Application of bangia fusco-purpurea coumarin compound or structural analogue of bangia fusco-purpurea coumarin compound in preparation of anti-brain cancer drugs or reagents
By using coumarin compounds such as liverin enol and syllolone hydrate, effective inhibition of brain cancer cells has been achieved and existing anti-antibody cancer is solved by using high heterogeneity of brain cancer and tumor stem cell characteristics. The problem of poor efficacy of brain cancer drugs provides new possibilities for treating brain cancer.
Patent Information
- Application Number
- CN202411113491.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-08-14
- Publication Date
- 2024-12-20
AI Technical Summary
Existing anti-brain cancer drugs are difficult to effectively target the high heterogeneity of brain cancer and tumor stem cell characteristics, resulting in poor treatment effects, and there are few researches on the natural products of erythromycosis in nervous system and brain cancer.
Using coumarin compounds such as liverin enol and syllolone hydrate as new drugs or reagents to reduce the survival rate of brain cancer cells, inhibit cell invasion and tumor stem cell properties, as potential treatments for the brain Cancer drugs.
These compounds show inhibitory effects on brain cancer cells, especially tumor stem cells, and have potential effects on brain cancer treatments, and even show significant effects at concentrations lower than the traditional drug temozolomide.
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Abstract
Description
Technical Field
[0001] The present invention relates to the field of medical technology, and in particular to use of a red hair algae coumarin compound or a structural analogue thereof in the preparation of an anti-brain cancer drug or reagent. Background Art
[0002] Brain cancer has a poor prognosis and a high mortality rate in both children and adults, so new treatments need to be developed. Therefore, the exploration and development of compounds with anti-brain cancer activity is an important topic.
[0003] Cancer is essentially a heterogeneous disease, so cancer treatment requires understanding its different degrees of heterogeneity and its origins, including the differences in cancers from different tissues caused by tissue-specific tumor types. Tumor heterogeneity increases the complexity and difficulty of cancer treatment, making it almost impossible to kill all types of cancer cells using only a single treatment. For example, propofol has been shown to have anti-tumor effects on lung cancer, but not on brain cancer (the experimental design of the study has excluded the possible factor that the compound cannot effectively pass through the blood-brain barrier). In addition, human cancer cell lines are one of the important models for the development of anticancer drugs. Although all cancer cell lines have some common biological characteristics, each cancer cell line still has its own unique genotype and phenotype, which may lead to differences in its response to the same drug. Brain cancer is a highly heterogeneous tumor. Therefore, it is not easy to infer the possible effect of the test compound on brain cancer based on the experimental results of the test compound in other types of cancer in previous studies.
[0004] On the other hand, the inhibition of tumor stem cell characteristics is also a difficult problem in cancer treatment. Glioblastoma (GBM) is one of the most common primary brain cancers and contains self-renewing, tumorigenic cancer stem cells (CSC), which contribute to tumor occurrence and resistance to treatment. Temozolomide (TMZ) is a chemotherapy drug that has been the first-line standard of care for the aggressive brain cancer GBM in the United States since 2005. In a 2021 in vitro model study of patient-derived GBM provided by biopsy, TMZ was shown to promote the formation of glioma stem cells. This shows that clinical drugs with anti-tumor effects may still lead to the formation of cells with tumor stem cell characteristics. Therefore, in addition to the traditional anti-tumor effects (three biological activities such as reducing the survival rate of cancer cells, inhibiting colony formation, and reducing cell invasion), the anti-tumor stem cell effect of the target compound should also be tested separately.
[0005] The exploration of possible new biological activities and applications of marine compounds and their structural analogs is a research direction that researchers in the field of natural products have long been concerned about. For example, many teams have long studied the possible biological activities of active substances derived from edible seaweed, especially red algae. In the 1990s, Putian, Fujian, China, began to cultivate a red algae, Bangia fuscopurpurea (also known as red hair vegetable, red hair moss, cow hair algae, red hair vegetable, etc.), and the area has become one of the main production areas of red hair algae. There are three research trends in the previous exploration of the biological activities of red hair algae and its natural products. First, the main research objects are natural products (proteins and polysaccharides) derived from its water extracts, while the number of studies related to its ethanol extracts is significantly smaller. Second, its biological activity research is mainly aimed at the peripheral system, and the number of biological activity studies directly related to the nervous system is still relatively small (especially central nervous system diseases). Third, its anti-cancer activity research is significantly less. At present, it is only known that the polysaccharides of red hair algae have been confirmed by previous studies to have anti-ovarian cancer and other effects, but it is still not entirely clear what the possible effects of natural products derived from red hair algae on other types of cancer may be.
