.41 – 0.65 (m, 3H). 13 C NMR (101 MHz, DO) δ 169.3, 156.6, 130.2, 123.4, 115.4, 54.5, 36.2.

[0054] Example 11: Preparation of serine amine-borane

[0055]

[0056] 105.1 mg of serine and 306.6 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 2 mL of 1,4-dioxane, and the mixture was stirred at 20 °C for 36 hours. A 1 M, 1.5 mL solution of boranetetrahydrofuran complex was added to the reaction mixture, and the mixture was stirred for another 24 hours. After the reaction was complete, 2 mL of water was added to quench the reaction, and the resulting serine amine-borane (compound 11) was isolated. The NMR spectrum of compound 11 is shown below: 1 H NMR (400 MHz, D2O) δ 3.95 (d,J= 3.7 Hz, 1H), 3.81 (d,J= 5.0 Hz, 1H), 3.20 (t,J= 4.4 Hz, 1H), 1.73 – 0.86 (m, 3H). 13 C NMR (101 MHz, DO) δ 172.8, 62.9, 58.4.

[0057] Example 12: Preparation of 4-dihydroxyboryl-L-phenylalanine (BPA)amine-borane

[0058]

[0059] 208.1 mg of 4-dihydroxyboryl-L-phenylalanine and 204.4 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of tetrahydrofuran and stirred at 25 °C for 24 hours to obtain a reaction mixture. A 1 M, 3 mL solution of a borane dimethyl sulfide complex was added to the reaction mixture, and stirring continued for 36 hours. After the reaction was complete, 2 mL of water was added to quench the reaction, and then 4-dihydroxyboryl-L-phenylalanine amine-borane (denoted as compound 12) was obtained. The NMR spectrum of compound 12 is shown below: 1 H NMR (400 MHz, D2O) δ 7.50 (d,J= 7.4 Hz, 2H), 7.13 (d,J= 7.5 Hz, 2H), 3.48 (t,J= 6.2 Hz, 1H), 2.71 – 2.51 (m, 2H), 1.26 – 0.97 (m, 3H). 13C NMR (101 MHz, DO) δ 172.1, 138.3, 136.2, 133.4, 130.5, 57.6, 36.3.

[0060] Example 13: Preparation of proline amine-borane

[0061]

[0062] 115.1 mg of proline and 306.6 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of 1,4-dioxane, and the mixture was stirred at 40 °C for 24 hours to obtain a reaction mixture. A 1 M, 2 mL solution of boranetetrahydrofuran complex was added to the reaction mixture, and the mixture was stirred for another 12 hours. After the reaction was complete, 2 mL of methanol was added to quench the reaction, and the proline amine-borane (denoted as compound 13) was then isolated. The NMR spectrum of compound 13 is shown below: 1 H NMR (400 MHz, D2O) δ 3.47-3.34 (m, 2H), 2.86-2.77 (m, 1H), 2.23-2.11 (m, 1H), 1.95-1.82 (m, 2H), 1.78-1.63 (m, 1H), 1.16 – 0.94 (m, 3H). 13 C NMR (101 MHz, DO) δ 173.4, 59.2, 46.7, 29.8, 24.6.

[0063] Example 14: Preparation of alanine amine-borane

[0064]

[0065] 89.1 mg of alanine and 306.6 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of N,N-dimethylformamide, and the mixture was stirred at 10 °C for 48 hours to obtain a reaction mixture. A 1 M, 2 mL solution of boranetetrahydrofuran complex was added to the reaction mixture, and the mixture was stirred for another 36 hours. After the reaction was complete, 2 mL of ethanol was added to quench the reaction, and then alanine amine-borane (denoted as compound 14) was obtained. The NMR spectrum of compound 14 is shown below: 1 H NMR (400 MHz, D2O) δ 3.09 (q,J= 7.1 Hz, 1H), 1.37 (d,J= 7.2 Hz, 3H), 1.18 – 0.93 (m, 3H). 13C NMR (101 MHz, DO) δ 176.4, 53.2, 17.6.

