A diamino pyrimidine macrocycle containing a crown ether fragment and its preparation method and application

Abstract

The application relates to the technical field of chemical medicines, in particular to a kind of dianilopyrimidine macrocycle containing crown ether fragment and its preparation method and application.A series of dianilopyrimidine macrocycle containing crown ether fragment is synthesized in the application, and the problem of poor clinical effect of drug in current brain glioma treatment is overcome.The series of compounds have good inhibitory activity on brain glioma, and are suitable for treating brain glioma and related drug research and development.Moreover, the drug is widely distributed in vivo and is rapidly cleared, so that the possibility of drug accumulation poisoning is avoided;oral bioavailability is moderate (F=18.7%);these findings highlight the advantages and challenges of the pharmacokinetic characteristics of compound 13, and lay a foundation for further optimizing and improving the in-vivo stability and bioavailability strategy.

Description

Technical Field

[0001] This application relates to the field of chemical pharmaceutical technology, specifically to a diaminopyrimidine macrocyclic compound containing a crown ether fragment, its preparation method, and its application. Background Technology

[0002] With the significant improvement in living standards, people are paying increasing attention to health. However, industrialization and the accompanying environmental pollution have made disease mechanisms more complex, and the situation regarding cancer prevention and treatment remains severe. Brain diseases, due to their unique location, have become one of the most challenging problems in clinical diagnosis and treatment. In particular, gliomas, the most malignant type of brain tumor, remain a difficult area to treat clinically due to their genetic instability, cellular heterogeneity, low survival rates after drug treatment, and difficulty in penetrating the blood-brain barrier. Since the FDA approved temozolomide in 1999, it has remained a first-line chemotherapy drug for gliomas; however, its mechanism of action as an alkylating agent leads to toxicity and drug resistance with long-term use. Therefore, developing novel and highly effective treatment strategies for gliomas remains an urgent and arduous task.

[0003] Literature review revealed that focal adhesion kinase (FAK) is highly expressed in gliomas, and previous studies have shown that FAK inhibitors have potential therapeutic effects on gliomas. Therefore, we designed and developed a novel class of small-molecule FAK kinase inhibitors containing crown ether fragments, aiming to exert anti-tumor effects by inhibiting FAK kinase activity and blocking downstream signaling pathways.

[0004] In addition, considering that gliomas originate in the central nervous system, the ability to penetrate the blood-brain barrier is crucial for the efficacy of the drug. Summary of the Invention

[0005] In order to solve the above-mentioned technical problems in the prior art, this application provides a diaminopyrimidine macrocyclic compound containing a crown ether fragment, its preparation method and application.

[0006] A diaminopyrimidine macrocyclic compound containing a crown ether fragment has the following structural formula (Ⅰ): Equation (Ⅰ).

[0007] Furthermore, R1 is selected from any one of Cl, CF3, H, and NO2.

[0008] Furthermore, R2 can be any of the following structures: .

[0009] Furthermore, specifically one of the following six structures: .

[0010] The specific steps for preparing the diaminopyrimidine macrocyclic compound containing the crown ether fragment are as follows: 2,4-Dichloro-5-substituted pyrimidines were subjected to nucleophilic substitution reactions with 2-amino-N-methylbenzamide to obtain compounds with the structure shown in formula (II). The compounds with the structure shown in formula (II) were then coupled with amines with the structure shown in R2 to obtain diaminopyrimidine macrocyclic compounds with crown ether fragments as shown in formula (III). Formula (II); Formula (III).

[0011] Furthermore, R1 is selected from any one of Cl, CF3, H, and NO2.

[0012] Furthermore, R2 can be any of the following structures: .

[0013] Further, the specific operation of the method is as follows: Different substituted pyrimidine compounds are added to a 100 mL round-bottom flask, dissolved in an appropriate amount of DMF, stirred in an ice bath, and NaH is slowly added. A large amount of heat is released. After stirring for another 5 minutes, 2-amino-nitromethylbenzamide is added to the reaction system. After stirring at room temperature for 6 hours, the reaction is quenched with cold water, and then stirred for 15 minutes, resulting in solid precipitation. The solid and liquid phases are separated by filtration to obtain the crude compound (II). After drying, a small amount of ethyl acetate is added to the crude compound and the mixture is slurried to obtain the pure compound (II).

[0014] 4-Nitrobenzene-1,2-diol and benzenesulfonates with different chain lengths were added to a 100 mL round-bottom flask, and an appropriate amount of DMF was added to dissolve them. K₂CO₃ was added to the reaction system, and the mixture was stirred at 80 °C for 6 hours under argon protection. After the reaction was completed, the mixture was cooled to room temperature. Then, 20 mL of saturated sodium chloride solution was added to the mixture to quench the reaction, and the mixture was stirred for 10 minutes. Subsequently, the mixture was extracted three times with DCM (15 mL each time), washed three times with saturated sodium chloride (10 mL each time), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography with a mobile phase ratio of dichloromethane / methanol = 100 / 1, yielding pure product R₂.