[0006] In 2023, researchers have successfully identified the structures of various polyphenolic compounds contained in the ethanol extract of red hair algae, including two coumarin compounds, namely hesperidinol (CAS: 1221-43-8) and hesperidinol (CAS: 23971-42-8). Although researchers have shown that these coumarin compounds have antiproliferative activity against a few peripheral cancer cell lines, it is still unclear how these compounds affect the survival rate of nervous system-related cancer cells (especially brain cancer), and whether these compounds may affect other important characteristics of cancer cells, such as colony formation ability, cell invasion ability, tumor stem cell characteristics, etc. Summary of the invention
[0007] The present invention aims to solve one of the technical problems in the related art at least to a certain extent. To this end, one object of the present invention is to provide a use of a coumarin compound of red hair algae or a structural analog thereof in the preparation of a drug or an agent, wherein the drug or agent is used for anti-brain cancer.
[0008] According to the use of the present invention, the red hair algae coumarin compounds or their structural analogs can reduce the survival rate of brain cancer cells, reduce the invasion of brain cancer cells, and inhibit the characteristics of tumor stem cells, so that they can be used to prepare drugs for treating brain cancer.
[0009] Optionally, the red hair algae coumarin compound is aurantium enol, and its structural analog is a structural analog of the red hair algae coumarin compound hesperidone, and the structural analog of hesperidone is hesperidone hydrate. Both coumarin compounds have an inhibitory effect on the tumor stem cell characteristics of brain cancer. The minimum effective concentration of hesperidone hydrate is the same as that of the clinical drug temozolomide, while aurantium enol is lower than temozolomide. The anti-brain cancer effect of aurantium enol also includes the effect of inhibiting brain cancer tumor stem cell markers.
[0010] In a second aspect of the present invention, an embodiment of the present invention provides a drug for anti-brain cancer, comprising a red hair algae coumarin compound or a structural analog thereof in a pharmaceutically acceptable carrier. The red hair algae coumarin compound is aurantium enol, and its structural analog is a structural analog of the red hair algae coumarin compound hesperidone, and the structural analog of hesperidone is hesperidone hydrate.
[0011] In the third aspect of the present invention, embodiments of the present invention provide the use of red hair algae coumarin compounds or structural analogs thereof in the preparation of anti-brain cancer agents.
[0012] Additional aspects and advantages of the present invention will be given in part in the following description and in part will be obvious from the following description, or will be learned through practice of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS
[0013] Figure 1 The effect of the compounds according to the examples of the present invention on the cell viability of GBM8401 cells;
[0014] Figure 2 The effect of the compounds according to the examples of the present invention on the cell viability of U87 cells;
[0015] Figure 3 Analysis of the effect of TMZ on the colony-forming ability of GBM8401 cells according to an embodiment of the present invention;
[0016] Figure 4 Analysis of the effect of Auraptenol on the colony-forming ability of GBM8401 cells according to an embodiment of the present invention;
[0017] Figure 5 This is an analysis of the effect of Meranzin on the colony-forming ability of GBM8401 cells according to an embodiment of the present invention;
[0018] Figure 6 Analysis of the effect of Auraptenol on the invasive ability of U87 cells according to an embodiment of the present invention;
[0019] Figure 7 Analysis of the effect of Meranzin on the invasive ability of U87 cells according to an embodiment of the present invention;
[0020] Figure 8 Analysis of the effect of TMZ on the spheroid formation ability of U87 cells according to an embodiment of the present invention;
[0021] Fig. 9 Analysis of the effect of Auraptenol on the spheroid formation ability of U87 cells according to an embodiment of the present invention;
[0022] Fig.10 Analysis of the effect of Meranzin on the spheroid formation ability of U87 cells according to an embodiment of the present invention;
[0023] Fig.11 This is an immunoblot analysis of the effect of Auraptenol on the expression of CD44 protein, a tumor stem cell marker, in GBM8401 cells according to an example of the present invention. DETAILED DESCRIPTION
[0024] The technical solution of the present invention is described below through specific examples. It should be understood that the one or more method steps mentioned in the present invention do not exclude the existence of other method steps before and after the combination step, or other method steps can be inserted between these explicitly mentioned steps; it should also be understood that these embodiments are only used to illustrate the present invention, and are not used to limit the scope of the present invention. Moreover, unless otherwise specified, the numbering of each method step is only a convenient tool for identifying each method step, and is not intended to limit the order of arrangement of each method step or to limit the scope of the present invention. Changes or adjustments in their relative relationships should also be regarded as the scope of the present invention without substantially changing the technical content.
[0025] In order to better understand the above technical scheme, the exemplary embodiments of the present invention are described in more detail below. Although exemplary embodiments of the present invention are shown, it should be understood that the present invention can be implemented in various forms and should not be limited by the embodiments set forth herein. On the contrary, these embodiments are provided in order to more thoroughly understand the present invention and to fully convey the scope of the present invention to those skilled in the art.
[0026] The test materials used in the present invention are all common commercial products, and can be purchased from the market of companies related to general experimental reagent sales.