[0066] Example 15: Preparation of cysteine ​​amine-borane

[0067]

[0068] 105.1 mg of cysteine ​​and 204.4 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of 1,2-dichloroethane and stirred at 60 °C for 24 hours to obtain a reaction mixture. A 1 M, 1 mL solution of boranetetrahydrofuran complex was added to the reaction mixture, and stirring continued for 36 hours. After the reaction was complete, 2 mL of water was added to quench the reaction, and then cysteine ​​amine-borane (denoted as compound 15) was obtained. The NMR spectrum of compound 15 is shown below: 1 H NMR (400 MHz, D2O) δ 4.63 (dt,J= 6.8, 5.2 Hz, 1H), 3.68 – 3.51 (m, 2H), 1.38 – 0.97 (m, 3H). 13 C NMR (101 MHz, DO) δ 171.8, 56.9, 54.4.

[0069] Example 16: Preparation of Aspartic Amine-Boronane

[0070]

[0071] 105.1 mg of aspartic acid and 408.8 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 3 mL of N-methylpyrrolidone, and the mixture was stirred at 25 °C for 36 hours to obtain a reaction mixture. A 1 M, 1 mL solution of borane dimethyl sulfide complex was added to the reaction mixture, and the mixture was stirred for another 24 hours. After the reaction was complete, 2 mL of methanol was added to quench the reaction, and then the aspartic acid amine-borane (denoted as compound 16) was obtained. The NMR spectrum of compound 16 is shown below: 1 H NMR (400 MHz, D2O) δ 3.89 (dd, J = 3.75, 8.69 Hz, 1H), 2.59 - 2.88 (m, 2H), 1.63 - 0.96 (m, 3H). 13 C NMR (101 MHz, DO) δ 175.1, 173.4, 54.4, 35.4.

[0072] Example 17: Preparation of N-methylphenylalanine amine-borane

[0073]

[0074] 179.1 mg of N-methylphenylalanine and 306.6 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 2 mL of tetrahydrofuran, and the mixture was stirred at 30 °C for 36 hours to obtain a reaction mixture. A 1 M, 1 mL solution of borane tetrahydrofuran complex was added to the reaction mixture, and the mixture was stirred for another 24 hours. After the reaction was complete, 2 mL of methanol was added to quench the reaction, and then N-methylphenylalanine amine-borane (denoted as compound 17) was obtained. The NMR spectrum of compound 17 is shown below: 1 H NMR (400 MHz, D2O) δ 7.42 – 7.23 (m, 5H), 3.38 (t,J= 6.5 Hz, 1H), 3.05 – 2.99 (m, 2H), 2.52 (s, 3H), 1.52 – 1.17 (m, 3H). 13 C NMR (101 MHz, DO) δ 172.2, 138.0, 129.3, 128.1, 125.6, 65.0, 37.3, 33.5.

[0075] Example 18: Preparation of N,N-dimethylphenylalanine amine-borane

[0076]

[0077] 179.1 mg of N-methylphenylalanine and 306.6 mg of N,O-bis(trimethylsilyl)acetamide were dissolved in 2 mL of tetrahydrofuran, and the mixture was stirred at 30 °C for 36 hours to obtain a reaction mixture. A 1 M, 1 mL solution of a borane tetrahydrofuran complex was added to the reaction mixture, and the mixture was stirred for another 36 hours. After the reaction was complete, 2 mL of methanol was added to quench the reaction, and then N,N-dimethylphenylalanine amine-borane (denoted as compound 18) was obtained. The NMR spectrum of compound 18 is shown below: 1 H NMR (400 MHz, D2O) δ 7.42 – 7.23 (m, 5H), 3.38 (t,J= 6.5 Hz, 1H), 3.05 – 2.99 (m, 2H), 2.50 (s, 6H), 1.72 – 1.25 (m, 3H). 13 C NMR (101 MHz, DO) δ 173.1, 137.6, 129.0, 127.9, 126.0, 64.6, 37.3, 35.9, 33.5.