[0015] Finally, the compound with the structure shown in formula (II) and formula (III) were subjected to a Buchwald-Hartwig amination reaction to construct a carbon-nitrogen (CN) bond. Compounds (II) and (III) were sequentially added to a 100 mL round-bottom flask, and 1,4-dioxane was added and stirred to dissolve. Palladium acetate and the phosphine ligand Xantphos were added while stirring, and finally cesium carbonate was added. The reaction was carried out at 100 °C for 12 hours under an argon atmosphere. After the reaction was completed, the mixture was allowed to stand at room temperature, and the reaction was quenched with saturated sodium bicarbonate. The mixture was extracted three times with DCM (15 mL each time), washed three times with saturated sodium chloride (10 mL each time), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography with a mobile phase ratio of DCM / MeOH = 100 / 1 to obtain the diaminopyrimidine compound (III) containing a crown ether fragment.

[0016] Furthermore, the equivalence ratio of each substituted pyrimidine to 2-amino-N-methylbenzamide in the nucleophilic substitution reaction was adjusted to (pyrimidine: aniline: sodium hydride = 1: 1.1: 2.5). The reaction time was adjusted according to the nucleophilicity of the substrate pyrimidine, generally from 30 min to 6 h, and the specific time could be determined by thin-layer chromatography. After the reaction was complete, the compound with the structure shown in formula (II) was preferentially separated by vacuum filtration.

[0017] Furthermore, the carbon-nitrogen bond construction in the key reaction requires subsequent protection under argon or other inert gases due to the presence of palladium on carbon. The organic solvent is selected from DMF, THF, and 1,4-dioxane, preferably 1,4-dioxane, based on the solubility of the reaction solvent and the size of the phosphine ligand. The catalysts commonly used in the Buchwald reaction are selected from Pd2(dba)3 and pd(OAc)2, preferably pd(OAc)2. The ligands commonly used in the Buchwald reaction are selected from P(t-Bu)3, BINAP, P(o-tolyl)3, and Xantphos, preferably Xantphos. The reaction system also includes a base, selected from NaOH, Cs2CO3, and Ca2CO3, preferably Cs2CO3. The temperature for the substitution reaction is 60℃~100℃, preferably 100℃. The time for the nucleophilic substitution reaction is 6 h~24 h, preferably 12 h. h; the molar ratio of the compound with the structure shown in formula (II), the compound with the structure shown in formula (III), pd(OAc)2, Xantphos, and Cs2CO3 is 1:(0.5~1.2):(1~4):(0.1~0.2):(1~2), preferably 1:1.1:0.15:1.5. After the reaction is complete, the target compound is purified by column chromatography. The application of the aforementioned diaminopyrimidine macrocyclic compounds containing crown ether fragments in the preparation of drugs.

[0018] Furthermore, the drug is an anti-tumor drug, especially an anti-glioma drug.

[0019] Compared with the prior art, the technical effects created by this application are reflected in: This application synthesizes a series of diaminopyrimidine macrocyclic compounds containing crown ether fragments, overcoming the current drug delivery and resistance problems in glioma treatment. This series of compounds exhibits good inhibitory activity against gliomas, making them suitable for glioma treatment and related drug development. Furthermore, the drug is widely distributed and rapidly cleared in vivo, avoiding the possibility of drug accumulation and toxicity; its oral bioavailability is moderate (F = 18.7%). These findings highlight the advantages and challenges of the pharmacokinetic properties of compound 13, laying the foundation for strategies to further optimize and improve its in vivo stability and bioavailability. Detailed Implementation

[0020] The technical solution of this application will be further defined below with reference to specific implementation methods, but the scope of protection is not limited to the description.

[0021] Example 12-((2,5-dichloropyrimidin-4-yl)amino)-N-methylbenzamide (1) 2,4,5-Trichloropyrimidine (1834 mg, 10 mmol, 1 eq) was dissolved in DMF (15 mL). Then, NaH (600 mg, 25 mmol, 2.5 eq) was added with stirring in an ice bath. After stirring the reaction mixture in an ice bath for 5 minutes, 2-amino-N-methylbenzamide (1650 mg, 11 mmol, 1.1 eq) was added. The reaction mixture was stirred at 80 °C for 6 hours under argon protection. After the reaction was complete, water (15 mL) was added to terminate the reaction, resulting in a large amount of solid precipitate. This precipitate was filtered through a Buchner funnel, washed with a large amount of water (200 mL), dried, and then slurried with a small amount of ethyl acetate to give a yellow pure product 1 in 68% yield.

