Coumarin derivative, preparation method and application thereof
By preparing the coumarin derivative NP845, the shortcomings of existing prostate cancer treatment methods have been addressed, achieving significant inhibition of prostate cancer cells and effective control of tumor proliferation, and providing an antagonistic drug solution for endothelin receptor A.
Patent Information
- Application Number
- CN202310479264.9
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2023-04-28
- Publication Date
- 2025-11-04
- Estimated Expiration
- 2043-04-28
AI Technical Summary
Current treatments for prostate cancer suffer from castration resistance in the later stages of hormone therapy, significant side effects of chemotherapy, and nursing problems associated with surgical treatment. There is an urgent need to develop effective drugs specifically targeting prostate cancer, especially antagonists targeting endothelin receptor A.
A coumarin derivative was prepared by synthesizing NP845 through a specific chemical reaction. This derivative was then used to prepare an anti-prostate cancer drug and an endothelin receptor A antagonist, which significantly inhibited the expression of endothelin receptor A in tumor tissue.
The coumarin derivative NP845 significantly inhibited the proliferation of prostate cancer cells LNCaP and reduced the expression of prostate-specific antigen in serum. When used in combination with docetaxel, it significantly inhibited tumor proliferation, showing good effects in both in vitro and in vivo experiments.
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Abstract
Description
TECHNICAL FIELD
[0001] The application belongs to the technical field of coumarin compounds, and particularly relates to a coumarin derivative as well as a preparation method and application thereof. BACKGROUND
[0002] Prostate cancer is an epithelial malignant tumor occurring in the prostate, and is the most common malignant tumor of the male urogenital system. With the continuous growth of the average life expectancy of Chinese people, the change of the dietary structure and the improvement of the diagnosis technology, the incidence of prostate cancer has shown a gradually increasing trend in recent years, which has brought a heavy burden to the society and the people.
[0003] The existing technology generally includes hormone therapy, chemotherapy or surgical therapy for the treatment of prostate cancer. The hormone therapy generally adopts anti-androgen drugs such as enzalutamide for treatment, however, the hormone therapy often leads to castration-resistant prostate cancer in the later stage; the chemotherapy generally mainly uses cytotoxic drugs such as methotrexate, which can cause bone marrow suppression and other side effects to the patient's body; and the surgical therapy faces a series of problems such as postoperative care. Therefore, it is urgent to develop effective therapeutic drugs specifically for prostate cancer. Researches show that endothelin receptor A (ETA) is overexpressed in the pathogenesis of prostate cancer, and antagonizing the expression of ETA has a positive significance for the treatment of prostate cancer. SUMMARY
[0004] The technical problem to be solved by the present application is to provide a coumarin derivative, a preparation method and application thereof, which can significantly inhibit the proliferation of prostate cancer cells LNCaP, can significantly inhibit the expression of endothelin receptor A in tumor tissues at a dosage of 20 mg / kg, and can obviously reduce the expression level of prostate-specific antigen in serum when combined with docetaxel, thereby playing an anti-tumor proliferation role.
[0005] To solve the above technical problems, the technical scheme adopted by the present application is as follows: a coumarin derivative, the structural formula of the coumarin derivative is
[0006] The present application also provides a method for preparing the above-mentioned coumarin derivative, which is as follows:
[0007] S1, coumarin, N,N-diisopropylethylamine and O-benzotriazole-tetramethyl urea hexafluorophosphate are dissolved in N,N-dimethylformamide, and reacted at room temperature for 0.5 h, then amino-functionalized polyethylene glycol is added, and reacted at room temperature for 12 h, and the intermediate product is obtained through column chromatography separation;
[0008] The structural formula of the amino-functionalized polyethylene glycol is The structural formula of the intermediate product is
[0009] S2, mixing the intermediate product obtained in S1 and DL-pyroglutamic acid, adding N,N-diisopropylethylamine and O-benzotriazole-tetramethyluronium hexafluorophosphate to react, separating by column chromatography to obtain the coumarin derivative; the coumarin derivative has a structural formula of
[0010] Preferably, the coumarin, N,N-diisopropylethylamine, O-benzotriazole-tetramethyluronium hexafluorophosphate, amino-functionalized polyethylene glycol and N,N-dimethylformamide in S1 are used in a ratio of 2.0 g:2.65 g:7.8 g:4.0 g:20 mL.
[0011] Preferably, the column chromatography in S1 is silica gel adsorption column chromatography.
[0012] Preferably, the coumarin, N,N-diisopropylethylamine, O-benzotriazole-tetramethyluronium hexafluorophosphate, amino-functionalized polyethylene glycol and N,N-dimethylformamide in S1 are used in a ratio of 2.0 g:2.65 g:7.8 g:4.0 g:20 mL.
[0013] Preferably, the column chromatography in S2 is silica gel adsorption column chromatography.