[0027] Compound name: Auraptenol, or CID 13343541 (CAS: 1221-43-8): Product number CFN99340, ≧98%, Chemface. The chemical structure of this compound is as follows:
[0028]
[0029] Compound name: Meranzin hydrate (hereinafter referred to as Meranzin in the present specification) (CAS: 5875-49-0): Product No. TN1922, 99.49%, TargetMol, USA. The chemical structure of this compound is as follows:
[0030]
[0031] Compound name: Temozolomide (TMZ): Product No. T1178, 99.87%, TargetMol.
[0032] This application uses in vitro brain cancer models: brain cancer cell lines GBM8401 and U-87MG (U87).
[0033] As primary antibodies, CD44 (santa cruz; sc-9960, 1:500) and β-actin (santa cruz; sc-47778, 1:500) were used.
[0034] All experimental results are expressed as mean ± standard deviation (SD). Unpaired t-test was used to compare and analyze the data between groups. *(P<0.05) and **(P<0.01) respectively indicate that the differences between groups are statistically significant. The chemical structures of the compounds in the specification of the present invention were displayed using InDraw structural editor (Shanghai Eagle Valley Information Technology Co., Ltd.).
[0035] The present invention is described below with reference to specific embodiments. It should be noted that these embodiments are merely illustrative and do not limit the present invention in any way.
[0036] Example 1 Effects of different concentrations of the test compound on the cytotoxicity of brain cancer cell lines GBM8401 and U87
[0037] Cells (3000 cells / well) were seeded into 96-well plates and incubated overnight at 37°C, 95% air and 5% CO2. After treating the cells with cell culture medium (RPMI1640, containing 10% FBS) supplemented with the test compound for 24 hours, Alamar Blue reagent (cell culture medium: Alamar Blue reagent = 10:1; Invitrogen) was added and the cells were cultured at 37°C for 3 hours. Blank solvent (vehicle; 0.1% dimethyl sulfoxide (DMSO)). The absorbance was measured at 570nm and 620nm using an ELISA reader (Dynex Technologies, Inc) (the absorbance at 570nm of the sample minus the absorbance at 620nm was used as the value of the sample to calculate the cell viability). Cell viability = (experimental treatment group / blank solvent group) × 100%.
[0038] This example selected Auraptenol, Meranzin and Temozolomide (TMZ) as test compounds.
[0039] The results are as follows Figure 1 As shown, Figure 1is the survival rate of GBM8401 cells. After 24 hours of compound treatment, TMZ (50 μM) can significantly reduce the survival rate of GBM8401 cells, while Auraptenol (50 μM) and Meranzin (50 μM) have no effect on the survival rate of GBM8401 cells.
[0040] like Figure 2 As shown, Figure 2 is the survival rate of U87 cells; after 24 hours of compound treatment, TMZ (0.5-50 μM) can significantly reduce the survival rate of U87 cells, Auraptenol (5-50 μM) and Meranzin (50 μM) can both significantly reduce the survival rate of U87 cells.
[0041] Example 2 Analysis of the effect of different concentrations of the test compound on GBM8401 cell colony formation
[0042] Cells (3000 cells / well) were seeded in 6-well plates and treated with test compounds in culture medium for 7 days. Cells were fixed with 4% paraformaldehyde (Sigma-Aldrich) and stained with crystal violet (Sigma-Aldrich), and colonies (more than 50 cells) were counted.
[0043] This example selected Auraptenol, Meranzin and Temozolomide (TMZ) as test compounds.
[0044] The results are as follows Figure 3 As shown, after 7 days of treatment, TMZ (5-50 μM) could significantly reduce the colony-forming ability of GBM8401.
[0045] like Figure 4 As shown, Auraptenol did not affect the colony-forming ability of GBM8401 after 7 days of treatment.
[0046] like Figure 5 As shown, Meranzin did not affect the colony-forming ability of GBM8401 after 7 days of treatment.
[0047] Example 3 Analysis of the invasion of U87 cells by test compounds at different concentrations
[0048] Cells were seeded in the upper chamber of the cell invasion assay device (1×10 5cells, 50 μL / well) and supplemented with serum-free medium containing the compound to be tested. The lower chamber was filled with 30 μL of medium containing 10% FBS serum. A polycarbonate membrane with a pore size of 8 μm (Nucleopore; Costar) was coated with Matrigel (BD Biosciences) to allow cell adhesion, and the upper and lower chambers were separated by the coated membrane. After incubation for 24 hours under 5% CO2 and 37°C, the cells that migrated from the upper side of the membrane to its lower side were fixed in anhydrous methanol and stained with 10% Giemsa solution (Sigma-Aldrich). Finally, the fixed cells were photographed and counted using a microscopic imaging system (Leica). For each well, five areas were randomly selected for quantification.