[0022] The characterization data of compound 1 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 12.21 (s, 1H), 8.87 (s, 1H), 8.50 (d, J = 24Hz, 2H), 7.81 (s, 1H), 7.61 (s, 1H), 7.22 (s, 1H), 2.81 (s, 3H). Example 22-((2-chloro-5-nitropyrimidin-4-yl)amino)-N-methylbenzamide (2) 2,4-Dichloro-5-nitropyrimidine (2910 mg, 15 mmol, 1 eq) was dissolved in DMF (15 mL). Then, NaH (1800 mg, 75 mmol, 5 eq) was added with stirring in an ice bath. After stirring the reaction mixture in an ice bath for 5 minutes, 2-amino-N-methylbenzamide (2480 mg, 16.5 mmol, 1.1 eq) was added. The reaction mixture was stirred at 80 °C for 6 hours under argon protection. After the reaction was complete, water (15 mL) was added to terminate the reaction, resulting in a large amount of solid precipitate. This precipitate was filtered through a Buchner funnel, washed with a large amount of water (200 mL), dried, and then slurried with a small amount of ethyl acetate to give a yellow pure product 2 in 76% yield.

[0023] The characterization data for compound 2 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 12.45 (s, 1H), 9.21 (s, 1H), 8.76 (d, J =4.8 Hz, 1H), 8.18 (dd, J = 8.4, 1.2 Hz, 1H), 7.71 (dd, J = 8.0, 1.6 Hz, 1H), 7.59(d, J = 7.2 Hz, 1H), 7.33 (t, J = 8.0 Hz, 1H), 2.79 (d, J = 4.4 Hz, 3H). Example 32-((2-chloro-5-(trifluoromethyl)pyrimidin-4-yl)amino)-N-methylbenzamide (3) 2,4-Dichloro-5-trifluoromethylpyrimidine (2170 mg, 10 mmol, 1 eq) was dissolved in DMF (15 mL). Then, NaH (600 mg, 25 mmol, 2.5 eq) was added with stirring in an ice bath. After stirring the reaction mixture in an ice bath for 5 minutes, 2-amino-N-methylbenzamide (1650 mg, 11 mmol, 1.1 eq) was added. The reaction mixture was stirred at 80 °C for 6 hours under argon protection. After the reaction was complete, water (15 mL) was added to terminate the reaction, resulting in a large amount of solid precipitate. This precipitate was filtered through a Buchner funnel, and the filter cake was washed with a large amount of water (200 mL), dried, and then slurried with a small amount of ethyl acetate to give a yellow pure product 3, with a yield of 49%.

[0024] The characterization data for compound 3 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 12.03 (s, 1H), 8.86 (d, J = 4.8 Hz, 1H),8.69 (s, 1H), 8.37 (d, J = 7.2 Hz, 1H), 7.78 (dd, J = 8.0, 1.6 Hz, 1H), 7.64 –7.55 (m, 1H), 7.26 (t, J = 8.0 Hz, 1H), 2.79 (d, J = 4.4 Hz, 3H). Example 42-((2-chloropyrimidin-4-yl)amino)-N-methylbenzamide (4) 2,4-Dichloropyrimidine (1490 mg, 10 mmol, 1 eq) was dissolved in DMF (15 mL). Then, NaH (600 mg, 25 mmol, 2.5 eq) was added with stirring in an ice bath. After stirring the reaction mixture in an ice bath for 5 minutes, 2-amino-N-methylbenzamide (1650 mg, 11 mmol, 1.1 eq) was added. The reaction mixture was stirred at 80 °C for 6 hours under argon protection. After the reaction was complete, water (15 mL) was added to terminate the reaction, resulting in a large solid precipitate. This precipitate was filtered through a Buchner funnel, washed with a large amount of water (200 mL), dried, and then slurried with a small amount of ethyl acetate to give a white pure product 4, with a yield of 32%. The characterization data for compound 4 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 10.89 (s, 1H), 8.63 (q, J = 4.8 Hz, 1H), 8.19 (d, J = 5.6 Hz, 1H), 