2-position amino acid ester estradiol derivatives, processes for their preparation and use

Abstract

The application discloses a 2-position amino acid ester estradiol derivative, and a preparation method and application thereof. The chemical structural formula of the 2-position amino acid ester estradiol derivative is as follows: wherein R is selected from R1=, R2=, R3=, R4= and R5=. The 2-position amino acid ester estradiol derivative provided by the application has a significant inhibiting effect on various tumor cells such as human breast cancer cells, human ovarian cancer cells, human breast ductal carcinoma cells and human cervical cancer cells, and is suitable for preparing an anti-tumor drug, and is especially suitable for treating the above cancers.

Description

Technical Field

[0001] This invention relates to the field of antitumor drug technology, and in particular to estradiol derivatives of amino acid esters at position 2, their preparation methods and applications. Background Technology

[0002] In recent years, steroidal compounds have attracted widespread attention in the field of antitumor drug development due to their unique core structure and diverse physiological activities. Studies have confirmed that structural modifications to the steroid core or side chains, such as the introduction of heteroatoms, nitrogen-containing groups, or amino acids, can significantly enhance their anti-inflammatory, antitumor, and antioxidant activities, and improve the pharmacokinetic properties of the parent compound. This has become an important direction for the development of highly effective and low-toxicity antitumor steroidal drugs.

[0003] 2-Methoxyestradiol is a commonly used anticancer steroid drug in clinical practice. Its core structural feature is the introduction of a methoxy functional group at the 2-position of estradiol. However, this drug suffers from rapid metabolism and low bioavailability, which seriously affects clinical treatment efficacy. Researchers have modified the structure of estrone analogs through sulfonation, esterification, and other methods. Although this has improved the stability of the compounds and overcome metabolic defects to some extent, problems such as narrow tumor cell inhibitory spectrum and insufficient activity still exist. There is an urgent need to develop new estrogenic antitumor compounds with novel structures.

[0004] Amino acids are the basic building blocks of human proteins, possessing excellent biocompatibility and in vivo affinity, and participating in energy metabolism and substance transport processes. Introducing amino acids into drug molecules for structural modification can effectively improve the shortcomings of poor solubility and low cell membrane permeability of the parent drug, while simultaneously enhancing its biological activity and targeting. This strategy has been widely applied in the design and development of anti-tumor drugs. Furthermore, the conjugation derivatives of steroidal compounds and amino acids have become a research hotspot. Studies have confirmed that amino acid ester derivatives of parent drugs such as α-tocopherol, phosphonates, and alizarin exhibit excellent tumor cell proliferation inhibitory activity, providing important references for amino acid modification of steroidal compounds.

[0005] Based on this, this invention uses estradiol as a lead compound and introduces a Boc-protected amino acid ester group at its 2-position to design and synthesize a series of 2-position amino acid ester-estradiol derivatives. The aim is to obtain novel antitumor steroidal compounds with high tumor cell inhibitory activity and broad inhibitory spectrum, and to provide a simple and efficient preparation method for them, providing new candidates and technical ideas for antitumor drug development. Summary of the Invention

[0006] One object of the present invention is to solve at least the above-mentioned problems and to provide at least the advantages that will be described later.

[0007] Another objective of this invention is to provide a 2-position amino acid ester-estrogenic derivative to address the problem of poor efficacy of the existing drug 2-methoxyestradiol.

[0008] Another object of the present invention is to provide a method for preparing the above-mentioned compound.

[0009] A further object of the present invention is to provide the use of the above-mentioned compounds in antitumor drugs.

[0010] To achieve the above objectives and other advantages, the present invention adopts the following approach: The chemical structure of the 2-position amino acid ester-estrogen derivative is as follows: ; Where R is selected from R1= R2= R3= R4= R5= One of them.

[0011] The method for preparing the 2-position amino acid ester-estrogenic derivative provided in this application includes the following steps: S1. Using representative amino acids that make up human proteins as raw materials, Boc protection of amino groups was performed to obtain intermediate compounds 2a, 2b, 2c, 2d, and 2e. S2. Using estrone as a raw material, its 2-C position is nitrated to obtain the fourth compound; S3. Under Pd / C catalysis and hydrogen conditions, the nitro group of the fourth compound is reduced to obtain the fifth compound containing an amino group at the 2-position carbon. S4. CDI was used to react with compounds 2a-2e to form acylimidazolium intermediates, which were then condensed with the fifth compound. After purification by column chromatography, 2-amino acid ester-estrogenic derivatives 6a-6e were obtained.

