Application of beta-elemonic acid in preparation of anti-breast cancer drugs

Abstract

The invention provides an application of beta-elemonic acid in preparation of an anti-breast cancer drug, the structural formula of the beta-elemonic acid is # imgabs0 #, the research finds that the beta-elemonic acid shows good anti-breast cancer activity in vivo and in vitro, can induce breast cancer cell death, can be used as a drug active component to prepare the anti-breast cancer drug, and can be used for preparing the anti-breast cancer drug. Good application prospects are realized in the field of breast cancer treatment.

Description

Technical Field

[0001] The invention relates to the technical field of biomedicine, and in particular to use of beta-elemenonic acid in preparing an anti-breast cancer drug. Background Art

[0002] Breast cancer is one of the most common malignant tumors in women. According to the 2024 global cancer statistics, the incidence and mortality of breast cancer in women ranked first and second respectively. Breast cancer can be roughly divided into: luminal A, luminal B, HER2 overexpression, triple-negative breast cancer, Normal like, etc. according to different analysis types.

[0003] At present, the treatment methods for breast cancer include surgery, chemotherapy, radiotherapy, immunotherapy, targeted therapy and traditional Chinese medicine. Western medicine has serious side effects and is harmful to the human body. my country has a long history of using traditional Chinese medicine, and traditional Chinese medicine has few side effects. Therefore, it is necessary to study small molecule drugs of traditional Chinese medicine for the treatment of breast cancer.

[0004] β-elemonic acid is a triterpenoid active ingredient extracted from the oleaginous plant frankincense tree or the colloidal resin frankincense in the stem of the same plant. Studies in recent years have shown that β-elemenonic acid has anti-inflammatory, anti-ulcer, anti-tumor effects and has important drug development value. At present, there is no research report on the anti-breast cancer activity of β-elemenonic acid. Summary of the invention

[0005] In view of the above-mentioned shortcomings of the prior art, the purpose of the present invention is to provide a use of β-elemenic acid in the preparation of anti-breast cancer drugs, which is used to solve the problem that the existing breast cancer treatment methods using Western medicines are very harmful to the human body.

[0006] To achieve the above purpose and other related purposes, the present invention provides a use of β-elemenonic acid as a pharmaceutical active ingredient in the preparation of an anti-breast cancer drug.

[0007] β-elemonic acid is a triterpenoid active ingredient extracted from the colloidal resin frankincense in the stem of the Oleaceae plant frankincense tree or the same plant. The inventor of the present application found through in-depth research and experiments that a triterpenoid β-elemenonic acid exhibits a strong killing effect on breast cancer tumor cells. The β-elemenonic acid of the present invention exhibits good anti-breast cancer activity in vivo and in vitro, can induce breast cancer cell death, and is a potential anti-breast cancer drug with good application prospects in the field of breast cancer treatment.

[0008] Preferably, the structural formula of the β-elemenic acid is:

[0009] β-elemenic acid is a triterpenoid compound extracted from frankincense produced by the stems of the Oleaceae plant Frankincense tree or plants of the same genus. It is insoluble in water but easily soluble in organic solvents. It is a colorless to light yellow solid at room temperature and has the smell of elemenine.

[0010] Preferably, the anti-breast cancer drug is a drug used for preventing and / or treating breast cancer.

[0011] Preferably, the dosage form of the anti-breast cancer drug is selected from any one of compressed tablets, dispersible tablets, enteric-coated tablets, capsules, granules, pellets, emulsions, powders, oral solutions and injections.

[0012] The present invention also provides an anti-breast cancer drug, comprising beta-elemenic acid and pharmaceutically acceptable excipients.

[0013] Preferably, the structural formula of the β-elemenic acid is:

[0014] Preferably, the dosage form of the anti-breast cancer drug is selected from any one of compressed tablets, dispersible tablets, enteric-coated tablets, capsules, granules, pellets, emulsions, powders, oral solutions and injections.