8.07 (d, J = 8.4 Hz, 1H), 7.67 (dd, J = 7.6, 1.6 Hz, 1H), 7.53 (t, J = 7.2 Hz, 1H), 7.20 (t, J = 7.6 Hz, 1H), 6.86 (d, J = 6.0 Hz, 1H), 2.77(d, J = 4.4 Hz, 3H). Example 512-Nitro-2,3,5,6,8,9-Hexahydrobenzo[b][1,4,7,10]tetraoxane-dodecene (5) 4-Nitrophenyl-1,2-diol (310 mg, 2 mmol, 1 eq) and (ethane-1,2-diylbis(oxy))bis(ethane-2,1-diyl)bis(4-methylbenzenesulfonate) (917 mg, 2 mmol, 1 eq) were dissolved in DMF (7 mL). K₂CO₃ (554 mg, 4 mmol, 2 eq) was then added to the mixture. The reaction mixture was stirred at 80 °C for 6 hours under an argon atmosphere. After the reaction was complete, the mixture was cooled to room temperature. The reaction was then quenched by adding 20 mL of saturated saline solution and stirring for 10 minutes. The mixture was then extracted three times with DCM (15 mL each time), washed three times with saturated sodium chloride (10 mL each time), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography with a mobile phase ratio of dichloromethane / methanol = 100 / 1, yielding a white solid compound 5 in 36% yield. The characterization data of compound 5 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 7.97 – 7.89 (m, 2H), 7.26 (d, J= 8.8 Hz,1H), 4.30 – 4.20 (m, 4H), 3.79 – 3.72 (m, 2H), 3.71 – 3.62 (m, 2H), 3.59 (s,4H). Example 62,3,5,6,8,9-Hexahydrobenzo[b][1,4,7,10]tetraoxane-dodecene-12-amine (6) Compound 1 (350 mg, 2 mmol, 1 eq) was dissolved in methanol. Then, Pd / C (35 mg, 10 wt%) was added to the mixture. The reaction mixture was stirred at 25 °C for 10 hours under a hydrogen atmosphere. After the reaction was complete, diatomaceous earth was added to a Buchner funnel, and the reaction mixture was poured into it to remove palladium on carbon. The funnel was washed repeatedly with methanol until the product was completely washed. The filtrate was then concentrated under reduced pressure and dried under vacuum to give compound 6 as a purple solid in 99% yield. The characterization data for compound 6 are as follows: 1 H NMR (400 MHz, DMSO-d6) δ 6.70 (d, J = 8.4 Hz, 1H), 6.24 (d, J = 2.4 Hz, 1H), 6.10 (dd, J = 8.4, 2.4 Hz, 1H), 4.76 (s, 2H), 3.99 – 3.94 (m, 2H), 3.95 –3.89 (m, 2H), 3.76 – 3.69 (m, 2H), 3.62 (d, J = 8.0 Hz, 6H). Example 72-((2-((2,3,5,6,8,9-hexahydrobenzo[b][1,4,7,10]tetraoxane-12-yl)amino)-5-(trifluoromethyl)pyrimidin-4-yl)amino)-N-methylbenzamide (7) Carbon-nitrogen (CN) bonds were constructed using the Buchwald-Hartwig amination reaction. Compound 6 (119 mg, 0.5 mmol, 1.1 eq) and a trifluoromethyl-containing pyrimidine derivative 3 (149 mg, 0.45 mmol, 1 eq) were dissolved in 10 mL of 1,4-dioxane. Palladium acetate (15 mg, 0.07 mmol, 0.15 eq) and the phosphine ligand Xantphos (78 mg, 0.14 mmol, 0.3 eq) were added with stirring, followed by the addition of cesium carbonate (220 mg, 0.68 mmol, 1.5 eq). The reaction was carried out at 100 °C for 12 h under argon protection. After the reaction was complete, the mixture was allowed to stand at room temperature, and the reaction was quenched by adding saturated sodium bicarbonate. The mixture was extracted three times with DCM (15 mL each time), washed three times with saturated sodium chloride (10 mL each time), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was purified by column chromatography with a mobile phase ratio of DCM / MeOH = 100 / 1, yielding a white solid 7 in 37% yield.

