KR102259781B1 - Novel use of Sedum middendorffianum Maxim extract - Google Patents

Abstract

The present invention relates to a novel use of a composition comprising an A. giraffe extract, and specifically, to a pharmaceutical composition for preventing, improving or treating cancer comprising an ethanol extract of A. giraffe extract, a health functional food composition, and a composition for photodynamic treatment As a result, the extract of Sedum middendorttianum Maxim of the present invention is excellent in inhibiting the viability of cancer cells, and can effectively prevent, improve and treat cancer as a pharmaceutical composition and a food composition using the same, specifically, the present invention ginseng ginseng extract is excellent in inhibiting tumors by inducing apoptosis in cancer cells and showing strong cytotoxicity to cancer cells. In addition, it was confirmed that the IC50 value was lowered by more than 3 times when irradiated with UVA of weak intensity that did not affect cell survival with A. giraffe extract alone, so it can be utilized for the prevention, improvement and treatment effect of cancer using photodynamic therapy. have. In addition, the extract of the ginseng plant of the present invention is a compound derived from nature and has no side effects, so it can be used in various ways as pharmaceutical compositions, health functional foods, and the like.

Description

Novel use of Sedum middendorffianum Maxim extract}

The present invention relates to a novel use of a composition comprising an A. giraffe extract, and more particularly, to a pharmaceutical composition for preventing, improving or treating cancer, a health functional food composition, and a composition for photodynamic treatment, including the A. giraffe extract. .

In general, it is known that cancer arises mainly from the uncontrolled growth of malignant cells due to an imbalance between cell growth and apoptosis. Apoptosis is one of the most well-known methods of programmed cell death, and inducing apoptosis in cancer cells is widely used as a therapeutic strategy to control the growth of cancer cells. Indeed, it is known that many conventional anticancer agents induce the death of tumor cells through the apoptotic pathway.

It is known that ovarian cancer is the main cause of death in gynecology cancer diseases, and most ovarian cancer patients are diagnosed with advanced disease (stage III or IV), and the survival rate is known to be only 10-30%. Conventionally, chemotherapeutic agents such as cisplatin, platinum, and taxane have been generally used for the treatment of ovarian cancer, but they exhibited a wide range of side effects such as nausea and vomiting (Ferrandina G, et al. Increased cyclooxygenase-2). (COX-2) expression is associated with chemotherapy resistance and outcome in ovarian cancer patients. Ann Oncol. 2002;13:1205-1211), and it has also been reported that resistance to chemotherapy appears repeatedly in most patients ( Kigawa J. New strategy for overcoming resistance to chemotherapy of ovarian cancer. Yonago Acta Med. 2013;56:43-50). Therefore, there is a need for the development of a new natural product-derived therapeutic agent for the prevention and treatment of ovarian cancer.

On the other hand, the ginseng giraffe is a perennial plant with thick stems and a height of about 20 cm. It is known for its uses as tonic and seonhyeol. However, there is no report on the selective apoptotic activity of the A. giraffe extract against cancer cells, and in particular, nothing is known about the photodynamic therapeutic efficacy of the A. giraffe extract.

As a result of the study for the prevention, improvement and treatment of ovarian cancer, the present inventors not only showed toxicity by selectively inducing apoptosis of cancer cells, but also confirmed the function as a photosensitizer when treated with UVA, The present invention was completed.

Accordingly, an object of the present invention is to provide a pharmaceutical composition for preventing or treating ovarian cancer, comprising an extract of Sedum middendorttianum Maxim.

In addition, another object of the present invention is to provide a health functional food composition for improving or preventing ovarian cancer, including an extract of Sedum middendorttianum Maxim.

In addition, another object of the present invention is to provide a composition for photodynamic therapy, comprising an extract of Sedum middendorttianum Maxim.

In order to achieve the object of the present invention as described above, the present invention provides a pharmaceutical composition for preventing or treating ovarian cancer, comprising an extract of Sedum middendorttianum Maxim.

As an embodiment of the present invention, the extract can be extracted using arabica ginseng plant water, C1 to C4 lower alcohol, or a mixed solvent thereof.

In another embodiment of the present invention, the pharmaceutical composition may have cytotoxicity to cancer cells.

