Application of flavonoid compound in preparation of medicine for treating breast cancer

Abstract

The invention provides an application of a flavonoid compound in preparation of a medicine for treating breast cancer. The compound can effectively inhibit proliferation of four types of triple-negative breast cancer cells, namely MDA-MB-231, MDA-MB-468, Cal51 and BT549.

Description

Technical Field

[0001] The present invention belongs to the technical field of pharmaceutical compound manufacturing, and particularly relates to an application of a flavonoid compound in preparing a drug for treating breast cancer. Background Art

[0002] Flavonoids are polyphenolic compounds, mainly including: isoflavonoids, flavanones, flavanols, flavonols, flavonoids and anthocyanins in various plants. Fruits, vegetables, and plant-based beverages (such as green tea, wine, and cocoa products) are the main dietary sources of flavonoids. Flavonoids have multiple anti-cancer effects, such as regulating the activity of reactive oxygen species (ROS) scavenging enzymes, participating in cell cycle arrest, inducing cell apoptosis, autophagy, and inhibiting cancer cell proliferation and invasion. Flavonoids have a dual effect on ROS homeostasis, acting as antioxidants under normal conditions and being effective pro-oxidants in cancer cells, which can trigger apoptotic pathways and downregulate pro-inflammatory signaling pathways.

[0003] Breast cancer is one of the most common malignant tumors in women. Breast cancer is a systemic disease. In the early and middle stages of the disease, breast cancer cells can enter the circulatory system and metastasize throughout the body. Currently, the main treatments for breast cancer are surgery, chemotherapy, radiotherapy, and endocrine therapy. However, the survival rate for patients with advanced breast cancer is low, and cure is difficult. Breast cancer pathological types include luminal A, luminal B, normal breast-like, human epidermal growth factor receptor 2 (HER-2)-overexpressing, and basal-like 2. Triple-negative breast cancer is a type of breast cancer that is negative for estrogen receptors, progesterone receptors, and HER-2. Overexpression and amplification of human epidermal growth factor receptor 2 (HER2) have been found in various solid tumors, including breast and gastric cancers. For patients who test HER2 positive, treatment with trastuzumab targeting HER2 combined with chemotherapy has shown significant results. However, for patients with HER2-negative advanced breast cancer who have failed endocrine therapy, chemotherapy remains the primary treatment. The "Chinese Breast Cancer Diagnosis and Treatment Guidelines (2018 Edition)" issued by the National Health Commission states that commonly used chemotherapy drugs for advanced breast cancer include anthracyclines (epirubicin, doxorubicin, and doxycycline), taxanes, vinorelbine, capecitabine, gemcitabine, and platinum-based chemotherapy. However, their severe side effects limit their clinical application, and patients often discontinue treatment due to intolerance to severe adverse reactions. Therefore, developing chemotherapy regimens with improved efficacy and reduced toxicity is crucial. Summary of the Invention

[0004] The present invention provides a compound of formula A (A) Application in the preparation of drugs for treating breast cancer.

[0005] The present invention provides a method for treating breast cancer, characterized in that a therapeutically effective amount of a compound of formula A is administered to a patient in need thereof.

[0006] The present invention also provides the application or use of the compound represented by formula A in treating breast cancer.

[0007] The present invention also provides a method for using the compound represented by formula A in treating breast cancer.

[0008] According to the present invention, the breast cancer is triple-negative breast cancer.

[0009] According to the present invention, the triple-negative breast cancer cells are MDA-MB-231, MDA-MB-468, Cal51 and BT549.

[0010] The drug further contains a pharmaceutically acceptable carrier.

[0011] The pharmaceutically acceptable carriers are various excipients commonly used or known in the pharmaceutical field, including but not limited to: diluents, binders, antioxidants, pH regulators, preservatives, lubricants, disintegrants, etc.

[0012] Examples of diluents include lactose, starch, cellulose derivatives, inorganic calcium salts, and sorbitol. Examples of binders include starch, gelatin, sodium carboxymethyl cellulose, and polyvinyl pyrrolidone. Examples of antioxidants include vitamin E, sodium bisulfite, sodium sulfite, and butylated hydroxyanisole. Examples of pH adjusters include hydrochloric acid, sodium hydroxide, citric acid, tartaric acid, Tris, acetic acid, sodium dihydrogen phosphate, and disodium hydrogen phosphate. Examples of preservatives include methyl parahydroxybenzoate, ethyl parahydroxybenzoate, m-cresol, and benzalkonium chloride. Examples of lubricants include magnesium stearate, micropowdered silica gel, and talc. Examples of disintegrants include starch, methyl cellulose, xanthan gum, and cross-linked sodium carboxymethyl cellulose.