[0025] The characterization data for compound 7 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 11.36 (s, 1H), 9.70 (s, 1H), 8.74 (q, J =4.4 Hz, 1H), 8.59 – 8.26 (m, 2H), 7.72 (dd, J = 8.0, 1.6 Hz, 1H), 7.45 (s, 1H), 7.37 (s, 1H), 7.16 (td, J = 7.6, 1.2 Hz, 2H), 6.95 (d, J = 8.8 Hz, 1H), 4.10 –4.03 (m, 2H), 3.91 (s, 2H), 3.72 – 3.66 (m, 4H), 3.63 (s, 4H), 2.79 (d, J = 4.4Hz, 3H); ESI-HRMS C 25 H 26 F3N5O5([M+H]+) calcd 534.1959, found 534.1963. HPLCpurity: 98.9%. Example 82-((5-chloro-2-((2,3,5,6,8,9-hexahydrobenzo[b][1,4,7,10]tetraoxocyclododecane-12-yl)amino)pyrimidin-4-yl)amino)-N-methylbenzamide (8) Carbon-nitrogen (CN) bonds were constructed using the Buchwald-Hartwig amination reaction. Compound 6 (84 mg, 0.35 mmol, 1.1 eq) and a trifluoromethyl-containing pyrimidine derivative 1 (96 mg, 0.32 mmol, 1 eq) were dissolved in 10 mL of 1,4-dioxane. Palladium acetate (14 mg, 0.06 mmol, 0.2 eq) and the phosphine ligand Xantphos (74 mg, 0.13 mmol, 0.4 eq) were added with stirring, followed by the addition of cesium carbonate (157 mg, 0.48 mmol, 1.5 eq). The reaction was carried out at 100 °C for 12 h under argon protection. After the reaction was completed, the mixture was allowed to stand at room temperature, and the reaction was quenched by adding saturated sodium bicarbonate. The mixture was extracted three times with DCM (15 mL each time), washed three times with saturated sodium chloride (10 mL each time), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography with a mobile phase ratio of DCM / MeOH = 100 / 1, yielding a white solid 8 in 37% yield. The characterization data for compound 8 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 11.60 (s, 1H), 9.31 (s, 1H), 8.78 – 8.72(m, 2H), 8.20 (s, 1H), 7.75 (d, J = 6.4 Hz, 1H), 7.52 – 7.44 (m, 1H), 7.37 (d, J = 2.4 Hz, 1H), 7.20 (dd, J = 8.8, 2.4 Hz, 1H), 7.14 (t, J = 7.2 Hz, 1H), 6.95 (d, J = 8.8 Hz, 1H), 4.09 – 4.03 (m, 2H), 3.98 (t, J = 4.4 Hz, 2H), 3.70 (q, J = 4.0Hz, 4H), 3.64 (s, 4H), 2.81 (d, J= 4.4 Hz, 3H); ESI-HRMS C 24 H 26 ClN5O5([M+H]+)calcd 500.1695, found 500.1694. Example 9 15-nitro-2,3,5,6,8,9,11,12-octahydrobenzo[b][1,4,7,10,13]pentoxacyclopentadecane (9) 4-Nitrobenzene-1,2-diol (465 mg, 3 mmol, 1 eq) and ((oxy-bridged (ethane-2,1-diyl))bis(oxy))bis(ethane-2,1-diyl)bis(4-methylbenzenesulfonate) (1508 mg, 3 mmol, 1 eq) were dissolved in DMF (10 mL). K₂CO₃ (831 mg, 6 mmol, 2 eq) was then added to the mixture. The reaction mixture was stirred at 80 °C for 8 hours under argon protection. After the reaction was complete, the mixture was cooled to room temperature. Then, 20 mL of saturated sodium bicarbonate solution was added to the mixture to terminate the reaction, and the mixture was stirred for 10 minutes. The mixture was extracted three times with DCM (15 mL each time), washed three times with saturated sodium chloride (10 mL each time), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography with DCM / MeOH = 100 / 1, yielding white compound 9 in 57% yield. The characterization data of compound 1 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 7.90 (dd, J = 8.8, 2.8 Hz, 1H), 7.73 (d, J =2.8 Hz, 1H), 7.15 (d, J = 9.2 Hz, 1H), 4.22 – 4.14 (m, 4H), 3.79 (q, J = 4.4 Hz, 4H), 3.62 (s, 8H). Example 102,3,5,6,8,9,11,12-octahydrobenzo[b][1,4,7,10,13]pentoxacyclopentane-15-amine (10) Compound 9 (450 mg, 2 mmol, 1 eq) was dissolved in methanol. Then, Pd / C (45 mg, 10% wt) was added. The reaction mixture was stirred at 25°C for 10 hours under a hydrogen atmosphere. After the reaction was complete, diatomaceous earth was added to a Buchner funnel, and the reaction mixture was poured into it to remove palladium on carbon. The funnel was washed repeatedly with methanol until thoroughly clean. Subsequently, the filtrate was concentrated under reduced pressure and dried under vacuum to give compound 10 in purple form, with a yield of 98%. The characterization data of compound 10 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 6.64 (d, J = 8.4 Hz, 1H), 6.22 (d, J = 2.4 Hz, 1H), 6.05 (dd, J = 8.4, 2.4 Hz, 1H), 4.67 (s, 2H), 3.98 – 3.91 (m, 2H), 3.89(t, J = 4.8 Hz, 2H), 3.78 – 3.71 (m, 2H), 3.71 (t, J = 5.2 Hz, 2H), 3.60 (s, 8H). 