[0015] The present invention also provides a use of the anti-breast cancer drug in preparing a drug for preventing and / or treating breast cancer.

[0016] As described above, the present invention has the following beneficial effects: β-elemenic acid exhibits good anti-breast cancer activity in vivo and in vitro, can induce breast cancer cell death, can be used as a pharmaceutical active ingredient to prepare anti-breast cancer drugs, and has good application prospects in the field of breast cancer treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 This is a graph showing the effect of β-elemenic acid on the growth and proliferation of breast cancer cells MCF-7, BT-474, MDA-MB-231, MDA-MB-453, KPL-4, and Bcap-37 detected by the CCK8 method in the present invention.

[0018] Figure 2 This is a graph showing the effect of β-elemenic acid on apoptosis of breast cancer cells Bcap37 and MDA231 detected by flow cytometry in the present invention.

[0019] Figure 3 This is a diagram showing the effect of β-elemenate on the growth and proliferation of breast cancer cells using a nude mouse subcutaneous tumor-bearing experiment in the present invention. DETAILED DESCRIPTION

[0020] The present invention is further described below in conjunction with specific embodiments, but the protection scope of the present invention is not limited thereto.

[0021] It should be noted that the process equipment or devices not specifically specified in the following embodiments are all conventional equipment or devices in the art.

[0022] Before further describing the specific embodiments of the present invention, it should be understood that the scope of protection of the present invention is not limited to the specific embodiments described below; it should also be understood that the terms used in the examples of the present invention are for describing the specific embodiments rather than for limiting the scope of protection of the present invention; in the present specification and claims, unless otherwise expressly stated herein, the singular forms "a", "an" and "the" include plural forms.

[0023] When the embodiments give numerical ranges, it should be understood that, unless otherwise specified in the present invention, both endpoints of each numerical range and any numerical value between the two endpoints can be selected. Unless otherwise defined, all technical and scientific terms used in the present invention have the same meaning as those generally understood by those skilled in the art. In addition to the specific methods, equipment, and materials used in the embodiments, according to the grasp of the prior art by those skilled in the art and the record of the present invention, any methods, equipment, and materials of the prior art similar or equivalent to the methods, equipment, and materials described in the embodiments of the present invention can also be used to realize the present invention.

[0024] Unless otherwise stated, the experimental methods, detection methods, and preparation methods disclosed in the present invention all adopt conventional techniques in the field of molecular biology, biochemistry, chromatin structure and analysis, analytical chemistry, cell culture, recombinant DNA technology, and related fields.

[0025] The β-elemenic acid used in the following examples of the present invention was purchased from Shanghai Yuanye Biotechnology Co., Ltd. (CAS No.: 28282-25-9).

[0026] The structural formula of the β-elemenic acid is:

[0027] The present application also provides an anti-breast cancer drug, comprising β-elemenic acid and a pharmaceutically acceptable excipient. The dosage form of the anti-breast cancer drug is not limited, for example, it can be any one of a compressed tablet, a dispersible tablet, an enteric-coated tablet, a capsule, a granule, a pill, an emulsion, a powder, an oral solution and an injection.

[0028] The present application also provides a use of an anti-breast cancer drug for preparing a drug for preventing and / or treating breast cancer.

[0029] Experimental methods The Cell Counting Kit 8 (CCK8) method was used to detect the effect of β-elemenic acid on breast cancer cell proliferation. (1) Breast cancer cells with good cell growth status and in the logarithmic growth phase were selected, digested with 0.25% trypsin, and terminated by adding the corresponding complete culture medium containing 10% FBS. After being blown into a cell suspension, it was transferred to an EP tube and centrifuged at 800 rpm for 5 minutes. The supernatant was discarded and complete culture medium was added to make a cell suspension.