[0012] Preferably, the representative amino acids mentioned in step S1 are glycine, L-valine, L-phenylalanine, L-leucine, and L-tryptophan; compounds 2a to 2e are respectively: 2a is Boc-glycine, 2b is Boc-L-valine, 2c is Boc-L-phenylalanine, 2d is Boc-L-leucine, and 2e is Boc-L-tryptophan, with the following specific chemical structural formulas: ; Where R corresponds to R1= R2= R3= R4= R5= .

[0013] Preferably, step S1 specifically includes: Dissolve 0.50-1.50 g of the representative amino acid in 20-40 mL of DMF, slowly add 3.92-5.88 mmol of Boc anhydride at 80 °C, stir for 10 h, and monitor the reaction endpoint by thin-layer chromatography. After the reaction is complete, remove the solvent by rotary evaporation under reduced pressure, extract with ethyl acetate 3 times, wash with water 3 times, combine the organic phases, wash twice with saturated sodium chloride solution, dry with anhydrous sodium sulfate, and purify by column chromatography after rotary evaporation under reduced pressure to obtain white solid compounds 2a-2e.

[0014] Preferably, step S2 specifically includes: 1.16 g of estrone was dissolved in 50 mL of ethyl acetate, and 4 mL of glacial acetic acid and 2 mL of concentrated nitric acid were slowly added dropwise. The mixture was stirred at room temperature for 0.5 h, and the reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, sodium hydroxide was added to adjust the pH to neutral, and the mixture was extracted with ethyl acetate. The organic phase was washed three times with saturated sodium bicarbonate solution, twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by column chromatography to obtain a bright yellow solid, compound IV, whose chemical formula is as follows: .

[0015] Preferably, step S3 specifically includes: 1.30 g of the fourth compound was dissolved in 50 mL of anhydrous methanol, and 0.42 g of Pd / C was added. The mixture was stirred at room temperature for 1 h under a hydrogen atmosphere, and the reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, Pd / C was removed by filtration, the solvent was evaporated, and the mixture was purified by column chromatography to obtain a white solid fifth compound, the chemical formula of which is as follows: .

[0016] Preferably, step S4 specifically includes: 1.00-2.00 g of compounds 2a-2e were dissolved in dehydrated DMF. 600-900 mg of CDI was added in portions under ice bath conditions, and the mixture was stirred at 0°C for 1 h to form acylimidazoles. 1.00-2.00 g of compound 5 was added, and the reaction was carried out overnight under ice bath conditions. The reaction endpoint was monitored by thin-layer chromatography. After the reaction was complete, the mixture was quenched with water, extracted three times with ethyl acetate, and the combined organic phases were washed once with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by column chromatography to obtain white solid 2-amino acid ester-estrogenic derivatives 6a-6e, with the following specific chemical structures: ; Where R corresponds to R1= R2= R3= R4= R5= .

[0017] This application also provides the use of the aforementioned 2-position amino acid ester-estrogenic derivative in the preparation of antitumor drugs.

[0018] The antitumor drug is a drug that targets human breast cancer cells, human ovarian cancer cells, human breast ductal carcinoma cells, and human cervical cancer cells. The dosage form of the antitumor drug is an injection, tablet, pill, capsule, suspension, or emulsion. The route of administration is oral, subcutaneous, intravenous, or intramuscular.

[0019] The present invention has at least the following beneficial effects: The 2-position amino acid ester-estrogenic derivative of this invention is a novel estrogenic compound. By introducing an amino acid ester group at the 2-position of estrone, it significantly improves the activity defects of traditional estrogenic antitumor drugs and has a good inhibitory effect on human breast cancer, ovarian cancer, breast ductal carcinoma, cervical cancer and other tumor cells. Some compounds have better inhibitory activity than commercially available 2-methoxyestradiol, providing new candidates for the development of antitumor drugs. The preparation method of this invention uses common human amino acids and estradiol as raw materials, which are readily available and inexpensive. The reaction steps are simple, requiring only four steps: amino protection, nitration, reduction, and condensation to obtain the target product. Each step is carried out under mild and easily controlled conditions, resulting in high product yields (intermediate yield over 90%, target product yield over 84%). The product has high purity after column chromatography purification, making it suitable for large-scale preparation. The derivative of this invention uses amino acids as modifying groups, exhibiting good biocompatibility and in vivo affinity, effectively improving drug solubility and permeability, laying the foundation for its in vivo application. Furthermore, this derivative can be formulated into various dosage forms, offering flexible administration routes and broad clinical application prospects. In vitro antitumor proliferation activity assays showed that the present invention has inhibitory effects on human breast cancer cells, human ovarian cancer cells, human cervical cancer cells, human liver cancer cells, and prostate cancer cells. Therefore, the 2-position amino acid ester-estrogenic derivative of the present invention can be used to prepare a drug for treating tumors. This drug can be formulated into injections, tablets, pills, capsules, suspensions, or emulsions, and can be administered orally, or via subcutaneous, intravenous, or intramuscular injection.