13 C NMR (100 MHz, DMSO-d6) δ 154.87, 148.55, 141.05, 118.33, 112.28, 108.22,71.05, 71.02, 70.05, 69.95, 69.23, 69.19, 68.92, 68.80. Example 11 N-methyl-2-((2-((2,3,5,6,8,9,11,12-octahydrobenzo[b][1,4,7,10,13]pentoxacyclopentadecene-15-yl)amino)pyrimidin-4-yl)amino)benzamide (11) Carbon-nitrogen (CN) bonds were constructed using the Buchwald-Hartwig amination reaction. Compound 10 (156 mg, 0.55 mmol, 1 eq) and pyrimidine derivative 4 (166.3 mg, 0.61 mmol, 1.1 eq) were dissolved in 10 mL of 1,4-dioxane. Palladium acetate (12.32 mg, 0.06 mmol, 0.1 eq) and the phosphine ligand Xantphos (64 mg, 0.11 mmol, 0.2 eq) were added with stirring, followed by the addition of cesium carbonate (269 mg, 0.83 mmol, 1.5 eq). The reaction was carried out at 100 °C for 12 h under argon protection. After the reaction was completed, the mixture was allowed to stand at room temperature, and the reaction was quenched by adding saturated sodium bicarbonate. The mixture was extracted three times with DCM (15 mL each time), washed three times with saturated sodium chloride (10 mL each time), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography with a mobile phase ratio of DCM / MeOH = 100 / 1, yielding a white solid 11 in 18% yield. The characterization data of compound 11 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 10.77 (s, 1H), 9.06 (s, 1H), 8.65 (q, J =4.8 Hz, 1H), 8.46 (d, J = 8.4 Hz, 1H), 8.05 (d, J = 5.6 Hz, 1H), 7.68 (dd, J = 8.0,1.6 Hz, 1H), 7.48 – 7.36 (m, 2H), 7.19 (dd, J = 8.8, 2.4 Hz, 1H), 7.07 (t, J =7.2 Hz, 1H), 6.86 (d, J = 8.8 Hz, 1H), 6.25 (d, J = 5.6 Hz, 1H), 4.05 – 3.98 (m,2H), 3.96 (t, J = 4.4 Hz, 2H), 3.79 – 3.72 (m, 4H), 3.62 (s, 8H), 2.79 (d, J =4.4 Hz, 3H). 13 C NMR (100 MHz, DMSO-d 6) δ 169.49, 160.15, 160.12, 157.39,149.04, 143.81, 140.36, 135.30, 131.91, 128.53, 121.69, 121.63, 121.48,115.25, 112.35, 107.25, 99.21, 70.82, 70.79, 70.44, 70.29, 69.72, 69.54,69.34, 68.81, 26.70. ESI-HRMS C 26 H 31 N5O6([M+Na]+) calcd 532.216655, found532.21709. Example 12N-methyl-2-((5-nitro-2-((2,3,5,6,8,9,11,12-octahydrobenzo[b][1,4,7,10,13]pentoxacyclopentadecane-15-yl)amino)pyrimidin-4-yl)amino)benzamide (12) Carbon-nitrogen (CN) bonds were constructed using the Buchwald-Hartwig amination reaction. Compound 10 (156 mg, 0.56 mmol, 1 eq) and the nitro-containing pyrimidine derivative 2 (188 mg, 0.60 mmol, 1.1 eq) were dissolved in 10 mL of 1,4-dioxane. Palladium acetate (12.32 mg, 0.06 mmol, 0.1 eq) and the phosphine ligand Xantphos (64 mg, 0.11 mmol, 0.2 eq) were added with stirring, followed by the addition of cesium carbonate (269 mg, 0.83 mmol, 1.5 eq). The reaction was carried out at 100 °C for 12 h under argon protection. After the reaction was completed, the mixture was allowed to stand at room temperature, and the reaction was quenched by adding saturated sodium bicarbonate. The mixture was extracted three times with DCM (15 mL each time), washed three times with saturated sodium chloride (10 mL each time), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography with a mobile phase ratio of DCM / MeOH = 100 / 1, yielding a white solid 12, with a yield of 19%. The characterization data of compound 12 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 12.11 (s, 1H), 10.34 (s, 1H), 9.10 (s,1H), 8.63 (q, J= 4.8 Hz, 1H), 8.15 (d, J = 8.4 Hz, 1H), 7.63 (d, J = 7.6 Hz, 1H), 7.41 (t, J = 7.6 Hz, 1H), 7.25 (d, J = 6.4 Hz, 2H), 7.11 (d, J = 9.6 Hz, 1H), 6.83(d, J = 8.8 Hz, 1H), 4.03 (t, J = 4.4 Hz, 2H), 3.76 (t, J = 4.4 Hz, 2H), 3.68 (s,4H), 3.63 – 3.59 (m, 8H), 2.78 (d, J = 4.4 Hz, 3H). 13 C NMR (100 MHz, CDCl3) δ173.09, 164.71, 163.11, 158.72, 153.57, 150.57, 141.21, 137.19, 135.60,133.00, 131.28, 130.27, 129.45, 126.14, 119.21, 119.07, 113.44, 75.61, 75.06,75.01, 74.12, 74.04, 73.99, 73.51, 31.38. ESI-HRMS C 26 H 30 N6O8([M+Na]+) calcd577.2017, found 577.2024. Example 13 N-methyl-2-((2-((2,3,5,6,8,9,11,12-octahydrobenzo[b][1,4,7,10,13]pentoxacyclopentadecane-15-yl)amino)-5-(trifluoromethyl)pyrimidin-4-yl)amino)benzamide (13) Carbon-nitrogen (CN) bonds were constructed using the Buchwald-Hartwig amination reaction. Compound 10 (470 mg, 1.67 mmol, 1 eq) and a trifluoromethyl-containing pyrimidine derivative 4 (524 mg, 1.6 mmol, 1 eq) were dissolved in 10 mL of 1,4-dioxane. Palladium acetate (72 mg, 0.32 mmol, 0.2 eq) and the phosphine ligand Xantphos (370 mg, 0.64 mmol, 0.4 eq) were added with stirring, followed by the addition of cesium carbonate (783 mg, 2.4 mmol, 1.5 eq). The reaction was carried out at 100 °C for 12 hours under argon protection. After the reaction was completed, the mixture was allowed to stand at room temperature, and the reaction was quenched by adding saturated sodium bicarbonate. The mixture was extracted three times