As another embodiment of the present invention, the pharmaceutical composition may induce apoptosis in cancer cells.

In addition, the present invention provides a health functional food composition for improving or preventing ovarian cancer, comprising an extract of Sedum middendorttianum Maxim.

In addition, the present invention provides a composition for photodynamic therapy, comprising an extract of Sedum middendorttianum Maxim.

In addition, the present invention provides a method for preventing, improving or treating ovarian cancer, comprising administering the composition to a subject.

In addition, the present invention provides the use of the composition for the prevention, improvement and treatment of ovarian cancer.

The extract of Sedum middendorttianum Maxim of the present invention is excellent in inhibiting the viability of cancer cells, and can effectively prevent, improve and treat cancer as a pharmaceutical composition and a food composition using the same.

Specifically, the extract of the present invention is excellent in inhibiting tumors by inducing apoptosis in cancer cells, thereby exhibiting strong cytotoxicity to cancer cells. In addition, by confirming that the IC50 value was lowered by more than three times when irradiated with UVA of weak intensity that did not affect cell survival with A. giraffe extract alone, it could be utilized for the prevention, improvement and treatment effect of cancer using photodynamic therapy. have. In addition, the extract of the ginseng root of the present invention is a compound derived from nature and has no side effects, so it can be used in various ways as pharmaceutical compositions, health functional foods, etc.

Figure 1 shows the effect of a giraffe extract on the cell viability of human normal ovarian cells (IOSE) and human ovarian cancer cells (A2780) through MTT assay.
Figure 2 shows the results of analyzing the apoptosis of the aphid ginseng extract through Annexin V-FITC assay.
Figure 3 shows the effect on the cell viability of human ovarian cancer cells (A2780) when treated with UVA to the A. giraffe through MTT assay.
Figure 4 shows the results of analysis of apoptosis induced by A. giraffe extract and UVA irradiation through Annexin V-FITC assay.

As a result of conducting research for the prevention, improvement and treatment of ovarian cancer, the present inventors confirmed that the A. giraffe extract selectively induces apoptosis in cancer cells and exhibits toxicity, and completed the present invention.

Accordingly, the present invention provides a pharmaceutical composition for preventing or treating ovarian cancer, comprising an extract of Sedum middendorttianum Maxim.

As used herein, the term “prevention” refers to any act of inhibiting or delaying the onset of ovarian cancer by administering the pharmaceutical composition according to the present invention.

As used herein, the term “treatment” refers to any action in which symptoms for ovarian cancer are improved or beneficially changed by administration of the pharmaceutical composition according to the present invention.

The term "extract" used in the present invention refers to an extract obtained by extraction treatment of a substance, a diluted or concentrated liquid of the extract, a dried product obtained by drying the extract, a prepared or purified product of the extract, or a mixture thereof, such as an extract It includes extracts of all formulations that can be formed by themselves and using extracts.

The method for preparing the extract of the present invention is not particularly limited, and may be extracted according to a method commonly used in the art. Non-limiting examples of the extraction method include hot water extraction method, ultrasonic extraction method, filtration method, reflux extraction method, immersion extraction method, high temperature and high pressure steam extraction method, etc., which may be performed alone or in combination of two or more methods. have. The type of extraction solvent used in the present invention is not particularly limited, and any solvent known in the art may be used, and C1 to C4 lower alcohols such as water, methanol, ethanol, butanol, etc., and a mixed solvent thereof, etc. It may be a variety of organic solvents including, preferably, ethanol may be used as the solvent, and more preferably 70% ethanol may be used.

In the present invention, the ovarian cancer prevention or treatment efficacy of the extract of A. giraffe extract was directly identified through various experiments.

More specifically, in one embodiment of the present invention, it was confirmed the effect of A. giraffe extract on the cell viability of human normal ovarian cells (IOSE) and human ovarian cancer cells (A2780) through MTT assay (see Example 2). , The apoptosis-inducing activity was confirmed for Cisplatin, a treatment for ovarian cancer, and extracts from A. giraffe extract through Annexin V-FITC assay (see Example 3).