[0020] Other advantages, objectives and features of the present invention will become apparent in part from the following description, and in part from those skilled in the art through study and practice of the invention. Detailed Implementation

[0021] The present invention will now be described in further detail so that those skilled in the art can implement it based on the description.

[0022] It should be understood that terms such as “having,” “comprising,” and “including” as used herein do not exclude the presence or addition of one or more other elements or combinations thereof.

[0023] It should be noted that, unless otherwise specified, the experimental methods described in the following implementation plan are all conventional methods, and the reagents and materials described are all commercially available unless otherwise specified.

[0024] The reaction route for the 2-position amino acid ester-estrogenic derivative of the present invention is as follows: .

[0025] Example 1 Preparation of compound 2 (a~e) (a) Preparation of compound 2a (Boc-glycine): R= ; 0.29 g g of glycine was dissolved in 30 mL of DMF, and 4.31 mmol of Boc anhydride was slowly added at 80 °C. The mixture was stirred for 10 h, and the reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, the mixture was evaporated under reduced pressure, the solvent was dried, and the mixture was extracted three times with ethyl acetate and washed three times with water. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, evaporated under reduced pressure, and purified by column chromatography to obtain a white solid with a yield of 90%.

[0026] (ii) Preparation of compound 2b (Boc-L-valine): R= ; 0.46 g of L-valine was dissolved in 30 mL of DMF, and 4.31 mmol of Boc anhydride was slowly added at 80 °C. The mixture was stirred for 10 h, and the reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, the solvent was evaporated under reduced pressure, and the mixture was extracted three times with ethyl acetate and washed three times with water. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, evaporated under reduced pressure, and purified by column chromatography to obtain a white solid with a yield of 93%.

[0027] (III) Preparation of compound 2c (Boc-L-phenylalanine): R= ; 0.65 g of L-phenylalanine was dissolved in 35 mL of DMF, and 4.31 mmol of Boc anhydride was slowly added at 80 °C. The mixture was stirred for 10 h, and the reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, the solvent was evaporated under reduced pressure, and the mixture was extracted three times with ethyl acetate and washed three times with water. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, evaporated under reduced pressure, and purified by column chromatography to obtain a white solid with a yield of 90%.

[0028] (iv) Preparation of compound 2d (Boc-L-leucine): R= ; 0.51 g of L-leucine was dissolved in 30 mL of DMF, and 4.31 mmol of Boc anhydride was slowly added at 80 °C. The mixture was stirred for 10 h, and the reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, the solvent was evaporated under reduced pressure, and the mixture was extracted three times with ethyl acetate and washed three times with water. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, evaporated under reduced pressure, and purified by column chromatography to obtain a white solid with a yield of 91%.

[0029] (v) Preparation of compound 2e (Boc-L-tryptophan): R= ; 0.80 g of L-tryptophan was dissolved in 35 mL of DMF, and 4.31 mmol of Boc anhydride was slowly added at 80 °C. The mixture was stirred for 10 h, and the reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, the mixture was evaporated under reduced pressure, the solvent was dried, and the mixture was extracted three times with ethyl acetate and washed three times with water. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, evaporated under reduced pressure, and purified by column chromatography to obtain a white solid with a yield of 91%.

[0030] Example 2 Synthesis of the fourth compound: ; 1.16 g of estradiol was dissolved in 50 mL of ethyl acetate, 4 mL of glacial acetic acid was slowly added dropwise, and 2 mL of concentrated nitric acid was added dropwise. The mixture was stirred at room temperature for 0.5 h. The reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, sodium hydroxide was added to adjust the pH to neutral. The mixture was extracted with ethyl acetate, washed three times with saturated sodium bicarbonate solution, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by column chromatography to obtain a bright yellow solid with a yield of 57%.