with DCM (15 mL each time), washed three times with saturated sodium chloride (10 mL each time), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography with a mobile phase ratio of DCM / MeOH = 100 / 1, yielding a white solid 13 in 33% yield. The characterization data of compound 13 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 11.38 (s, 1H), 9.65 (s, 1H), 8.74 (q, J =4.8 Hz, 1H), 8.41 (s, 1H), 7.71 (dd, J = 8.0, 1.6 Hz, 1H), 7.41 (s, 1H), 7.26(s, 1H), 7.18 – 7.10 (m, 2H), 6.87 (d, J = 8.8 Hz, 1H), 4.03 (t, J = 4.4 Hz, 2H), 3.90 – 3.73 (m, 4H), 3.71 (s, 2H), 3.61 (d, J = 2.4 Hz, 8H), 2.79 (d, J = 4.4 Hz, 3H); 13 C NMR (100 MHz, DMSO- d6) δ 169.31, 161.40, 156.52, 156.33, 149.00,145.05, 139.26, 133.62, 131.69, 128.26, 126.48, 123.80, 123.14, 122.85,122.16, 114.88, 114.05, 108.66, 70.85, 70.40, 70.31, 69.48, 69.30, 68.87,26.69. ESI-HRMS C 27 H 30 F3N5O6([M+Na]+) calcd 600.2040, found 600.2037. HPLCpurity: 99.5%. Example 142-((5-chloro-2-((2,3,5,6,8,9,12,13-octahydro-11H-benzo[k][1,4,7,10]tetraoxocyclopentadecane-15-yl)amino)pyrimidin-4-yl)amino)-N-methylbenzamide (14) Carbon-nitrogen (CN) bonds were constructed using the Buchwald-Hartwig amination reaction. Compound 10 (156 mg, 0.6 mmol, 1 eq) and a chlorinated pyrimidine derivative 1 (180 mg, 0.61 mmol, 1.1 eq) were dissolved in 10 mL of 1,4-dioxane. Palladium acetate (12.32 mg, 0.06 mmol, 0.1 eq) and the phosphine ligand Xantphos (64 mg, 0.11 mmol, 0.2 eq) were added with stirring, followed by the addition of cesium carbonate (269 mg, 0.83 mmol, 1.5 eq). The reaction was carried out at 100 °C for 12 h under argon protection. After the reaction was completed, the mixture was allowed to stand at room temperature, and the reaction was quenched by adding saturated sodium bicarbonate. The mixture was extracted three times with DCM (15 mL each time), washed three times with saturated sodium chloride (10 mL each time), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography with a mobile phase ratio of DCM / MeOH = 100 / 1, yielding a white solid 14 in 25% yield. The characterization data of compound 14 are as follows: 1 H NMR (400 MHz, DMSO- d 6) δ 11.61 (s, 1H), 9.26 (s, 1H), 8.81 – 8.69 (m, 2H), 8.19 (s, 1H), 7.75 (d,J = 8.0 Hz, 1H), 7.46 (t, J = 8.0 Hz, 1H), 7.28 –7.24 (m, 1H), 7.14 (dt, J = 15.2, 8.0 Hz, 2H), 6.88 (d, J = 8.8 Hz, 1H), 4.03 (t, J = 4.4 Hz, 2H), 3.95 – 3.90 (m, 2H), 3.80 – 3.71 (m, 4H), 3.62 (s, 8H), 2.81(d, J = 4.4 Hz, 3H). 13 C NMR (100 MHz, DMSO- d 6) δ 169.41, 158.37, 155.37, 155.10,149.02, 144.32, 139.87, 134.62, 132.01, 128.44, 122.27, 121.63, 120.96,114.99, 113.00, 107.77, 105.11, 70.82, 70.40, 70.29, 69.60, 69.50, 69.30,68.82, 26.79. ESI-HRMS C 26 H 30 ClN5O6([M+Na]+) calcd 566.1777, found 566.1782.HPLC purity: 98.8%. Example 15: In vitro activity testing methods and results To determine the cellular activity of the crown ether-containing diaminopyrimidine macrocyclic inhibitor, we used the MTT assay to test the antiproliferative activity of the macrocyclic compound in different cell lines. Cells grown to the logarithmic growth phase were washed with PBS, digested with trypsin containing EDTA, centrifuged, and counted. Cells were then divided into groups at a density of 5 x 10⁻⁶ cells / cells. 4The samples were seeded in 96-well plates with 5 replicates to first test whether the activity of the positive control was within the range reported in the literature. 24 h post-inoculation, the positive control was administered at concentration gradients of 50 μM – 5 μM – 0.5 μM – 0.05 μM – 0.005 μM. After 72 h of incubation, the drug-containing medium was discarded, and basal medium containing 10% MTT was added. After 4 h of incubation, the supernatant was discarded, and 150 μM DMSO was added to each well to dissolve the blue-purple crystalline formazan. The plates were then shaken for 10 min, and the absorbance at 490 nm was measured using a microplate reader. Results were processed in GraphPad, and the IC50 of the positive control was calculated. 50 Once the values ​​are consistent with the range reported in the literature, drug activity testing can be performed. The in vitro activity data of the corresponding compounds are shown in Table 1. The macrocyclic inhibitors containing crown ether fragments showed good anti-proliferation activity against glioblastoma cell lines, with significantly better effects than the lead compound TAE-226, and exhibited high selectivity compared to normal cells.