From the above examples, it was confirmed that the pharmaceutical composition of the present invention may be selectively cytotoxic to cancer cells, and it was confirmed that the cytotoxicity to the cancer cells is the effect of inducing apoptosis, but the pharmaceutical composition The efficacy of the red composition is not limited thereto.

As used herein, the term “apoptosis” refers to the death of cancer cells due to genetic properties, and for the purpose of the present invention, apoptosis refers to the death of ovarian cancer cells.

In another embodiment of the present invention, to confirm the effect on the cell viability of human ovarian cancer cells (A2780) when UVA is treated to the A. giraffe extract through MTT assay (see Example 4), Annexin V-FITC assay Apoptosis-inducing activity on cancer cells was confirmed when UVA was treated to the A. giraffe extract through (see Example 5).

From the above example, the function as a photosensitizer of the A. giraffe extract of the present invention is confirmed, and the extract may be selectively cytotoxic to cancer cells as a composition for photodynamic therapy, and exhibiting cytotoxicity to the cancer cells. It has been confirmed that this is the efficacy of inducing apoptosis, but the efficacy of the photodynamic composition is not limited thereto.

Accordingly, as another aspect of the present invention, the present invention provides a composition for photodynamic therapy, comprising an extract of Sedum middendorttianum Maxim.

The term "photodynamic therapy" used in the present invention refers to a treatment method having a therapeutic effect by selectively accumulating light in diseased cells when a light of a specific wavelength is applied to the skin after a photosensitizer that responds to light is applied. Specifically, it refers to a method of treating ovarian cancer using an extract of A. giraffe extract as a photosensitizer, wherein the photosensitizer is excited when irradiated with light of a specific wavelength and then emits a fluorescence signal. A substance that generates or reacts with a surrounding substrate or oxygen to generate a reactive oxygen species. The reactive oxygen species has an effect of apoptosis or necrosis of surrounding tumor cells, and for the purpose of the present invention, photodynamic therapy is It means to kill ovarian cancer by using an extract of ginseng root as a sensory agent.

The prophylactic or therapeutic composition according to the present invention includes an extract of ginseng plant as an active ingredient, and may further include a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier is commonly used in formulation, and includes, but is not limited to, saline, sterile water, Ringer's solution, buffered saline, cyclodextrin, dextrose solution, maltodextrin solution, glycerol, ethanol, liposome, and the like. It does not, and may further include other conventional additives such as antioxidants and buffers, if necessary. In addition, diluents, dispersants, surfactants, binders, lubricants, etc. may be additionally added to form an injectable formulation such as an aqueous solution, suspension, emulsion, etc., pill, capsule, granule or tablet. Regarding suitable pharmaceutically acceptable carriers and formulations, formulations can be preferably made according to each component using the method disclosed in Remington's literature. The pharmaceutical composition of the present invention is not particularly limited in the formulation, but may be formulated as an injection, an inhalant, or an external preparation for skin.

The prophylactic or therapeutic composition of the present invention may be administered orally or parenterally (eg, intravenously, subcutaneously, intraperitoneally or locally applied) according to a desired method, but preferably may be administered orally, The dosage varies depending on the condition and weight of the patient, the severity of the disease, the drug form, the route and time of administration, but may be appropriately selected by those skilled in the art.

The prophylactic or therapeutic composition of the present invention is administered in a pharmaceutically effective amount. In the present invention, "pharmaceutically effective amount" means an amount sufficient to treat or diagnose a disease with a reasonable benefit/risk ratio applicable to medical treatment or diagnosis, and the effective dose level is determined by the patient's disease type, severity, drug activity, sensitivity to drugs, administration time, administration route and excretion rate, duration of treatment, factors including concomitant drugs, and other factors well known in the medical field. The pharmaceutical composition according to the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered single or multiple. It is important to administer an amount capable of obtaining the maximum effect in a minimum amount without side effects in consideration of all the above factors, and this can be easily determined by a person skilled in the art.

As another aspect of the present invention, the present invention provides a health functional food composition for improving or preventing ovarian cancer, comprising an extract of Sedum middendorttianum Maxim.