[0013] The amount of the compound of formula A contained in the medicine is 0.1-1000 mg, preferably 1-500 mg, more preferably 5-100 mg.

[0014] The pharmaceutical agent may be in the form of an oral dosage form, such as a tablet, capsule, pill, powder, granule, suspension, syrup, etc.; or in the form of an injectable dosage form, such as an injection solution, powder injection, etc., which is administered by intravenous, intraperitoneal, subcutaneous, or intramuscular injection. All dosage forms used are well known to those skilled in the pharmaceutical field.

[0015] Routes of administration of the drug include, but are not limited to: oral; buccal; sublingual; transdermal; pulmonary; rectal; parenteral, e.g., by injection, including subcutaneous, intradermal, intramuscular, intravenous; by implanted reservoir or reservoir.

[0016] The dosage of the compound of formula A will depend on the age, health and weight of the recipient, the type of concomitant medication, the frequency of treatment, the route of administration, etc. The drug can be administered as a single daily dose, once a day, once every two days, once every three days, once every four days, or the total daily dose can be administered in divided doses of two, three or four times a day. The dosage of the compound of formula A is 0.01-100 mg / kg / day, preferably 0.1-10 mg / kg / day, for example 0.5 mg / kg / day, 1 mg / kg / day, 2 mg / kg / day, 5 mg / kg / day, etc.

[0017] The compound of formula A or a drug containing the compound of formula A can be used in combination with other drugs for treating breast cancer.

[0018] According to the present invention, other drugs for treating breast cancer include but are not limited to: chemotherapy drugs, such as anthracyclines (epirubicin, doxorubicin, doxorubicin), cyclophosphamide, taxanes, vinorelbine, capecitabine, gemcitabine, platinums, etc.; hormone drugs, such as exemestane, letrozole, tamoxifen, etc.; anti-tumor angiogenesis drugs, such as apatinib, anlotinib, etc.; monoclonal antibodies, such as trastuzumab, etc.

[0019] The present invention also provides a method for preparing a compound of formula A, wherein compound 1 is used as a starting material to synthesize intermediate 6 through multiple steps of reaction, and then intermediate 6 is synthesized through two steps of reaction to obtain compound A. The reaction scheme is as follows:

[0020]

[0021] Intermediates 1-6 can be prepared according to synthetic methods known in the art, for example: Structure-Activity Relationships of Baicalein and its Analogs as Novel TSLP Inhibitors, Scientific Reports, 2019, 9, 1-21; Structural characterization of chrysinderivatives, Journal of Chemical and Pharmaceutical Research, 2020, 12, 24-27; Synthesis and Biological Evaluation of Acetylcholinesterase Inhibitor Macakurzin C and Its Derivatives, Synlett, 2015, 26, 1131-1134. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 Compound A inhibits the proliferation of MDA-MB-231 cells

[0023] Figure 2 Compound A inhibits the proliferation of MDA-MB-468 cells

[0024] Figure 3 Compound A inhibits the proliferation of Cal51 cells

[0025] Figure 4 Compound A inhibits the proliferation of BT549 cells DETAILED DESCRIPTION

[0026] The technical solutions of the present invention will be described in further detail below with reference to specific embodiments. It should be understood that the following embodiments are merely illustrative and explanations of the present invention and should not be construed as limiting the scope of protection of the present invention. All technologies implemented based on the above content of the present invention are encompassed within the scope of protection that the present invention is intended to protect.

[0027] Unless otherwise specified, the raw materials and reagents used in the following examples are commercially available or can be prepared by known methods.

[0028] CH3I: iodomethane; K2CO3: potassium carbonate; THF: tetrahydrofuran; DMSO: dimethyl sulfoxide; BBr3: boron tribromide; CuI: cuprous iodide; DMF: N,N-dimethylformamide; DEAD: diethyl azodicarboxylate; PPh3: triphenylphosphine; Eu(fod)3: tris(6,6,7,7,8,8,8-heptafluoro-2,2-dimethyl-3,5-octanedionato)europium; Tpp: thiamine pyrophosphate; DMAP: 4-dimethylaminopyridine; TEA: triethylamine; Ac2O: acetic anhydride.