[0031] Example 3 Synthesis of the fifth compound: ; Take 1.30 g of the fourth compound, dissolve it in 50 mL of anhydrous methanol, add 0.42 g of Pd / C, stir at room temperature for 1 h under hydrogen atmosphere, monitor the reaction endpoint by thin-layer chromatography, after the reaction is complete, filter Pd / C, evaporate the solvent, and purify by column chromatography to obtain a white solid as the fifth compound, with a yield of 96%.

[0032] The fifth compound can be prepared multiple times using the above method or by increasing the amount of each raw material to obtain a higher yield.

[0033] Example 4 Synthetic 2-amino acid ester-estrogenic derivatives 6a~6e (a) Synthesis of 2-position amino acid ester-estrogen derivative 6a: R= ; 0.68 g of compound 2a was dissolved in dehydrated DMF, and 699 mg of CDI was added in portions under ice bath conditions. The mixture was stirred at 0°C for 1 h, and then 1.24 g of compound 5 was added. The reaction was carried out overnight under ice bath conditions. The reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, the mixture was extracted with ethyl acetate, washed three times with saturated sodium bicarbonate solution, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by column chromatography to give a white solid with a melting point of 151-152 °C and a yield of 84%. 1 H NMR (400 MHz, Chloroform- d ) d 8.83 (s, 1H), 8.62 (s, 1H), 7.12 (s, 1H), 6.67 (s, 1H), 5.71 (s, 1H), 3.98(s,2H), 1.46 (s, 9H), 0.87 (s, 3H). 13 C NMR (100 MHz, CDCl3) d 169.33, 156.61,146.00, 135.10, 131.84, 123.04, 119.32, 118.31, 80.81, 60.48, 50.36, 48.00,43.76, 38.16, 35.90, 31.51,28.87, 28.37, 28.30, 26.46, 25.83, 21.55, 21.06,14.18, 13.84. HRMS(ESI + ) calcd. for C 25 H 34 N₂O₅[M+Na] + ]465.2365, found 465.2368. (ii) Synthesis of 2-position amino acid ester-estrogenic derivative 6b: R = ; 0.85 g of compound 2b was dissolved in dehydrated DMF. 699 mg of CDI was added in portions under ice bath conditions, and the mixture was stirred at 0 °C for 1 h. Then, 1.24 g of compound 5 was added, and the mixture was reacted overnight under ice bath conditions. The reaction endpoint was monitored by thin-layer chromatography. After the reaction was complete, the mixture was extracted with ethyl acetate, washed three times with saturated sodium bicarbonate solution, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by column chromatography to give a white solid with a melting point of 115-116 °C and a yield of 90%. 1 H NMR (400 MHz, Chloroform- d ) d 9.15 (s, 1H), 8.95 (s, 1H), 6.81 (s, 1H), 6.62 (s, 1H), 5.43 (d, J = 8.5 Hz,1H), 4.25 (s, 1H), 1.43 (s, 9H), 1.06 (s, 3H), 1.04 (s, 3H), 0.84 (s, 3H). 13 CNMR (100 MHz, CDCl3) d 221.25, 171.97, 156.71, 146.00, 134.92, 131.74, 123.23,119.36, 118.84, 80.65,60.70, 50.35, 47.99, 43.62, 38.18, 35.93, 31.52, 31.02,29.70, 29.67, 28.93, 28.36, 26.53, 25.67, 21.58, 19.35, 18.49, 13.87. HRMS(ESI + ) calcd. for C 28 H 40 N2O5[M+H + ]485.3015, found 485.3008. (III) Synthesis of 2-position amino acid ester-estrogen derivative 6c: R = ; 1.04 g of compound 2c was dissolved in dehydrated DMF. 699 mg of CDI was added in portions under ice bath conditions. The mixture was stirred at 0 °C for 1 h. Then, 1.24 g of compound 5 was added, and the mixture was reacted overnight under ice bath conditions. The reaction endpoint was monitored by thin-layer chromatography. After the reaction was complete, the mixture was extracted with ethyl acetate, washed three times with saturated sodium bicarbonate solution, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by column chromatography to give a white solid with a melting point of 115-116 °C and a yield of 87%.1 H NMR (400 MHz, Chloroform- d ) d 8.85 (s, 1H), 8.69 (s, 1H), 7.29 (d, J = 6.2 Hz, 2H), 7.25 (d, J = 6.6 Hz, 1H),7.23 - 7.20 (m, 2H), 6.66 (s, 1H), 6.63 (s, 1H), 5.41 (d, J = 7.8 Hz, 1H), 4.72(s, 1H), 3.17 (dd, J = 13.9, 6.5 Hz, 1H), 3.06 (dd, J = 13.9, 7.9 Hz, 1H), 1.39(s, 9H), 0.85 (s, 3H). 