As used in the present invention, the term "health functional food" is the same term as food for specific health purposes, and refers to foods with high medical and medical effects that are processed to efficiently exhibit bioregulatory functions in addition to nutritional supply. . In some cases, the terms functional food, health food, and health supplement may be used interchangeably, and the food may be prepared in various forms such as tablets, capsules, powders, granules, liquids, pills, etc. to obtain useful effects.

The health functional food of the present invention may include additional ingredients that are commonly used in food compositions to improve odor, taste, vision, and the like. For example, vitamins A, C, D, E, B1, B2, B6, B12, niacin, biotin, folate, panthotenic acid, and the like may be included. In addition, it may include minerals such as zinc (Zn), iron (Fe), calcium (Ca), chromium (Cr), magnesium (Mg), manganese (Mn), copper (Cu). In addition, amino acids such as lysine, tryptophan, cysteine, and valine may be included. In addition, preservatives (potassium sorbate, sodium benzoate, salicylic acid, sodium dihydroacetate, etc.), disinfectants (bleaching powder and high bleaching powder, sodium hypochlorite, etc.), antioxidants (butylhydroxyanisole (BHA), butylhydroxytoluene (BHT) ), colorant (tar pigment, etc.), color developer (sodium nitrite, sodium nitrite, etc.), bleach (sodium sulfite), seasoning (MSG sodium glutamate, etc.), sweetener (dulcin, cyclimate, saccharin, sodium, etc.), Food additives such as flavorings (vanillin, lactones, etc.), expanding agents (alum, D-potassium hydrogen tartrate, etc.), strengthening agents, emulsifiers, thickeners (flavors), film agents, gum base agents, foam inhibitors, solvents, and improving agents can be added. The additive may be selected according to the type of food and used in an appropriate amount.

When the health functional food of the present invention is used as a food additive, it may be added as it is or used together with other foods or food ingredients, and may be appropriately used according to a conventional method.

In the health functional food of the present invention, the content of the extract is not particularly limited, and may be variously changed depending on the condition of the administration target, the type of specific disease, the degree of progression, and the like. If necessary, it may also be included in the total content of the food.

As another aspect of the present invention, the present invention provides a method for preventing, improving or treating ovarian cancer, comprising administering to an individual a composition comprising a giraffe extract.

As another aspect of the present invention, the present invention provides the use of the composition for preventing, improving and treating ovarian cancer.

In the present invention, "individual" means a subject in need of a method for preventing, controlling or treating a disease, and more specifically, a human or non-human primate, mouse, rat, dog, cat , refers to mammals such as horses and cattle.

Hereinafter, the present invention will be described in more detail through examples. However, these examples are for illustrative purposes only, and the scope of the present invention is not limited to these examples.

[Example]

Example 1. Preparation of extracts from A. giraffe extract

10 times (g/L) of dry weight (g/L) 70% (v/v) ethanol aqueous solution was added to the shade-dried baby giraffe vinegar 　 (Sedum middendorffianum Maxim., 　 provided by Gyeonggi-do, Korea 　 Hantaek Botanical Garden), immersed at room temperature for 7 days, and the filtrate was taken. Extraction was repeated 　 once 　 in the same way. The extracted 　 filtrate was concentrated under reduced pressure at a temperature of 　 35 ° C. and then freeze-dried to obtain an ethanol extract of 70% 　 　 　 Aeggi ginseng.

Example 2. Confirmation of cytotoxicity against cancer cells of the extract of ginseng plant

The effect of A. giraffe extract on the cell viability of human normal ovarian cells (IOSE) and human ovarian cancer cells (A2780) was investigated through MTT assay. ^{Briefly, cells (5×10 4} ) were seeded into each well of a 96-well plate containing 50 μL of RPMI medium per well. After 24 hours, different concentrations of extracts were added. After 48 hours, 25 μL of MTT (5 mg/mL stock solution) was added, and then the plate was incubated for an additional 4 hours. Then, after removing the medium, formazan blue formed in the cells was dissolved in 50 μL DMSO. Optical density was measured at 540 nm using a microplate spectrophotometer (Spectra Max; Molecular Devices, Sunnyvale, CA, USA).

As a result, as shown in FIG. 1, the ethanol extract of A. giraffe had no effect on the cell growth of normal ovarian cells (IOSE) (IC50>200ug/ml), but significantly inhibited the viability of ovarian cancer cells (A2780). (IC50 = 50.25ug/ml), it was confirmed that cancer cell-selective cytotoxicity was shown.