[0029] Example 1 Synthesis of Compound A

[0030] 1) Synthesis of Intermediate 6

[0031] Compound 1 is used as the starting material, which reacts with iodomethane to produce intermediate 2; then intermediate 3 is produced under the action of benzaldehyde; it then reacts with elemental iodine and boron tribromide to produce intermediate 4; intermediate 4 reacts with cuprous iodide and potassium carbonate to produce intermediate 5; finally, intermediate 5 reacts with N,N-diethylaniline at a high temperature of 250°C to produce intermediate 6.

[0032]

[0033] a) CH3I, K2CO3, THF, 89%; b) 4M NaOH, Benzaldehyde, THF, 86%; c) 1) I2, DMSO; 2) BBr3, DCM, two-step yield 63%; d) K2CO3, CuI, DMF, 75%; e) N,N-Diethylaniline, 250°C, 46%.

[0034] Intermediate 6: 1 H NMR (400MHz, CDCl3) δ13.02(d,J=1.5Hz,1H),7.98–7.77(m,2H),7.51(dd,J=7.4,1.7Hz,3H),7.26(d,J=1.6Hz ,1H),6.72(dd,J=10.0,1.6Hz,1H),6.42(d,J=1.6Hz,1H),5.62(dd,J=10.1,1.6Hz,1H),1.48(d,J=1.7Hz,6H). 13 C NMR (100MHz, CDCl3) δ182.56,163.75,159.61,157.15,156.45,131.78,131.37, 129.09,128.18,126.26,115.50,105.73,105.67,105.60,95.12,78.05,28.32.

[0035] 2) Synthesis of Intermediate 7

[0036]

[0037] Under argon, intermediate 6 (50 mg, 0.156 mmol) and 3-methylbut-2-en-1-ol (20.17 mg, 0.234 mmol) were dissolved in anhydrous tetrahydrofuran and placed in a 50 mL round-bottom flask with stirring. DEAD (41 mg, 0.234 mmol) and triphenylphosphine (61 mg, 0.234 mmol) were added under ice-cooling. After stirring for 30 minutes, the reaction system was allowed to continue at room temperature. The reaction was monitored by TLC. Upon completion, 200 mL of ethyl acetate was added to the system. The reaction solution was washed and extracted with saturated sodium chloride solution (100 mL x 3). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by flash silica gel chromatography to yield compound 7 (32 mg, 80%) as a white solid.

[0038] 1H NMR (400MHz, Acetone-d6) δ8.04(dd,J=5.5,3.0Hz,2H),7.58(dd,J=5.7,2.8Hz,3H),6.80(d,J=2.7Hz,1H),6.73(dt,J=10.1,2.2Hz,1H),6.66 (d,J=2.6Hz,1H),5.86(dd,J=10.0,2.8Hz,1H),5.60(d,J=7.6Hz,1H),4.60(d,J=7.4Hz,2H),1.75(s,3H),1.67(s,3H),1.47(t,J=2.3Hz,6H). 13 C NMR (100MHz, Acetone-d6) δ175.74,160.46,158.67,157.93,154.01,137.63,131.58,131.30,130.59, 129.05,125.99,120.70,116.47,113.69,112.83,108.04,100.54,77.53,71.47,27.48,25.09,17.19.

[0039] 3) Preparation of Compound A

[0040]

[0041] Compound 7 (30 mg, 0.077 mmol) and Eu(fod)3 (4 mg, 0.004 mmol) were dissolved in chloroform and placed in a 50 mL round-bottom flask under reflux at 60°C. The reaction was monitored by TLC. Upon completion, 200 mL of ethyl acetate was added to the system. The reaction solution was washed and extracted with saturated sodium chloride solution (100 mL x 3). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by flash silica gel chromatography to obtain intermediate 8 (24 mg, 76%) as a white solid.

[0042] After dissolving intermediate 8 (35 mg, 0.090 mmol) in anhydrous tetrahydrofuran, the mixture was placed in a 50 mL round-bottom flask and stirred. Methyl iodide (0.01 mL, 0.180 mmol) and potassium carbonate (37 mg, 0.270 mmol) were slowly added dropwise under an ice bath. After stirring for 30 minutes, the reaction system was placed at room temperature to continue the reaction. The reaction was monitored by TLC. After the reaction was complete, 200 mL of ethyl acetate was added to the system. The reaction solution was washed and extracted with saturated sodium chloride solution (100 mL × 3). The organic phase was collected, treated with anhydrous sodium sulfate as a desiccant, and then filtered and concentrated. The crude product was purified by flash silica gel chromatography to obtain compound A (82%) as a white solid. Intermediate 8:

[0043] 1 H NMR (400MHz, Acetone-d6) δ13.21(s,1H),8.16–7.95(m,2H),7.72–7.50(m,3H),6.79(s,1H),6.66(d,J=10. 0Hz,1H),5.77(d,J=10.0Hz,1H),5.24(m,1H),3.53(d,J=7.1Hz,2H),1.84(s,3H),1.66(s,3H),1.48(s,6H). 13 C NMR (100MHz, Acetone-d6) δ182.78,163.74,156.80,154.47,154.42,131.88,131.58,131.28,129.19, 128.40,126.38,122.30,115.23,107.59,105.23,105.20,105.02,77.91,27.46,24.99,21.32,17.37.

[0044] Compound A: 1 H NMR (400MHz, Acetone-d6) δ8.14–7.92(m,2H),7.70–7.46(m,3H),6.74(d,J=10.1Hz,1H),6.66(s,1H),5.89(d ,J=10.1Hz,1H),5.32–5.22(m,1H),3.84(s,3H),3.61(d,J=7.0Hz,2H),1.85(s,3H),1.67(s,3H),1.49(s,6H). 13 C NMR (100 MHz, Acetone-d6) δ

[0045] 175.94,160.49,156.05,155.11,153.14,131.94,131.65,131.28,130.91,129.11,126.02,121.98,116.04,113.43,112.75,112.45,107.94,77.54,61.91,27.44,25.01,21.88,17.45.

[0046] Example 2 Anti-triple negative breast cancer cell proliferation experiment - CCK-8 detection

[0047] 1) Experimental Materials

[0048] Cells: Human breast cancer cells MDA-MB-231, MDA-MB-468, BT549, Cal51

[0049] Cell proliferation detection kit: Cell Counting Kit-8 (CCK8) kit (TargetMol)

[0050] Solvent: dimethyl sulfoxide (DMSO) (Solarbio)

[0051] 2) Experimental steps

[0052] Cell culture: MDA-MB-231, MDA-MB-468, Cal51, and BT549 cells were revived in T75 cell culture flasks and cultured in a 37°C 5% CO2 incubator. When the cell density in the cell culture flask reached above 90%, the cells were passaged for at least 2 times. The cells were then plated in 96-well cell culture plates at a cell density of 1×10 4 Culture overnight to allow cells to fully adhere to the wall.

[0053] Drug Treatment: Compound A was added to the experimental group cells to a final concentration of 10 μM. The negative control group (control) cells were treated with only DMSO, the solvent used to dissolve the compound. The positive control group was treated with 5-fluorouracil (5-FU). The blank group was treated with only culture medium without cells. The cells were incubated at 37°C in a 5% CO2 incubator for 48 hours.

[0054] Detection: After 48 hours, 10 μL of CCK-8 solution was added to each well of the experimental group, control group, and blank group and incubated for 1-2 hours. The absorbance at 450 nm was measured using a microplate reader to calculate the cell survival rate.

[0055] Cell survival rate = [(As-Ab) / (Ac-Ab)] × 100%

[0056] As: experimental well; Ac: negative control well; Ab: blank well

[0057] 3) Experimental results

[0058] The experimental results are as follows Figure 1-4 As shown, the experiment showed that compound A could significantly inhibit the proliferation of four triple-negative breast cancer cells: MDA-MB-231, MDA-MB-468, Cal51, and BT549.

[0059] The above describes the embodiments of the present invention. However, the present invention is not limited to the above embodiments. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention shall be included in the scope of protection of the present invention.

Claims

1. A compound of formula A, its stereoisomers, pharmaceutically acceptable salts or polymorphs: (A), Application in the preparation of drugs for treating breast cancer.

2. A method for treating breast cancer as claimed in claim 1, characterized in that: A therapeutically effective amount of a compound of formula A is administered to a patient in need thereof.

3. The method according to claim 1-2, wherein the breast cancer is triple-negative breast cancer.

4. As described in claim 3, the triple-negative breast cancer cells are MDA-MB-231, MDA-MB-468, Cal51 and BT549.

5. A pharmaceutical composition comprising the compound of formula A according to claim 1, its stereoisomers, pharmaceutically acceptable salts or polymorphs.

6. The pharmaceutical composition according to claim 5, characterized in that Further comprising other therapeutic drugs, the The other therapeutic drugs are selected from cancer therapeutic drugs.

7. The pharmaceutical composition according to claim 5-6, characterized in that It can be used to treat triple-negative breast cancer.

Images