[0014] The application further provides the coumarin derivative for use in the preparation of an anti-prostate cancer drug.
[0015] The application further provides the coumarin derivative for use in the preparation of an endothelin receptor A antagonist drug.
[0016] Compared with the prior art, the application has the following advantages:
[0017] The coumarin derivative prepared by the application can be used in the preparation of an anti-prostate cancer drug, and the coumarin derivative prepared by the application can significantly inhibit the expression of endothelin receptor A in tumor tissues, and can be used in the preparation of an endothelin receptor A antagonist drug.
[0018] The application will be further described in detail below with reference to the drawings and examples. BRIEF DESCRIPTION OF DRAWINGS
[0019] Figure 1 The coumarin derivative prepared in Example 1 is shown in the structure characterization mass spectrum.
[0020] Figure 2 The coumarin derivative prepared in Example 1 is shown in the structure characterization mass spectrum.
[0021] Figure 3 The coumarin derivative prepared in Example 1 is shown in the structure characterization mass spectrum.
[0022] Figure 4 Figure for the LNCaP cell proliferation inhibition effect of the coumarin derivative of Example 2.
[0023] Figure 5 Figure for the ETA inhibition effect in LNCaP cells of the coumarin derivative of Example 3.
[0024] Figure 6 Figure for the administration mode of the LNCaP cell transplanted tumor nude mice of Example 4.
[0025] Figure 7 Figure for the tumor volume index and tumor specific antigen index of the mice of Example 4. DETAILED DESCRIPTION
[0026] Example 1
[0027] The coumarin derivative of the present example has the structural formula of
[0028] The present example also provides a method for preparing the above-mentioned coumarin derivative, which is as follows:
[0029] S1, dissolve 2.0 g of coumarin, 2.65 g of N,N-diisopropylethylamine and 7.8 g of O-benzotriazol-tetramethylurea hexafluorophosphate in 20 mL of N,N-dimethylformamide, react at room temperature for 0.5 h, then add 4.0 g of amino-functionalized polyethylene glycol, react at room temperature for 12 h, separate by silica gel adsorption column chromatography (dichloromethane:methanol = 20:1) to obtain an intermediate product;
[0030] The amino-functionalized polyethylene glycol has the structural formula of The intermediate product has the structural formula of
[0031] S2, mix 2.0 g of the intermediate product obtained in S1 and 0.93 g of DL-pyroglutamic acid, then add 1.4 g of N,N-diisopropylethylamine and 4.1 g of O-benzotriazol-tetramethylurea hexafluorophosphate for reaction, separate by silica gel adsorption column chromatography (dichloromethane:methanol = 30:1) to obtain a coumarin derivative, denoted as NP845; the coumarin derivative has the structural formula of
[0032] Figure 1 is the structural characterization mass spectrum of the coumarin derivative prepared in the present example, Figure 2 is the structural characterization hydrogen spectrum, Figure 3 is the structural characterization carbon spectrum.
[0033] Example 2
[0034] This example is the application of the coumarin derivative (NP845) prepared in Example 1 for the preparation of an anti-prostate cancer drug.
[0035] This example uses LNCaP cell line as the research object, and the cell line is purchased from Wuhan Punsen Life Science and Technology Co., Ltd.
[0036] The cell counting method (CCK8 method) was used to determine the proliferation inhibition effect of NP845 on LNCaP: 10% fetal bovine serum and 100 U / mL penicillin-streptomycin (1:1) were added to RPMI 1640 medium, LNCaP cells were inoculated in a 96-well plate, and cultured at 37°C in 5% carbon dioxide for 24 hours. The cells were observed to be well attached and growing under a microscope. Different concentrations of NP845 were added and cultured for 12 hours. 1 / 10 volume of cell counting reagent (CCK8) was added and cultured for another 2 hours. The absorbance at 450 nm was read using an enzyme marker.
[0037] Figure 4 The schematic diagram of the inhibition effect of NP845 on the proliferation of LNCaP cells is shown in Figure 1. Figure 4 It can be seen that the ETA positive tool drug atrasentan can significantly inhibit the proliferation of LNCaP cells, and the IC 50 50 of NP845 on LNCaP cell proliferation is 47.71 μM, which is in the same order of magnitude as atrasentan, indicating that NP845 has good in vitro pharmacological activity.
[0038] The experiment shows that the coumarin derivative NP845 can significantly inhibit the proliferation of prostate cancer cells LNCaP in vitro, and the IC 50 50 is 47.71 μM.
[0039] Example 3
[0040] This example is the application of the coumarin derivative (NP845) prepared in Example 1 for the preparation of an anti-prostate cancer drug.