13 C NMR (100 MHz, CDCl3) d 221.24, 171.62, 156.21, 146.21,136.07, 135.28, 131.82, 129.22, 128.86, 127.16, 122.95, 119.50, 118.94,80.94, 56.29,50.36, 47.99, 43.68, 38.67, 38.17, 35.93, 31.67, 31.55, 29.70,28.93, 28.31, 26.51, 25.75, 21.59, 13.89. + ) calcd. for C 32 H 40 N2O5[M+H + ]533.2937, found 533.3019. (iv) Synthesis of 2-position amino acid ester-estrogenic derivative 6d: R= ; 0.91 g of compound 2d was dissolved in dehydrated DMF, and 699 mg of CDI was added in portions under ice bath conditions. The mixture was stirred at 0°C for 1 h, and then 1.24 g of compound 5 was added. The reaction was carried out overnight under ice bath conditions. The reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, the mixture was extracted with ethyl acetate, washed three times with saturated sodium bicarbonate solution, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by column chromatography to give a white solid with a melting point of 142-143 °C and a yield of 85%. 1 H NMR (400 MHz, Chloroform- d ) d 9.40 (s, 1H), 9.02 (s, 1H), 6.69 (s, 1H), 6.59 (s, 1H), 5.40 (d, J = 8.0 Hz, 1H), 4.54 (t, J = 7.8 Hz, 1H), 1.43 (s, 9H), 0.98 (d, J = 6.5 Hz, 3H), 0.94 (d, J =6.4 Hz, 3H), 0.84 (s, 3H). 13 C NMR (100 MHz, CDCl3) d 221.20, 173.18, 156.69,145.86, 134.76, 131.69, 123.36, 119.46, 118.83, 80.80, 53.67, 50.37, 47.98,43.56, 41.29,38.13, 35.94, 31.52, 29.67, 28.95, 28.38, 28.33, 26.56, 25.62,24.77, 24.74, 23.12, 21.58, 21.40, 13.87. + ) calcd. for C 29 H 42 N2O5[M+H + ]499.3172, found 499.3170. (v) Synthesis of 2-position amino acid ester-estrogenic derivative 6e: R= ; 1.19 g of compound 2e was dissolved in dehydrated DMF, and 699 mg of CDI was added in portions under ice bath conditions. The mixture was stirred at 0 °C for 1 h, and then 1.24 g of compound 5 was added. The reaction was carried out overnight under ice bath conditions. The reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, the mixture was extracted with ethyl acetate, washed three times with saturated sodium bicarbonate solution, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by column chromatography to give a white solid with a melting point of 153-154 °C and a yield of 86%. 1 H NMR (400 MHz, Chloroform- d ) d 8.65 (s, 1H), 8.58 - 8.43 (m, 1H), 8.38 (s, 1H), 7.62 (d, J = 7.9 Hz, 1H), 7.33(d, J = 8.1 Hz, 1H), 7.21 - 7.14 (m, 1H), 7.08 (t, J = 7.4 Hz, 1H), 7.03 (d, J =2.4 Hz, 1H), 6.65 (s, 1H), 6.53 (s, 1H), 5.35 (d, J = 7.3 Hz, 1H), 4.76 (s,1H), 3.30 (t, J = 6.0 Hz, 2H), 2.81 - 2.70 (m, 2H), 1.40 (s, 9H), 0.85 (s, 3H). 13 C NMR (100 MHz, CDCl3) d 221.24, 172.07, 156.03, 146.44, 136.31, 135.49,131.86, 127.26, 123.49, 122.84, 122.33, 119.89, 119.57, 119.04, 118.61,111.41, 109.77, 80.92, 60.46, 50.36, 47.98, 43.65, 38.13, 35.90, 31.51,29.71, 28.87, 28.29, 26.50, 25.75, 21.57, 13.86. HRMS(ESI + ) calcd. forC 34 H 41 N3O5[M+H + ] 572.3124, found 572.3127. Comparative test <Comparative Example 1> Commercially available 2-methoxyestradiol The MTT assay was used to test in vitro antitumor activity and determine IC50. 50 (Half-maximal inhibitory concentration) value. 4 × 10⁻⁶ cells of good growth... 4 Cells were incubated in 96-well plates (cells / mL) overnight. Samples were added according to a concentration gradient as experimental groups (containing the 2-amino acid ester-estrogenic derivatives (6a-6e) from the embodiments of this invention and the commercially available drug 2-methoxyestradiol as comparative examples), the control group was culture medium containing cells but no drug, and the blank group was culture medium without cells or drug. Control wells contained cells without any compound, and each condition was tested in six parallel Wells wells. After a period of culture, 20 μL MTT (5 mg / mL) was added to each well, and the plates were cultured for another 4 hours. The supernatant was discarded, and 200 μL DMSO was added to each well. The plates were shaken for 10 min, and the OD value was measured using a Bio-Tek ELX 800 analyzer. Cell inhibition rate = [(OD positive - OD negative) - (OD test - OD positive)] / (OD positive - OD negative) × 100%. The IC50 of the compound was calculated using GraphPadPrism 8 software. 50 The values ​​were determined, and statistical difference analysis was performed. IC 50 The values ​​are shown in Table 1.