[0049] In this example, Auraptenol and Meranzin were selected as test compounds.
[0050] The results are as follows Figure 6 As shown, Auraptenol (5-50 μM) significantly reduced the cell invasion ability of U87 cells after 24 hours of treatment.
[0051] like Figure 7 As shown, after 24 hours of treatment, Meranzin (0.5-50 μM) could significantly reduce the cell invasion ability of U87.
[0052] Example 4 Effects of different concentrations of the test compound on spheroid formation of U87 cells
[0053] The cells were suspended in serum-free DMEM / F12 (Gibco) medium containing B-27 (Gibco), 20 ng / mL EGF (PeproTech) and 20 ng / mL bFGF (PeproTech). U87 cells were seeded on ultra-low attachment 6-well plates (Corning Life Sciences) and treated with the test compounds. After 7 days, spheres were observed and counted under an optical microscope (Leica).
[0054] This example selected Auraptenol, Meranzin and Temozolomide (TMZ) as test compounds.
[0055] The results are as follows Figure 8 As shown, TMZ (5-50 μM) significantly reduced the sphere-forming ability of U87 cells after 7 days of treatment.
[0056] like Fig. 9 As shown, Auraptenol (1-50 μM) significantly reduced the sphere-forming ability of U87 cells after 7 days of treatment.
[0057] like Fig.10 As shown, Meranzin (5-50 μM) significantly reduced the sphere-forming ability of U87 cells after 7 days of treatment.
[0058] Example 5 Immunoblotting analysis of GBM8401 cells with different concentrations of the test compound
[0059] After GBM8401 cells were treated with the test compound for 24 hours, proteins were extracted and purified from each group of cells. The protein concentration of each group of samples was determined using a BCA protein assay kit (Pierce Biotechnology). SDS-PAGE electrophoresis was performed, and the proteins separated by electrophoresis were transferred to a PVDF membrane. The PVDF membrane was treated with primary antibodies and corresponding secondary antibodies targeting two targets (CD44 and β-actin), and the results were detected and recorded for analysis to evaluate the effect of the test compound on the expression of the target protein.
[0060] In this example, Auraptenol was selected as the test compound.
[0061] The results are as follows Fig.11 As shown, after 24 hours of treatment, Auraptenol (0.5-50 μM) reduced the protein expression of CD44, a tumor stem cell marker of GBM8401 (but did not significantly affect the protein expression of the internal reference β-actin).
[0062] In the description of this specification, the description with reference to the terms "one embodiment", "some embodiments", "example", "specific example", or "some examples" means that the specific features, structures, materials or characteristics described in conjunction with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic representation of the above terms should not be understood as necessarily referring to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described may be combined in any one or more embodiments or examples in a suitable manner. In addition, those skilled in the art may combine and combine different embodiments or examples described in this specification.
[0063] Although the embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and cannot be understood as limiting the present invention. Those skilled in the art may change, modify, replace and modify the above embodiments within the scope of the present invention.
Claims
1. Use of red hair algae coumarin compounds or structural analogs thereof in the preparation of medicines, wherein the medicines are used for combating brain cancer.
2. The use according to claim 1, It is characterized in that The red hair algae coumarin compound is aurantium enol, the structural analogue of the red hair algae coumarin compound is the structural analogue of the red hair algae coumarin compound hesperidone, and the structural analogue of hesperidone is hesperidone hydrate.
3. The use according to claim 1, It is characterized in that The red hair algae coumarin compound or its structural analogue can reduce the survival rate of brain cancer cells, reduce the invasion of brain cancer cells, and inhibit the characteristics of tumor stem cells.
4. The use according to claim 3, It is characterized in that The red hair algae coumarin compound can inhibit the expression of tumor stem cell markers.
5. The use according to claim 4, It is characterized in that The tumor stem cell marker is CD44 protein.
6. A drug for fighting brain cancer, It is characterized in that The invention comprises a pharmaceutically acceptable carrier of a red hair algae coumarin compound or a structural analogue thereof.
7. The drug according to claim 6, It is characterized in that The red hair algae coumarin compound is aurantium enol, the structural analogue of the red hair algae coumarin compound is the structural analogue of the red hair algae coumarin compound hesperidone, and the structural analogue of hesperidone is hesperidone hydrate.
8. Use of red hair algae coumarin compounds or their structural analogs in the preparation of anti-brain cancer agents.
9. The use according to claim 8, It is characterized in that The red hair algae coumarin compound is aurantium enol, the structural analogue of the red hair algae coumarin compound is the structural analogue of the red hair algae coumarin compound hesperidone, and the structural analogue of hesperidone is hesperidone hydrate.
10. The use according to claim 8, It is characterized in that The anti-brain cancer agent acts as a chemical or composition required in a cell experiment or an animal experiment as one of the experimental objectives.