[0030] (2) Count the cells using a cell counting plate. Inoculate 4,000 cells in a 96-well plate with 6 replicate wells per group of cells at a volume of 100 μL per well. Add an equal amount of phosphate buffered saline (PBS solution) to the wells on the edges to prevent edge effects.

[0031] (3) Incubate the cells in a 37°C, 5% CO2 incubator overnight with conventional culture medium. Remove the liquid culture medium from each well and add drugs of corresponding concentrations prepared in the dark to the wells for continued culture.

[0032] (4) After 24 h of incubation, remove the culture medium from the plate in a dark environment, prepare the CCK8 solution required for the experiment, add 100 μL of solution to each well (avoid the generation of bubbles during the addition process to avoid inaccurate measurement by the microplate reader), and incubate in an incubator for 1-2 h.

[0033] (5) Adjust the wavelength of the automatic microplate reader to 450 nm and measure the OD value of each well.

[0034] (6) The growth curve of each group of cells was drawn according to the OD value, and Graphpad 8.0 was used to calculate and analyze the cell proliferation.

[0035] Effect of β-elemenic acid on apoptosis of breast cancer cells detected by flow cytometry (1) Select breast cancer cells Bcap37 and MDA231 that are in good growth condition and in the logarithmic growth phase. After trypsin digestion, resuspend the cells in culture medium and count them using a hemocytometer. 5 Cells were seeded in six-well plates and cultured overnight.

[0036] (2) When the cell density reaches about 50%, the original culture medium is removed, the cells are washed twice with PBS, and a certain concentration of β-elemenate is added to each well in a dark environment. The cells are then cultured in an incubator for 24 h.

[0037] (3) Collect the culture supernatant, wash twice with pre-cooled PBS, and collect the cells in a 15 mL centrifuge tube after trypsin digestion. Add complete medium to terminate digestion, pipette evenly to form a cell suspension, and transfer to a 1.5 mL centrifuge tube. Centrifuge at 1500 rpm for 5 min, discard the supernatant, add 100 μL of 1× binding buffer to resuspend, mix the cells thoroughly, add 3 μL of Annexin V-FITC and 3 μL of PI dye to each treatment group, and transfer to a flow cytometry tube and incubate in the dark at room temperature for 15 min.

[0038] (4) Add 300 μL of 1× binding buffer to each sample tube under light-proof conditions, and then use flow cytometry to detect the apoptosis rate of cells in each group.

[0039] The nude mouse subcutaneous tumor model was used to detect the effect of β-elemenate on the growth and proliferation of breast cancer cells. (1) The purchased nude mice were placed in an SPF-level animal room and kept in a balanced manner for 2 weeks.

[0040] (2) Take triple-negative breast cancer cells MDA-MB-231 in the logarithmic growth phase, digest them with trypsin, resuspend them in PBS and count them until the cell density reaches 5×10 7 / mL, mix the cell suspension and matrix gel at a ratio of 1:1 and place on ice for inoculation.

[0041] (3) Use iodine to disinfect the skin on the left side of the nude mouse's mid-abdomen. Use a 1 mL syringe to draw up 100 μL of the cell suspension and inject it horizontally into the subcutaneous tissue of the nude mouse. Be careful to avoid bleeding at the inoculation site and outflow of cell fluid.

[0042] (4) Observe the tumor formation of nude mice every three days until the subcutaneous tumor volume grows to 500 mm 3 The solid tumor is divided into several pieces of approximately 1 mm 3 The tumor was implanted subcutaneously in nude mice.

[0043] (5) After about a week of observation, when the nude mouse flank is about 50 mm 3 When the tumor mass was found, the mice were randomly divided into groups, 6 in each group. The groups were: blank solvent group (injection solvent: 1% DMSO + 4% Tween 80 + 95% saline containing 0.05% sodium carboxymethyl cellulose), low-dose β-elemenonic acid administration group (50 mg / kg), medium-dose β-elemenonic acid administration group (100 mg / kg) and high-dose β-elemenonic acid administration group (200 mg / kg).