[0026] Table 1. In vitro activity data for Examples 7, 8, and 11-14

[0027] Example 16: In vivo PK test method and results In addition, the pharmacokinetic (PK) characteristics of the crown ether-containing diaminopyrimidine macrocyclic inhibitor 13 in ICR mice were investigated to determine its optimal route of administration in vivo. Experimental mice were purchased from Beijing Vita River Laboratory Animal Technology Co., Ltd., animal license number SCXK (Beijing) 2021-0006. Six ICR mice were randomly assigned to two groups. Mice were fasted for 12 hours prior to the experiment. Compound 16c was administered intravenously at a dose of 2 mg / kg and orally at a dose of 10 mg / kg, respectively. Approximately 0.05 mL of blood was collected from the cheek at 5 minutes, 15 minutes, 30 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 8 hours, and 24 hours after administration. Blood collection was performed using EDTA-K2 anticoagulant tubes and kept on wet ice for at least 15 minutes. Plasma was separated by centrifugation at 6000 rpm for 3 minutes and stored at -20°C until analysis. The concentration of compound 13 in mouse plasma samples was determined using LC-MS / MS (Triple Quad 5500), and pharmacokinetic parameters were calculated using WinNonlin software.

[0028] As shown in Table 2, compound 13 was administered intravenously (IV) at a dose of 2 mg / kg and orally via gastric sac (IG) at a dose of 10 mg / kg. The half-life (T) after IG administration is as follows: 1 / 2 = 1.90 hours) and average stay (MRT) 0-∞= 1.25 hours) significantly longer than the intravenous injection group (T = 1.25 hours) 1 / 2 = 0.361 hours, MRT 0-∞ = 0.277 hours), indicating that the IG pathway is more suitable for prolonging the duration of drug effect in vivo. The apparent volume of distribution (Vz) of the intravenous injection group was 2457 mL / kg, and the clearance (Cl) was as high as 4814 mL / hr / kg. Combined with its short half-life, this indicates that the drug is widely distributed and rapidly cleared in vivo, avoiding the possibility of drug accumulation and toxicity. The oral bioavailability was moderate (F = 18.7%). These findings highlight the advantages and challenges of the pharmacokinetic properties of compound 13, laying the foundation for strategies to further optimize and improve its in vivo stability and bioavailability.

[0029] Table 2. In vivo pharmacokinetic studies of compound 13

[0030] All data are expressed as mean ± SD from three independent experiments. C max : Peak Plasma Concentration, T max : Time to Peak Concentration,T 1 / 2 : elimination half-life period, AUC: area under curve, Vz: Apparent Volume of Distribution, Cl: Clearance, MRT: Mean Residence Time, F: oralbioavailability All data are expressed as mean ± standard deviation of three independent experiments; Cmax: peak plasma concentration; Tmax: time to peak concentration; T1 / 2: elimination half-life; AUC: area under the curve; Vz: apparent volume of distribution; Cl: clearance rate; MRT: mean residence time; F: oral bioavailability.

[0031] Finally, it should be noted that the above embodiments are merely representative examples of this application. Obviously, the technical solutions of this application are not limited to the above embodiments, and many variations are possible. All variations that can be directly derived or conceived by those skilled in the art from the content disclosed in this application should be considered within the scope of protection of this application.

Claims

1. A diaminopyrimidine macrocyclic compound containing a crown ether fragment, characterized in that, Its specific structural formula is shown in equation (Ⅰ) below: Equation (Ⅰ).

2. The compound according to claim 1, characterized in that, R1 is selected from any one of Cl, CF3, H, and NO2.

3. The compound according to claim 1, characterized in that, R2 can be any of the following structures: 。 4. The compound according to claim 1, characterized in that, Specifically, it is one of the following six structures: 。 5. The method for preparing the diaminopyrimidine macrocyclic compound containing a crown ether fragment as described in claim 1, characterized in that, The specific steps are as follows: 2,4-Dichloro-5-substituted pyrimidines were subjected to nucleophilic substitution reactions with 2-amino-N-methylbenzamide to obtain compounds with the structure shown in formula (II). The compounds with the structure shown in formula (II) were then coupled with amines with the structure shown in R2 to obtain diaminopyrimidine macrocyclic compounds with crown ether fragments as shown in formula (III). Formula (II); Formula (III).

6. The preparation method according to claim 5, characterized in that, R1 is selected from any one of Cl, CF3, H, and NO2.

7. The preparation method according to claim 5, characterized in that, R2 can be any of the following structures: 。 8. The use of the diaminopyrimidine macrocyclic compound containing a crown ether fragment as described in claim 1 in the preparation of pharmaceuticals.

9. The application according to claim 8, characterized in that, The drug in question is an anti-tumor drug.

10. The application according to claim 8, characterized in that, The aforementioned anti-tumor drug is an anti-glioma drug.

Images