Example 3. Confirmation of apoptosis-inducing efficacy against cancer cells of the A. giraffe extract

Annexin V-FITC assay (double staining of Annexin V and PI) was performed in the following way to confirm the apoptosis-inducing efficacy of A. giraffe extract against cancer cells. ^{Cells were aliquoted at 1x10 5} cells/dish in a 60 mm dish and stabilized for 24 hours. After the addition of Giraffe extract and cisplatin and culturing for 48 hours, cells were recruited. For Annexin V and PI double staining, cells were suspended in 100 μL of binding buffer (10 mM HEPES/NaOH, 140 mM NaCl, 2.5 mM CaCl 2 , pH 7.4) and 1.25 μL of FITC-conjugated Annexin V and PI ( 50 mg/ml). The mixture was incubated for 15 minutes at room temperature in the dark and analyzed on a FACS Carter-Plus flow cytometer.

As a result, as shown in FIG. 2 , it was confirmed that apoptosis was induced according to the concentration of the ethanol extract of A. giraffe, and it was confirmed from the results that the toxicity of cancer cells expressed in A. giraffe was through apoptotic cell death.

Example 4. Confirmation of photodynamic treatment efficacy against cancer cells of Kirin root extract

In order to confirm the photodynamic treatment efficacy of the A. giraffe extract against cancer cells, the effect of UVA treatment on the cell viability of human ovarian cancer cells (A2780) was investigated using MTT assay. After dispensing cells into each well of a 96-well plate, the cells were stabilized for 24 hours. After the addition of Giraffe extract and cisplatin, 750mJ/cm ² UVA was irradiated. After 48 hours, MTT assay was performed to confirm the IC50 value.

^{As a result, as shown in FIG. 3, when irradiated with UVA of weak intensity (750mJ/cm 2} ) without affecting cell viability alone to the ethanol extract of A. giraffe extract, the IC50 value was 43.37ug/ml to 12.7ug/ml. It was confirmed that it was lowered by more than 3 times. That is, from the above results, it could be confirmed that the A. giraffe extract can be used as a photodynamic therapy (PDT, photodynamic therapy) for cancer treatment. In the case of cisplatin used for ovarian cancer treatment, UVA irradiation was performed in the same manner as above. It was confirmed that there was no change in cytotoxicity.

Example 5. Confirmation of efficacy of inducing apoptosis against cancer cells when UVA irradiated with A. giraffe extract

In order to confirm the efficacy of inducing apoptosis on cancer cells when UVA was applied to the extracts of Kirinifera aphids, human ovarian cancer cells (A2780) were aliquoted in a 60mm dish at 1x10 ⁵ cells/dish and stabilized for 24 hours. After adding the extract of Kirinchiae aphids ^{and irradiating 750mJ/cm 2} UVA and culturing for 48 hours, the cells were recruited. Annexin V and PI double staining was performed to confirm whether apoptosis was present.

As a result, as shown in FIG. 4, the apoptosis induction efficacy was increased by about 2 times by UVA treatment, and from the above results, it was confirmed that cancer cell toxicity induced when the ethanol extract of A. giraffe was treated with UVA was through apoptotic cell death. .

The description of the present invention stated above is for illustration, and those of ordinary skill in the art to which the present invention pertains can understand that it can be easily modified into other specific forms without changing the technical spirit or essential features of the present invention. There will be. Therefore, it should be understood that the embodiments described above are illustrative and non-limiting in all respects.

Claims (12)

delete delete delete delete delete delete delete delete A composition for photodynamic treatment of ovarian cancer, comprising an extract of Sedum middendorttianum Maxim. The composition for photodynamic treatment of ovarian cancer according to claim 9, wherein the extract is characterized in that the extract is extracted from A. giraffe root using water, a C1 to C4 lower alcohol, or a mixed solvent thereof. The composition for photodynamic therapy of ovarian cancer according to claim 9, wherein the composition has cytotoxicity against cancer cells. The composition for photodynamic treatment of ovarian cancer according to claim 10, wherein the composition induces apoptosis in cancer cells.

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