[0078] II. Verification of the water solubility, safety, and efficacy of amino acid amine-borane compounds

[0079] 1. Determination of water solubility of amino acid amine-borane prepared in Examples 1-3

[0080] 30.0 mg of each compound (Examples 1-3) was added to 200.0 μL of distilled water and mixed thoroughly to obtain suspensions. Each suspension was sonicated for 2 h, then centrifuged at 5000 rpm for 10 min. 100 μL of the supernatant was collected and diluted to 5 mL. The boron content in the supernatant was determined by inductively coupled plasma mass spectrometry (ICP-MS), repeated six times, and the average value was taken. Commercially available boron drug BPA was used as a control. The results showed that the water solubility of compound 1 was 52.36 g / L, compound 2 was 100.93 g / L, and compound 3 was 88.25 g / L, while the water solubility of the control group, commercially available boron drug BPA, was 1.46 g / L. The results indicate that compounds 1-3 have good water solubility, reaching up to 69.1 times that of commercially available boron drug BPA.

[0081] 2. Cytotoxicity test of the amino acid amine-borane prepared in Examples 1 to 8

[0082] This experiment used mouse fibroblasts (L929 cells, purchased from the Shanghai Cell Bank of the Chinese Academy of Sciences) for cytotoxicity testing. L929 cells were cultured in DMEM (Dulbecco's Modified Eagle Medium) containing 10% (v / v) fetal bovine serum, 1% (v / v) penicillin, and streptomycin. The cells were placed in a cell culture incubator at 37°C and 5% CO2, and passaged 3-5 times before use. The viability of L929 cells was evaluated using the Almar Blue assay kit; generally, a cell viability >75% was considered to indicate that the compound was not cytotoxic. The compounds obtained in Examples 1-8 were dissolved at a concentration of 10 mM (millimoles per liter) in sterile fetal bovine serum (FBS) buffer, filtered through a microporous membrane for sterilization, and then further sterilized at 60°C for 24 hours before being stored under aseptic conditions. The 10 mM FBS solutions of the compounds from Examples 1 to 8 were diluted with DMEM culture medium to obtain 0.5 mM, 1 mM, 2.5 mM, and 5 mM DMEM culture media of compounds 1 to 8 from Examples 1 to 8. L929 cells were seeded at a density of 1500 cells / well in 96-well plates and cultured for 24 h under the above conditions. After washing off the DMEM culture medium, DMEM culture medium containing different concentrations (0.5 mM, 1 mM, 2.5 mM, and 5 mM) of the compounds prepared in Examples 1 to 8 was added. Cells cultured in DMEM culture medium without the added compounds served as a blank control. Cells were co-cultured in an incubator for 24 h. Cell viability was then assessed using an Almar Blue assay kit, and the fluorescence intensity of the culture medium at an excitation wavelength of 530 nm and an emission wavelength of 600 nm was measured using a microplate reader to evaluate cell viability. The experimental results are shown in Figures 16 to 23. As shown in Figure 16, at lower culture concentrations of compound 1 (1 mM and 0.5 mM), the survival rate of L929 cells reached over 80%, indicating that compound 1 is essentially non-cytotoxic. As shown in Figure 17, at all culture concentrations of compound 2, the survival rate of L929 cells reached over 75%, indicating that compound 2 is non-cytotoxic at all concentrations. As shown in Figure 18, at all culture concentrations of compound 3, the survival rate of L929 cells reached over 75%, indicating that compound 3 is non-cytotoxic at all concentrations. As shown in Figure 19, at culture concentrations of compound 4 (2.5 mM, 1 mM, and 0.5 mM), the survival rate of L929 cells reached over 80%, indicating that compound 4 is essentially non-cytotoxic.As shown in Figure 20, at culture concentrations of 2.5 mM, 1 mM, and 0.5 mM of compound 5, the survival rate of L929 cells was not significantly different from, and even higher than, that of the control group, indicating that compound 5 has essentially no cytotoxicity. As shown in Figure 21, at lower culture concentrations of compound 6 (1 mM and 0.5 mM), the survival rate of L929 cells was not significantly different from, and even higher than, that of the control group, indicating that compound 6 has essentially no cytotoxicity. As shown in Figure 22, at a low culture concentration of compound 7 (0.5 mM), the survival rate of L929 cells was greater than 75%, indicating that compound 7 has essentially no cytotoxicity. As shown in Figure 23, at culture concentrations of 2.5 mM, 1 mM, and 0.5 mM of compound 8, the viability of L929 cells reached over 80%, indicating that compound 8 has essentially no cytotoxicity. In conclusion, the various essential amino acid amines-boranes prepared by this method are essentially non-cytotoxic.