[0034] Table 1: In vitro antitumor proliferative activity of 2-position amino acid ester-estrogen derivatives (IC50, unit: μmol / L) The in vitro antitumor proliferation activity assays shown in Table 1 above demonstrate that the 2-position amino acid ester-estrogenic derivative of this invention exhibits inhibitory effects on different cancer cells. Overall, the synthesized target compounds showed good in vitro antitumor activity against MCF-7, T47D, and HeLa cells.

[0035] IC 50 The smaller the value, the stronger the antitumor proliferative activity of the compound. Compound 6e showed the strongest inhibitory effect on MCF-7, T47D, and HeLa cells, with an IC50 value of [missing value]. 50 The values ​​were 2.41 μM, 2.68 μM, and 6.62 μM, respectively, significantly superior to other derivatives and positive control drugs, especially in terms of inhibitory activity against MCF-7 and T47D cells compared to 2-methoxyestradiol (IC50). 50 The significant increases in concentrations (>80 μM and 13.79 μM) indicate its promising potential against breast and cervical cancer. Compound 6d also exhibited good inhibitory activity against MCF-7 and HeLa cells (IC50 >80 μM and 13.79 μM), suggesting good anti-breast and anti-cervical cancer potential. 50The concentrations were 15.60 μM and 11.25 μM, respectively, but they had no significant inhibitory effect on SKOV-3 cells (IC50). 50 >80μM), indicating that it has a certain selectivity for different tumor cells. Compound 6c has a more prominent inhibitory effect on T47D cells (IC50). 50 The concentration (7.38 μM) showed relatively weak inhibitory effects on other tumor cells. The introduction of different amino acid residues significantly affected the antitumor activity of the compounds. Compound 6e, containing tryptophan residues, exhibited the strongest activity in multiple cell lines, possibly related to the enhanced affinity of tryptophan's aromatic heterocyclic structure for tumor cell targets. Compound 6d, containing leucine residues, showed some selective inhibition against MCF-7 and HeLa cells. Derivatives containing glycine, valine, and phenylalanine (6a-c) showed relatively weaker activity, indicating that the structure of the amino acid side chains plays a crucial regulatory role in activity. Compared to the clinically commonly used drug 2-methoxyestradiol, compound 6e showed significantly better inhibitory activity in MCF-7 and T47D cells than the control.

[0036] Although the embodiments of the present invention have been disclosed above, they are not limited to the applications listed in the specification and embodiments. They can be applied to various fields suitable for the present invention. For those skilled in the art, other modifications can be easily made. Therefore, the present invention is not limited to the specific details.