[0044] (6) The drug was administered by regular intraperitoneal injection every two days. The nude mice were weighed and the tumor diameter was measured before each administration to observe the physical condition of the nude mice and the growth of the tumor.

[0045] (7) After the tumor had grown for about 18 days, the mice were killed by spinal dislocation, and the tumor mass was removed with scissors and forceps. The mass was weighed and recorded, and then photographed with a camera.

[0046] 2. Experimental results Figure 1 The results of the present invention using CCK8 method to detect the effect of β-elemenic acid on the growth and proliferation of different breast cancer cells. The results show that β-elemenic acid has a greater effect on the survival rate of breast cancer cells MCF-7, BT-474, MDA-MB-231, MDA-MB-453, KPL-4, and Bcap-37. 50 They are 106.9μM, 78.49μM, 102.7μM, 129.6μM, 152.1μM and 79.16μM respectively.

[0047] Figure 2 The present invention uses flow cytometry to detect the effect of β-elemenic acid on breast cancer cell apoptosis. The results show that under 100μM, 120μM, and 160μM β-elemenic acid treatment for 24 hours, the apoptosis rates of MDA-MB-231 cells were 6.85%, 53.52%, and 84.59%, respectively; under 80μM, 120μM, and 160μM β-elemenic acid treatment for 24 hours, the apoptosis rates of Bcap37 cells were 19.39%, 39.02%, and 60.82%, respectively.

[0048] Figure 3 The figure is the result of the present invention using a nude mouse subcutaneous tumor model to detect the effect of β-elemenic acid on the growth and proliferation of breast cancer cells. The results show that the tumor growth rate in the β-elemenic acid administration group is significantly lower than that in the control group, and the growth rate of the high-dose group is less than that of the medium-dose group and less than that of the low-dose group. The tumor mass results at the final treatment node show that β-elemenic acid treatment can significantly inhibit the growth of mouse tumors.

[0049] In summary, the present invention has found that β-elemenic acid exhibits good anti-breast cancer activity in vitro and in vivo, can induce breast cancer cell death, can be used as a pharmaceutical active ingredient to prepare anti-breast cancer drugs, and has good application prospects in the field of breast cancer treatment. Therefore, the present invention effectively overcomes the various shortcomings of the prior art and has a high industrial utilization value.

[0050] The above embodiments are merely illustrative of the principles and effects of the present invention, and are not intended to limit the present invention. Anyone familiar with the art may modify or alter the above embodiments without departing from the spirit and scope of the present invention. Therefore, all equivalent modifications or alterations made by a person of ordinary skill in the art without departing from the spirit and technical concept disclosed by the present invention shall still be covered by the claims of the present invention.

Claims

1. Use of β-elemenic acid as a pharmaceutical active ingredient in the preparation of anti-breast cancer drugs.

2. The use according to claim 1, characterized in that: The structural formula of the β-elemenic acid is:

3. The use according to claim 1, characterized in that: The anti-breast cancer drug is a drug used to prevent and / or treat breast cancer.

4. The use according to claim 1, characterized in that: The dosage form of the anti-breast cancer drug is selected from any one of compressed tablets, dispersible tablets, enteric-coated tablets, capsules, granules, pellets, emulsions, powders, oral solutions and injections.

5. An anti-breast cancer drug, characterized in that: The invention comprises beta-elemenic acid and pharmaceutically acceptable auxiliary materials.

6. The anti-breast cancer drug according to claim 5, characterized in that: The structural formula of the β-elemenic acid is:

7. The anti-breast cancer drug according to claim 5, characterized in that: The dosage form of the anti-breast cancer drug is selected from any one of compressed tablets, dispersible tablets, enteric-coated tablets, capsules, granules, pellets, emulsions, powders, oral solutions and injections.

8. Use of the anti-breast cancer drug according to any one of claims 5 to 7 in the preparation of a drug for preventing and / or treating breast cancer.

Images