[0083] 3. Experiment on boron atom uptake in 4T1 cells of amino acid amine-borane prepared in Examples 1 to 7.

[0084] This experiment used mouse breast cancer cells (4T1 cells, purchased from the Shanghai Cell Bank of the Chinese Academy of Sciences) to perform a boron uptake assay. 4T1 cells were cultured in 1640 medium containing 10% (v / v) fetal bovine serum and placed in a cell culture incubator at 37°C and 5% CO2. Cells were cultured and passaged 3-5 times before use. 4T1 cells in logarithmic growth phase with a cell content of 80%-85% were seeded into 6-well plates (1 × 10⁶ cells per well). 6 (cells). After cell adhesion, cells were treated with 1 mM 1,4-dihydroxyboronylphenylalanine (BPA) and the compounds prepared in Examples 1-7 above for 24 h, respectively, with cells cultured in DMEM medium without the added compounds used as a blank control. The culture medium was discarded, and the cells were washed three times with PBS. After washing with PBS, 0.25% trypsin was added and digested at 37°C for 3 min. Digestion was stopped by adding regular culture medium, and the cell suspension was transferred to centrifuge tubes. After repeated pipetting to form a single-cell suspension, the cells were counted, centrifuged at 1500 rpm for 5 min, and the supernatant was aspirated. The cells were then centrifuged with concentrated nitric acid (per 10 mM). 6 Cells were digested with 0.5 mL of solution for 2 hours, then diluted to 10 mL with ultrapure water and filtered through a microporous membrane. The zero point was adjusted using a sample that had not been incubated with boron-containing culture medium, and the boron concentration in the solution was determined by ICP-MS to calculate the intracellular boron concentration. Three samples were taken, and the average value was calculated. The experimental results are shown in Figure 24. In 4T1 cells, the intracellular boron concentration obtained from the control group BPA was 5.00 μg / 10⁻¹⁰. 6 The intracellular boron concentration obtained from compound 2 was 133.85 μg / 10 cells. 6The number of cells was 26.8 times that of the control group BPA. The intracellular boron concentration obtained with compound 1 was 91.65 μg / 102 cells. 6 The number of cells was 18.3 times that of the control group BPA. The intracellular boron concentration obtained with compound 7 was 35.67 μg / 102. 6 The concentration of boron in cells was 7.1 times that of the control group (BPA). Compound 6 yielded an intracellular boron concentration of 22.33 μg / 1010 cells. 6 The concentration of boron in cells obtained by compound 3 was 4.5 times that of the control group BPA. The intracellular boron concentration obtained by compound 3 was 20.76 μg / 10 cells. 6 The number of cells was 4.2 times that of the control group BPA. The intracellular boron concentration obtained with compound 4 was 13.00 μg / 102 cells. 6 The concentration of boron in cells was 2.6 times that of the control group (BPA). Compound 5 yielded an intracellular boron concentration of 1.67 μg / 1010 cells. 6 In 4T1 cells, the boron uptake of the control group BPA was 3.0 times that of BPA. In summary, except for isoleucine and lysine, the boron uptake results of the essential amino acid amine borane in 4T1 cells were superior to those of BPA, reaching a maximum of 26.8 times that of BPA.