Claims

1. 2-amino acid ester-estrogenic derivative, characterized in that, The chemical structural formula is as follows: ； Where R is selected from R1= R2= R3= R4= R5= One of them. A method for preparing 2,2-position amino acid ester-estrogenic derivatives, characterized in that, Includes the following steps: S1. Using representative amino acids that make up human proteins as raw materials, Boc protection of amino groups was performed to obtain intermediate compounds 2a, 2b, 2c, 2d, and 2e. S2. Using estrone as a raw material, its 2-C position is nitrated to obtain the fourth compound; S3. Under Pd / C catalysis and hydrogen conditions, the nitro group of the fourth compound is reduced to obtain the fifth compound containing an amino group at the 2-position carbon. S4. CDI was used to react with compounds 2a-2e to form acylimidazolium intermediates, which were then condensed with the fifth compound. After purification by column chromatography, 2-amino acid ester-estrogenic derivatives 6a-6e were obtained.

3. The preparation method according to claim 2, characterized in that, The representative amino acids mentioned in step S1 are glycine, L-valine, L-phenylalanine, L-leucine, and L-tryptophan; compounds 2a to 2e are respectively: 2a is Boc-glycine, 2b is Boc-L-valine, 2c is Boc-L-phenylalanine, 2d is Boc-L-leucine, and 2e is Boc-L-tryptophan, with the following specific chemical structural formulas: ； Where R corresponds to R1= R2= R3= R4= R5= .

4. The preparation method according to claim 2, characterized in that, Step S1 specifically includes: Dissolve 0.20-1.50 g of the representative amino acid in 20-40 mL of DMF, slowly add 3.92-5.88 mmol of Boc anhydride at 80 °C, stir for 10 h, and monitor the reaction endpoint by thin-layer chromatography. After the reaction is complete, remove the solvent by rotary evaporation under reduced pressure, extract with ethyl acetate 3 times, wash with water 3 times, combine the organic phases, wash twice with saturated sodium chloride solution, dry with anhydrous sodium sulfate, and purify by column chromatography after rotary evaporation under reduced pressure to obtain white solid compounds 2a-2e.

5. The preparation method according to claim 2, characterized in that, Step S2 specifically includes: 1.16 g of estrone was dissolved in 50 mL of ethyl acetate, and 4 mL of glacial acetic acid and 2 mL of concentrated nitric acid were slowly added dropwise. The mixture was stirred at room temperature for 0.5 h, and the reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, sodium hydroxide was added to adjust the pH to neutral, and the mixture was extracted with ethyl acetate. The organic phase was washed three times with saturated sodium bicarbonate solution, twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by column chromatography to obtain a bright yellow solid, compound IV, whose chemical formula is as follows: 。 6. The preparation method according to claim 2, characterized in that, Step S3 specifically includes: 1.30 g of the fourth compound was dissolved in 50 mL of anhydrous methanol, and 0.42 g of Pd / C was added. The mixture was stirred at room temperature for 1 h under a hydrogen atmosphere, and the reaction endpoint was monitored by thin-layer chromatography. After the reaction was completed, Pd / C was removed by filtration, the solvent was evaporated, and the mixture was purified by column chromatography to obtain a white solid fifth compound, the chemical formula of which is as follows: 。 7. The preparation method according to claim 2, characterized in that, Step S4 specifically includes: 0.50-2.00 g of compounds 2a-2e were dissolved in dehydrated DMF. 600-900 mg of CDI was added in portions under ice bath conditions, and the mixture was stirred at 0°C for 1 h to form acylimidazoles. 1.00-2.00 g of the fifth compound was added, and the reaction was carried out overnight under ice bath conditions. The reaction endpoint was monitored by thin-layer chromatography. After the reaction was complete, the mixture was quenched with water, extracted with ethyl acetate, washed three times with saturated sodium bicarbonate solution, and the combined organic phases were washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by column chromatography to obtain white solid 2-amino acid ester-estrogenic derivatives 6a-6e, with the specific chemical structures shown below. ； Where R corresponds to R1= R2= R3= R4= R5= .

8. The use of the 2-position amino acid ester-estrogenic derivative of claim 1 in the preparation of an antitumor drug.

9. The application according to claim 8, characterized in that, The antitumor drug is a drug that targets human breast cancer cells, human ovarian cancer cells, human breast ductal carcinoma cells, and human cervical cancer cells. The dosage form of the antitumor drug is an injection, tablet, pill, capsule, suspension, or emulsion. The route of administration is oral, subcutaneous, intravenous, or intramuscular.

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