[0041] Western blot was used to determine the inhibitory effect of NP845 on ETA in LNCaP cells: LNCaP cells were inoculated in 96-well plates in RPMI 1640 medium containing 10% fetal bovine serum and 100 U / mL penicillin-streptomycin (1:1) and cultured at 37°C in 5% carbon dioxide for 24 hours. The cells were observed under a microscope to be well adhered and in good condition. NP845 was added to a concentration of 5, 10, 25, 50, and 100 μM, respectively, and cultured for 12 hours. The cells were treated with RIPA lysis buffer, and equal amounts of cells were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The gel was transferred to a nitrocellulose membrane, which was blocked with 3% bovine serum albumin and then incubated with 1:1000 diluted ETA primary antibody at room temperature for 2 hours. Horseradish peroxidase-conjugated secondary antibody was added and incubated overnight.
[0042] Figure 5 Western blot was used to analyze the inhibitory effect of NP845 on ETA in LNCaP cells. Figure 5 It was found that NP845 inhibited the expression of ETA in LNCaP cells in a dose-dependent manner, indicating that NP845 is a specific inhibitor of ETA.
[0043] Example 4
[0044] This example describes the application of the coumarin derivative (NP845) prepared in Example 1 to the inhibition of LNCaP transplanted tumors in nude mice.
[0045] Figure 6 Schematic diagram of the administration method for LNCaP cell transplanted tumor nude mice. LNCaP cells were suspended in Matrigel to a concentration of (5 x 10 7 The nude mouse model was divided into four groups, with eight mice in each group. During the 21-day administration period, docetaxel was administered only on days 1, 4, 7, and 9 at a dose of 5 mg / Kg via tail vein injection, while NP845 was administered continuously for 21 days via intraperitoneal injection.
[0046] The tumor was collected 12 hours after the last administration, and the tumor volume was measured along two orthogonal axes using a caliper. In vivo mouse experiments showed that NP845 at a dose of 20 mg / kg in combination with docetaxel significantly inhibited the proliferation rate of the tumor. The concentration of specific antigens in the tumor tissue of each group of nude mice was determined by enzyme-linked immunosorbent assay.
[0047] Figure 7Fig. 6 is a graph showing the tumor volume and serum prostate cancer specific antigen index of nude mice, wherein (A) is a graph showing the tumor volume index of LNCaP cell transplanted tumors treated with NP845, and (B) is a graph showing the tumor specific antigen index. The results show that NP845 can significantly inhibit the proliferation of tumor cells and reduce the concentration of specific antigen in tumor tissue. Figure 7 It can be seen that NP845 can significantly inhibit the proliferation of tumor cells and reduce the concentration of specific antigen in tumor tissue at a dose of 20 mg / kg, which indicates that NP845 can be used for the treatment of prostate cancer.
[0048] The above description is only the preferred embodiments of the present application, and does not limit the present application in any way. Any simple modification, change and equivalent change made according to the technical essence of the present application to the above embodiments are still within the protection scope of the technical solution of the present application.
Claims
1. A coumarin derivative, characterized by, The structural formula of the coumarin derivative is 2. A process for the preparation of the coumarin derivative according to claim 1, characterized in that, The method comprises: S1, dissolving coumarin, N,N-diisopropylethylamine and O-benzotriazole-tetramethyl urea hexafluorophosphate in N,N-dimethylformamide, reacting at room temperature for 0.5 h, then adding amino-functionalized polyethylene glycol, reacting at room temperature for 12 h, and separating by column chromatography to obtain an intermediate product; The amino-functionalized polyethylene glycol has a structure of The intermediate product has a structure of S2, mixing the intermediate product obtained in S1 with DL-pyroglutamic acid, adding N,N-diisopropylethylamine and O-benzotriazole-tetramethyluronium hexafluorophosphate to react, and separating by column chromatography to obtain the coumarin derivative; the structural formula of the coumarin derivative is 3. The method of claim 2, wherein, The amount ratio of the coumarin, N,N-diisopropylethylamine, O-benzotriazole-tetramethyl urea hexafluorophosphate, amino-functionalized polyethylene glycol and N,N-dimethylformamide in S1 is 2.0 g:2.65 g:7.8 g:4.0 g:20 mL.
4. The method of claim 2, wherein, The column chromatography in S1 is silica gel adsorption column chromatography.
5. The method of claim 2, wherein, The mass ratio of the intermediate product, DL-pyroglutamic acid, N,N-diisopropylethylamine and O-benzotriazole-tetramethyl urea hexafluorophosphate in S2 is 2.0:0.93:1.4:4.
1.
6. The method of claim 2, wherein, The column chromatography in S2 is silica gel adsorption column chromatography.
7. Use of a coumarin derivative according to claim 1, characterized in that, The coumarin derivative is used for preparing an anti-prostate cancer drug.
8. Use of a coumarin derivative according to claim 1, characterized in that, The coumarin derivative is used for preparing an endothelin receptor A antagonist drug.