[0085] The above content is only for illustrating the technical concept of the present invention and should not be construed as limiting the scope of protection of the present invention. Any modifications made to the technical solution based on the technical concept proposed in this invention shall fall within the scope of protection of the claims of this invention.

Claims

1. An amino acid amine-borane compound, characterized in that, Having a compound of formula (I) or a pharmaceutically acceptable salt thereof, and stereoisomers, isotopic products and derivatives of the compound of formula (I) or a pharmaceutically acceptable salt thereof: Wherein, R is a hydrogen atom, alkyl, aryl, heteroaryl, alkyl containing a substituent, aryl containing a substituent, or heteroaryl containing a substituent; R′ is a hydrogen atom, alkyl, or alkyl containing a substituent.

2. The amino acid amine-borane compound according to claim 1, characterized in that, The pharmaceutically acceptable salts of the compounds of formula (I) are their ammonium salts, lithium salts, sodium salts, potassium salts, rubidium salts, cesium salts, magnesium salts, or calcium salts.

3. The amino acid amine-borane compound according to claim 1 or 2, characterized in that, All enantiomers of a compound of formula (I) or a pharmaceutically acceptable salt thereof, and mixtures of enantiomers in any proportion.

4. The amino acid amine-borane compound according to claim 1, characterized in that, Isotopic products having enantiomers of compounds of formula (I) or pharmaceutically acceptable salts thereof contain 10 B. 11 B. 12 C 13 C 18 O or 2 H atom.

5. The method for preparing the amino acid amine-borane compound according to any one of claims 1 to 4, characterized in that, Includes the following steps: 1) Dissolve amino acids and N,O-bis(trimethylsilyl)acetamide in an organic solvent and react to obtain silyl-protected amino acids; 2) Add a borane complex to the silicon-protected amino acid obtained in step 1) to obtain an amino acid amine-borane compound.

6. The method for preparing the amino acid amine-borane compound according to claim 5, characterized in that, The equivalent ratio of the amino acid, N,O-bis(trimethylsilyl)acetamide and borane complex is 1:(1~5):(1~5).

7. The use of the amino acid amine-borane compound according to any one of claims 1 to 4 in the preparation of a drug or preparation for treating tumors, or in the preparation of a drug or preparation for inhibiting the progression of tumors.

8. The application according to claim 7, characterized in that, The tumors mentioned are glioblastoma multiforme, malignant meningioma, intramedullary spinal cord glioma, advanced or recurrent head and neck cancer, thyroid cancer, malignant melanoma, recurrent breast cancer, metastatic liver cancer, malignant brain tumor, osteosarcoma, lung cancer, squamous cell carcinoma of the skin, or nasopharyngeal carcinoma.

9. The application according to claim 7, characterized in that, The pharmaceutical preparation is an injectable formulation made by mixing an amino acid amine-borane compound with physiological saline, phosphate buffer, sterile water, or glucose solution.

10. The use of the amino acid amine-borane compound according to any one of claims 1 to 4 in the preparation of a drug formulation for the treatment of tumors by boron neutron capture.

11. The application according to claim 10, characterized in that, The tumors mentioned are glioblastoma multiforme, malignant meningioma, intramedullary spinal cord glioma, advanced or recurrent head and neck cancer, thyroid cancer, malignant melanoma, recurrent breast cancer, metastatic liver cancer, malignant brain tumor, osteosarcoma, lung cancer, squamous cell carcinoma of the skin, or nasopharyngeal carcinoma.

12. The application according to claim 10, characterized in that, The pharmaceutical preparation is an injectable formulation made by mixing an amino acid amine-borane compound with physiological saline, phosphate buffer solution, sterile water or glucose solution.

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